Multiscale Modeling of Virus Entry via Receptor-Mediated Endocytosis

NASA Astrophysics Data System (ADS)

Liu, Jin

2012-11-01

Virus infections are ubiquitous and remain major threats to human health worldwide. Viruses are intracellular parasites and must enter host cells to initiate infection. Receptor-mediated endocytosis is the most common entry pathway taken by viruses, the whole process is highly complex and dictated by various events, such as virus motions, membrane deformations, receptor diffusion and ligand-receptor reactions, occurring at multiple length and time scales. We develop a multiscale model for virus entry through receptor-mediated endocytosis. The binding of virus to cell surface is based on a mesoscale three dimensional stochastic adhesion model, the internalization (endocytosis) of virus and cellular membrane deformation is based on the discretization of Helfrich Hamiltonian in a curvilinear space using Monte Carlo method. The multiscale model is based on the combination of these two models. We will implement this model to study the herpes simplex virus entry into B78 cells and compare the model predictions with experimental measurements.

3D-QSAR comparative molecular field analysis on opioid receptor antagonists: pooling data from different studies.

PubMed

Peng, Youyi; Keenan, Susan M; Zhang, Qiang; Kholodovych, Vladyslav; Welsh, William J

2005-03-10

Three-dimensional quantitative structure-activity relationship (3D-QSAR) models were constructed using comparative molecular field analysis (CoMFA) on a series of opioid receptor antagonists. To obtain statistically significant and robust CoMFA models, a sizable data set of naltrindole and naltrexone analogues was assembled by pooling biological and structural data from independent studies. A process of "leave one data set out", similar to the traditional "leave one out" cross-validation procedure employed in partial least squares (PLS) analysis, was utilized to study the feasibility of pooling data in the present case. These studies indicate that our approach yields statistically significant and highly predictive CoMFA models from the pooled data set of delta, mu, and kappa opioid receptor antagonists. All models showed excellent internal predictability and self-consistency: q(2) = 0.69/r(2) = 0.91 (delta), q(2) = 0.67/r(2) = 0.92 (mu), and q(2) = 0.60/r(2) = 0.96 (kappa). The CoMFA models were further validated using two separate test sets: one test set was selected randomly from the pooled data set, while the other test set was retrieved from other published sources. The overall excellent agreement between CoMFA-predicted and experimental binding affinities for a structurally diverse array of ligands across all three opioid receptor subtypes gives testimony to the superb predictive power of these models. CoMFA field analysis demonstrated that the variations in binding affinity of opioid antagonists are dominated by steric rather than electrostatic interactions with the three opioid receptor binding sites. The CoMFA steric-electrostatic contour maps corresponding to the delta, mu, and kappa opioid receptor subtypes reflected the characteristic similarities and differences in the familiar "message-address" concept of opioid receptor ligands. Structural modifications to increase selectivity for the delta over mu and kappa opioid receptors have been predicted on the basis of the CoMFA contour maps. The structure-activity relationships (SARs) together with the CoMFA models should find utility for the rational design of subtype-selective opioid receptor antagonists.

Studying Neutrophil Migration In Vivo Using Adoptive Cell Transfer.

PubMed

Miyabe, Yoshishige; Kim, Nancy D; Miyabe, Chie; Luster, Andrew D

2016-01-01

Adoptive cell transfer experiments can be used to study the roles of cell trafficking molecules on the migratory behavior of specific immune cell populations in vivo. Chemoattractants and their G protein-coupled seven-transmembrane-spanning receptors regulate migration of cells in vivo, and dysregulated expression of chemoattractants and their receptors is implicated in autoimmune and inflammatory diseases. Inflammatory arthritides, such as rheumatoid arthritis (RA), are characterized by the recruitment of inflammatory cells into joints. The K/BxN serum transfer mouse model of inflammatory arthritis shares many similar features with RA. In this autoantibody-induced model of arthritis, neutrophils are the critical immune cells necessary for the development of joint inflammation and damage. We have used adoptive neutrophil transfer to define the contributions of chemoattractant receptors, cytokines, and activation receptors expressed on neutrophils that critically regulate their entry into the inflamed joint. In this review, we describe the procedure of neutrophil adoptive transfer to study the influence of neutrophil-specific receptors or mediators upon the their recruitment into the joint using the K/BxN model of inflammatory arthritis as a model of how adoptive cell transfer studies can be used to study immune cell migration in vivo.

Docking-based Screening of Ficus religiosa Phytochemicals as Inhibitors of Human Histamine H2 Receptor.

PubMed

Chaudhary, Amit; Yadav, Birendra Singh; Singh, Swati; Maurya, Pramod Kumar; Mishra, Alok; Srivastva, Shweta; Varadwaj, Pritish Kumar; Singh, Nand Kumar; Mani, Ashutosh

2017-10-01

Ficus religiosa L. is generally known as Peepal and belongs to family Moraceae . The tree is a source of many compounds having high medicinal value. In gastrointestinal tract, histamine H2 receptors have key role in histamine-stimulated gastric acid secretion. Their over stimulation causes its excessive production which is responsible for gastric ulcer. This study aims to screen the range of phytochemicals present in F. religiosa for binding with human histamine H2 and identify therapeutics for a gastric ulcer from the plant. In this work, a 3D-structure of human histamine H2 receptor was modeled by using homology modeling and the predicted model was validated using PROCHECK. Docking studies were also performed to assess binding affinities between modeled receptor and 34 compounds. Molecular dynamics simulations were done to identify most stable receptor-ligand complexes. Absorption, distribution, metabolism, excretion, and screening was done to evaluate pharmacokinetic properties of compounds. The results suggest that seven ligands, namely, germacrene, bergaptol, lanosterol, Ergost-5-en-3beta-ol, α-amyrin acetate, bergapten, and γ-cadinene showed better binding affinities. Among seven phytochemicals, lanosterol and α-amyrin acetate were found to have greater stability during simulation studies. These two compounds may be a suitable therapeutic agent against histamine H2 receptor. This study was performed to screen antiulcer compounds from F. religiosa . Molecular modeling, molecular docking and MD simulation studies were performed with selected phytochemicals from F. religiosa . The analysis suggests that Lanosterol and α-amyrin may be a suitable therapeutic agent against histamine H2 receptor. This study facilitates initiation of the herbal drug discovery process for the antiulcer activity. Abbreviations used: ADMET: Absorption, distribution, metabolism, excretion and toxicity, DOPE: Discrete Optimized Potential Energy, OPLS: Optimized potential for liquid simulations, RMSD: Root-mean-square deviation, HOA: Human oral absorption, MW: Molecular weight, SP: Standard-precision, XP: Extra-precision, GPCRs: G protein-coupled receptors, SASA: Solvent accessible surface area, Rg: Radius of gyration, NHB: Number of hydrogen bond.

Purinergic P2X receptors: structural models and analysis of ligand-target interaction.

PubMed

Dal Ben, Diego; Buccioni, Michela; Lambertucci, Catia; Marucci, Gabriella; Thomas, Ajiroghene; Volpini, Rosaria

2015-01-07

The purinergic P2X receptors are ligand-gated cation channels activated by the endogenous ligand ATP. They assemble as homo- or heterotrimers from seven cloned subtypes (P2X1-7) and all trimer subunits present a common topology consisting in intracellular N- and C- termini, two transmembrane domains and a large extracellular domain. These membrane proteins are present in virtually all mammalian tissues and regulate a large variety of responses in physio- and pathological conditions. The development of ligands that selectively activate or block specific P2X receptor subtypes hence represents a promising strategy to obtain novel pharmacological tools for the treatment of pain, cancer, inflammation, and neurological, cardiovascular, and endocrine diseases. The publication of the crystal structures of zebrafish P2X4 receptor in inactive and ATP-bound active forms provided structural data for the analysis of the receptor structure, the interpretation of mutagenesis data, and the depiction of ligand binding and receptor activation mechanism. In addition, the availability of ATP-competitive ligands presenting selectivity for P2X receptor subtypes supports the design of new potent and selective ligands with possibly improved pharmacokinetic profiles, with the final aim to obtain new drugs. This study describes molecular modelling studies performed to develop structural models of the human and rat P2X receptors in inactive and active states. These models allowed to analyse the role of some non-conserved residues at ATP binding site and to study the receptor interaction with some non-specific or subtype selective agonists and antagonists. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

The heterodimeric sweet taste receptor has multiple potential ligand binding sites.

PubMed

Cui, Meng; Jiang, Peihua; Maillet, Emeline; Max, Marianna; Margolskee, Robert F; Osman, Roman

2006-01-01

The sweet taste receptor is a heterodimer of two G protein coupled receptors, T1R2 and T1R3. This discovery has increased our understanding at the molecular level of the mechanisms underlying sweet taste. Previous experimental studies using sweet receptor chimeras and mutants show that there are at least three potential binding sites in this heterodimeric receptor. Receptor activity toward the artificial sweeteners aspartame and neotame depends on residues in the amino terminal domain of human T1R2. In contrast, receptor activity toward the sweetener cyclamate and the sweet taste inhibitor lactisole depends on residues within the transmembrane domain of human T1R3. Furthermore, receptor activity toward the sweet protein brazzein depends on the cysteine rich domain of human T1R3. Although crystal structures are not available for the sweet taste receptor, useful homology models can be developed based on appropriate templates. The amino terminal domain, cysteine rich domain and transmembrane helix domain of T1R2 and T1R3 have been modeled based on the crystal structures of metabotropic glutamate receptor type 1, tumor necrosis factor receptor, and bovine rhodopsin, respectively. We have used homology models of the sweet taste receptors, molecular docking of sweet ligands to the receptors, and site-directed mutagenesis of the receptors to identify potential ligand binding sites of the sweet taste receptor. These studies have led to a better understanding of the structure and function of this heterodimeric receptor, and can act as a guide for rational structure-based design of novel non-caloric sweeteners, which can be used in the fighting against obesity and diabetes.

Mu-opiate receptors measured by positron emission tomography are increased in temporal lobe epilepsy.

PubMed

Frost, J J; Mayberg, H S; Fisher, R S; Douglass, K H; Dannals, R F; Links, J M; Wilson, A A; Ravert, H T; Rosenbaum, A E; Snyder, S H

1988-03-01

Neurochemical studies in animal models of epilepsy have demonstrated the importance of multiple neurotransmitters and their receptors in mediating seizures. The role of opiate receptors and endogenous opioid peptides in seizure mechanisms is well developed and is the basis for measuring opiate receptors in patients with epilepsy. Patients with complex partial seizures due to unilateral temporal seizure foci were studied by positron emission tomography using 11C-carfentanil to measure mu-opiate receptors and 18F-fluoro-deoxy-D-glucose to measure glucose utilization. Opiate receptor binding is greater in the temporal neocortex on the side of the electrical focus than on the opposite side. Modeling studies indicate that the increase in binding is due to an increase in affinity or the number of unoccupied receptors. No significant asymmetry of 11C-carfentanil binding was detected in the amygdala or hippocampus. Glucose utilization correlated inversely with 11C-carfentanil binding in the temporal neocortex. Increased opiate receptors in the temporal neocortex may represent a tonic anticonvulsant system that limits the spread of electrical activity from other temporal lobe structures.

Molecular modeling on structure-function analysis of human progesterone receptor modulators.

PubMed

Pal, Ria; Islam, Md Ataul; Hossain, Tabassum; Saha, Achintya

2011-01-01

Considering the significance of progesterone receptor (PR) modulators, the present study is explored to envisage the biophoric signals for binding to selective PR subtype-A using ligand-based quantitative structure activity relationship (QSAR) and pharmacophore space modeling studies on nonsteroidal substituted quinoline and cyclocymopol monomethyl ether derivatives. Consensus QSAR models (Training set (Tr): n(Tr)=100, R(2) (pred)=0.702; test set (Ts): n(Ts)=30, R(2) (pred)=0.705, R(2) (m)=0.635; validation set (Vs): n(Vs)=40, R(2) (pred)=0.715, R(2) (m)=0.680) suggest that molecular topology, atomic polarizability and electronegativity, atomic mass and van der Waals volume of the ligands have influence on the presence of functional atoms (F, Cl, N and O) and consequently contribute significant relations on ligand binding affinity. Receptor independent space modeling study (Tr: n(Tr)=26, Q(2)=0.927; Ts: n(Ts)=60, R(2) (pred)=0.613, R(2) (m)=0.545; Vs: n(Vs)=84, R(2) (pred)=0.611, R(2) (m)=0.507) indicates the importance of aromatic ring, hydrogen bond donor, molecular hydrophobicity and steric influence for receptor binding. The structure-function characterization is adjudged with the receptor-based docking study, explaining the significance of the mapped molecular attributes for ligand-receptor interaction in the catalytic cleft of PR-A.

Homology Modeling of Class A G Protein-Coupled Receptors

PubMed Central

Costanzi, Stefano

2012-01-01

G protein-coupled receptors (GPCRs) are a large superfamily of membrane bound signaling proteins that hold great pharmaceutical interest. Since experimentally elucidated structures are available only for a very limited number of receptors, homology modeling has become a widespread technique for the construction of GPCR models intended to study the structure-function relationships of the receptors and aid the discovery and development of ligands capable of modulating their activity. Through this chapter, various aspects involved in the constructions of homology models of the serpentine domain of the largest class of GPCRs, known as class A or rhodopsin family, are illustrated. In particular, the chapter provides suggestions, guidelines and critical thoughts on some of the most crucial aspect of GPCR modeling, including: collection of candidate templates and a structure-based alignment of their sequences; identification and alignment of the transmembrane helices of the query receptor to the corresponding domains of the candidate templates; selection of one or more templates receptor; election of homology or de novo modeling for the construction of specific extracellular and intracellular domains; construction of the three-dimensional models, with special consideration to extracellular regions, disulfide bridges, and interhelical cavity; validation of the models through controlled virtual screening experiments. PMID:22323225

Molecular modeling of ligand-receptor interactions in the OR5 olfactory receptor.

PubMed

Singer, M S; Shepherd, G M

1994-06-02

Olfactory receptors belong to the superfamily of seven transmembrane domain, G protein-coupled receptors. In order to begin analysis of mechanisms of receptor activation, a computer model of the OR5 olfactory receptor has been constructed and compared with other members of this superfamily. We have tested docking of the odor molecule lyral, which is known to activate the OR5 receptor. The results point to specific ligand-binding residues on helices III through VII that form a binding pocket in the receptor. Some of these residues occupy sequence positions identical to ligand-binding residues conserved among other superfamily members. The results provide new insights into possible molecular mechanisms of odor recognition and suggest hypotheses to guide future experimental studies using site-directed mutagenesis.

Mode of Action and Human Relevance Analysis for Nuclear Receptor-Mediated Liver Toxicity: A Case Study with Phenobarbital as a Model Constitutive Androstane Receptor (CAR) Activator

EPA Science Inventory

The constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are key nuclear receptors involved in the regulation of cellular responses. to exposure to many xenobiotics and various physiological processes. Phenobarbital (PB) is a non­ genotoxic i...

Molecular characterization of the receptor binding structure-activity relationships of influenza B virus hemagglutinin.

PubMed

Carbone, V; Kim, H; Huang, J X; Baker, M A; Ong, C; Cooper, M A; Li, J; Rockman, S; Velkov, T

2013-01-01

Selectivity of α2,6-linked human-like receptors by B hemagglutinin (HA) is yet to be fully understood. This study integrates binding data with structure-recognition models to examine the impact of regional-specific sequence variations within the receptor-binding pocket on selectivity and structure activity relationships (SAR). The receptor-binding selectivity of influenza B HAs corresponding to either B/Victoria/2/1987 or the B/Yamagata/16/88 lineages was examined using surface plasmon resonance, solid-phase ELISA and gel-capture assays. Our SAR data showed that the presence of asialyl sugar units is the main determinant of receptor preference of α2,6 versus α2,3 receptor binding. Changes to the type of sialyl-glycan linkage present on receptors exhibit only a minor effect upon binding affinity. Homology-based structural models revealed that structural properties within the HA pocket, such as a glyco-conjugate at Asn194 on the 190-helix, sterically interfere with binding to avian receptor analogs by blocking the exit path of the asialyl sugars. Similarly, naturally occurring substitutions in the C-terminal region of the 190-helix and near the N-terminal end of the 140-loop narrows the horizontal borders of the binding pocket, which restricts access of the avian receptor analog LSTa. This study helps bridge the gap between ligand structure and receptor recognition for influenza B HA; and provides a consensus SAR model for the binding of human and avian receptor analogs to influenza B HA.

Molecular modeling and docking studies of human 5-hydroxytryptamine 2A (5-HT2A) receptor for the identification of hotspots for ligand binding.

PubMed

Kanagarajadurai, Karuppiah; Malini, Manoharan; Bhattacharya, Aditi; Panicker, Mitradas M; Sowdhamini, Ramanathan

2009-12-01

The serotonergic system has been implicated in emotional and cognitive function. In particular, 5-HT(2A) (5-hydroxytrytamine receptor 2A) is attributed to a number of disorders like schizophrenia, depression, eating disorders and anxiety. 5-HT(2A), being a GPCR (G-protein coupled receptor), is important in the pharmaceutical industry as a proven target for these disorders. Despite their extensive clinical importance, the structural studies of this protein is lacking due to difficulties in determining its crystal structure. We have performed sequence analysis and molecular modeling of 5-HT(2A) that has revealed a set of conserved residues and motifs considered to play an important role in maintaining structural integrity and function of the receptor. The analysis also revealed a set of residues specific to the receptor which distinguishes them from other members of the subclass and their orthologs. Further, starting from the model structure of human 5-HT(2A) receptor, docking studies were attempted to envisage how it might interact with eight of its ligands (such as serotonin, dopamine, DOI, LSD, haloperidol, ketanserin, risperidone and clozapine). The binding studies of dopamine to 5-HT(2A) receptor can bring up better understanding in the etiology of a number of neurological disorders involving both these two receptors. Our sequence analysis and study of interactions of this receptor with other ligands reveal additional residue hotspots such as Asn 363 and Tyr 370. The function of these residues can be further analyzed by rational design of site-directed mutagenesis. Two distinct binding sites are identified which could play important roles in ligand binding and signaling.

SOURCE APPORTIONMENT OF EXPOSURES TO VOLATILE ORGANIC COMPOUNDS: II. APPLICATION OF RECEPTOR MODELS TO TEAM STUDY DATA. (R826788)

EPA Science Inventory

Four receptor-oriented source apportionment models were applied to personal exposure measurements for toxic volatile organic compounds (VOCs). The measurements are from the total exposure assessment methodology studies conducted from 1980 to 1984 in New Jersey (NJ) and Califor...

Fluctuation correlation models for receptor immobilization

NASA Astrophysics Data System (ADS)

Fourcade, B.

2017-12-01

Nanoscale dynamics with cycles of receptor diffusion and immobilization by cell-external-or-internal factors is a key process in living cell adhesion phenomena at the origin of a plethora of signal transduction pathways. Motivated by modern correlation microscopy approaches, the receptor correlation functions in physical models based on diffusion-influenced reaction is studied. Using analytical and stochastic modeling, this paper focuses on the hybrid regime where diffusion and reaction are not truly separable. The time receptor autocorrelation functions are shown to be indexed by different time scales and their asymptotic expansions are given. Stochastic simulations show that this analysis can be extended to situations with a small number of molecules. It is also demonstrated that this analysis applies when receptor immobilization is coupled to environmental noise.

Role of spatial inhomogenity in GPCR dimerisation predicted by receptor association-diffusion models

NASA Astrophysics Data System (ADS)

Deshpande, Sneha A.; Pawar, Aiswarya B.; Dighe, Anish; Athale, Chaitanya A.; Sengupta, Durba

2017-06-01

G protein-coupled receptor (GPCR) association is an emerging paradigm with far reaching implications in the regulation of signalling pathways and therapeutic interventions. Recent super resolution microscopy studies have revealed that receptor dimer steady state exhibits sub-second dynamics. In particular the GPCRs, muscarinic acetylcholine receptor M1 (M1MR) and formyl peptide receptor (FPR), have been demonstrated to exhibit a fast association/dissociation kinetics, independent of ligand binding. In this work, we have developed a spatial kinetic Monte Carlo model to investigate receptor homo-dimerisation at a single receptor resolution. Experimentally measured association/dissociation kinetic parameters and diffusion coefficients were used as inputs to the model. To test the effect of membrane spatial heterogeneity on the simulated steady state, simulations were compared to experimental statistics of dimerisation. In the simplest case the receptors are assumed to be diffusing in a spatially homogeneous environment, while spatial heterogeneity is modelled to result from crowding, membrane micro-domains and cytoskeletal compartmentalisation or ‘corrals’. We show that a simple association-diffusion model is sufficient to reproduce M1MR association statistics, but fails to reproduce FPR statistics despite comparable kinetic constants. A parameter sensitivity analysis is required to reproduce the association statistics of FPR. The model reveals the complex interplay between cytoskeletal components and their influence on receptor association kinetics within the features of the membrane landscape. These results constitute an important step towards understanding the factors modulating GPCR organisation.

Pain-relieving prospects for adenosine receptors and ectonucleotidases

PubMed Central

Zylka, Mark J.

2010-01-01

Adenosine receptor agonists have potent antinociceptive effects in diverse preclinical models of chronic pain. In contrast, the efficacy of adenosine or adenosine receptor agonists at treating pain in humans is unclear. Two ectonucleotidases that generate adenosine in nociceptive neurons were recently identified. When injected spinally, these enzymes have long-lasting adenosine A1 receptor (A1R)-dependent antinociceptive effects in inflammatory and neuropathic pain models. Furthermore, recent findings indicate that spinal adenosine A2A receptor activation can enduringly inhibit neuropathic pain symptoms. Collectively, these studies suggest the possibility of treating chronic pain in humans by targeting specific adenosine receptor subtypes in anatomically defined regions with agonists or with ectonucleotidases that generate adenosine. PMID:21236731

The innovative concept of three-dimensional hybrid receptor modeling

NASA Astrophysics Data System (ADS)

Stojić, A.; Stanišić Stojić, S.

2017-09-01

The aim of this study was to improve the current understanding of air pollution transport processes at regional and long-range scale. For this purpose, three-dimensional (3D) potential source contribution function and concentration weighted trajectory models, as well as new hybrid receptor model, concentration weighted boundary layer (CWBL), which uses a two-dimensional grid and a planetary boundary layer height as a frame of reference, are presented. The refined approach to hybrid receptor modeling has two advantages. At first, it considers whether each trajectory endpoint meets the inclusion criteria based on planetary boundary layer height, which is expected to provide a more realistic representation of the spatial distribution of emission sources and pollutant transport pathways. Secondly, it includes pollutant time series preprocessing to make hybrid receptor models more applicable for suburban and urban locations. The 3D hybrid receptor models presented herein are designed to identify altitude distribution of potential sources, whereas CWBL can be used for analyzing the vertical distribution of pollutant concentrations along the transport pathway.

Shifting physician prescribing to a preferred histamine-2-receptor antagonist. Effects of a multifactorial intervention in a mixed-model health maintenance organization.

PubMed

Brufsky, J W; Ross-Degnan, D; Calabrese, D; Gao, X; Soumerai, S B

1998-03-01

This study was undertaken to determine whether a program of education, therapeutic reevaluation of eligible patients, and performance feedback could shift prescribing to cimetidine from other histamine-2 receptor antagonists, which commonly are used in the management of ulcers and reflux, and reduce costs without increasing rates of ulcer-related hospital admissions. This study used an interrupted monthly time series with comparison series in a large mixed-model health maintenance organization. Physicians employed in health centers (staff model) and physicians in independent medical groups contracting to provide health maintenance organization services (group model) participated. The comparative percentage prescribed of specific histamine-2 receptor antagonists (market share), total histamine-2 receptor antagonist prescribing, cost per histamine-2 receptor antagonist prescription, and the rate of hospitalization for gastrointestinal illness were assessed. In the staff model, therapeutic reevaluation resulted in a sudden increase in market share of the preferred histamine-2 receptor antagonist cimetidine (+53.8%) and a sudden decrease in ranitidine (-44.7%) and famotidine (-4.8%); subsequently, cimetidine market share grew by 1.1% per month. In the group model, therapeutic reevaluation resulted in increased cimetidine market share (+9.7%) and decreased prescribing of other histamine-2 receptor antagonists (ranitidine -11.6%; famotidine -1.2%). Performance feedback did not result in further changes in prescribing in either setting. Use of omeprazole, an expensive alternative, essentially was unchanged by the interventions, as were overall histamine-2 receptor antagonist prescribing and hospital admissions for gastrointestinal illnesses. This intervention, which cost approximately $60,000 to implement, resulted in estimated annual savings in histamine-2 receptor antagonist expenditures of $1.06 million. Annual savings in histamine-2 receptor antagonist expenditures after this multifaceted intervention were more than implementation costs, with no discernible effects on numbers of hospitalizations. The magnitude of effect and cost savings were much greater in the staff model; organizational factors and economic incentives may have contributed to these differences. More research is needed to determine the generalizability of this approach to other technologies and managed care settings.

Homology Modeling of Dopamine D2 and D3 Receptors: Molecular Dynamics Refinement and Docking Evaluation

PubMed Central

Platania, Chiara Bianca Maria; Salomone, Salvatore; Leggio, Gian Marco; Drago, Filippo; Bucolo, Claudio

2012-01-01

Dopamine (DA) receptors, a class of G-protein coupled receptors (GPCRs), have been targeted for drug development for the treatment of neurological, psychiatric and ocular disorders. The lack of structural information about GPCRs and their ligand complexes has prompted the development of homology models of these proteins aimed at structure-based drug design. Crystal structure of human dopamine D3 (hD3) receptor has been recently solved. Based on the hD3 receptor crystal structure we generated dopamine D2 and D3 receptor models and refined them with molecular dynamics (MD) protocol. Refined structures, obtained from the MD simulations in membrane environment, were subsequently used in molecular docking studies in order to investigate potential sites of interaction. The structure of hD3 and hD2L receptors was differentiated by means of MD simulations and D3 selective ligands were discriminated, in terms of binding energy, by docking calculation. Robust correlation of computed and experimental Ki was obtained for hD3 and hD2L receptor ligands. In conclusion, the present computational approach seems suitable to build and refine structure models of homologous dopamine receptors that may be of value for structure-based drug discovery of selective dopaminergic ligands. PMID:22970199

The two-state dimer receptor model: a general model for receptor dimers.

PubMed

Franco, Rafael; Casadó, Vicent; Mallol, Josefa; Ferrada, Carla; Ferré, Sergi; Fuxe, Kjell; Cortés, Antoni; Ciruela, Francisco; Lluis, Carmen; Canela, Enric I

2006-06-01

Nonlinear Scatchard plots are often found for agonist binding to G-protein-coupled receptors. Because there is clear evidence of receptor dimerization, these nonlinear Scatchard plots can reflect cooperativity on agonist binding to the two binding sites in the dimer. According to this, the "two-state dimer receptor model" has been recently derived. In this article, the performance of the model has been analyzed in fitting data of agonist binding to A(1) adenosine receptors, which are an example of receptor displaying concave downward Scatchard plots. Analysis of agonist/antagonist competition data for dopamine D(1) receptors using the two-state dimer receptor model has also been performed. Although fitting to the two-state dimer receptor model was similar to the fitting to the "two-independent-site receptor model", the former is simpler, and a discrimination test selects the two-state dimer receptor model as the best. This model was also very robust in fitting data of estrogen binding to the estrogen receptor, for which Scatchard plots are concave upward. On the one hand, the model would predict the already demonstrated existence of estrogen receptor dimers. On the other hand, the model would predict that concave upward Scatchard plots reflect positive cooperativity, which can be neither predicted nor explained by assuming the existence of two different affinity states. In summary, the two-state dimer receptor model is good for fitting data of binding to dimeric receptors displaying either linear, concave upward, or concave downward Scatchard plots.

Cooperative ethylene receptor signaling

PubMed Central

Liu, Qian; Wen, Chi-Kuang

2012-01-01

The gaseous plant hormone ethylene is perceived by a family of five ethylene receptor members in the dicotyledonous model plant Arabidopsis. Genetic and biochemical studies suggest that the ethylene response is suppressed by ethylene receptor complexes, but the biochemical nature of the receptor signal is unknown. Without appropriate biochemical measures to trace the ethylene receptor signal and quantify the signal strength, the biological significance of the modulation of ethylene responses by multiple ethylene receptors has yet to be fully addressed. Nevertheless, the ethylene receptor signal strength can be reflected by degrees in alteration of various ethylene response phenotypes and in expression levels of ethylene-inducible genes. This mini-review highlights studies that have advanced our understanding of cooperative ethylene receptor signaling. PMID:22827938

Receptors as a master key for synchronization of rhythms

NASA Astrophysics Data System (ADS)

Nagano, Seido

2004-03-01

A simple, but general scheme to achieve synchronization of rhythms was derived. The scheme has been inductively generalized from the modelling study of cellular slime mold. It was clarified that biological receptors work as apparatuses that can convert external stimulus to the form of nonlinear interaction within individual oscillators. Namely, the mathematical model receptor works as a nonlinear coupling apparatus between nonlinear oscillators. Thus, synchronization is achieved as a result of competition between two kinds of non-linearities, and to achieve synchronization, even a small external stimulation via model receptors can change the characteristics of individual oscillators significantly. The derived scheme is very simple mathematically, but it is a very powerful scheme as numerically demonstrated. The biological receptor scheme should significantly help understanding of synchronization phenomena in biology since groups of limit cycle oscillators and receptors are ubiquitous in biological systems. Reference: S. Nagano, Phys Rev. E67, 056215(2003)

Development of 7TM receptor-ligand complex models using ligand-biased, semi-empirical helix-bundle repacking in torsion space: application to the agonist interaction of the human dopamine D2 receptor.

PubMed

Malo, Marcus; Persson, Ronnie; Svensson, Peder; Luthman, Kristina; Brive, Lars

2013-03-01

Prediction of 3D structures of membrane proteins, and of G-protein coupled receptors (GPCRs) in particular, is motivated by their importance in biological systems and the difficulties associated with experimental structure determination. In the present study, a novel method for the prediction of 3D structures of the membrane-embedded region of helical membrane proteins is presented. A large pool of candidate models are produced by repacking of the helices of a homology model using Monte Carlo sampling in torsion space, followed by ranking based on their geometric and ligand-binding properties. The trajectory is directed by weak initial restraints to orient helices towards the original model to improve computation efficiency, and by a ligand to guide the receptor towards a chosen conformational state. The method was validated by construction of the β1 adrenergic receptor model in complex with (S)-cyanopindolol using bovine rhodopsin as template. In addition, models of the dopamine D2 receptor were produced with the selective and rigid agonist (R)-N-propylapomorphine ((R)-NPA) present. A second quality assessment was implemented by evaluating the results from docking of a library of 29 ligands with known activity, which further discriminated between receptor models. Agonist binding and recognition by the dopamine D2 receptor is interpreted using the 3D structure model resulting from the approach. This method has a potential for modeling of all types of helical transmembrane proteins for which a structural template with sequence homology sufficient for homology modeling is not available or is in an incorrect conformational state, but for which sufficient empirical information is accessible.

Association of protein tyrosine phosphatase, non-receptor type 22 +1858C→T polymorphism and susceptibility to vitiligo: Systematic review and meta-analysis.

PubMed

Agarwal, Silky; Changotra, Harish

2017-01-01

Protein tyrosine phosphatase, non-receptor type 22 gene, which translates to lymphoid tyrosine phosphatase, is considered to be a susceptibility gene marker associated with several autoimmune diseases. Several studies have demonstrated the association of protein tyrosine phosphatase, non-receptor type 22 +1858C→T polymorphism with vitiligo. However, these studies showed conflicting results. Meta-analysis of the same was conducted earlier that included fewer number of publications in their study. We performed a meta-analysis of a total of seven studies consisting of 2094 cases and 3613 controls to evaluate the possible association of protein tyrosine phosphatase, non-receptor type 22 +1858C>T polymorphism with vitiligo susceptibility. We conducted a literature search in PubMed, Google Scholar and Dogpile for all published paper on protein tyrosine phosphatase, non-receptor type 22 +1858C→T polymorphism and vitiligo risk till June 2016. Data analysis was performed by RevMan 5.3 and comprehensive meta-analysis v3.0 software. Meta-analysis showed an overall significant association of protein tyrosine phosphatase, non- receptor type 22 +1858C→T polymorphism with vitiligo in all models (allelic model [T vs. C]: odds ratio = 1.50, 95% confidence interval [1.32-1.71], P< 0.001; dominant model [TT + CT vs. CC]: odds ratio = 1.61, 95% confidence interval [1.16-2.24], P = 0.004; recessive model [TT vs. CT + CC]: odds ratio = 4.82, 95% confidence interval [1.11-20.92], P = 0.04; homozygous model [TT vs. CC]: odds ratio = 5.34, 95% confidence interval [1.23-23.24], P = 0.03; co-dominant model [CT vs. CC]: odds ratio = 1.52, 95% confidence interval [1.09-2.13], P = 0.01). No publication bias was detected in the funnel plot study. Limited ethnic-based studies, unable to satisfy data by gender or vitiligo-type are some limitations of the present meta-analysis. Stratifying data by ethnicity showed an association of protein tyrosine phosphatase, non-receptor type 22 +1858C→T with vitiligo in European population (odds ratio = 1.53, 95% confidence interval [1.34-1.75], P< 0.001) but not in Asian population (odds ratio = 0.59, 95% confidence interval [0.26-1.32], P = 0.2). In conclusion, protein tyrosine phosphatase, non-receptor type 22 +1858 T allele predisposes European individuals to vitiligo.

Aberrant dopamine D2-like receptor function in a rodent model of schizophrenia.

PubMed

Perez, Stephanie M; Lodge, Daniel J

2012-11-01

Based on the observation that antipsychotic medications display antagonist properties at dopamine D2-like receptors, aberrant dopamine signaling has been proposed to underlie psychosis in patients with schizophrenia. Thus, it is not surprising that considerable research has been devoted to understanding the mechanisms involved in the antipsychotic action of these compounds. It is important to note that the majority of these studies have been performed in "normal" experimental animals. Given that these animals do not possess the aberrant neuronal information processing typically associated with schizophrenia, the aim of the current study was to examine the dopamine D2 receptor system in a rodent model of schizophrenia. Here, we demonstrate that methylazoxymethanol acetate (MAM)-treated rats display an enhanced effect of quinpirole on dopamine neuron activity and an aberrant locomotor response to D2-like receptor activation, suggesting changes in postsynaptic D2-like receptor function. To better understand the mechanisms underlying the enhanced response to D2-like ligands in MAM-treated rats, we examined the expression of D2, D3, and dopamine transporter mRNA in the nucleus accumbens and ventral tegmental area by quantitative reverse transcription-polymerase chain reaction. MAM-treated rats displayed a significant increase in dopamine D3 receptor mRNA expression in the nucleus accumbens with no significant changes in the expression of the D2 receptor. Taken together, these data demonstrate robust alterations in dopamine D2-like receptor function in a rodent model of schizophrenia and provide evidence that preclinical studies examining the mechanisms of antipsychotic drug action should be performed in animal models that mirror aspects of the abnormal neuronal transmission thought to underlie symptoms of schizophrenia.

A competitive binding model predicts the response of mammalian olfactory receptors to mixtures

NASA Astrophysics Data System (ADS)

Singh, Vijay; Murphy, Nicolle; Mainland, Joel; Balasubramanian, Vijay

Most natural odors are complex mixtures of many odorants, but due to the large number of possible mixtures only a small fraction can be studied experimentally. To get a realistic understanding of the olfactory system we need methods to predict responses to complex mixtures from single odorant responses. Focusing on mammalian olfactory receptors (ORs in mouse and human), we propose a simple biophysical model for odor-receptor interactions where only one odor molecule can bind to a receptor at a time. The resulting competition for occupancy of the receptor accounts for the experimentally observed nonlinear mixture responses. We first fit a dose-response relationship to individual odor responses and then use those parameters in a competitive binding model to predict mixture responses. With no additional parameters, the model predicts responses of 15 (of 18 tested) receptors to within 10 - 30 % of the observed values, for mixtures with 2, 3 and 12 odorants chosen from a panel of 30. Extensions of our basic model with odorant interactions lead to additional nonlinearities observed in mixture response like suppression, cooperativity, and overshadowing. Our model provides a systematic framework for characterizing and parameterizing such mixing nonlinearities from mixture response data.

Constitutive dimerization of the G-protein coupled receptor, neurotensin receptor 1, reconstituted into phospholipid bilayers.

PubMed

Harding, Peter J; Attrill, Helen; Boehringer, Jonas; Ross, Simon; Wadhams, George H; Smith, Eleanor; Armitage, Judith P; Watts, Anthony

2009-02-01

Neurotensin receptor 1 (NTS1), a Family A G-protein coupled receptor (GPCR), was expressed in Escherichia coli as a fusion with the fluorescent proteins eCFP or eYFP. A fluorophore-tagged receptor was used to study the multimerization of NTS1 in detergent solution and in brain polar lipid bilayers, using fluorescence resonance energy transfer (FRET). A detergent-solubilized receptor was unable to form FRET-competent complexes at concentrations of up to 200 nM, suggesting that the receptor is monomeric in this environment. When reconstituted into a model membrane system at low receptor density, the observed FRET was independent of agonist binding, suggesting constitutive multimer formation. In competition studies, decreased FRET in the presence of untagged NTS1 excludes the possibility of fluorescent protein-induced interactions. A simulation of the experimental data indicates that NTS1 exists predominantly as a homodimer, rather than as higher-order multimers. These observations suggest that, in common with several other Family A GPCRs, NTS1 forms a constitutive dimer in lipid bilayers, stabilized through receptor-receptor interactions in the absence of other cellular signaling components. Therefore, this work demonstrates that well-characterized model membrane systems are useful tools for the study of GPCR multimerization, allowing fine control over system composition and complexity, provided that rigorous control experiments are performed.

Integrated Experimental and Model-based Analysis Reveals the Spatial Aspects of EGFR Activation Dynamics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shankaran, Harish; Zhang, Yi; Chrisler, William B.

2012-10-02

The epidermal growth factor receptor (EGFR) belongs to the ErbB family of receptor tyrosine kinases, and controls a diverse set of cellular responses relevant to development and tumorigenesis. ErbB activation is a complex process involving receptor-ligand binding, receptor dimerization, phosphorylation, and trafficking (internalization, recycling and degradation), which together dictate the spatio-temporal distribution of active receptors within the cell. The ability to predict this distribution, and elucidation of the factors regulating it, would help to establish a mechanistic link between ErbB expression levels and the cellular response. Towards this end, we constructed mathematical models for deconvolving the contributions of receptor dimerizationmore » and phosphorylation to EGFR activation, and to examine the dependence of these processes on sub-cellular location. We collected experimental datasets for EGFR activation dynamics in human mammary epithelial cells, with the specific goal of model parameterization, and used the data to estimate parameters for several alternate models. Model-based analysis indicated that: 1) signal termination via receptor dephosphorylation in late endosomes, prior to degradation, is an important component of the response, 2) less than 40% of the receptors in the cell are phosphorylated at any given time, even at saturating ligand doses, and 3) receptor dephosphorylation rates at the cell surface and early endosomes are comparable. We validated the last finding by measuring EGFR dephosphorylation rates at various times following ligand addition both in whole cells, and in endosomes using ELISAs and fluorescent imaging. Overall, our results provide important information on how EGFR phosphorylation levels are regulated within cells. Further, the mathematical model described here can be extended to determine receptor dimer abundances in cells co-expressing various levels of ErbB receptors. This study demonstrates that an iterative cycle of experiments and modeling can be used to gain mechanistic insight regarding complex cell signaling networks.« less

Air Pollution Source/receptor Relationships in South Coast Air Basin, CA

NASA Astrophysics Data System (ADS)

Gao, Ning

This research project includes the application of some existing receptor models to study the air pollution source/receptor relationships in the South Coast Air Basin (SoCAB) of southern California, the development of a new receptor model and the testing and the modifications of some existing models. These existing receptor models used include principal component factor analysis (PCA), potential source contribution function (PSCF) analysis, Kohonen's neural network combined with Prim's minimal spanning tree (TREE-MAP), and direct trilinear decomposition followed by a matrix reconstruction. The ambient concentration measurements used in this study are a subset of the data collected during the 1987 field exercise of Southern California Air Quality Study (SCAQS). It consists of a number of gaseous and particulate pollutants analyzed from samples collected by SCAQS samplers at eight sampling sites, Anaheim, Azusa, Burbank, Claremont, Downtown Los Angeles, Hawthorne, Long Beach, and Rubidoux. Based on the information of emission inventories, meteorology and ambient concentrations, this receptor modeling study has revealed mechanisms that influence the air quality in SoCAB. Some of the mechanisms affecting the air quality in SoCAB that were revealed during this study include the following aspects. The SO_2 collected at sampling sites is mainly contributed by refineries in the coastal area and the ships equipped with oil-fired boilers off shore. Combustion of fossil fuel by automobiles dominates the emission of NO_{rm x} that is subsequently transformed and collected at sampling sites. Electric power plants also contribute HNO_3 to the sampling sites. A large feedlot in the eastern region of SoCAB has been identified as the major source of NH_3. Possible contributions from other industrial sources such as smelters and incinerators were also revealed. The results of this study also suggest the possibility of DMS (dimethylsulfide) and NH_3 emissions from off-shore sediments that have been contaminated by waste sludge disposal. The study also discovered that non-anthropogenic sources account for the observation of many chemical components being brought to the sampling sites, such as seasalt particles, soil particles, and Cl emission from Mojave Desert. The potential and limitation of the receptor models have been evaluated and some modifications have been made to improve the value of the models. A source apportionment method has been developed based on the application results of the potential source contribution function (PSCF) analysis.

The effects of oxotremorine, epibatidine, atropine, mecamylamine and naloxone in the tail-flick, hot-plate, and formalin tests in the naked mole-rat (Heterocephalus glaber).

PubMed

Dulu, Thomas D; Kanui, Titus I; Towett, Philemon K; Maloiy, Geoffrey M; Abelson, Klas S P

2014-01-01

The naked mole-rat (Heterocephalus glaber) is a promising animal model for the study of pain mechanisms, therefore a thorough characterization of this species is essential. The aim of the present study was to establish the naked mole-rat as a model for studying the cholinergic receptor system in antinociception by investigating the involvement of muscarinic, nicotinic and opioid receptors in nociceptive tests in this species. The effects of systemic administration of the muscarinic receptor agonist oxotremorine and the nicotinic receptor agonist epibatidine were investigated in the tail-flick, the hot-plate, and the formalin tests. The effects of co-administration of the muscarinic receptor antagonist atropine, the nicotinic receptor antagonist mecamylamine, and the opioid receptor antagonist naloxone were also investigated. Oxotremorine and epibatidine induced a significant, dose-dependent antinociceptive effect in the tail-flick, hot-plate, and formalin tests, respectively. The effects of oxotremorine and epibatidine were blocked by atropine and mecamylamine, respectively. In all three nociceptive tests, naloxone in combination with oxotremorine or epibatidine enhanced the antinociceptive effects of the drugs. The present study demonstrated that stimulation of muscarinic and nicotinic receptors produces antinociceptive effects in the naked-mole rat. The reversal effect of atropine and mecamylamine suggests that this effect is mediated by cholinergic receptors. As naloxone increases the antinociceptive effects of cholinergic agonists, it is suggested that the cholinergic antinociception acts via a gateway facilitated by opioid receptor blockage; however, the precise interaction between these receptor systems needs further investigation.

Gamma-Aminobutyric acid and benzodiazepine receptors in the kindling model of epilepsy: a quantitative radiohistochemical study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shin, C.; Pedersen, H.B.; McNamara, J.O.

1985-10-01

Quantitative radiohistochemistry was utilized to study alterations of gamma-aminobutyric acid (GABA) and benzodiazepine receptors in the kindling model of epilepsy. The radioligands used for GABA and benzodiazepine receptors were (TH) muscimol and (TH)flunitrazepam, respectively. GABA receptor binding was increased by 22% in fascia dentata of the hippocampal formation but not in neocortex or substantia nigra of kindled rats. Within fascia dentata, GABA receptor binding was increased to an equivalent extent in stratum granulosum and throughout stratum moleculare; no increase was found in dentate hilus or stratum lacunosummoleculare or stratum radiatum of CA1. The increased binding was present at 24 hrmore » but not at 28 days after the last kindled seizure. The direction, anatomic distribution, and time course of the increased GABA receptor binding were paralleled by increased benzodiazepine receptor binding. The anatomic distribution of the increased GABA receptor binding is consistent with a localization to somata and dendritic trees of dentate granule cells. The authors suggest that increased GABA and benzodiazepine receptor binding may contribute to enhanced inhibition of dentate granule cells demonstrated electrophysiologically in kindled animals.« less

The 5-HT7 receptor in learning and memory. Importance of the hippocampus

PubMed Central

Roberts, Amanda J.; Hedlund, Peter B.

2011-01-01

The 5-HT7 receptor is a more recently discovered G-protein-coupled receptor for serotonin. The functions and possible clinical relevance of this receptor are not yet fully understood. The present paper reviews to what extent the use of animal models of learning and memory and other techniques have implicated the 5-HT7 receptor in such processes. The studies have used a combination of pharmacological and genetic tools targeting the receptor to evaluate effects on behavior and cellular mechanisms. In tests such as the Barnes maze, contextual fear conditioning and novel location recognition that involve spatial learning and memory there is a considerable amount of evidence supporting an involvement of the 5-HT7 receptor. Supporting evidence has also been obtained in studies of mRNA expression and cellular signaling as well as in electrophysiological experiments. Especially interesting are the subtle but distinct effects observed in hippocampus-dependent models of place learning where impairments have been described in mice lacking the 5-HT7 receptor or after administration of a selective antagonist. While more work is required, it appears that 5-HT7 receptors are particularly important in allocentric representation processes. In instrumental learning tasks both procognitive effects and impairments in memory have been observed using pharmacological tools targeting the 5-HT7 receptor. In conclusion, the use of pharmacological and genetic tools in animal studies of learning and memory suggest a potentially important role for the 5-HT7 receptor in cognitive processes. PMID:21484935

Molecular modelling studies on the ORL1-receptor and ORL1-agonists

NASA Astrophysics Data System (ADS)

Bröer, Britta M.; Gurrath, Marion; Höltje, Hans-Dieter

2003-11-01

The ORL1 ( opioid receptor like 1)- receptor is a member of the family of rhodopsin-like G protein-coupled receptors (GPCR) and represents an interesting new therapeutical target since it is involved in a variety of biomedical important processes, such as anxiety, nociception, feeding, and memory. In order to shed light on the molecular basis of the interactions of the GPCR with its ligands, the receptor protein and a dataset of specific agonists were examined using molecular modelling methods. For that purpose, the conformational space of a very potent non-peptide ORL1-receptor agonist (Ro 64-6198) with a small number of rotatable bonds was analysed in order to derive a pharmacophoric arrangement. The conformational analyses yielded a conformation that served as template for the superposition of a set of related analogues. Structural superposition was achieved by employing the program FlexS. Using the experimental binding data and the superposition of the ligands, a 3D-QSAR analysis applying the GRID/GOLPE method was carried out. After the ligand-based modelling approach, a 3D model of the ORL1-receptor has been constructed using homology modelling methods based on the crystal structure of bovine rhodopsin. A representative structure of the model taken from molecular dynamics simulations was used for a manual docking procedure. Asp-130 and Thr-305 within the ORL1-receptor model served as important hydrophilic interaction partners. Furthermore, a hydrophobic cavity was identified stabilizing the agonists within their binding site. The manual docking results were supported using FlexX, which identified the same protein-ligand interaction points.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kufareva, Irina; Gustavsson, Martin; Zheng, Yi

Chemokines and their cell surface G protein–coupled receptors are critical for cell migration, not only in many fundamental biological processes but also in inflammatory diseases and cancer. Recent X-ray structures of two chemokines complexed with full-length receptors provided unprecedented insight into the atomic details of chemokine recognition and receptor activation, and computational modeling informed by new experiments leverages these insights to gain understanding of many more receptor:chemokine pairs. In parallel, chemokine receptor structures with small molecules reveal the complicated and diverse structural foundations of small molecule antagonism and allostery, highlight the inherent physicochemical challenges of receptor:chemokine interfaces, and suggest novelmore » epitopes that can be exploited to overcome these challenges. The structures and models promote unique understanding of chemokine receptor biology, including the interpretation of two decades of experimental studies, and will undoubtedly assist future drug discovery endeavors.« less

A DFT and semiempirical model-based study of opioid receptor affinity and selectivity in a group of molecules with a morphine structural core.

PubMed

Bruna-Larenas, Tamara; Gómez-Jeria, Juan S

2012-01-01

We report the results of a search for model-based relationships between mu, delta, and kappa opioid receptor binding affinity and molecular structure for a group of molecules having in common a morphine structural core. The wave functions and local reactivity indices were obtained at the ZINDO/1 and B3LYP/6-31G(∗∗) levels of theory for comparison. New developments in the expression for the drug-receptor interaction energy expression allowed several local atomic reactivity indices to be included, such as local electronic chemical potential, local hardness, and local electrophilicity. These indices, together with a new proposal for the ordering of the independent variables, were incorporated in the statistical study. We found and discussed several statistically significant relationships for mu, delta, and kappa opioid receptor binding affinity at both levels of theory. Some of the new local reactivity indices incorporated in the theory appear in several equations for the first time in the history of model-based equations. Interaction pharmacophores were generated for mu, delta, and kappa receptors. We discuss possible differences regulating binding and selectivity in opioid receptor subtypes. This study, contrarily to the statistically backed ones, is able to provide a microscopic insight of the mechanisms involved in the binding process.

Antagonism of Lateral Amygdala Alpha1-Adrenergic Receptors Facilitates Fear Conditioning and Long-Term Potentiation

ERIC Educational Resources Information Center

Lazzaro, Stephanie C.; Hou, Mian; Cunha, Catarina; LeDoux, Joseph E.; Cain, Christopher K.

2010-01-01

Norepinephrine receptors have been studied in emotion, memory, and attention. However, the role of alpha1-adrenergic receptors in fear conditioning, a major model of emotional learning, is poorly understood. We examined the effect of terazosin, an alpha1-adrenergic receptor antagonist, on cued fear conditioning. Systemic or intra-lateral amygdala…

Computational study of the heterodimerization between μ and δ receptors

NASA Astrophysics Data System (ADS)

Liu, Xin; Kai, Ming; Jin, Lian; Wang, Rui

2009-06-01

A growing body of evidence indicated that the G protein coupled receptors exist as homo- or hetero-dimers in the living cell. The heterodimerization between μ and δ opioid receptors has attracted researchers' particular interests, it is reported to display novel pharmacological and signalling regulation properties. In this study, we construct the full-length 3D-model of μ and δ opioid receptors using the homology modelling method. Threading program was used to predict the possible templates for the N- and C-terminus domains. Then, a 30 ns molecular dynamics simulations was performed with each receptor embedded in an explicit membrane-water environment to refine and explore the conformational space. Based on the structures extracted from the molecular dynamics, the likely interface of μ-δ heterodimer was investigated through the analysis of protein-protein docking, cluster, shape complementary and interaction energy. The computational modelling works revealed that the most likely interface of heterodimer was formed between the transmembrane1,7 (TM1,7) domains of μ receptor and the TM(4,5) domains of δ receptor, with emphasis on μ-TM1 and δ-TM4, the next likely interface was μ(TM6,7)-δ(TM4,5), with emphasis on μ-TM6 and δ-TM4. Our results were consistent with previous reports.

Delineation of the peptide binding site of the human galanin receptor.

PubMed Central

Kask, K; Berthold, M; Kahl, U; Nordvall, G; Bartfai, T

1996-01-01

Galanin, a neuroendocrine peptide of 29 amino acids, binds to Gi/Go-coupled receptors to trigger cellular responses. To determine which amino acids of the recently cloned seven-transmembrane domain-type human galanin receptor are involved in the high-affinity binding of the endogenous peptide ligand, we performed a mutagenesis study. Mutation of the His264 or His267 of transmembrane domain VI to alanine, or of Phe282 of transmembrane domain VII to glycine, results in an apparent loss of galanin binding. The substitution of Glu271 to serine in the extracellular loop III of the receptor causes a 12-fold loss in affinity for galanin. We combined the mutagenesis results with data on the pharmacophores (Trp2, Tyr9) of galanin and with molecular modelling of the receptor using bacteriorhodopsin as a model. Based on these studies, we propose a binding site model for the endogenous peptide ligand in the galanin receptor where the N-terminus of galanin hydrogen bonds with Glu271 of the receptor, Trp2 of galanin interacts with the Zn2+ sensitive pair of His264 and His267 of transmembrane domain VI, and Tyr9 of galanin interacts with Phe282 of transmembrane domain VII, while the C-terminus of galanin is pointing towards the N-terminus of th Images PMID:8617199

Quantitative in vivo receptor binding. III. Tracer kinetic modeling of muscarinic cholinergic receptor binding

DOE Office of Scientific and Technical Information (OSTI.GOV)

Frey, K.A.; Hichwa, R.D.; Ehrenkaufer, R.L.

1985-10-01

A tracer kinetic method is developed for the in vivo estimation of high-affinity radioligand binding to central nervous system receptors. Ligand is considered to exist in three brain pools corresponding to free, nonspecifically bound, and specifically bound tracer. These environments, in addition to that of intravascular tracer, are interrelated by a compartmental model of in vivo ligand distribution. A mathematical description of the model is derived, which allows determination of regional blood-brain barrier permeability, nonspecific binding, the rate of receptor-ligand association, and the rate of dissociation of bound ligand, from the time courses of arterial blood and tissue tracer concentrations.more » The term ''free receptor density'' is introduced to describe the receptor population measured by this method. The technique is applied to the in vivo determination of regional muscarinic acetylcholine receptors in the rat, with the use of (TH)scopolamine. Kinetic estimates of free muscarinic receptor density are in general agreement with binding capacities obtained from previous in vivo and in vitro equilibrium binding studies. In the striatum, however, kinetic estimates of free receptor density are less than those in the neocortex--a reversal of the rank ordering of these regions derived from equilibrium determinations. A simplified model is presented that is applicable to tracers that do not readily dissociate from specific binding sites during the experimental period.« less

Losartan suppresses the kainate-induced changes of angiotensin AT1 receptor expression in a model of comorbid hypertension and epilepsy.

PubMed

Atanasova, Dimitrinka; Tchekalarova, Jana; Ivanova, Natasha; Nenchovska, Zlatina; Pavlova, Ekaterina; Atanassova, Nina; Lazarov, Nikolai

2018-01-15

Experimental and clinical studies have demonstrated that components of renin-angiotensin system are elevated in the hippocampus in epileptogenic conditions. In the present work, we explored the changes in the expression of angiotensin II receptor, type 1 (AT 1 receptor) in limbic structures, as well as the effect of the AT1 receptor antagonist losartan in a model of comorbid hypertension and epilepsy. The expression of AT 1 receptors was compared between spontaneously hypertensive rats (SHRs) and Wistar rats by using immunohistochemistry in the kainate (KA) model of temporal lobe epilepsy (TLE). The effect of losartan was studied on AT 1 receptor expression in epileptic rats that were treated for a period of 4weeks after status epilepticus. The naive and epileptic SHRs were characterized by stronger protein expression of AT 1 receptor than normotensive Wistar rats in the CA1, CA3a, CA3b, CA3c field and the hilus of the dentate gyrus of the dorsal hippocampus but fewer cells were immunostained in the piriform cortex. Increased AT 1 immunostaining was observed in the basolateral amygdala of epileptic SHRs but not of epileptic Wistar rats. Losartan exerted stronger and structure-dependent suppression of AT 1 receptor expression in SHRs compared to Wistar rats. Our results confirm the important role of AT 1 receptor in epilepsy and suggest that the AT 1 receptor antagonists could be used as a therapeutic strategy for treatment of comorbid hypertension and epilepsy. Copyright © 2017 Elsevier Inc. All rights reserved.

The CCK(-like) receptor in the animal kingdom: functions, evolution and structures.

PubMed

Staljanssens, Dorien; Azari, Elnaz Karimian; Christiaens, Olivier; Beaufays, Jérôme; Lins, Laurence; Van Camp, John; Smagghe, Guy

2011-03-01

In this review, the cholecystokinin (CCK)(-like) receptors throughout the animal kingdom are compared on the level of physiological functions, evolutionary basis and molecular structure. In vertebrates, the CCK receptor is an important member of the G-protein coupled receptors as it is involved in the regulation of many physiological functions like satiety, gastrointestinal motility, gastric acid secretion, gall bladder contraction, pancreatic secretion, panic, anxiety and memory and learning processes. A homolog for this receptor is also found in nematodes and arthropods, called CK receptor and sulfakinin (SK) receptor, respectively. These receptors seem to have evolved from a common ancestor which is probably still closely related to the nematode CK receptor. The SK receptor is more closely related to the CCK receptor and seems to have similar functions. A molecular 3D-model for the CCK receptor type 1 has been built together with the docking of the natural ligands for the CCK and SK receptors in the CCK receptor type 1. These molecular models can help to study ligand-receptor interactions, that can in turn be useful in the development of new CCK(-like) receptor agonists and antagonists with beneficial health effects in humans or potential for pest control. Copyright © 2010 Elsevier Inc. All rights reserved.

Validation and pharmacological characterisation of MK-801-induced locomotor hyperactivity in BALB/C mice as an assay for detection of novel antipsychotics.

PubMed

Bradford, Andrea M; Savage, Kevin M; Jones, Declan N C; Kalinichev, Mikhail

2010-10-01

We evaluated locomotor hyperactivity induced in BALB/C mice by an N-methyl-D-aspartate receptor antagonist MK-801 as an assay for the detection of antipsychotic drugs. We assessed the effects of antipsychotic drugs to validate the assay (study 1), selective dopamine and serotonin ligands for pharmacological characterisation of the model (study 2) and a number of compounds with efficacy in models of schizophrenia to understand the predictive validity of the model (study 3). Adult males (n  = 9/group) were pretreated with a test compound, habituated to locomotor activity cages before receiving MK-801 (0.32 mg/kg) and activity recorded for a further 75 or 120 min. In study 1, we tested haloperidol, clozapine, olanzapine, risperidone, ziprasidone, aripiprazole, sertindole and quetiapine. In study 2, we tested SCH23390 (D(1) antagonist), sulpiride (D(2)/D(3) antagonist), raclopride (D(2)/D(3) antagonist), SB-277011 (D(3) antagonist), L-745,870 (D(4) antagonist), WAY100635 (5-HT(1A) antagonist), 8-OH-DPAT (5-HT(1A) agonist), ketanserin (5-HT(2A)/5-HT(2C) antagonist) and SB-242084 (5-HT(2C) antagonist). In study 3, we tested xanomeline (M(1)/M(4) receptor agonist), LY379268 (mGluR2/3 receptor agonist), diazepam (GABA(A) modulator) and thioperamide (H(3) receptor antagonist). All antipsychotics suppressed MK-801-induced hyperactivity in a dose-dependent and specific manner. The effects of antipsychotics appear to be mediated via dopamine D(1), D(2) and 5-HT(2) receptors. Xanomeline, LY379268 and diazepam were active in this assay while thioperamide was not. MK-801-induced hyperactivity in BALB/C mice model of positive symptoms has shown predictive validity with novel compounds acing at M(1)/M(4), mGluR2/3 and GABA(A) receptors and can be used as a screening assay for detection of novel pharmacotherapies targeting those receptors.

Kinetic operational models of agonism for G-protein-coupled receptors.

PubMed

Hoare, Samuel R J; Pierre, Nicolas; Moya, Arturo Gonzalez; Larson, Brad

2018-06-07

The application of kinetics to research and therapeutic development of G-protein-coupled receptors has become increasingly valuable. Pharmacological models provide the foundation of pharmacology, providing concepts and measurable parameters such as efficacy and potency that have underlain decades of successful drug discovery. Currently there are few pharmacological models that incorporate kinetic activity in such a way as to yield experimentally-accessible drug parameters. In this study, a kinetic model of pharmacological response was developed that provides a kinetic descriptor of efficacy (the transduction rate constant, k τ ) and allows measurement of receptor-ligand binding kinetics from functional data. The model assumes: (1) receptor interacts with a precursor of the response ("Transduction potential") and converts it to the response. (2) The response can decay. Familiar response vs time plots emerge, depending on whether transduction potential is depleted and/or response decays. These are the straight line, the "association" exponential curve, and the rise-and-fall curve. Convenient, familiar methods are described for measuring the model parameters and files are provided for the curve-fitting program Prism (GraphPad Software) that can be used as a guide. The efficacy parameter k τ is straightforward to measure and accounts for receptor reserve; all that is required is measurement of response over time at a maximally-stimulating concentration of agonist. The modular nature of the model framework allows it to be extended. Here this is done to incorporate antagonist-receptor binding kinetics and slow agonist-receptor equilibration. In principle, the modular framework can incorporate other cellular processes, such as receptor desensitization. The kinetic response model described here can be applied to measure kinetic pharmacological parameters than can be used to advance the understanding of GPCR pharmacology and optimize new and improved therapeutics. Copyright © 2018 Elsevier Ltd. All rights reserved.

A comparison of progestin and androgen receptor binding using the CoMFA technique

NASA Astrophysics Data System (ADS)

Loughney, Deborah A.; Schwender, Charles F.

1992-12-01

A series of 48 steroids has been studied with the SYBYL QSAR module using Relative Binding Affinities (RBAs) to progesterone and androgen receptors obtained from the literature. Models for the progesterone and androgen data were developed. Both models show regions where sterics and electrostatics correlate to binding affinity but are different for androgen and progesterone which suggests differences possibly important for receptor selectivity. The progesterone model is more predictive than the androgen (predictive r2 of 0.725 vs. 0.545 for progesterone and androgen, respectively).

Molecular docking, molecular modeling, and molecular dynamics studies of azaisoflavone as dual COX-2 inhibitors and TP receptor antagonists.

PubMed

Hadianawala, Murtuza; Mahapatra, Amarjyoti Das; Yadav, Jitender K; Datta, Bhaskar

2018-02-26

Designed multi-target ligand (DML) is an emerging strategy for the development of new drugs and involves the engagement of multiple targets with the same moiety. In the context of NSAIDs it has been suggested that targeting the thromboxane prostanoid (TP) receptor along with cyclooxygenase-2 (COX-2) may help to overcome cardiovascular (CVS) complications associated with COXIBs. In the present work, azaisoflavones were studied for their COX-2 and TP receptor binding activities using structure based drug design (SBDD) techniques. Flavonoids were selected as a starting point based on their known COX-2 inhibitory and TP receptor antagonist activity. Iterative design and docking studies resulted in the evolution of a new class scaffold replacing the benzopyran-4-one ring of flavonoids with quinolin-4-one. The docking and binding parameters of these new compounds are found to be promising in comparison to those of selective COX-2 inhibitors, such as SC-558 and celecoxib. Owing to the lack of structural information, a model for the TP receptor was generated using a threading base alignment method with loop optimization performed using an ab initio method. The model generated was validated against known antagonists for TP receptor using docking/MMGBSA. Finally, the molecules that were designed for selective COX-2 inhibition were docked into the active site of the TP receptor. Iterative structural modifications and docking on these molecules generated a series which displays optimum docking scores and binding interaction for both targets. Molecular dynamics studies on a known TP receptor antagonist and a designed molecule show that both molecules remain in contact with protein throughout the simulation and interact in similar binding modes. Graphical abstract ᅟ.

An assessment of the effects of serotonin 6 (5-HT6) receptor antagonists in rodent models of learning.

PubMed

Lindner, Mark D; Hodges, Donald B; Hogan, John B; Orie, Anitra F; Corsa, Jason A; Barten, Donna M; Polson, Craig; Robertson, Barbara J; Guss, Valerie L; Gillman, Kevin W; Starrett, John E; Gribkoff, Valentin K

2003-11-01

Antagonists of serotonin 6 (5-HT6) receptors have been reported to enhance cognition in animal models of learning, although this finding has not been universal. We have assessed the therapeutic potential of the specific 5-HT6 receptor antagonists 4-amino-N-(2,6-bis-methylamino-pyrimidin-4-yl)-benzenesulfonamide (Ro 04-6790) and 5-chloro-N-(4-methoxy-3-piperazin-1-yl-phenyl)-3-methyl-2-benzothiophenesulfonamide (SB-271046) in rodent models of cognitive function. Although mice express the 5-HT6 receptor and the function of this receptor has been investigated in mice, all reports of activity with 5-HT6 receptor antagonists have used rat models. In the present study, receptor binding revealed that the pharmacological properties of the mouse receptor are different from the rat and human receptor: Ro 04-6790 does not bind to the mouse 5-HT6 receptor, so all in vivo testing included in the present report was conducted in rats. We replicated previous reports that 5-HT6 receptor antagonists produce a stretching syndrome previously shown to be mediated through cholinergic mechanisms, but Ro 04-6790 and SB-271046 failed to attenuate scopolamine-induced deficits in a test of contextual fear conditioning. We also failed to replicate the significant effects reported previously in both an autoshaping task and in a version of the Morris water maze. The results of our experiments are not consistent with previous reports that suggested that 5-HT6 antagonists might have therapeutic potential for cognitive disorders.

Insights into the structure and pharmacology of the human trace amine-associated receptor 1 (hTAAR1): homology modelling and docking studies.

PubMed

Cichero, Elena; Espinoza, Stefano; Gainetdinov, Raul R; Brasili, Livio; Fossa, Paola

2013-04-01

Trace amine-associated receptor 1 (TAAR1) is a G protein-coupled receptor that belongs to the family of TAAR receptors and responds to a class of compounds called trace amines, such as β-phenylethylamine (β-PEA) and 3-iodothyronamine (T(1)AM). The receptor is known to have a very rich pharmacology and could be also activated by other classes of compounds, including adrenergic and serotonergic ligands. It is expected that targeting TAAR1 could provide a novel pharmacological approach to correct monoaminergic dysfunctions found in several brain disorders, such as schizophrenia, depression, attention deficit hyperactivity disorder and Parkinson's disease. Only recently, the first selective TAAR1 agonist RO5166017 has been identified. To explore the molecular mechanisms of protein-agonist interaction and speed up the identification of new chemical entities acting on this biomolecular target, we derived a homology model for the hTAAR1. The putative protein-binding site has been explored by comparing the hTAAR1 model with the β(2)-adrenoreceptor binding site, available by X-ray crystallization studies, and with the homology modelled 5HT(1A) receptor. The obtained results, in tandem with docking studies performed with RO5166017, β-PEA and T(1)AM, provided an opportunity to reasonably identify the hTAAR1 key residues involved in ligand recognition and thus define important starting points to design new agonists. © 2012 John Wiley & Sons A/S.

Computational Insight Into the Structural Organization of Full-Length Toll-Like Receptor 4 Dimer in a Model Phospholipid Bilayer

PubMed Central

Patra, Mahesh Chandra; Kwon, Hyuk-Kwon; Batool, Maria; Choi, Sangdun

2018-01-01

Toll-like receptors (TLRs) are a unique category of pattern recognition receptors that recognize distinct pathogenic components, often utilizing the same set of downstream adaptors. Specific molecular features of extracellular, transmembrane (TM), and cytoplasmic domains of TLRs are crucial for coordinating the complex, innate immune signaling pathway. Here, we constructed a full-length structural model of TLR4—a widely studied member of the interleukin-1 receptor/TLR superfamily—using homology modeling, protein–protein docking, and molecular dynamics simulations to understand the differential domain organization of TLR4 in a membrane-aqueous environment. Results showed that each functional domain of the membrane-bound TLR4 displayed several structural transitions that are biophysically essential for plasma membrane integration. Specifically, the extracellular and cytoplasmic domains were partially immersed in the upper and lower leaflets of the membrane bilayer. Meanwhile, TM domains tilted considerably to overcome the hydrophobic mismatch with the bilayer core. Our analysis indicates an alternate dimerization or a potential oligomerization interface of TLR4-TM. Moreover, the helical properties of an isolated TM dimer partly agree with that of the full-length receptor. Furthermore, membrane-absorbed or solvent-exposed surfaces of the toll/interleukin-1 receptor domain are consistent with previous X-ray crystallography and biochemical studies. Collectively, we provided a complete structural model of membrane-bound TLR4 that strengthens our current understanding of the complex mechanism of receptor activation and adaptor recruitment in the innate immune signaling pathway. PMID:29593733

Expression profiling of G-protein-coupled receptors in human urothelium and related cell lines.

PubMed

Ochodnický, Peter; Humphreys, Sian; Eccles, Rachel; Poljakovic, Mirjana; Wiklund, Peter; Michel, Martin C

2012-09-01

What's known on the subject? and What does the study add? Urothelium emerged as a crucial integrator of sensory inputs and outputs in the bladder wall, and urothelial G-protein-coupled receptors (GPCRs) may represent plausible targets for treatment of various bladder pathologies. Urothelial cell lines provide a useful tool to study urothelial receptor function, but their validity as models for native human urothelium remains unclear. We characterize the mRNA expression of genes coding for GPCRs in human freshly isolated urothelium and compare the expression pattern with those in human urothelial cell lines. To characterize the mRNA expression pattern of genes coding for G-protein-coupled receptors (GPCRs) in human freshly isolated urothelium. To compare GPCR expression in human urothelium-derived cell lines to explore the suitability of these cell lines as model systems to study urothelial function. Native human urothelium (commercially sourced) and human urothelium-derived non-cancer (UROtsa and TERT-NHUC) and cancer (J82) cell lines were used. For mRNA expression profiling we used custom-designed real-time polymerase chain reaction array for 40 receptors and several related genes. Native urothelium expressed a wide variety of GPCRs, including α(1A), α(1D) and all subtypes of α(2) and β adrenoceptors. In addition, M(2) and M(3) cholinergic muscarinic receptors, angiotensin II AT(1) receptor, serotonin 5-HT(2A) receptor and all subtypes of bradykinin, endothelin, cannabinoid, tachykinin and sphingosine-1-phosphate receptors were detected. Nerve growth factor and both its low- and high-affinity receptors were also expressed in urothelium. In all cell lines expression of most GPCRs was markedly downregulated, with few exceptions. In UROtsa cells, but much less in other cell lines, the expression of β(2) adrenoceptors, M(3) muscarinic receptors, B(1) and B(2) bradykinin receptors, ET(B) endothelin receptors and several subtypes of sphingosine-1-phosphate receptors was largely retained. Human urothelium expresses a wide range of receptors which enables sensing and integration of various extracellular signals. Human urothelium-derived cell lines, especially UROtsa cells, show comparable mRNA expression to native tissue for several physiologically relevant GPCRs, but lose expression of many other receptors. The use of cell lines as model systems of human urothelium requires careful validation of suitability for the genes of interest. © 2012 BJU INTERNATIONAL.

Receptor model comparisons and wind direction analyses of volatile organic compounds and submicrometer particles in an arid, binational, urban air shed.

PubMed

Mukerjee, Shaibal; Norris, Gary A; Smith, Luther A; Noble, Christopher A; Neas, Lucas M; Ozkaynak, A Halûk; Gonzales, Melissa

2004-04-15

The relationship between continuous measurements of volatile organic compounds sources and particle number was evaluated at a Photochemical Assessment Monitoring Station Network (PAMS) site located near the U.S.-Mexico Border in central El Paso, TX. Sources of volatile organic compounds (VOCs) were investigated using the multivariate receptor model UNMIX and the effective variance least squares receptor model known as Chemical Mass Balance (CMB, Version 8.0). As expected from PAMS measurements, overall findings from data screening as well as both receptor models confirmed that mobile sources were the major source of VOCs. Comparison of hourly source contribution estimates (SCEs) from the two receptor models revealed significant differences in motor vehicle exhaust and evaporative gasoline contributions. However, the motor vehicle exhaust contributions were highly correlated with each other. Motor vehicle exhaust was also correlated with the ultrafine and accumulation mode particle count, which suggests that motor vehicle exhaust is a source of these particles at the measurement site. Wind sector analyses were performed using the SCE and pollutant data to assess source location of VOCs, particle count, and criteria pollutants. Results from this study have application to source apportionment studies and mobile source emission control strategies that are ongoing in this air shed.

Integrated experimental and model-based analysis reveals the spatial aspects of EGFR activation dynamics

PubMed Central

Shankaran, Harish; Zhang, Yi; Chrisler, William B.; Ewald, Jonathan A.; Wiley, H. Steven; Resat, Haluk

2012-01-01

The epidermal growth factor receptor (EGFR) belongs to the ErbB family of receptor tyrosine kinases, and controls a diverse set of cellular responses relevant to development and tumorigenesis. ErbB activation is a complex process involving receptor-ligand binding, receptor dimerization, phosphorylation, and trafficking (internalization, recycling and degradation), which together dictate the spatio-temporal distribution of active receptors within the cell. The ability to predict this distribution, and elucidation of the factors regulating it, would help to establish a mechanistic link between ErbB expression levels and the cellular response. Towards this end, we constructed mathematical models to determine the contributions of receptor dimerization and phosphorylation to EGFR activation, and to examine the dependence of these processes on sub-cellular location. We collected experimental datasets for EGFR activation dynamics in human mammary epithelial cells, with the specific goal of model parameterization, and used the data to estimate parameters for several alternate models. Model-based analysis indicated that: 1) signal termination via receptor dephosphorylation in late endosomes, prior to degradation, is an important component of the response, 2) less than 40% of the receptors in the cell are phosphorylated at any given time, even at saturating ligand doses, and 3) receptor phosphorylation kinetics at the cell surface and early endosomes are comparable. We validated the last finding by measuring the EGFR dephosphorylation rates at various times following ligand addition both in whole cells and in endosomes using ELISAs and fluorescent imaging. Overall, our results provide important information on how EGFR phosphorylation levels are regulated within cells. This study demonstrates that an iterative cycle of experiments and modeling can be used to gain mechanistic insight regarding complex cell signaling networks. PMID:22952062

Receptor-based 3D QSAR analysis of estrogen receptor ligands - merging the accuracy of receptor-based alignments with the computational efficiency of ligand-based methods

NASA Astrophysics Data System (ADS)

Sippl, Wolfgang

2000-08-01

One of the major challenges in computational approaches to drug design is the accurate prediction of binding affinity of biomolecules. In the present study several prediction methods for a published set of estrogen receptor ligands are investigated and compared. The binding modes of 30 ligands were determined using the docking program AutoDock and were compared with available X-ray structures of estrogen receptor-ligand complexes. On the basis of the docking results an interaction energy-based model, which uses the information of the whole ligand-receptor complex, was generated. Several parameters were modified in order to analyze their influence onto the correlation between binding affinities and calculated ligand-receptor interaction energies. The highest correlation coefficient ( r 2 = 0.617, q 2 LOO = 0.570) was obtained considering protein flexibility during the interaction energy evaluation. The second prediction method uses a combination of receptor-based and 3D quantitative structure-activity relationships (3D QSAR) methods. The ligand alignment obtained from the docking simulations was taken as basis for a comparative field analysis applying the GRID/GOLPE program. Using the interaction field derived with a water probe and applying the smart region definition (SRD) variable selection, a significant and robust model was obtained ( r 2 = 0.991, q 2 LOO = 0.921). The predictive ability of the established model was further evaluated by using a test set of six additional compounds. The comparison with the generated interaction energy-based model and with a traditional CoMFA model obtained using a ligand-based alignment ( r 2 = 0.951, q 2 LOO = 0.796) indicates that the combination of receptor-based and 3D QSAR methods is able to improve the quality of the underlying model.

Localization and regulation of glucagon receptors in the chick eye and preproglucagon and glucagon receptor expression in the mouse eye.

PubMed

Feldkaemper, Marita P; Burkhardt, Eva; Schaeffel, Frank

2004-09-01

Myopia is a condition in which the eye is too long for the focal length of cornea and lens. Analysis of the messengers that are released by the retina to control axial eye growth in the animal model of the chicken revealed that glucagon-immunoreactive amacrine cells are involved in the retinal image processing that controls the growth of the sclera. It was found that the amount of retinal glucagon mRNA increased during treatment with positive lenses and pharmacological studies supported the idea that glucagon may act as a stop signal for eye growth. Glucagon exerts its regulatory effects by binding to a single type of glucagon receptor. In this study, we have sequenced the chicken glucagon receptor and compared its DNA and amino acid sequence with the human and mouse homologues. After sequencing about 80% of the receptor, we found a homology between 79.4 and 75.6% on cDNA level. At the protein level, about 73% of the amino acids were identical. Moreover, the cellular localization and regulation of the glucagon receptor in the chick retina was studied. In situ hybridization studies showed that many cells in the ganglion cell layer and inner nuclear layer, and some cells in the outer nuclear layer, express the receptor mRNA. Injection of the glucagon agonist Lys17,18,Glu21-glucagon induced a down-regulation of glucagon receptor mRNA content. Since the mouse would be an attractive mammalian model to study the biochemical and genetic basis of myopia, and because recent studies have demonstrated that form deprivation myopia can be induced, the expression of preproglucagon and glucagon receptor genes were also studied in the mouse retina and were found to be expressed.

Comparison of Kinetic Models for Dual-Tracer Receptor Concentration Imaging in Tumors

PubMed Central

Hamzei, Nazanin; Samkoe, Kimberley S; Elliott, Jonathan T; Holt, Robert W; Gunn, Jason R; Hasan, Tayyaba; Pogue, Brian W; Tichauer, Kenneth M

2014-01-01

Molecular differences between cancerous and healthy tissue have become key targets for novel therapeutics specific to tumor receptors. However, cancer cell receptor expression can vary within and amongst different tumors, making strategies that can quantify receptor concentration in vivo critical for the progression of targeted therapies. Recently a dual-tracer imaging approach capable of providing quantitative measures of receptor concentration in vivo was developed. It relies on the simultaneous injection and imaging of receptor-targeted tracer and an untargeted tracer (to account for non-specific uptake of the targeted tracer). Early implementations of this approach have been structured on existing “reference tissue” imaging methods that have not been optimized for or validated in dual-tracer imaging. Using simulations and mouse tumor model experimental data, the salient findings in this study were that all widely used reference tissue kinetic models can be used for dual-tracer imaging, with the linearized simplified reference tissue model offering a good balance of accuracy and computational efficiency. Moreover, an alternate version of the full two-compartment reference tissue model can be employed accurately by assuming that the K1s of the targeted and untargeted tracers are similar to avoid assuming an instantaneous equilibrium between bound and free states (made by all other models). PMID:25414912

Is the Acute NMDA Receptor Hypofunction a Valid Model of Schizophrenia?

PubMed Central

Adell, Albert; Jiménez-Sánchez, Laura; López-Gil, Xavier; Romón, Tamara

2012-01-01

Several genetic, neurodevelopmental, and pharmacological animal models of schizophrenia have been established. This short review examines the validity of one of the most used pharmacological model of the illness, ie, the acute administration of N-methyl-D-aspartate (NMDA) receptor antagonists in rodents. In some cases, data on chronic or prenatal NMDA receptor antagonist exposure have been introduced for comparison. The face validity of acute NMDA receptor blockade is granted inasmuch as hyperlocomotion and stereotypies induced by phencyclidine, ketamine, and MK-801 are regarded as a surrogate for the positive symptoms of schizophrenia. In addition, the loss of parvalbumin-containing cells (which is one of the most compelling finding in postmortem schizophrenia brain) following NMDA receptor blockade adds construct validity to this model. However, the lack of changes in glutamic acid decarboxylase (GAD67) is at variance with human studies. It is possible that changes in GAD67 are more reflective of the neurodevelopmental condition of schizophrenia. Finally, the model also has predictive validity, in that its behavioral and transmitter activation in rodents are responsive to antipsychotic treatment. Overall, although not devoid of drawbacks, the acute administration of NMDA receptor antagonists can be considered as a good model of schizophrenia bearing a satisfactory degree of validity. PMID:21965469

Aromatic interactions impact ligand binding and function at serotonin 5-HT2C G protein-coupled receptors: receptor homology modelling, ligand docking, and molecular dynamics results validated by experimental studies

NASA Astrophysics Data System (ADS)

Córdova-Sintjago, Tania; Villa, Nancy; Fang, Lijuan; Booth, Raymond G.

2014-02-01

The serotonin (5-hydroxytryptamine, 5-HT) 5-HT2 G protein-coupled receptor (GPCR) family consists of types 2A, 2B, and 2C that share ∼75% transmembrane (TM) sequence identity. Agonists for 5-HT2C receptors are under development for psychoses; whereas, at 5-HT2A receptors, antipsychotic effects are associated with antagonists - in fact, 5-HT2A agonists can cause hallucinations and 5-HT2B agonists cause cardiotoxicity. It is known that 5-HT2A TM6 residues W6.48, F6.51, and F6.52 impact ligand binding and function; however, ligand interactions with these residues at the 5-HT2C receptor have not been reported. To predict and validate molecular determinants for 5-HT2C-specific activation, results from receptor homology modelling, ligand docking, and molecular dynamics simulation studies were compared with experimental results for ligand binding and function at wild type and W6.48A, F6.51A, and F6.52A point-mutated 5-HT2C receptors.

Prediction of in vitro and in vivo oestrogen receptor activity using hierarchical clustering

EPA Science Inventory

In this study, hierarchical clustering classification models were developed to predict in vitro and in vivo oestrogen receptor (ER) activity. Classification models were developed for binding, agonist, and antagonist in vitro ER activity and for mouse in vivo uterotrophic ER bindi...

Role of physicochemical properties in the activation of peroxisome proliferator-activated receptor δ.

PubMed

Maltarollo, Vinícius G; Homem-de-Mello, Paula; Honorio, Káthia M

2011-10-01

Current researches on treatments for metabolic diseases involve a class of biological receptors called peroxisome proliferator-activated receptors (PPARs), which control the metabolism of carbohydrates and lipids. A subclass of these receptors, PPARδ, regulates several metabolic processes, and the substances that activate them are being studied as new drug candidates for the treatment of diabetes mellitus and metabolic syndrome. In this study, several PPARδ agonists with experimental biological activity were selected for a structural and chemical study. Electronic, stereochemical, lipophilic and topological descriptors were calculated for the selected compounds using various theoretical methods, such as density functional theory (DFT). Fisher's weight and principal components analysis (PCA) methods were employed to select the most relevant variables for this study. The partial least squares (PLS) method was used to construct the multivariate statistical model, and the best model obtained had 4 PCs, q ( 2 ) = 0.80 and r ( 2 ) = 0.90, indicating a good internal consistency. The prediction residues calculated for the compounds in the test set had low values, indicating the good predictive capability of our PLS model. The model obtained in this study is reliable and can be used to predict the biological activity of new untested compounds. Docking studies have also confirmed the importance of the molecular descriptors selected for this system.

A Review of the Updated Pharmacophore for the Alpha 5 GABA(A) Benzodiazepine Receptor Model

PubMed Central

Clayton, Terry; Poe, Michael M.; Rallapalli, Sundari; Biawat, Poonam; Savić, Miroslav M.; Rowlett, James K.; Gallos, George; Emala, Charles W.; Kaczorowski, Catherine C.; Stafford, Douglas C.; Arnold, Leggy A.; Cook, James M.

2015-01-01

An updated model of the GABA(A) benzodiazepine receptor pharmacophore of the α5-BzR/GABA(A) subtype has been constructed prompted by the synthesis of subtype selective ligands in light of the recent developments in both ligand synthesis, behavioral studies, and molecular modeling studies of the binding site itself. A number of BzR/GABA(A) α5 subtype selective compounds were synthesized, notably α5-subtype selective inverse agonist PWZ-029 (1) which is active in enhancing cognition in both rodents and primates. In addition, a chiral positive allosteric modulator (PAM), SH-053-2′F-R-CH3 (2), has been shown to reverse the deleterious effects in the MAM-model of schizophrenia as well as alleviate constriction in airway smooth muscle. Presented here is an updated model of the pharmacophore for α5β2γ2 Bz/GABA(A) receptors, including a rendering of PWZ-029 docked within the α5-binding pocket showing specific interactions of the molecule with the receptor. Differences in the included volume as compared to α1β2γ2, α2β2γ2, and α3β2γ2 will be illustrated for clarity. These new models enhance the ability to understand structural characteristics of ligands which act as agonists, antagonists, or inverse agonists at the Bz BS of GABA(A) receptors. PMID:26682068

NRP-1 Receptor Expression Mismatch in Skin of Subjects with Experimental and Diabetic Small Fiber Neuropathy

PubMed Central

Van Acker, Nathalie; Ragé, Michael; Vermeirsch, Hilde; Schrijvers, Dorien; Nuydens, Rony; Byttebier, Geert; Timmers, Maarten; De Schepper, Stefanie; Streffer, Johannes; Andries, Luc; Plaghki, Léon; Cras, Patrick; Meert, Theo

2016-01-01

The in vivo cutaneous nerve regeneration model using capsaicin is applied extensively to study the regenerative mechanisms and therapeutic efficacy of disease modifying molecules for small fiber neuropathy (SFN). Since mismatches between functional and morphological nerve fiber recovery are described for this model, we aimed at determining the capability of the capsaicin model to truly mimic the morphological manifestations of SFN in diabetes. As nerve and blood vessel growth and regenerative capacities are defective in diabetes, we focused on studying the key regulator of these processes, the neuropilin-1 (NRP-1)/semaphorin pathway. This led us to the evaluation of NRP-1 receptor expression in epidermis and dermis of subjects presenting experimentally induced small fiber neuropathy, diabetic polyneuropathy and of diabetic subjects without clinical signs of small fiber neuropathy. The NRP-1 receptor was co-stained with CD31 vessel-marker using immunofluorescence and analyzed with Definiens® technology. This study indicates that capsaicin application results in significant loss of epidermal NRP-1 receptor expression, whereas diabetic subjects presenting small fiber neuropathy show full epidermal NRP-1 expression in contrast to the basal expression pattern seen in healthy controls. Capsaicin induced a decrease in dermal non-vascular NRP-1 receptor expression which did not appear in diabetic polyneuropathy. We can conclude that the capsaicin model does not mimic diabetic neuropathy related changes for cutaneous NRP-1 receptor expression. In addition, our data suggest that NRP-1 might play an important role in epidermal nerve fiber loss and/or defective regeneration and that NRP-1 receptor could change the epidermal environment to a nerve fiber repellant bed possibly through Sem3A in diabetes. PMID:27598321

The influence of synaptic size on AMPA receptor activation: a Monte Carlo model.

PubMed

Montes, Jesus; Peña, Jose M; DeFelipe, Javier; Herreras, Oscar; Merchan-Perez, Angel

2015-01-01

Physiological and electron microscope studies have shown that synapses are functionally and morphologically heterogeneous and that variations in size of synaptic junctions are related to characteristics such as release probability and density of postsynaptic AMPA receptors. The present article focuses on how these morphological variations impact synaptic transmission. We based our study on Monte Carlo computational simulations of simplified model synapses whose morphological features have been extracted from hundreds of actual synaptic junctions reconstructed by three-dimensional electron microscopy. We have examined the effects that parameters such as synaptic size or density of AMPA receptors have on the number of receptors that open after release of a single synaptic vesicle. Our results indicate that the maximum number of receptors that will open after the release of a single synaptic vesicle may show a ten-fold variation in the whole population of synapses. When individual synapses are considered, there is also a stochastical variability that is maximal in small synapses with low numbers of receptors. The number of postsynaptic receptors and the size of the synaptic junction are the most influential parameters, while the packing density of receptors or the concentration of extrasynaptic transporters have little or no influence on the opening of AMPA receptors.

The Influence of Synaptic Size on AMPA Receptor Activation: A Monte Carlo Model

PubMed Central

Montes, Jesus; Peña, Jose M.; DeFelipe, Javier; Herreras, Oscar; Merchan-Perez, Angel

2015-01-01

Physiological and electron microscope studies have shown that synapses are functionally and morphologically heterogeneous and that variations in size of synaptic junctions are related to characteristics such as release probability and density of postsynaptic AMPA receptors. The present article focuses on how these morphological variations impact synaptic transmission. We based our study on Monte Carlo computational simulations of simplified model synapses whose morphological features have been extracted from hundreds of actual synaptic junctions reconstructed by three-dimensional electron microscopy. We have examined the effects that parameters such as synaptic size or density of AMPA receptors have on the number of receptors that open after release of a single synaptic vesicle. Our results indicate that the maximum number of receptors that will open after the release of a single synaptic vesicle may show a ten-fold variation in the whole population of synapses. When individual synapses are considered, there is also a stochastical variability that is maximal in small synapses with low numbers of receptors. The number of postsynaptic receptors and the size of the synaptic junction are the most influential parameters, while the packing density of receptors or the concentration of extrasynaptic transporters have little or no influence on the opening of AMPA receptors. PMID:26107874

Delta-opioid receptors as targets for migraine therapy.

PubMed

Charles, Andrew; Pradhan, Amynah A

2016-06-01

The purpose of this review is to contrast the properties of the δ-opioid receptor with those of the μ-opioid receptor, which is the primary target of most currently available opioid analgesics. We also discuss preclinical evidence that indicates the potential efficacy of δ-opioid receptor agonists as migraine therapy. The use of currently available opioid analgesics is highly problematic for patients with migraine. Delta-opioid receptors have key differences from μ receptors; these differences make the δ receptor an attractive therapeutic target for migraine. Delta-opioid receptors are expressed in both the peripheral and central nervous system in anatomical regions and cell types that are believed to play a role in migraine. Delta-receptor agonists have also shown promising effects in multiple migraine models, including nitroglycerin evoked hyperalgesia and conditioned place aversion, and cortical spreading depression. Evidence from animal models indicates that activation of δ receptors is less likely to cause tolerance and dependence, and less likely to cause hyperalgesia. In addition, δ receptors may have antidepressant and anxiolytic properties that are distinct from those of μ receptors. In human studies investigating other conditions, δ-receptor agonists have been generally safe and well tolerated. Delta-opioid receptor agonists have promising potential as acute and/or preventive migraine therapies, without the problems associated with currently used opioid analgesics.

A computational study of the chemokine receptor CXCR1 bound with interleukin-8

NASA Astrophysics Data System (ADS)

Wang, Yang; Severin Lupala, Cecylia; Wang, Ting; Li, Xuanxuan; Yun, Ji-Hye; Park, Jae-hyun; Jin, Zeyu; Lee, Weontae; Tan, Leihan; Liu, Haiguang

2018-03-01

CXCR1 is a G-protein coupled receptor, transducing signals from chemokines, in particular the interleukin-8 (IL8) molecules. This study combines homology modeling and molecular dynamics simulation methods to study the structure of CXCR1-IL8 complex. By using CXCR4-vMIP-II crystallography structure as the homologous template, CXCR1-IL8 complex structure was constructed, and then refined using all-atom molecular dynamics simulations. Through extensive simulations, CXCR1-IL8 binding poses were investigated in detail. Furthermore, the role of the N-terminal of CXCR1 receptor was studied by comparing four complex models differing in the N-terminal sequences. The results indicate that the receptor N-terminal affects the binding of IL8 significantly. With a shorter N-terminal domain, the binding of IL8 to CXCR1 becomes unstable. The homology modeling and simulations also reveal the key receptor-ligand residues involved in the electrostatic interactions known to be vital for complex formation. Project supported by the National Natural Science Foundation of China (Grant Nos. 11575021, U1530401, and U1430237) and the National Research Foundation of Korea (Grant Nos. NRF-2017R1A2B2008483 and NRF-2016R1A6A3A04010213).

Hybrid approach to structure modeling of the histamine H3 receptor: Multi-level assessment as a tool for model verification.

PubMed

Jończyk, Jakub; Malawska, Barbara; Bajda, Marek

2017-01-01

The crucial role of G-protein coupled receptors and the significant achievements associated with a better understanding of the spatial structure of known receptors in this family encouraged us to undertake a study on the histamine H3 receptor, whose crystal structure is still unresolved. The latest literature data and availability of different software enabled us to build homology models of higher accuracy than previously published ones. The new models are expected to be closer to crystal structures; and therefore, they are much more helpful in the design of potential ligands. In this article, we describe the generation of homology models with the use of diverse tools and a hybrid assessment. Our study incorporates a hybrid assessment connecting knowledge-based scoring algorithms with a two-step ligand-based docking procedure. Knowledge-based scoring employs probability theory for global energy minimum determination based on information about native amino acid conformation from a dataset of experimentally determined protein structures. For a two-step docking procedure two programs were applied: GOLD was used in the first step and Glide in the second. Hybrid approaches offer advantages by combining various theoretical methods in one modeling algorithm. The biggest advantage of hybrid methods is their intrinsic ability to self-update and self-refine when additional structural data are acquired. Moreover, the diversity of computational methods and structural data used in hybrid approaches for structure prediction limit inaccuracies resulting from theoretical approximations or fuzziness of experimental data. The results of docking to the new H3 receptor model allowed us to analyze ligand-receptor interactions for reference compounds.

Interaction Studies of Withania Somnifera's Key Metabolite Withaferin A with Different Receptors Assoociated with Cardiovascular Disease.

PubMed

Ravindran, Rekha; Sharma, Nitika; Roy, Sujata; Thakur, Ashoke R; Ganesh, Subhadra; Kumar, Sriram; Devi, Jamuna; Rajkumar, Johanna

2015-01-01

Withania somnifera commonly known as Ashwagandha in India is used in many herbal formulations to treat various cardiovascular diseases. The key metabolite of this plant, Withaferin A was analyzed for its molecular mechanism through docking studies on different targets of cardiovascular disease. Six receptor proteins associated with cardiovascular disease were selected and interaction studies were performed with Withaferin A using AutoDock Vina. CORINA was used to model the small molecules and HBAT to compute the hydrogen bonding. Among the six targets, β1- adrenergic receptors, HMG-CoA and Angiotensinogen-converting enzyme showed significant interaction with Withaferin A. Pharmacophore modeling was done using PharmaGist to understand the pharmacophoric potential of Withaferin A. Clustering of Withaferin A with different existing drug molecules for cardiovascular disease was performed with ChemMine based on structural similarity and physicochemical properties. The ability of natural active component, Withaferin A to interact with different receptors associated with cardiovascular disease was elucidated with various modeling techniques. These studies conclusively revealed Withaferin A as a potent lead compound against multiple targets associated with cardiovascular disease.

Coupled stochastic spatial and non-spatial simulations of ErbB1 signaling pathways demonstrate the importance of spatial organization in signal transduction.

PubMed

Costa, Michelle N; Radhakrishnan, Krishnan; Wilson, Bridget S; Vlachos, Dionisios G; Edwards, Jeremy S

2009-07-23

The ErbB family of receptors activates intracellular signaling pathways that control cellular proliferation, growth, differentiation and apoptosis. Given these central roles, it is not surprising that overexpression of the ErbB receptors is often associated with carcinogenesis. Therefore, extensive laboratory studies have been devoted to understanding the signaling events associated with ErbB activation. Systems biology has contributed significantly to our current understanding of ErbB signaling networks. However, although computational models have grown in complexity over the years, little work has been done to consider the spatial-temporal dynamics of receptor interactions and to evaluate how spatial organization of membrane receptors influences signaling transduction. Herein, we explore the impact of spatial organization of the epidermal growth factor receptor (ErbB1/EGFR) on the initiation of downstream signaling. We describe the development of an algorithm that couples a spatial stochastic model of membrane receptors with a nonspatial stochastic model of the reactions and interactions in the cytosol. This novel algorithm provides a computationally efficient method to evaluate the effects of spatial heterogeneity on the coupling of receptors to cytosolic signaling partners. Mathematical models of signal transduction rarely consider the contributions of spatial organization due to high computational costs. A hybrid stochastic approach simplifies analyses of the spatio-temporal aspects of cell signaling and, as an example, demonstrates that receptor clustering contributes significantly to the efficiency of signal propagation from ligand-engaged growth factor receptors.

Empirically-based modeling and mapping to consider the co-occurrence of ecological receptors and stressors

EPA Science Inventory

Part of the ecological risk assessment process involves examining the potential for environmental stressors and ecological receptors to co-occur across a landscape. In this study, we introduce a Bayesian joint modeling framework for use in evaluating and mapping the co-occurrence...

Human IgG subclass cross-species reactivity to mouse and cynomolgus monkey Fcγ receptors.

PubMed

Derebe, Mehabaw G; Nanjunda, Rupesh K; Gilliland, Gary L; Lacy, Eilyn R; Chiu, Mark L

2018-05-01

In therapeutic antibody discovery and early development, mice and cynomolgus monkey are used as animal models to assess toxicity, efficacy and other properties of candidate molecules. As more candidate antibodies are based on human immunoglobulin (IgG) subclasses, many strategies are pursued to simulate the human system in the test animal. However, translation rate from a successful preclinical trial to an approved drug is extremely low. This may partly be due to differences in interaction of human IgG based candidate molecules to endogenous Fcγ receptors of model animals in comparison to those of human Fcγ receptors. In this study, we compare binding characteristics of human IgG subclasses commonly used in drug development (IgG1, IgG2, IgG4) and their respective Fc silent versions (IgG1σ, IgG2σ, IgG4 PAA) to human, mouse, and cynomolgus monkey Fcγ receptors. To control interactions between Fab and Fc domains, the test IgGs all have the same variable region sequences. We found distinct variations of interaction of human IgG subclasses to model animal Fcγ receptors in comparison to their human counterparts. Particularly, cynomolgus monkey Fcγ receptors showed consistently tighter binding to human IgGs than human Fcγ receptors. Moreover, the presumably Fc silent human IgG4 PAA framework bound to cynomolgus monkey FcγRI with nanomolar affinity while only very weak binding was observed for the human FcγRI. Our results highlighted the need for a thorough in vitro affinity characterization of candidate IgGs against model animal Fcγ receptors and careful design of preclinical studies. Copyright © 2018. Published by Elsevier B.V.

[The potential of group II metabotropic glutamate receptor antagonists as a novel antidepressant].

PubMed

Chaki, Shigeyuki

2012-08-01

Recently, abnormalities of glutamatergic transmission have been implicated in the pathophysiology of depression. Moreover, both ketamine, an NMDA receptor antagonist, and riluzole, a modulator of glutamatergic, transmission have been reported to be effective for the treatment of patients with treatment-refractory depression. Based on these findings, extensive studies to develop agents acting on glutamatergic transmission have been conducted. Glutamate receptors are divided into two main subtypes, ionotropic glutamate receptors and metabotropic glutamate (mGlu) receptors, both of which have subtypes. Of these, much attention has been paid to mGlu2/3 receptors. mGlu2/3 receptor antagonists such as MGS0039 and LY341495 have been reported to exert antidepressant effects in animal models of depression including the forced swim test, tail suspension test, learned helplessness paradigm, olfactory bulmectomy model and isolation rearing model, and to enhance serotonin release in the prefrontal cortex and dopamine release in the nucleus accumbens. Moreover, activation of AMPA receptor and mTOR signaling have been suggested to be involved in the antidepressant effects of mGlu2/3 receptor antagonists, as demonstrated in the actions of ketamine. Thus, mGlu2/3 receptor antagonists may share some neural networks with ketamine in exerting their antidepressant effects. In addition, the potential of other agents targeting glutamatergic transmission for novel antidepressants is being investigated.

Designing peptide inhibitor of insulin receptor to induce diabetes mellitus type 2 in animal model Mus musculus.

PubMed

Permatasari, Galuh W; Utomo, Didik H; Widodo

2016-10-01

A designing peptide as agent for inducing diabetes mellitus type 2 (T2DM) in an animal model is challenging. The computational approach provides a sophisticated tool to design a functional peptide that may block the insulin receptor activity. The peptide that able to inhibit the binding between insulin and insulin receptor is a warrant for inducing T2DM. Therefore, we designed a potential peptide inhibitor of insulin receptor as an agent to generate T2DM animal model by bioinformatics approach. The peptide has been developed based on the structure of insulin receptor binding site of insulin and then modified it to obtain the best properties of half life, hydrophobicity, antigenicity, and stability binding into insulin receptor. The results showed that the modified peptide has characteristics 100h half-life, high-affinity -95.1±20, and high stability 28.17 in complex with the insulin receptor. Moreover, the modified peptide has molecular weight 4420.8g/Mol and has no antigenic regions. Based on the molecular dynamic simulation, the complex of modified peptide-insulin receptor is more stable than the commercial insulin receptor blocker. This study suggested that the modified peptide has the promising performance to block the insulin receptor activity that potentially induce diabetes mellitus type 2 in mice. Copyright © 2016 Elsevier Ltd. All rights reserved.

Retrieval-induced NMDA receptor-dependent Arc expression in two models of cocaine-cue memory.

PubMed

Alaghband, Yasaman; O'Dell, Steven J; Azarnia, Siavash; Khalaj, Anna J; Guzowski, John F; Marshall, John F

2014-12-01

The association of environmental cues with drugs of abuse results in persistent drug-cue memories. These memories contribute significantly to relapse among addicts. While conditioned place preference (CPP) is a well-established paradigm frequently used to examine the modulation of drug-cue memories, very few studies have used the non-preference-based model conditioned activity (CA) for this purpose. Here, we used both experimental approaches to investigate the neural substrates of cocaine-cue memories. First, we directly compared, in a consistent setting, the involvement of cortical and subcortical brain regions in cocaine-cue memory retrieval by quantifying activity-regulated cytoskeletal-associated (Arc) protein expression in both the CPP and CA models. Second, because NMDA receptor activation is required for Arc expression, we investigated the NMDA receptor dependency of memory persistence using the CA model. In both the CPP and CA models, drug-paired animals showed significant increases in Arc immunoreactivity in regions of the frontal cortex and amygdala compared to unpaired controls. Additionally, administration of a NMDA receptor antagonist (MK-801 or memantine) immediately after cocaine-CA memory reactivation impaired the subsequent conditioned locomotion associated with the cocaine-paired environment. The enhanced Arc expression evident in a subset of corticolimbic regions after retrieval of a cocaine-context memory, observed in both the CPP and CA paradigms, likely signifies that these regions: (i) are activated during retrieval of these memories irrespective of preference-based decisions, and (ii) undergo neuroplasticity in order to update information about cues previously associated with cocaine. This study also establishes the involvement of NMDA receptors in maintaining memories established using the CA model, a characteristic previously demonstrated using CPP. Overall, these results demonstrate the utility of the CA model for studies of cocaine-context memory and suggest the involvement of an NMDA receptor-dependent Arc induction pathway in drug-cue memory interference. Copyright © 2014 Elsevier Inc. All rights reserved.

Retrieval-induced NMDA receptor-dependent Arc expression in two models of cocaine-cue memory

PubMed Central

Alaghband, Yasaman; O'Dell, Steven J.; Azarnia, Siavash; Khalaj, Anna J.; Guzowski, John F.; Marshall, John F.

2014-01-01

The association of environmental cues with drugs of abuse results in persistent drug-cue memories. These memories contribute significantly to relapse among addicts. While conditioned place preference (CPP) is a well-established paradigm frequently used to examine the modulation of drug-cue memories, very few studies have used the non-preference-based model conditioned activity (CA) for this purpose. Here, we used both experimental approaches to investigate the neural substrates of cocaine-cue memories. First, we directly compared, in a consistent setting, the involvement of cortical and subcortical brain regions in cocaine-cue memory retrieval by quantifying activity-regulated cytoskeletal associated gene (Arc) protein expression in both the CPP and CA models. Second, because NMDA receptor activation is required for Arc expression, we investigated the NMDA receptor dependency of memory persistence using the CA model. In both the CPP and CA models, drug-paired animals showed significant increases in Arc immunoreactivity in regions of the frontal cortex and amygdala compared to unpaired controls. Additionally, administration of a NMDA receptor antagonist (MK-801 or memantine) immediately after cocaine-CA memory reactivation impaired the subsequent conditioned locomotion associated with the cocaine-paired environment. The enhanced Arc expression evident in a subset of corticolimbic regions after retrieval of a cocaine-context memory, observed in both the CPP and CA paradigms, likely signifies that these regions: (i) are activated during retrieval of these memories irrespective of preference-based decisions, and (ii) undergo neuroplasticity in order to update information about cues previously associated with cocaine. This study also establishes the involvement of NMDA receptors in maintaining memories established using the CA model, a characteristic previously demonstrated using CPP. Overall, these results demonstrate the utility of the CA model for studies of cocaine-context memory and suggest the involvement of an NMDA receptor-dependent Arc induction pathway in drug-cue memory interference. PMID:25225165

Role of dopamine D2 receptors in human reinforcement learning.

PubMed

Eisenegger, Christoph; Naef, Michael; Linssen, Anke; Clark, Luke; Gandamaneni, Praveen K; Müller, Ulrich; Robbins, Trevor W

2014-09-01

Influential neurocomputational models emphasize dopamine (DA) as an electrophysiological and neurochemical correlate of reinforcement learning. However, evidence of a specific causal role of DA receptors in learning has been less forthcoming, especially in humans. Here we combine, in a between-subjects design, administration of a high dose of the selective DA D2/3-receptor antagonist sulpiride with genetic analysis of the DA D2 receptor in a behavioral study of reinforcement learning in a sample of 78 healthy male volunteers. In contrast to predictions of prevailing models emphasizing DA's pivotal role in learning via prediction errors, we found that sulpiride did not disrupt learning, but rather induced profound impairments in choice performance. The disruption was selective for stimuli indicating reward, whereas loss avoidance performance was unaffected. Effects were driven by volunteers with higher serum levels of the drug, and in those with genetically determined lower density of striatal DA D2 receptors. This is the clearest demonstration to date for a causal modulatory role of the DA D2 receptor in choice performance that might be distinct from learning. Our findings challenge current reward prediction error models of reinforcement learning, and suggest that classical animal models emphasizing a role of postsynaptic DA D2 receptors in motivational aspects of reinforcement learning may apply to humans as well.

Role of Dopamine D2 Receptors in Human Reinforcement Learning

PubMed Central

Eisenegger, Christoph; Naef, Michael; Linssen, Anke; Clark, Luke; Gandamaneni, Praveen K; Müller, Ulrich; Robbins, Trevor W

2014-01-01

Influential neurocomputational models emphasize dopamine (DA) as an electrophysiological and neurochemical correlate of reinforcement learning. However, evidence of a specific causal role of DA receptors in learning has been less forthcoming, especially in humans. Here we combine, in a between-subjects design, administration of a high dose of the selective DA D2/3-receptor antagonist sulpiride with genetic analysis of the DA D2 receptor in a behavioral study of reinforcement learning in a sample of 78 healthy male volunteers. In contrast to predictions of prevailing models emphasizing DA's pivotal role in learning via prediction errors, we found that sulpiride did not disrupt learning, but rather induced profound impairments in choice performance. The disruption was selective for stimuli indicating reward, whereas loss avoidance performance was unaffected. Effects were driven by volunteers with higher serum levels of the drug, and in those with genetically determined lower density of striatal DA D2 receptors. This is the clearest demonstration to date for a causal modulatory role of the DA D2 receptor in choice performance that might be distinct from learning. Our findings challenge current reward prediction error models of reinforcement learning, and suggest that classical animal models emphasizing a role of postsynaptic DA D2 receptors in motivational aspects of reinforcement learning may apply to humans as well. PMID:24713613

Stimulation of postsynapse adrenergic α2A receptor improves attention/cognition performance in an animal model of attention deficit hyperactivity disorder.

PubMed

Kawaura, Kazuaki; Karasawa, Jun-ichi; Chaki, Shigeyuki; Hikichi, Hirohiko

2014-08-15

A 5-trial inhibitory avoidance test using spontaneously hypertensive rat (SHR) pups has been used as an animal model of attention deficit hyperactivity disorder (ADHD). However, the roles of noradrenergic systems, which are involved in the pathophysiology of ADHD, have not been investigated in this model. In the present study, the effects of adrenergic α2 receptor stimulation, which has been an effective treatment for ADHD, on attention/cognition performance were investigated in this model. Moreover, neuronal mechanisms mediated through adrenergic α2 receptors were investigated. We evaluated the effects of both clonidine, a non-selective adrenergic α2 receptor agonist, and guanfacine, a selective adrenergic α2A receptor agonist, using a 5-trial inhibitory avoidance test with SHR pups. Juvenile SHR exhibited a shorter transfer latency, compared with juvenile Wistar Kyoto (WKY) rats. Both clonidine and guanfacine significantly prolonged the transfer latency of juvenile SHR. The effects of clonidine and guanfacine were significantly blocked by pretreatment with an adrenergic α2A receptor antagonist. In contrast, the effect of clonidine was not attenuated by pretreatment with an adrenergic α2B receptor antagonist, or an adrenergic α2C receptor antagonist, while it was attenuated by a non-selective adrenergic α2 receptor antagonist. Furthermore, the effects of neither clonidine nor guanfacine were blocked by pretreatment with a selective noradrenergic neurotoxin. These results suggest that the stimulation of the adrenergic α2A receptor improves the attention/cognition performance of juvenile SHR in the 5-trial inhibitory avoidance test and that postsynaptic, rather than presynaptic, adrenergic α2A receptor is involved in this effect. Copyright © 2014 Elsevier B.V. All rights reserved.

Structural interpretation of P2X receptor mutagenesis studies on drug action

PubMed Central

Evans, Richard J

2010-01-01

P2X receptors for ATP are ligand gated cation channels that form from the trimeric assembly of subunits with two transmembrane segments, a large extracellular ligand binding loop, and intracellular amino and carboxy termini. The receptors are expressed throughout the body, involved in functions ranging from blood clotting to inflammation, and may provide important targets for novel therapeutics. Mutagenesis based studies have been used to develop an understanding of the molecular basis of their pharmacology with the aim of developing models of the ligand binding site. A crystal structure for the zebra fish P2X4 receptor in the closed agonist unbound state has been published recently, which provides a major advance in our understanding of the receptors. This review gives an overview of mutagenesis studies that have led to the development of a model of the ATP binding site, as well as identifying residues contributing to allosteric regulation and antagonism. These studies are discussed with reference to the crystal to provide a structural interpretation of the molecular basis of drug action. PMID:20977449

A Randomized, Double-Blind, Crossover Comparison of MK-0929 and Placebo in the Treatment of Adults with ADHD

ERIC Educational Resources Information Center

Rivkin, Anna; Alexander, Robert C.; Knighton, Jennifer; Hutson, Pete H.; Wang, Xiaojing J.; Snavely, Duane B.; Rosah, Thomas; Watt, Alan P.; Reimherr, Fred W.; Adler, Lenard A.

2012-01-01

Objective: Preclinical models, receptor localization, and genetic linkage data support the role of D4 receptors in the etiology of ADHD. This proof-of-concept study was designed to evaluate MK-0929, a selective D4 receptor antagonist as treatment for adult ADHD. Method: A randomized, double-blind, placebo-controlled, crossover study was conducted…

Kinetic modeling of benzodiazepine receptor binding with PET and high specific activity [(11)C]Iomazenil in healthy human subjects.

PubMed

Bremner, J D; Horti, A; Staib, L H; Zea-Ponce, Y; Soufer, R; Charney, D S; Baldwin, R

2000-01-01

Quantitation of the PET benzodiazepine receptor antagonist, [(11)C]Iomazenil, using low specific activity radioligand was recently described. The purpose of this study was to quantitate benzodiazepine receptor binding in human subjects using PET and high specific activity [(11)C]Iomazenil. Six healthy human subjects underwent PET imaging following a bolus injection of high specific activity (>100 Ci/mmol) [(11)C]iomazenil. Arterial samples were collected at multiple time points after injection for measurement of unmetabolized total and nonprotein-bound parent compound in plasma. Time activity curves of radioligand concentration in brain and plasma were analyzed using two and three compartment model. Kinetic rate constants of transfer of radioligand between plasma, nonspecifically bound brain tissue, and specifically bound brain tissue compartments were fitted to the model. Values for fitted kinetic rate constants were used in the calculation of measures of benzodiazepine receptor binding, including binding potential (the ratio of receptor density to affinity), and product of BP and the fraction of free nonprotein-bound parent compound (V(3)'). Use of the three compartment model improved the goodness of fit in comparison to the two compartment model. Values for kinetic rate constants and measures of benzodiazepine receptor binding, including BP and V(3)', were similar to results obtained with the SPECT radioligand [(123)I]iomazenil, and a prior report with low specific activity [(11)C]Iomazenil. Kinetic modeling using the three compartment model with PET and high specific activity [(11)C]Iomazenil provides a reliable measure of benzodiazepine receptor binding. Synapse 35:68-77, 2000. Published 2000 Wiley-Liss, Inc.

A novel urotensin II receptor antagonist, KR-36996, improved cardiac function and attenuated cardiac hypertrophy in experimental heart failure.

PubMed

Oh, Kwang-Seok; Lee, Jeong Hyun; Yi, Kyu Yang; Lim, Chae Jo; Park, Byung Kil; Seo, Ho Won; Lee, Byung Ho

2017-03-15

Urotensin II and its receptor are thought to be involved in various cardiovascular diseases such as heart failure, pulmonary hypertension and atherosclerosis. Since the regulation of the urotensin II/urotensin II receptor offers a great potential for therapeutic strategies related to the treatment of cardiovascular diseases, the study of selective and potent antagonists for urotensin II receptor is more fascinating. This study was designed to determine the potential therapeutic effects of a newly developed novel urotensin II receptor antagonist, N-(1-(3-bromo-4-(piperidin-4-yloxy)benzyl)piperidin-4-yl)benzo[b]thiophene-3-carboxamide (KR-36996), in experimental models of heart failure. KR-36996 displayed a high binding affinity (Ki=4.44±0.67nM) and selectivity for urotensin II receptor. In cell-based study, KR-36996 significantly inhibited urotensin II-induced stress fiber formation and cellular hypertrophy in H9c2 UT cells. In transverse aortic constriction-induced cardiac hypertrophy model in mice, the daily oral administration of KR-36996 (30mg/kg) for 14 days significantly decreased left ventricular weight by 40% (P<0.05). In myocardial infarction-induced chronic heart failure model in rats, repeated echocardiography and hemodynamic measurements demonstrated remarkable improvement of the cardiac performance by KR-36996 treatment (25 and 50mg/kg/day, p.o.) for 12 weeks. Moreover, KR-36996 decreased interstitial fibrosis and cardiomyocyte hypertrophy in the infarct border zone. These results suggest that potent and selective urotensin II receptor antagonist could efficiently attenuate both cardiac hypertrophy and dysfunction in experimental heart failure. KR-36996 may be useful as an effective urotensin II receptor antagonist for pharmaceutical or clinical applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Interaction of Plant Extracts with Central Nervous System Receptors

PubMed Central

Lundstrom, Kenneth; Pham, Huyen Thanh; Dinh, Long Doan

2017-01-01

Background: Plant extracts have been used in traditional medicine for the treatment of various maladies including neurological diseases. Several central nervous system receptors have been demonstrated to interact with plant extracts and components affecting the pharmacology and thereby potentially playing a role in human disease and treatment. For instance, extracts from Hypericum perforatum (St. John’s wort) targeted several CNS receptors. Similarly, extracts from Piper nigrum, Stephania cambodica, and Styphnolobium japonicum exerted inhibition of agonist-induced activity of the human neurokinin-1 receptor. Methods: Different methods have been established for receptor binding and functional assays based on radioactive and fluorescence-labeled ligands in cell lines and primary cell cultures. Behavioral studies of the effect of plant extracts have been conducted in rodents. Plant extracts have further been subjected to mood and cognition studies in humans. Results: Mechanisms of action at molecular and cellular levels have been elucidated for medicinal plants in support of standardization of herbal products and identification of active extract compounds. In several studies, plant extracts demonstrated affinity to a number of CNS receptors in parallel indicating the complexity of this interaction. In vivo studies showed modifications of CNS receptor affinity and behavioral responses in animal models after treatment with medicinal herbs. Certain plant extracts demonstrated neuroprotection and enhanced cognitive performance, respectively, when evaluated in humans. Noteworthy, the penetration of plant extracts and their protective effect on the blood-brain-barrier are discussed. Conclusion: The affinity of plant extracts and their isolated compounds for CNS receptors indicates an important role for medicinal plants in the treatment of neurological disorders. Moreover, studies in animal and human models have confirmed a scientific basis for the application of medicinal herbs. However, additional investigations related to plant extracts and their isolated compounds, as well as their application in animal models and the conducting of clinical trials, are required. PMID:28930228

New insights into the stereochemical requirements of the bradykinin B2 receptor antagonists binding

NASA Astrophysics Data System (ADS)

Lupala, Cecylia S.; Gomez-Gutierrez, Patricia; Perez, Juan J.

2016-01-01

Bradykinin (BK) is a member of the kinin family, released in response to inflammation, trauma, burns, shock, allergy and some cardiovascular diseases, provoking vasodilatation and increased vascular permeability among other effects. Their actions are mediated through at least two G-protein coupled receptors, B1 a receptor up-regulated during inflammation episodes or tissue trauma and B2 that is constitutively expressed in a variety of cell types. The goal of the present work is to carry out a structure-activity study of BK B2 antagonism, taking into account the stereochemical features of diverse non-peptide antagonists and the way these features translate into ligand anchoring points to complementary regions of the receptor, through the analysis of the respective ligand-receptor complex. For this purpose an atomistic model of the BK B2 receptor was built by homology modeling and subsequently refined embedded in a lipid bilayer by means of a 600 ns molecular dynamics trajectory. The average structure from the last hundred nanoseconds of the molecular dynamics trajectory was energy minimized and used as model of the receptor for docking studies. For this purpose, a set of compounds with antagonistic profile, covering maximal diversity were selected from the literature. Specifically, the set of compounds include Fasitibant, FR173657, Anatibant, WIN64338, Bradyzide, CHEMBL442294, and JSM10292. Molecules were docked into the BK B2 receptor model and the corresponding complexes analyzed to understand ligand-receptor interactions. The outcome of this study is summarized in a 3D pharmacophore that explains the observed structure-activity results and provides insight into the design of novel molecules with antagonistic profile. To prove the validity of the pharmacophore hypothesized a virtual screening process was also carried out. The pharmacophore was used as query to identify new hits using diverse databases of molecules. The results of this study revealed a set of new hits with structures not connected to the molecules used for pharmacophore development. A few of these structures were purchased and tested. The results of the binding studies show about a 33 % success rate with a correlation between the number of pharmacophore points fulfilled and their antagonistic potency. Some of these structures are disclosed in the present work.

Human urothelial cell lines as potential models for studying cannabinoid and excitatory receptor interactions in the urinary bladder.

PubMed

Bakali, Evangelia; Elliott, Ruth A; Taylor, Anthony H; Lambert, David G; Willets, Jonathon M; Tincello, Douglas G

2014-06-01

To characterize human urothelial cell lines' cannabinoid receptor expression and evaluate their possible use for studying signalling interactions with purinergic and muscarinic receptor activation. PCR was used to detect cannabinoid (CB), muscarinic and purinergic receptor transcripts in HCV29 and UROtsa cells, whilst immunofluorescence evaluated protein expression and localization of cannabinoid receptors. The effect of CB1 agonist (ACEA) on carbachol- and ATP-induced changes in intracellular calcium ([Ca(2+)]i) levels was measured using fluorimetry. The ability of ACEA to reduce intracellular cAMP was investigated in HCV29 cells. CB1 and GPR55 receptor transcripts were detected in HCV29 and UROtsa cells, respectively. Immunofluorescence showed positive staining for CB1 in the HCV29 cells. Both cell lines expressed transcript levels for muscarinic receptors, but carbachol did not raise [Ca(2+)]i levels indicating a lack or low expression of G(q)-coupled muscarinic receptors. Transcripts for purinergic receptors were detected; ATP significantly increased [Ca(2+)]i in HCV29 and UROtsa cells by 395 ± 61 and 705 ± 100 nM (mean ± SEM, n = 6), respectively. ACEA did not alter ATP-induced [Ca(2+)]i or cAMP levels in HCV29 cells. Whilst HCV29 cells expressed CB1 and UROtsa cells expressed GPR55 receptors, these were not functionally coupled to the existing purinergic-driven increase in Ca2+ as such they do not represent a good model to study signalling interactions.

A simulation study to quantify the impacts of exposure ...

EPA Pesticide Factsheets

A simulation study to quantify the impacts of exposure measurement error on air pollution health risk estimates in copollutant time-series models The National Exposure Research Laboratory (NERL) Computational Exposure Division (CED) develops and evaluates data, decision-support tools, and models to be applied to media-specific or receptor-specific problem areas. CED uses modeling-based approaches to characterize exposures, evaluate fate and transport, and support environmental diagnostics/forensics with input from multiple data sources. It also develops media- and receptor-specific models, process models, and decision support tools for use both within and outside of EPA.

Modeling the Effects of β1-Adrenergic Receptor Blockers and Polymorphisms on Cardiac Myocyte Ca2+ Handling

PubMed Central

Amanfu, Robert K.

2014-01-01

β-Adrenergic receptor blockers (β-blockers) are commonly used to treat heart failure, but the biologic mechanisms governing their efficacy are still poorly understood. The complexity of β-adrenergic signaling coupled with the influence of receptor polymorphisms makes it difficult to intuit the effect of β-blockers on cardiac physiology. While some studies indicate that β-blockers are efficacious by inhibiting β-adrenergic signaling, other studies suggest that they work by maintaining β-adrenergic responsiveness. Here, we use a systems pharmacology approach to test the hypothesis that in ventricular myocytes, these two apparently conflicting mechanisms for β-blocker efficacy can occur concurrently. We extended a computational model of the β1-adrenergic pathway and excitation-contraction coupling to include detailed receptor interactions for 19 ligands. Model predictions, validated with Ca2+ and Förster resonance energy transfer imaging of adult rat ventricular myocytes, surprisingly suggest that β-blockers can both inhibit and maintain signaling depending on the magnitude of receptor stimulation. The balance of inhibition and maintenance of β1-adrenergic signaling is predicted to depend on the specific β-blocker (with greater responsiveness for metoprolol than carvedilol) and β1-adrenergic receptor Arg389Gly polymorphisms. PMID:24867460

Molecular recognition of a model globular protein apomyoglobin by synthetic receptor cyclodextrin: effect of fluorescence modification of the protein and cavity size of the receptor in the interaction.

PubMed

Saha, Ranajay; Rakshit, Surajit; Pal, Samir Kumar

2013-11-01

Labelling of proteins with some extrinsic probe is unavoidable in molecular biology research. Particularly, spectroscopic studies in the optical region require fluorescence modification of native proteins by attaching polycyclic aromatic fluoroprobe with the proteins under investigation. Our present study aims to address the consequence of the attachment of a fluoroprobe at the protein surface in the molecular recognition of the protein by selectively small model receptor. A spectroscopic study involving apomyoglobin (Apo-Mb) and cyclodextrin (CyD) of various cavity sizes as model globular protein and synthetic receptors, respectively, using steady-state and picosecond-resolved techniques, is detailed here. A study involving Förster resonance energy transfer, between intrinsic amino acid tryptophan (donor) and N, N-dimethyl naphthalene moiety of the extrinsic dansyl probes at the surface of Apo-Mb, precisely monitor changes in donor acceptor distance as a consequence of interaction of the protein with CyD having different cavity sizes (β and γ variety). Molecular modelling studies on the interaction of tryptophan and dansyl probe with β-CyD is reported here and found to be consistent with the experimental observations. In order to investigate structural aspects of the interacting protein, we have used circular dichroism spectroscopy. Temperature-dependent circular dichroism studies explore the change in the secondary structure of Apo-Mb in association with CyD, before and after fluorescence modification of the protein. Overall, the study well exemplifies approaches to protein recognition by CyD as a synthetic receptor and offers a cautionary note on the use of hydrophobic fluorescent labels for proteins in biochemical studies involving recognition of molecules. Copyright © 2013 John Wiley & Sons, Ltd.

The high-affinity receptor for IgG, FcγRI, of humans and non-human primates.

PubMed

Chenoweth, Alicia M; Trist, Halina M; Tan, Peck-Szee; Wines, Bruce D; Hogarth, P Mark

2015-11-01

Non-human primate (NHP) models, especially involving macaques, are considered important models of human immunity and have been essential in preclinical testing for vaccines and therapeutics. Despite this, much less characterization of macaque Fc receptors has occurred compared to humans or mice. Much of the characterization of macaque Fc receptors so far has focused on the low-affinity Fc receptors, particularly FcγRIIIa. From these studies, it is clear that there are distinct differences between the human and macaque low-affinity receptors and their interaction with human IgG. Relatively little work has been performed on the high-affinity IgG receptor, FcγRI, especially in NHPs. This review will focus on what is currently known of how FcγRI interacts with IgG, from mutation studies and recent crystallographic studies of human FcγRI, and how amino acid sequence differences in the macaque FcγRI may affect this interaction. Additionally, this review will look at the functional consequences of differences in the amino acid sequences between humans and macaques. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A Time-course Study with the Androgen Receptor Antagonist Flutamide in Fish

EPA Science Inventory

Flutamide, a drug registered to treat some types of prostate cancer in humans, has been used for many years as a model androgen receptor (AR) antagonist in studies aimed at characterizing disruption of the vertebrate hypothalamic-pituitary-gonadal (HPG) axis. Various studies hav...

Synthesis, in vitro and in vivo studies, and molecular modeling of N-alkylated dextromethorphan derivatives as non-competitive inhibitors of α3β4 nicotinic acetylcholine receptor.

PubMed

Jozwiak, Krzysztof; Targowska-Duda, Katarzyna M; Kaczor, Agnieszka A; Kozak, Joanna; Ligeza, Agnieszka; Szacon, Elzbieta; Wrobel, Tomasz M; Budzynska, Barbara; Biala, Grazyna; Fornal, Emilia; Poso, Antti; Wainer, Irving W; Matosiuk, Dariusz

2014-12-15

9 N-alkylated derivatives of dextromethorphan are synthesized and studied as non-competitive inhibitors of α3β4 nicotinic acetylcholine receptors (nAChRs). In vitro activity towards α3β4 nicotinic acetylcholine receptor is determined using a patch-clamp technique and is in the micromolar range. Homology modeling, molecular docking and molecular dynamics of ligand-receptor complexes in POPC membrane are used to find the mode of interactions of N-alkylated dextromethorphan derivatives with α3β4 nAChR. The compounds, similarly as dextromethorphan, interact with the middle portion of α3β4 nAChR ion channel. Finally, behavioral tests confirmed potential application of the studied compounds for the treatment of addiction. Copyright © 2014 Elsevier Ltd. All rights reserved.

A 3D QSAR CoMFA study of non-peptide angiotensin II receptor antagonists

NASA Astrophysics Data System (ADS)

Belvisi, Laura; Bravi, Gianpaolo; Catalano, Giovanna; Mabilia, Massimo; Salimbeni, Aldo; Scolastico, Carlo

1996-12-01

A series of non-peptide angiotensin II receptor antagonists was investigated with the aim of developing a 3D QSAR model using comparative molecular field analysis descriptors and approaches. The main goals of the study were dictated by an interest in methodologies and an understanding of the binding requirements to the AT1 receptor. Consistency with the previously derived activity models was always checked to contemporarily test the validity of the various hypotheses. The specific conformations chosen for the study, the procedures invoked to superimpose all structures, the conditions employed to generate steric and electrostatic field values and the various PCA/PLS runs are discussed in detail. The effect of experimental design techniques to select objects (molecules) and variables (descriptors) with respect to the predictive power of the QSAR models derived was especially analysed.

Formation of the long range Dpp morphogen gradient.

PubMed

Schwank, Gerald; Dalessi, Sascha; Yang, Schu-Fee; Yagi, Ryohei; de Lachapelle, Aitana Morton; Affolter, Markus; Bergmann, Sven; Basler, Konrad

2011-07-01

The TGF-β homolog Decapentaplegic (Dpp) acts as a secreted morphogen in the Drosophila wing disc, and spreads through the target tissue in order to form a long range concentration gradient. Despite extensive studies, the mechanism by which the Dpp gradient is formed remains controversial. Two opposing mechanisms have been proposed: receptor-mediated transcytosis (RMT) and restricted extracellular diffusion (RED). In these scenarios the receptor for Dpp plays different roles. In the RMT model it is essential for endocytosis, re-secretion, and thus transport of Dpp, whereas in the RED model it merely modulates Dpp distribution by binding it at the cell surface for internalization and subsequent degradation. Here we analyzed the effect of receptor mutant clones on the Dpp profile in quantitative mathematical models representing transport by either RMT or RED. We then, using novel genetic tools, experimentally monitored the actual Dpp gradient in wing discs containing receptor gain-of-function and loss-of-function clones. Gain-of-function clones reveal that Dpp binds in vivo strongly to the type I receptor Thick veins, but not to the type II receptor Punt. Importantly, results with the loss-of-function clones then refute the RMT model for Dpp gradient formation, while supporting the RED model in which the majority of Dpp is not bound to Thick veins. Together our results show that receptor-mediated transcytosis cannot account for Dpp gradient formation, and support restricted extracellular diffusion as the main mechanism for Dpp dispersal. The properties of this mechanism, in which only a minority of Dpp is receptor-bound, may facilitate long-range distribution.

Role of the Prostaglandin E2 EP1 Receptor in Traumatic Brain Injury

PubMed Central

Glushakov, Alexander V.; Fazal, Jawad A.; Narumiya, Shuh; Doré, Sylvain

2014-01-01

Brain injuries promote upregulation of so-called proinflammatory prostaglandins, notably prostaglandin E2 (PGE2), leading to overactivation of a class of its cognate G-protein-coupled receptors, including EP1, which is considered a promising target for treatment of ischemic stroke. However, the role of the EP1 receptor is complex and depends on the type of brain injury. This study is focused on the investigation of the role of the EP1 receptor in a controlled cortical impact (CCI) model, a preclinical model of traumatic brain injury (TBI). The therapeutic effects of post-treatments with a widely studied EP1 receptor antagonist, SC-51089, were examined in wildtype and EP1 receptor knockout C57BL/6 mice. Neurological deficit scores (NDS) were assessed 24 and 48 h following CCI or sham surgery, and brain immunohistochemical pathology was assessed 48 h after surgery. In wildtype mice, CCI resulted in an obvious cortical lesion and localized hippocampal edema with an associated significant increase in NDS compared to sham-operated animals. Post-treatments with the selective EP1 receptor antagonist SC-51089 or genetic knockout of EP1 receptor had no significant effects on cortical lesions and hippocampal swelling or on the NDS 24 and 48 h after CCI. Immunohistochemistry studies revealed CCI-induced gliosis and microglial activation in selected ipsilateral brain regions that were not affected by SC-51089 or in the EP1 receptor-deleted mice. This study provides further clarification on the respective contribution of the EP1 receptor in TBI and suggests that, under this experimental paradigm, the EP1 receptor would have limited effects in modulating acute neurological and anatomical pathologies following contusive brain trauma. Findings from this protocol, in combination with previous studies demonstrating differential roles of EP1 receptor in ischemic, neurotoxic, and hemorrhagic conditions, provide scientific background and further clarification of potential therapeutic application of prospective prostaglandin G-protein-coupled receptor drugs in the clinic for treatment of TBI and other acute brain injuries. PMID:25426930

Prediction of breast cancer risk by genetic risk factors, overall and by hormone receptor status.

PubMed

Hüsing, Anika; Canzian, Federico; Beckmann, Lars; Garcia-Closas, Montserrat; Diver, W Ryan; Thun, Michael J; Berg, Christine D; Hoover, Robert N; Ziegler, Regina G; Figueroa, Jonine D; Isaacs, Claudine; Olsen, Anja; Viallon, Vivian; Boeing, Heiner; Masala, Giovanna; Trichopoulos, Dimitrios; Peeters, Petra H M; Lund, Eiliv; Ardanaz, Eva; Khaw, Kay-Tee; Lenner, Per; Kolonel, Laurence N; Stram, Daniel O; Le Marchand, Loïc; McCarty, Catherine A; Buring, Julie E; Lee, I-Min; Zhang, Shumin; Lindström, Sara; Hankinson, Susan E; Riboli, Elio; Hunter, David J; Henderson, Brian E; Chanock, Stephen J; Haiman, Christopher A; Kraft, Peter; Kaaks, Rudolf

2012-09-01

There is increasing interest in adding common genetic variants identified through genome wide association studies (GWAS) to breast cancer risk prediction models. First results from such models showed modest benefits in terms of risk discrimination. Heterogeneity of breast cancer as defined by hormone-receptor status has not been considered in this context. In this study we investigated the predictive capacity of 32 GWAS-detected common variants for breast cancer risk, alone and in combination with classical risk factors, and for tumours with different hormone receptor status. Within the Breast and Prostate Cancer Cohort Consortium, we analysed 6009 invasive breast cancer cases and 7827 matched controls of European ancestry, with data on classical breast cancer risk factors and 32 common gene variants identified through GWAS. Discriminatory ability with respect to breast cancer of specific hormone receptor-status was assessed with the age adjusted and cohort-adjusted concordance statistic (AUROC(a)). Absolute risk scores were calculated with external reference data. Integrated discrimination improvement was used to measure improvements in risk prediction. We found a small but steady increase in discriminatory ability with increasing numbers of genetic variants included in the model (difference in AUROC(a) going from 2.7% to 4%). Discriminatory ability for all models varied strongly by hormone receptor status. Adding information on common polymorphisms provides small but statistically significant improvements in the quality of breast cancer risk prediction models. We consistently observed better performance for receptor-positive cases, but the gain in discriminatory quality is not sufficient for clinical application.

Long-Term Disease-Free Survival of Non-Metastatic Breast Cancer Patients in Iran: A Survival Model with Competing Risks Taking Cure Fraction and Frailty into Account

PubMed

Ghavami, Vahid; Mahmoudi, Mahmood; Rahimi Foroushani, Abbas; Baghishani, Hossein; Homaei Shandiz, Fatemeh; Yaseri, Mehdi

2017-10-26

Introduction: Survival modeling is a very important tool to detect risk factors and provide a basis for health care planning. However, cancer data may have properties leading to distorted results with routine methods. Therefore, this study aimed to cover specific factors (competing risk, cure fraction and heterogeneity) with a real dataset of Iranian breast cancer patients using a competing risk-cure-frailty model. Materials and methods: For this historical cohort study, information for 550 Iranian breast cancer patients who underwent surgery for tumor removal from 2001 to 2007 and were followed up to March 2017, was analyzed using R 3.2 software. Results: In contrast to T-stage and N-stage, hormone receptor status did not have any significant effect on the cure fraction (long-term disease-free survival). However, T-stage, N-stage and hormone receptor status all had a significant effect on short-term disease-free survival so that the hazard of loco-regional relapse or distant metastasis in cases positive for a hormone receptor was only 0.3 times that for their negative hormone receptor counterparts. The likelihood of locoregional relapse in the first quartile of follow up was nearly twice that of other quartiles. The least cumulative incidence of time to locoregional relapse was for cases with a positive hormone receptor, low N stage and low T stage. The effect of frailty term was significant in this study and a model with frailty appeared more appropriate than a model without, based on the Akaike information criterion (AIC); values for the frailty model and one without the frailty parameter were 1370.39 and 1381.46, respectively. Conclusions: The data from this study indicate ae necessity to consider competing risk, cure fraction and heterogeneity in survival modeling. The competing risk-cure-frailty model can cover complex situations with survival data. Creative Commons Attribution License

Buprenorphine-elicited alteration of adenylate cyclase activity in human embryonic kidney 293 cells coexpressing κ-, μ-opioid and nociceptin receptors

PubMed Central

Wang, Pei-Chen; Ho, Ing-Kang; Lee, Cynthia Wei-Sheng

2015-01-01

Buprenorphine, a maintenance drug for heroin addicts, exerts its pharmacological function via κ- (KOP), μ-opioid (MOP) and nociceptin/opioid receptor-like 1 (NOP) receptors. Previously, we investigated its effects in an in vitro model expressing human MOP and NOP receptors individually or simultaneously (MOP, NOP, and MOP+NOP) in human embryonic kidney 293 cells. Here, we expanded this cell model by expressing human KOP, MOP and NOP receptors individually or simultaneously (KOP, KOP+MOP, KOP+NOP and KOP+MOP+NOP). Radioligand binding with tritium-labelled diprenorphine confirmed the expression of KOP receptors. Immunoblotting and immunocytochemistry indicated that the expressed KOP, MOP and NOP receptors are N-linked glycoproteins and colocalized in cytoplasmic compartments. Acute application of the opioid receptor agonists— U-69593, DAMGO and nociceptin— inhibited adenylate cyclase (AC) activity in cells expressing KOP, MOP and NOP receptors respectively. Buprenorphine, when applied acutely, inhibited AC activity to ~90% in cells expressing KOP+MOP+NOP receptors. Chronic exposure to buprenorphine induced concentration-dependent AC superactivation in cells expressing KOP+NOP receptors, and the level of this superactivation was even higher in KOP+MOP+NOP-expressing cells. Our study demonstrated that MOP receptor could enhance AC regulation in the presence of coexpressed KOP and NOP receptors, and NOP receptor is essential for concentration-dependent AC superactivation elicited by chronic buprenorphine exposure. PMID:26153065

Identification of key residues for the binding of glucagon to the N-terminal domain of its receptor: an alanine scan and modeling study.

PubMed

Prévost, M; Vertongen, P; Waelbroeck, M

2012-10-01

Glucagon plays an essential role in the glycemia maintenance during fasting, but also aggravates hyperglycemia in diabetic patients. A series of analogues of glucagon were synthesized replacing each amino acid of the C-terminal region (residues 15-29) with alanine. The residues affecting the binding to the glucagon receptor are found to be located on one face of the glucagon helix. Several 3-dimensional models of the N-terminal domain of the glucagon receptor in complex with its ligand peptide were built and used to analyze the peptide-receptor interface in terms of the nature of the peptide residues and the interactions they form with the receptor. The models suggest that glucagon keeps its native helical structure upon binding, and that a large part of the interface formed with the receptor is hydrophobic. We find that in the C-terminal region, F22, V23, M27, and D15 are the most important residues for peptide binding. They bury a large portion of their solvent accessible surface area and make numerous interactions with the receptor mainly of the hydrophobic type. © Georg Thieme Verlag KG Stuttgart · New York.

G Protein-Coupled Estrogen Receptor (GPER) Agonist Dual Binding Mode Analyses toward Understanding of its Activation Mechanism: A Comparative Homology Modeling Approach.

PubMed

Arnatt, Christopher K; Zhang, Yan

2013-07-01

G protein-coupled estrogen receptor (GPER) has been shown to be important in several disease states such as estrogen sensitive cancers. While several selective ligands have been identified for the receptor, little is known about how they interact with GPER and how their structures influence their activity. Specifically, within one series of ligands, whose structure varied only at one position, the replacement of a hydrogen atom with an acetyl group changed a potent antagonist into a potent agonist. In this study, two GPER homology models were constructed based on the x-ray crystal structures of both the active and inactive β 2 -adrenergic receptors (β 2 AR) in an effort to characterize the differences of binding modes between agonists and antagonists to the receptor, and to understand their activity in relation to their structures. The knowledge attained in this study is expected to provide valuable information on GPER ligands structure activity relationship to benefit future rational design of potent agonists and antagonists of the receptor for potential therapeutic applications.

G Protein-Coupled Estrogen Receptor (GPER) Agonist Dual Binding Mode Analyses toward Understanding of its Activation Mechanism: A Comparative Homology Modeling Approach

PubMed Central

Arnatt, Christopher K.; Zhang, Yan

2015-01-01

G protein-coupled estrogen receptor (GPER) has been shown to be important in several disease states such as estrogen sensitive cancers. While several selective ligands have been identified for the receptor, little is known about how they interact with GPER and how their structures influence their activity. Specifically, within one series of ligands, whose structure varied only at one position, the replacement of a hydrogen atom with an acetyl group changed a potent antagonist into a potent agonist. In this study, two GPER homology models were constructed based on the x-ray crystal structures of both the active and inactive β2-adrenergic receptors (β2AR) in an effort to characterize the differences of binding modes between agonists and antagonists to the receptor, and to understand their activity in relation to their structures. The knowledge attained in this study is expected to provide valuable information on GPER ligands structure activity relationship to benefit future rational design of potent agonists and antagonists of the receptor for potential therapeutic applications. PMID:26229572

Dynamic conformational switching in the chemokine ligand is essential for G-protein-coupled receptor activation

PubMed Central

Joseph, Prem Raj B.; Sawant, Kirti V.; Isley, Angela; Pedroza, Mesias; Garofalo, Roberto P.; Richardson, Ricardo M.; Rajarathnam, Krishna

2014-01-01

Chemokines mediate diverse functions from organogenesis to mobilizing leucocytes, and are unusual agonists for class-A GPCRs (G-protein-coupled receptors) because of their large size and multi-domain structure. The current model for receptor activation, which involves interactions between chemokine N-loop and receptor N-terminal residues (Site-I) and between chemokine N-terminal and receptor extracellular loop/transmembrane residues (Site-II), fails to describe differences in ligand/receptor selectivity and the activation of multiple signalling pathways. In the present study, we show in neutrophil-activating chemokine CXCL8 that the highly conserved GP (glycine-proline) motif located distal to both N-terminal and N-loop residues couples Site-I and Site-II interactions. Mutations in the GP motif caused various differences from native-like function to complete loss of activity that could not be correlated with the specific mutation, receptor affinity or subtype, or a specific signalling pathway. NMR studies indicated that the GP motif does not influence Site-I interactions, but molecular dynamics simulations suggested that this motif dictates substates of the CXCL8 conformational ensemble. We conclude that the GP motif enables diverse receptor functions by controlling cross-talk between Site-I and Site-II, and further propose that the repertoire of chemokine functions is best described by a conformational ensemble model in which a network of long-range coupled indirect interactions mediate receptor activity. PMID:24032673

Ligand-binding pocket shape differences between S1P1 and S1P3 determine efficiency of chemical probe identification by uHTS

PubMed Central

Schürer, Stephan C.; Brown, Steven J.; Cabrera, Pedro Gonzales; Schaeffer, Marie-Therese; Chapman, Jacqueline; Jo, Euijung; Chase, Peter; Spicer, Tim; Hodder, Peter; Rosen, Hugh

2008-01-01

We have studied the Sphingosine 1-phosphate (S1P) receptor system to better understand why certain molecular targets within a closely related family are much more tractable when identifying compelling chemical leads. Five medically important G protein-coupled receptors for S1P regulate heart rate, coronary artery caliber, endothelial barrier integrity, and lymphocyte trafficking. Selective S1P receptor agonist probes would be of great utility to study receptor subtype-specific function. Through systematic screening of the same libraries, we identified novel selective agonists chemotypes for each of the S1P1 and S1P3 receptors. uHTS for S1P1 was more effective than for S1P3, with many selective, low nanomolar hits of proven mechanism emerging for. Receptor structure modeling and ligand docking reveal differences between the receptor binding pockets, which are the basis for sub-type selectivity. Novel selective agonists interact primarily in the hydrophobic pocket of the receptor in the absence of head-group interactions. Chemistry-space and shape-based analysis of the screening libraries in combination with the binding models explain the observed differential hit rates and enhanced efficiency for lead discovery for S1P1 vs. S1P3 in this closely related receptor family. PMID:18590333

Agonist Binding to Chemosensory Receptors: A Systematic Bioinformatics Analysis

PubMed Central

Fierro, Fabrizio; Suku, Eda; Alfonso-Prieto, Mercedes; Giorgetti, Alejandro; Cichon, Sven; Carloni, Paolo

2017-01-01

Human G-protein coupled receptors (hGPCRs) constitute a large and highly pharmaceutically relevant membrane receptor superfamily. About half of the hGPCRs' family members are chemosensory receptors, involved in bitter taste and olfaction, along with a variety of other physiological processes. Hence these receptors constitute promising targets for pharmaceutical intervention. Molecular modeling has been so far the most important tool to get insights on agonist binding and receptor activation. Here we investigate both aspects by bioinformatics-based predictions across all bitter taste and odorant receptors for which site-directed mutagenesis data are available. First, we observe that state-of-the-art homology modeling combined with previously used docking procedures turned out to reproduce only a limited fraction of ligand/receptor interactions inferred by experiments. This is most probably caused by the low sequence identity with available structural templates, which limits the accuracy of the protein model and in particular of the side-chains' orientations. Methods which transcend the limited sampling of the conformational space of docking may improve the predictions. As an example corroborating this, we review here multi-scale simulations from our lab and show that, for the three complexes studied so far, they significantly enhance the predictive power of the computational approach. Second, our bioinformatics analysis provides support to previous claims that several residues, including those at positions 1.50, 2.50, and 7.52, are involved in receptor activation. PMID:28932739

Computer-aided structure-affinity relationships in a set of piperazine and 3,8-diazabicyclo[3.2.1]octane derivatives binding to the μ-opioid receptor

NASA Astrophysics Data System (ADS)

Barlocco, Daniela; Cignarella, Giorgio; Greco, Giovanni; Novellino, Ettore

1993-10-01

Molecular modeling studies were carried out on a set of piperazine and 3,8-diazabicyclo[3.2.1]octane derivatives with the aim to highlight the main factors modulating their affinity for the μ-opioid receptor. Structure-affinity relationships were developed with the aid of molecular mechanics and semiempirical quantum-mechanics methods. According to our proposed pharmacodynamic model, the binding to the μ-receptor is promoted by the following physico-chemical features: the presence of hydrocarbon fragments on the nitrogen ring frame capable of interacting with one of two hypothesized hydrophobic receptor pockets; a `correct' orientation of an N-propionyl side chain so as to avoid a sterically hindered region of the receptor; the possibility of accepting a hydrogen bond from a receptor site complementary to the morphine phenol oxygen.

The modulatory role of spinally located histamine receptors in the regulation of the blood glucose level in d-glucose-fed mice.

PubMed

Sim, Yun-Beom; Park, Soo-Hyun; Kim, Sung-Su; Kim, Chea-Ha; Kim, Su-Jin; Lim, Su-Min; Jung, Jun-Sub; Ryu, Ohk-Hyun; Choi, Moon-Gi; Suh, Hong-Won

2014-02-01

The possible roles of spinal histamine receptors in the regulation of the blood glucose level were studied in ICR mice. Mice were intrathecally (i.t.) treated with histamine 1 (H1) receptor agonist (2-pyridylethylamine) or antagonist (cetirizine), histamine 2 (H2) receptor agonist (dimaprit) or antagonist (ranitidine), histamine 3 (H3) receptor agonist (α-methylhistamine) or antagonist (carcinine) and histamine 4 (H4) receptor agonist (VUF 8430) or antagonist (JNJ 7777120), and the blood glucose level was measured at 30, 60 and 120 min after i.t. administration. The i.t. injection with α-methylhistamine, but not carcinine slightly caused an elevation of the blood glucose level. In addition, histamine H1, H2, and H4 receptor agonists and antagonists did not affect the blood glucose level. In D-glucose-fed model, i.t. pretreatment with cetirizine enhanced the blood glucose level, whereas 2-pyridylethylamine did not affect. The i.t. pretreatment with dimaprit, but not ranitidine, enhanced the blood glucose level in D-glucose-fed model. In addition, α-methylhistamine, but not carcinine, slightly but significantly enhanced the blood glucose level D-glucose-fed model. Finally, i.t. pretreatment with JNJ 7777120, but not VUF 8430, slightly but significantly increased the blood glucose level. Although histamine receptors themselves located at the spinal cord do not exert any effect on the regulation of the blood glucose level, our results suggest that the activation of spinal histamine H2 receptors and the blockade of spinal histamine H1 or H3 receptors may play modulatory roles for up-regulation and down-regulation, respectively, of the blood glucose level in D-glucose fed model.

Ozanimod (RPC1063) is a potent sphingosine-1-phosphate receptor-1 (S1P1 ) and receptor-5 (S1P5 ) agonist with autoimmune disease-modifying activity.

PubMed

Scott, F L; Clemons, B; Brooks, J; Brahmachary, E; Powell, R; Dedman, H; Desale, H G; Timony, G A; Martinborough, E; Rosen, H; Roberts, E; Boehm, M F; Peach, R J

2016-06-01

Sphingosine1-phosphate (S1P) receptors mediate multiple events including lymphocyte trafficking, cardiac function, and endothelial barrier integrity. Stimulation of S1P1 receptors sequesters lymphocyte subsets in peripheral lymphoid organs, preventing their trafficking to inflamed tissue sites, modulating immunity. Targeting S1P receptors for treating autoimmune disease has been established in clinical studies with the non-selective S1P modulator, FTY720 (fingolimod, Gilenya™). The purpose of this study was to assess RPC1063 for its therapeutic utility in autoimmune diseases. The specificity and potency of RPC1063 (ozanimod) was evaluated for all five S1P receptors, and its effect on cell surface S1P1 receptor expression, was characterized in vitro. The oral pharmacokinetic (PK) parameters and pharmacodynamic effects were established in rodents, and its activity in three models of autoimmune disease (experimental autoimmune encephalitis, 2,4,6-trinitrobenzenesulfonic acid colitis and CD4(+) CD45RB(hi) T cell adoptive transfer colitis) was assessed. RPC1063 was specific for S1P1 and S1P5 receptors, induced S1P1 receptor internalization and induced a reversible reduction in circulating B and CCR7(+) T lymphocytes in vivo. RPC1063 showed high oral bioavailability and volume of distribution, and a circulatory half-life that supports once daily dosing. Oral RPC1063 reduced inflammation and disease parameters in all three autoimmune disease models. S1P receptor selectivity, favourable PK properties and efficacy in three distinct disease models supports the clinical development of RPC1063 for the treatment of relapsing multiple sclerosis and inflammatory bowel disease, differentiates RPC1063 from other S1P receptor agonists, and could result in improved safety outcomes in the clinic. © 2016 The British Pharmacological Society.

Impaired receptor-mediated catabolism of low density lipoprotein in the WHHL rabbit, an animal model of familial hypercholesterolemia

PubMed Central

Bilheimer, David W.; Watanabe, Yoshio; Kita, Toru

1982-01-01

The homozygous WHHL (Watanabe heritable hyperlipidemic) rabbit displays either no or only minimal low density lipoprotein (LDL) receptor activity on cultured fibroblasts and liver membranes and has therefore been proposed as an animal model for human familial hypercholesterolemia. To assess the impact of this mutation on LDL metabolism in vivo, we performed lipoprotein turnover studies in normal and WHHL rabbits using both native rabbit LDL and chemically modified LDL (i.e., methyl-LDL) that does not bind to LDL receptors. The total fractional catabolic rate (FCR) for LDL in the normal rabbit was 3.5-fold greater than in the WHHL rabbit. Sixty-seven percent of the total FCR for LDL in the normal rabbit was due to LDL receptor-mediated clearance and 33% was attributable to receptor-independent processes; in the WHHL rabbit, essentially all of the LDL was catabolized via receptor-independent processes. Despite a 17.5-fold elevated plasma pool size of LDL apoprotein (apo-LDL) in WHHL as compared to normal rabbits, the receptor-independent FCR—as judged by the turnover of methyl-LDL—was similar in the two strains. Thus, the receptor-independent catabolic processes are not influenced by the mutation affecting the LDL receptor. The WHHL rabbits also exhibited a 5.6-fold increase in the absolute rate of apo-LDL synthesis and catabolism. In absolute terms, the WHHL rabbit cleared 19-fold more apo-LDL via receptor-independent processes than did the normal rabbit and cleared virtually none by the receptor-dependent pathway. These results indicate that the homozygous WHHL rabbit shares a number of metabolic features in common with human familial hypercholesterolemia and should serve as a useful model for the study of altered lipoprotein metabolism associated with receptor abnormalities. We also noted that the in vivo metabolic behavior of human and rabbit LDL in the normal rabbit differed such that the mean total FCR for human LDL was only 64% of the mean total FCR for rabbit LDL, whereas human and rabbit methyl-LDL were cleared at identical rates. Thus, if human LDL and methyl-LDL had been used in these studies, the magnitude of both the total and receptor-dependent FCR would have been underestimated. PMID:6285345

Mechanism-based pharmacokinetic-pharmacodynamic modeling of the antinociceptive effect of buprenorphine in healthy volunteers.

PubMed

Yassen, Ashraf; Olofsen, Erik; Romberg, Raymonda; Sarton, Elise; Danhof, Meindert; Dahan, Albert

2006-06-01

The objective of this investigation was to characterize the pharmacokinetic-pharmacodynamic relation of buprenorphine's antinociceptive effect in healthy volunteers. Data on the time course of the antinociceptive effect after intravenous administration of 0.05-0.6 mg/70 kg buprenorphine in healthy volunteers was analyzed in conjunction with plasma concentrations by nonlinear mixed-effects analysis. A three-compartment pharmacokinetic model best described the concentration time course. Four structurally different pharmacokinetic-pharmacodynamic models were evaluated for their appropriateness to describe the time course of buprenorphine's antinociceptive effect: (1) E(max) model with an effect compartment model, (2) "power" model with an effect compartment model, (3) receptor association-dissociation model with a linear transduction function, and (4) combined biophase equilibration/receptor association-dissociation model with a linear transduction function. The latter pharmacokinetic-pharmacodynamic model described the time course of effect best and was used to explain time dependencies in buprenorphine's pharmacodynamics. The model converged, yielding precise estimation of the parameters characterizing hysteresis and the relation between relative receptor occupancy and antinociceptive effect. The rate constant describing biophase equilibration (k(eo)) was 0.00447 min(-1) (95% confidence interval, 0.00299-0.00595 min(-1)). The receptor dissociation rate constant (k(off)) was 0.0785 min(-1) (95% confidence interval, 0.0352-0.122 min(-1)), and k(on) was 0.0631 ml . ng(-1) . min(-1) (95% confidence interval, 0.0390-0.0872 ml . ng(-1) . min(-1)). This is consistent with observations in rats, suggesting that the rate-limiting step in the onset and offset of the antinociceptive effect is biophase distribution rather than slow receptor association-dissociation. In the dose range studied, no saturation of receptor occupancy occurred explaining the lack of a ceiling effect for antinociception.

Metformin sensitizes triple-negative breast cancer to proapoptotic TRAIL receptor agonists by suppressing XIAP expression.

PubMed

Strekalova, Elena; Malin, Dmitry; Rajanala, Harisha; Cryns, Vincent L

2017-06-01

Despite robust antitumor activity in diverse preclinical models, TNF-related apoptosis-inducing ligand (TRAIL) receptor agonists have not demonstrated efficacy in clinical trials, underscoring the need to identify agents that enhance their activity. We postulated that the metabolic stress induced by the diabetes drug metformin would sensitize breast cancer cells to TRAIL receptor agonists. Human triple (estrogen receptor, progesterone receptor, and HER2)-negative breast cancer (TNBC) cell lines were treated with TRAIL receptor agonists (monoclonal antibodies or TRAIL peptide), metformin, or the combination. The effects on cell survival, caspase activation, and expression of TRAIL receptors and the antiapoptotic protein XIAP were determined. In addition, XIAP was silenced by RNAi in TNBC cells and the effects on sensitivity to TRAIL were determined. The antitumor effects of metformin, TRAIL, or the combination were evaluated in an orthotopic model of metastatic TNBC. Metformin sensitized diverse TNBC cells to TRAIL receptor agonists. Metformin selectively enhanced the sensitivity of transformed breast epithelial cells to TRAIL receptor agonist-induced caspase activation and apoptosis with little effect on untransformed breast epithelial cells. These effects of metformin were accompanied by robust reductions in the protein levels of XIAP, a negative regulator of TRAIL-induced apoptosis. Silencing XIAP in TNBC cells mimicked the TRAIL-sensitizing effects of metformin. Metformin also enhanced the antitumor effects of TRAIL in a metastatic murine TNBC model. Our findings indicate that metformin enhances the activity of TRAIL receptor agonists, thereby supporting the rationale for additional translational studies combining these agents.

Enhanced effects of amphetamine but reduced effects of the hallucinogen, 5-MeO-DMT, on locomotor activity in 5-HT(1A) receptor knockout mice: implications for schizophrenia.

PubMed

van den Buuse, Maarten; Ruimschotel, Emma; Martin, Sally; Risbrough, Victoria B; Halberstadt, Adam L

2011-01-01

Serotonin-1A (5-HT(1A)) receptors may play a role in schizophrenia and the effects of certain antipsychotic drugs. However, the mechanism of interaction of 5-HT(1A) receptors with brain systems involved in schizophrenia, remains unclear. Here we show that 5-HT(1A) receptor knockout mice display enhanced locomotor hyperactivity to acute treatment with amphetamine, a widely used animal model of hyperdopaminergic mechanisms in psychosis. In contrast, the effect of MK-801 on locomotor activity, modeling NMDA receptor hypoactivity, was unchanged in the knockouts. The effect of the hallucinogen 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) was markedly reduced in 5-HT(1A) receptor knockout mice. There were no changes in apomorphine-induced disruption of PPI, a model of sensory gating deficits seen in schizophrenia. Similarly, there were no major changes in density of dopamine transporters (DAT) or dopamine D(1) or D(2) receptors which could explain the behavioural changes observed in 5-HT(1A) receptor knockout mice. These results extend our insight into the possible role of these receptors in aspects of schizophrenia. As also suggested by previous studies using agonist and antagonist drugs, 5-HT(1A) receptors may play an important role in hallucinations and to modulate dopaminergic activity in the brain. Copyright © 2011 Elsevier Ltd. All rights reserved.

Inter-comparison of receptor models for PM source apportionment: Case study in an industrial area

NASA Astrophysics Data System (ADS)

Viana, M.; Pandolfi, M.; Minguillón, M. C.; Querol, X.; Alastuey, A.; Monfort, E.; Celades, I.

2008-05-01

Receptor modelling techniques are used to identify and quantify the contributions from emission sources to the levels and major and trace components of ambient particulate matter (PM). A wide variety of receptor models are currently available, and consequently the comparability between models should be evaluated if source apportionment data are to be used as input in health effects studies or mitigation plans. Three of the most widespread receptor models (principal component analysis, PCA; positive matrix factorization, PMF; chemical mass balance, CMB) were applied to a single PM10 data set (n=328 samples, 2002-2005) obtained from an industrial area in NE Spain, dedicated to ceramic production. Sensitivity and temporal trend analyses (using the Mann-Kendall test) were applied. Results evidenced the good overall performance of the three models (r2>0.83 and α>0.91×between modelled and measured PM10 mass), with a good agreement regarding source identification and high correlations between input (CMB) and output (PCA, PMF) source profiles. Larger differences were obtained regarding the quantification of source contributions (up to a factor of 4 in some cases). The combined application of different types of receptor models would solve the limitations of each of the models, by constructing a more robust solution based on their strengths. The authors suggest the combined use of factor analysis techniques (PCA, PMF) to identify and interpret emission sources, and to obtain a first quantification of their contributions to the PM mass, and the subsequent application of CMB. Further research is needed to ensure that source apportionment methods are robust enough for application to PM health effects assessments.

Co-targeting the HER and IGF/insulin receptor axis in breast cancer, with triple targeting with endocrine therapy for hormone-sensitive disease.

PubMed

Chakraborty, Ashok; Hatzis, Christos; DiGiovanna, Michael P

2017-05-01

Interactions between HER2, estrogen receptor (ER), and insulin-like growth factor I receptor (IGF1R) are implicated in resistance to monotherapies targeting these receptors. We have previously shown in pre-clinical studies synergistic anti-tumor effects for co-targeting each pairwise combination of HER2, IGF1R, and ER. Strikingly, synergy for HER2/IGF1R targeting occurred not only in a HER2+ model, but also in a HER2-normal model. The purpose of the current study was therefore to determine the generalizability of synergistic anti-tumor effects of co-targeting HER2/IGF1R, the anti-tumor activity of triple-targeting HER2/IGF1R/ER in hormone-dependent cell lines, and the effect of using the multi-targeting drugs neratinib (pan-HER) and BMS-754807 (dual IGF1R/insulin receptor). Proliferation and apoptosis assays were performed in a large panel of cell lines representing varying receptor expression levels. Mechanistic effects were studied using phospho-protein immunoblotting. Analyses of drug interaction effects were performed using linear mixed-effects regression models. Enhanced anti-proliferative effects of HER/IGF-insulin co-targeting were seen in most, though not all, cell lines, including HER2-normal lines. For ER+ lines, triple targeting with inclusion of anti-estrogen generally resulted in the greatest anti-tumor effects. Double or triple targeting generally resulted in marked increases in apoptosis in the sensitive lines. Mechanistic studies demonstrated that the synergy between drugs was correlated with maximal inhibition of Akt and ERK pathway signaling. Dual HER/IGF-insulin targeting, and triple targeting with inclusion of anti-estrogen drugs, shows striking anti-tumor activity across breast cancer types, and drugs with broader receptor specificity may be more effective than single receptor selective drugs, particularly for ER- cells.

Refinement of the conformation of a critical region of charge-charge interaction between cholecystokinin and its receptor.

PubMed

Ding, Xi-Qin; Pinon, Delia I; Furse, Kristina E; Lybrand, Terry P; Miller, Laurence J

2002-05-01

Insight into the molecular basis of cholecystokinin (CCK) binding to its receptor has come from receptor mutagenesis and photoaffinity labeling studies, with both contributing to the current hypothesis that the acidic Tyr-sulfate-27 residue within the peptide is situated adjacent to basic Arg(197) in the second loop of the receptor. Here, we refine our understanding of this region of interaction by examining a structure-activity series of these positions within both ligand and receptor and by performing three-dimensional molecular modeling of key pairs of modified ligand and receptor constructs. The important roles of Arg(197) and Tyr-sulfate-27 were supported by the marked negative impact on binding and biological response with their natural partner molecule when the receptor residue was replaced by acidic Asp or Glu and when the peptide residue was replaced by basic Arg, Lys, p-amino-Phe, p-guanidino-Phe, or p-methylamino-Phe. Complementary ligand-receptor charge-exchange experiments were unable to regain the lost function. This was supported by the molecular modeling, which demonstrated that the charge-reversed double mutants could not form a good interaction without extensive rearrangement of receptor conformation. The models further predicted that R197D and R197E mutations would lead to conformational changes in the extracellular domain, and this was experimentally supported by data showing that these mutations decreased peptide agonist and antagonist binding and increased nonpeptidyl antagonist binding. These receptor constructs also had increased susceptibility to trypsin degradation relative to the wild-type receptor. In contrast, the relatively conservative R197K mutation had modest negative impact on peptide agonist binding, again consistent with the modeling demonstration of loss of a series of stabilizing inter- and intramolecular bonds. The strong correlation between predicted and experimental results support the reported refinement in the three-dimensional structure of the CCK-occupied receptor.

Comparison of source apportionments of PM2.5 using three receptor models (CMB, PMF, and SMP) in Seoul, Korea

NASA Astrophysics Data System (ADS)

Heo, J.; Kim, J. Y.; Kim, S. W.

2017-12-01

We compared source apportionments of PM2.5 in Seoul, Korea by three receptor models, Chemical Mass Balance (CMB), Positive Matrix Factorization (PMF), and Solver for Mixture Problem (SMP). The CMB model can estimate source apportionment with suitable source profiles of emissions, but it is difficult to find location-specific source profiles. In contrary, the multivariate receptor model does not need source profiles, but fundamental natural physical constraints (FNPCs) required for aerosol source apportionment are different in PMF and SMP. Ninety-six PM2.5 daily samples collected at Korea Institute of Science and Technology (KIST) in Seoul, Korea from October 2012 to September 2013 were analyzed in this study. The average PM2.5 mass concentration over the study period was 41.5 ± 27.7 mg m-3 and secondary inorganic species and organic matter were the main chemical species occupying about 73.7% - 87.9% of the PM2.5 mass concentration in all seasons. Secondary sulfate (18.0% - 26.1%), secondary nitrate (12.1% - 28.5%), vehicle (2.9% - 32.9%), biomass burning (13.2% - 21.3%) were identified by all three receptor models as the major sources accounting for approximately 76.3%-82.7% of the total PM2.5 and contributions of main sources represented their seasonality. However, three receptor models showed significant differences, especially for vehicle emission due to their measured/estimated source profiles. In this presentation, more detailed comparisons among CMB, PMF and SMP models will be presented focusing on the source profiles and contributions.

5D-QSAR for spirocyclic sigma1 receptor ligands by Quasar receptor surface modeling.

PubMed

Oberdorf, Christoph; Schmidt, Thomas J; Wünsch, Bernhard

2010-07-01

Based on a contiguous and structurally as well as biologically diverse set of 87 sigma(1) ligands, a 5D-QSAR study was conducted in which a quasi-atomistic receptor surface modeling approach (program package Quasar) was applied. The superposition of the ligands was performed with the tool Pharmacophore Elucidation (MOE-package), which takes all conformations of the ligands into account. This procedure led to four pharmacophoric structural elements with aromatic, hydrophobic, cationic and H-bond acceptor properties. Using the aligned structures a 3D-model of the ligand binding site of the sigma(1) receptor was obtained, whose general features are in good agreement with previous assumptions on the receptor structure, but revealed some novel insights since it represents the receptor surface in more detail. Thus, e.g., our model indicates the presence of an H-bond acceptor moiety in the binding site as counterpart to the ligands' cationic ammonium center, rather than a negatively charged carboxylate group. The presented QSAR model is statistically valid and represents the biological data of all tested compounds, including a test set of 21 ligands not used in the modeling process, with very good to excellent accuracy [q(2) (training set, n=66; leave 1/3 out) = 0.84, p(2) (test set, n=21)=0.64]. Moreover, the binding affinities of 13 further spirocyclic sigma(1) ligands were predicted with reasonable accuracy (mean deviation in pK(i) approximately 0.8). Thus, in addition to novel insights into the requirements for binding of spirocyclic piperidines to the sigma(1) receptor, the presented model can be used successfully in the rational design of new sigma(1) ligands. Copyright (c) 2010 Elsevier Masson SAS. All rights reserved.

Translational PK-PD modelling of molecular target modulation for the AMPA receptor positive allosteric modulator Org 26576.

PubMed

Bursi, Roberta; Erdemli, Gul; Campbell, Robert; Hutmacher, Matthew M; Kerbusch, Thomas; Spanswick, David; Jeggo, Ross; Nations, Kari R; Dogterom, Peter; Schipper, Jacques; Shahid, Mohammed

2011-12-01

The α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor potentiator Org 26576 represents an interesting pharmacological tool to evaluate the utility of glutamatergic enhancement towards the treatment of psychiatric disorders. In this study, a rat-human translational pharmacokinetic-pharmacodynamic (PK-PD) model of AMPA receptor modulation was used to predict human target engagement and inform dose selection in efficacy clinical trials. Modelling and simulation was applied to rat plasma and cerebrospinal fluid (CSF) pharmacokinetic and pharmacodynamic measurements to identify a target concentration (EC(80)) for AMPA receptor modulation. Human plasma pharmacokinetics was determined from 33 healthy volunteers and eight major depressive disorder patients. From four out of these eight patients, CSF PK was also determined. Simulations of human CSF levels were performed for several doses of Org 26576. Org 26576 (0.1-10 mg/kg, i.v.) potentiated rat hippocampal AMPA receptor responses in an exposure-dependant manner. The rat plasma and CSF PK data were fitted by one-compartment model each. The rat CSF PK-PD model yielded an EC(80) value of 593 ng/ml (90% confidence interval 406.8, 1,264.1). The human plasma and CSF PK data were simultaneously well described by a two-compartment model. Simulations showed that in humans at 100 mg QD, CSF levels of Org 26576 would exceed the EC(80) target concentration for about 2 h and that 400 mg BID would engage AMPA receptors for 24 h. The modelling approach provided useful insight on the likely human dose-molecular target engagement relationship for Org 26576. Based on the current analysis, 100 and 400 mg BID would be suitable to provide 'phasic' and 'continuous' AMPA receptor engagement, respectively.

mGlu₅-GABAB interplay in animal models of positive, negative and cognitive symptoms of schizophrenia.

PubMed

Wierońska, Joanna M; Kłeczek, Natalia; Woźniak, Monika; Gruca, Piotr; Łasoń-Tyburkiewicz, Magdalena; Papp, Mariusz; Brański, Piotr; Burnat, Grzegorz; Pilc, Andrzej

2015-09-01

Diverse preclinical studies exploiting the modulation of the GABAergic and/or glutamatergic system in brain via metabotropic receptors suggest their potential therapeutic utility. GS39783 and CDPPB, positive allosteric modulators of GABAB and mGlu5 receptors, were previously shown to reverse behavioral phenotypes in animal models to mimic selected (predominantly positive) symptoms of schizophrenia. In the present study we investigated the activity of selected GABAB (GS39783 and CGP7930) and mGlu5 (CDPPB) positive allosteric modulators. We focused mainly on the aspects of their efficacy in the models of negative and cognitive symptoms of schizophrenia. We used modified swim test, social interactions (models of negative symptoms) and novel object recognition (model of cognitive disturbances). The activity of the compounds was also tested in haloperidol-induced catalepsy test. The mutual interaction between GABAB/mGlu5 ligands was investigated as well. In the second part of the study, DHPG-induced PI hydrolysis in the presence of GABAB receptor antagonist (SKF97541), and SKF97541-induced inhibition of cAMP formation in the presence of DHPG, was performed. Both mGlu5 and GABAB receptor modulators effectively reversed MK-801-induced deficits in behavioral models of schizophrenia. Moreover, the concomitant administration of sub-effective doses of CDPPB and GS39783 induced a clear antipsychotic-like effect in all the procedures used, except DOI-induced head twitches. The concomitant administration of group I mGlu and GABAB agonists did not displayed any synergistic effects in vitro. Summing up, an activation of both types of receptor may be a promising mechanism for the development of novel antipsychotic drugs, efficacious toward positive, negative and cognitive symptoms. Copyright © 2015 Elsevier Ltd. All rights reserved.

Theoretical and Computational Studies of Peptides and Receptors of the Insulin Family

PubMed Central

Vashisth, Harish

2015-01-01

Synergistic interactions among peptides and receptors of the insulin family are required for glucose homeostasis, normal cellular growth and development, proliferation, differentiation and other metabolic processes. The peptides of the insulin family are disulfide-linked single or dual-chain proteins, while receptors are ligand-activated transmembrane glycoproteins of the receptor tyrosine kinase (RTK) superfamily. Binding of ligands to the extracellular domains of receptors is known to initiate signaling via activation of intracellular kinase domains. While the structure of insulin has been known since 1969, recent decades have seen remarkable progress on the structural biology of apo and liganded receptor fragments. Here, we review how this useful structural information (on ligands and receptors) has enabled large-scale atomically-resolved simulations to elucidate the conformational dynamics of these biomolecules. Particularly, applications of molecular dynamics (MD) and Monte Carlo (MC) simulation methods are discussed in various contexts, including studies of isolated ligands, apo-receptors, ligand/receptor complexes and intracellular kinase domains. The review concludes with a brief overview and future outlook for modeling and computational studies in this family of proteins. PMID:25680077

Progestogen treatments for cycle management in a sheep model of assisted conception affect the growth patterns, the expression of luteinizing hormone receptors, and the progesterone secretion of induced corpora lutea.

PubMed

Letelier, Claudia; García-Fernández, Rosa Ana; Contreras-Solis, Ignacio; Sanchez, María Angeles; Garcia-Palencia, Pilar; Sanchez, Belen; Gonzalez-Bulnes, Antonio; Flores, Juana María

2010-03-01

To determine, in a sheep model, the effect of a short-term progestative treatment on growth dynamics and functionality of induced corpora lutea. Observational, model study. Public university. Sixty adult female sheep. Synchronization and induction of ovulation with progestogens and prostaglandin analogues; ovarian ultrasonography, blood sampling, and ovariectomy. Determination of pituitary function and morphologic characteristics, expression of luteinizing hormone (LH) receptors, and progesterone secretion of corpora lutea. The use of progestative pretreatments for assisted conception affect the growth patterns, the expression of LH receptors, and the progesterone secretion of induced corpora lutea. The current study indicates, in a sheep model, the existence of deleterious effects from progestogens on functionality of induced corpora lutea. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Homology-based Modeling of Rhodopsin-like Family Members in the Inactive State: Structural Analysis and Deduction of Tips for Modeling and Optimization.

PubMed

Pappalardo, Matteo; Rayan, Mahmoud; Abu-Lafi, Saleh; Leonardi, Martha E; Milardi, Danilo; Guccione, Salvatore; Rayan, Anwar

2017-08-01

Modeling G-Protein Coupled Receptors (GPCRs) is an emergent field of research, since utility of high-quality models in receptor structure-based strategies might facilitate the discovery of interesting drug candidates. The findings from a quantitative analysis of eighteen resolved structures of rhodopsin family "A" receptors crystallized with antagonists and 153 pairs of structures are described. A strategy termed endeca-amino acids fragmentation was used to analyze the structures models aiming to detect the relationship between sequence identity and Root Mean Square Deviation (RMSD) at each trans-membrane-domain. Moreover, we have applied the leave-one-out strategy to study the shiftiness likelihood of the helices. The type of correlation between sequence identity and RMSD was studied using the aforementioned set receptors as representatives of membrane proteins and 98 serine proteases with 4753 pairs of structures as representatives of globular proteins. Data analysis using fragmentation strategy revealed that there is some extent of correlation between sequence identity and global RMSD of 11AA width windows. However, spatial conservation is not always close to the endoplasmic side as was reported before. A comparative study with globular proteins shows that GPCRs have higher standard deviation and higher slope in the graph with correlation between sequence identity and RMSD. The extracted information disclosed in this paper could be incorporated in the modeling protocols while using technique for model optimization and refinement. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

wFReDoW: A Cloud-Based Web Environment to Handle Molecular Docking Simulations of a Fully Flexible Receptor Model

PubMed Central

De Paris, Renata; Frantz, Fábio A.; Norberto de Souza, Osmar; Ruiz, Duncan D. A.

2013-01-01

Molecular docking simulations of fully flexible protein receptor (FFR) models are coming of age. In our studies, an FFR model is represented by a series of different conformations derived from a molecular dynamic simulation trajectory of the receptor. For each conformation in the FFR model, a docking simulation is executed and analyzed. An important challenge is to perform virtual screening of millions of ligands using an FFR model in a sequential mode since it can become computationally very demanding. In this paper, we propose a cloud-based web environment, called web Flexible Receptor Docking Workflow (wFReDoW), which reduces the CPU time in the molecular docking simulations of FFR models to small molecules. It is based on the new workflow data pattern called self-adaptive multiple instances (P-SaMIs) and on a middleware built on Amazon EC2 instances. P-SaMI reduces the number of molecular docking simulations while the middleware speeds up the docking experiments using a High Performance Computing (HPC) environment on the cloud. The experimental results show a reduction in the total elapsed time of docking experiments and the quality of the new reduced receptor models produced by discarding the nonpromising conformations from an FFR model ruled by the P-SaMI data pattern. PMID:23691504

Modeling and simulation studies of human β3 adrenergic receptor and its interactions with agonists.

PubMed

Sahi, Shakti; Tewatia, Parul; Malik, Balwant K

2012-12-01

β3 adrenergic receptor (β3AR) is known to mediate various pharmacological and physiological effects such as thermogenesis in brown adipocytes, lipolysis in white adipocytes, glucose homeostasis and intestinal smooth muscle relaxation. Several efforts have been made in this field to understand their function and regulation in different human tissues and they have emerged as potential attractive targets in drug discovery for the treatment of diabetes, depression, obesity etc. Although the crystal structures of Bovine Rhodopsin and β2 adrenergic receptor have been resolved, to date there is no three dimensional structural information on β3AR. Our aim in this study was to model 3D structure of β3AR by various molecular modeling and simulation techniques. In this paper, we describe a refined predicted model of β3AR using different algorithms for structure prediction. The structural refinement and minimization of the generated 3D model of β3AR were done by Schrodinger suite 9.1. Docking studies of β3AR model with the known agonists enabled us to identify specific residues, viz, Asp 117, Ser 208, Ser 209, Ser 212, Arg 315, Asn 332, within the β3AR binding pocket, which might play an important role in ligand binding. Receptor ligand interaction studies clearly indicated that these five residues showed strong hydrogen bonding interactions with the ligands. The results have been correlated with the experimental data available. The predicted ligand binding interactions and the simulation studies validate the methods used to predict the 3D-structure.

Selective CB2 receptor agonists. Part 2: Structure-activity relationship studies and optimization of proline-based compounds.

PubMed

Riether, Doris; Zindell, Renee; Wu, Lifen; Betageri, Raj; Jenkins, James E; Khor, Someina; Berry, Angela K; Hickey, Eugene R; Ermann, Monika; Albrecht, Claudia; Ceci, Angelo; Gemkow, Mark J; Nagaraja, Nelamangala V; Romig, Helmut; Sauer, Achim; Thomson, David S

2015-02-01

Through a ligand-based pharmacophore model (S)-proline based compounds were identified as potent cannabinoid receptor 2 (CB2) agonists with high selectivity over the cannabinoid receptor 1 (CB1). Structure-activity relationship investigations for this compound class lead to oxo-proline compounds 21 and 22 which combine an impressive CB1 selectivity profile with good pharmacokinetic properties. In a streptozotocin induced diabetic neuropathy model, 22 demonstrated a dose-dependent reversal of mechanical hyperalgesia. Copyright © 2014 Elsevier Ltd. All rights reserved.

Genetic background can result in a marked or minimal effect of gene knockout (GPR55 and CB2 receptor) in experimental autoimmune encephalomyelitis models of multiple sclerosis.

PubMed

Sisay, Sofia; Pryce, Gareth; Jackson, Samuel J; Tanner, Carolyn; Ross, Ruth A; Michael, Gregory J; Selwood, David L; Giovannoni, Gavin; Baker, David

2013-01-01

Endocannabinoids and some phytocannabinoids bind to CB1 and CB2 cannabinoid receptors, transient receptor potential vanilloid one (TRPV1) receptor and the orphan G protein receptor fifty-five (GPR55). Studies using C57BL/10 and C57BL/6 (Cnr2 (tm1Zim)) CB2 cannabinoid receptor knockout mice have demonstrated an immune-augmenting effect in experimental autoimmune encephalomyelitis (EAE) models of multiple sclerosis. However, other EAE studies in Biozzi ABH mice often failed to show any treatment effect of either CB2 receptor agonism or antagonism on inhibition of T cell autoimmunity. The influence of genetic background on the induction of EAE in endocannabinoid system-related gene knockout mice was examined. It was found that C57BL/6.GPR55 knockout mice developed less severe disease, notably in female mice, following active induction with myelin oligodendrocyte glycoprotein 35-55 peptide. In contrast C57BL/6.CB2 (Cnr2 (Dgen)) receptor knockout mice developed augmented severity of disease consistent with the genetically and pharmacologically-distinct, Cnr2 (tm1Zim) mice. However, when the knockout gene was bred into the ABH mouse background and EAE induced with spinal cord autoantigens the immune-enhancing effect of CB2 receptor deletion was lost. Likewise CB1 receptor and transient receptor potential vanilloid one knockout mice on the ABH background demonstrated no alteration in immune-susceptibility, in terms of disease incidence and severity of EAE, in contrast to that reported in some C57BL/6 mouse studies. Furthermore the immune-modulating influence of GPR55 was marginal on the ABH mouse background. Whilst sedative doses of tetrahydrocannabinol could induce immunosuppression, this was associated with a CB1 receptor rather than a CB2 receptor-mediated effect. These data support the fact that non-psychoactive doses of medicinal cannabis have a marginal influence on the immune response in MS. Importantly, it adds a note of caution for the translational value of some transgenic/gene knockout and other studies on low-EAE susceptibility backgrounds with inconsistent disease course and susceptibility.

Genetic Background Can Result in a Marked or Minimal Effect of Gene Knockout (GPR55 and CB2 Receptor) in Experimental Autoimmune Encephalomyelitis Models of Multiple Sclerosis

PubMed Central

Jackson, Samuel J.; Tanner, Carolyn; Ross, Ruth A.; Michael, Gregory J.; Selwood, David L.; Giovannoni, Gavin; Baker, David

2013-01-01

Endocannabinoids and some phytocannabinoids bind to CB1 and CB2 cannabinoid receptors, transient receptor potential vanilloid one (TRPV1) receptor and the orphan G protein receptor fifty-five (GPR55). Studies using C57BL/10 and C57BL/6 (Cnr2 tm1Zim) CB2 cannabinoid receptor knockout mice have demonstrated an immune-augmenting effect in experimental autoimmune encephalomyelitis (EAE) models of multiple sclerosis. However, other EAE studies in Biozzi ABH mice often failed to show any treatment effect of either CB2 receptor agonism or antagonism on inhibition of T cell autoimmunity. The influence of genetic background on the induction of EAE in endocannabinoid system-related gene knockout mice was examined. It was found that C57BL/6.GPR55 knockout mice developed less severe disease, notably in female mice, following active induction with myelin oligodendrocyte glycoprotein 35-55 peptide. In contrast C57BL/6.CB2 (Cnr2 Dgen) receptor knockout mice developed augmented severity of disease consistent with the genetically and pharmacologically-distinct, Cnr2 tm1Zim mice. However, when the knockout gene was bred into the ABH mouse background and EAE induced with spinal cord autoantigens the immune-enhancing effect of CB2 receptor deletion was lost. Likewise CB1 receptor and transient receptor potential vanilloid one knockout mice on the ABH background demonstrated no alteration in immune-susceptibility, in terms of disease incidence and severity of EAE, in contrast to that reported in some C57BL/6 mouse studies. Furthermore the immune-modulating influence of GPR55 was marginal on the ABH mouse background. Whilst sedative doses of tetrahydrocannabinol could induce immunosuppression, this was associated with a CB1 receptor rather than a CB2 receptor-mediated effect. These data support the fact that non-psychoactive doses of medicinal cannabis have a marginal influence on the immune response in MS. Importantly, it adds a note of caution for the translational value of some transgenic/gene knockout and other studies on low-EAE susceptibility backgrounds with inconsistent disease course and susceptibility. PMID:24130809

Design, synthesis and screening studies of potent thiazol-2-amine derivatives as fibroblast growth factor receptor 1 inhibitors.

PubMed

Kumar, B V S Suneel; Lakshmi, Narasu; Kumar, M Ravi; Rambabu, Gundla; Manjashetty, Thimmappa H; Arunasree, Kalle M; Sriram, Dharmarajan; Ramkumar, Kavya; Neamati, Nouri; Dayam, Raveendra; Sarma, J A R P

2014-01-01

Fibroblast growth factor receptor 1 (FGFR1) a tyrosine kinase receptor, plays important roles in angiogenesis, embryonic development, cell proliferation, cell differentiation, and wound healing. The FGFR isoforms and their receptors (FGFRs) considered as a potential targets and under intense research to design potential anticancer agents. Fibroblast growth factors (FGF's) and its growth factor receptors (FGFR) plays vital role in one of the critical pathway in monitoring angiogenesis. In the current study, quantitative pharmacophore models were generated and validated using known FGFR1 inhibitors. The pharmacophore models were generated using a set of 28 compounds (training). The top pharmacophore model was selected and validated using a set of 126 compounds (test set) and also using external validation. The validated pharmacophore was considered as a virtual screening query to screen a database of 400,000 virtual molecules and pharmacophore model retrieved 2800 hits. The retrieved hits were subsequently filtered based on the fit value. The selected hits were subjected for docking studies to observe the binding modes of the retrieved hits and also to reduce the false positives. One of the potential hits (thiazole-2-amine derivative) was selected based the pharmacophore fit value, dock score, and synthetic feasibility. A few analogues of the thiazole-2-amine derivative were synthesized. These compounds were screened for FGFR1 activity and anti-proliferative studies. The top active compound showed 56.87% inhibition of FGFR1 activity at 50 µM and also showed good cellular activity. Further optimization of thiazole-2-amine derivatives is in progress.

Structural interpretation of P2X receptor mutagenesis studies on drug action.

PubMed

Evans, Richard J

2010-11-01

P2X receptors for ATP are ligand gated cation channels that form from the trimeric assembly of subunits with two transmembrane segments, a large extracellular ligand binding loop, and intracellular amino and carboxy termini. The receptors are expressed throughout the body, involved in functions ranging from blood clotting to inflammation, and may provide important targets for novel therapeutics. Mutagenesis based studies have been used to develop an understanding of the molecular basis of their pharmacology with the aim of developing models of the ligand binding site. A crystal structure for the zebra fish P2X4 receptor in the closed agonist unbound state has been published recently, which provides a major advance in our understanding of the receptors. This review gives an overview of mutagenesis studies that have led to the development of a model of the ATP binding site, as well as identifying residues contributing to allosteric regulation and antagonism. These studies are discussed with reference to the crystal to provide a structural interpretation of the molecular basis of drug action. © 2010 The Author. British Journal of Pharmacology © 2010 The British Pharmacological Society.

3D-QSAR and docking studies on 4-anilinoquinazoline and 4-anilinoquinoline epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors

NASA Astrophysics Data System (ADS)

Assefa, Haregewein; Kamath, Shantaram; Buolamwini, John K.

2003-08-01

The overexpression and/or mutation of the epidermal growth factor receptor (EGFR) tyrosine kinase has been observed in many human solid tumors, and is under intense investigation as a novel anticancer molecular target. Comparative 3D-QSAR analyses using different alignments were undertaken employing comparative molecular field analysis (CoMFA) and comparative molecular similarity analysis (CoMSIA) for 122 anilinoquinazoline and 50 anilinoquinoline inhibitors of EGFR kinase. The SYBYL multifit alignment rule was applied to three different conformational templates, two obtained from a MacroModel Monte Carlo conformational search, and one from the bound conformation of erlotinib in complex with EGFR in the X-ray crystal structure. In addition, a flexible ligand docking alignment obtained with the GOLD docking program, and a novel flexible receptor-guided consensus dynamics alignment obtained with the DISCOVER program in the INSIGHTII modeling package were also investigated. 3D-QSAR models with q2 values up to 0.70 and r2 values up to 0.97 were obtained. Among the 4-anilinoquinazoline set, the q2 values were similar, but the ability of the different conformational models to predict the activities of an external test set varied considerably. In this regard, the model derived using the X-ray crystallographically determined bioactive conformation of erlotinib afforded the best predictive model. Electrostatic, hydrophobic and H-bond donor descriptors contributed the most to the QSAR models of the 4-anilinoquinazolines, whereas electrostatic, hydrophobic and H-bond acceptor descriptors contributed the most to the 4-anilinoquinoline QSAR, particularly the H-bond acceptor descriptor. A novel receptor-guided consensus dynamics alignment has also been introduced for 3D-QSAR studies. This new alignment method may incorporate to some extent ligand-receptor induced fit effects into 3D-QSAR models.

Role of σ1 Receptors in Learning and Memory and Alzheimer's Disease-Type Dementia.

PubMed

Maurice, Tangui; Goguadze, Nino

2017-01-01

The present chapter will review the role of σ 1 receptor in learning and memory and neuroprotection , against Alzheimer's type dementia. σ 1 Receptor agonists have been tested in a variety of pharmacological and pathological models of learning impairments in rodents these last past 20 years. Their anti-amnesic effects have been explained by the wide-range modulatory role of σ 1 receptors on Ca 2+ mobilizations, neurotransmitter responses, and particularly glutamate and acetylcholine systems, and neurotrophic factors. Recent observations from genetic and pharmacological studies have shown that σ 1 receptor can also be targeted in neurodegenerative diseases, and particularly Alzheimer's disease . Several compounds, acting partly through the σ 1 receptor, have showed effective neuroprotection in transgenic mouse models of Alzheimer's disease . We will review the data and discuss the possible mechanisms of action, particularly focusing on oxidative stress and mitochondrial integrity, trophic factors and a novel hypothesis suggesting a functional interaction between the σ 1 receptor and α 7 nicotinic acetylcholine receptor. Finally, we will discuss the pharmacological peculiarities of non-selective σ 1 receptor ligands, now developed as neuroprotectants in Alzheimer's disease , and positive modulators, recently described and that showed efficacy against learning and memory deficits.

Reassessment of the Unique Mode of Binding between Angiotensin II Type 1 Receptor and Their Blockers

PubMed Central

Matsuo, Yoshino; Saku, Keijiro; Karnik, Sadashiva S.

2013-01-01

While the molecular structures of angiotensin II (Ang II) type 1 (AT1) receptor blockers (ARBs) are very similar, they are also slightly different. Although each ARB has been shown to exhibit a unique mode of binding to AT1 receptor, different positions of the AT1 receptor have been analyzed and computational modeling has been performed using different crystal structures for the receptor as a template and different kinds of software. Therefore, we systematically analyzed the critical positions of the AT1 receptor, Tyr113, Tyr184, Lys199, His256 and Gln257 using a mutagenesis study, and subsequently performed computational modeling of the binding of ARBs to AT1 receptor using CXCR4 receptor as a new template and a single version of software. The interactions between Tyr113 in the AT1 receptor and the hydroxyl group of olmesartan, between Lys199 and carboxyl or tetrazole groups, and between His256 or Gln257 and the tetrazole group were studied. The common structure, a tetrazole group, of most ARBs similarly bind to Lys199, His256 and Gln257 of AT1 receptor. Lys199 in the AT1 receptor binds to the carboxyl group of EXP3174, candesartan and azilsartan, whereas oxygen in the amidecarbonyl group of valsartan may bind to Lys199. The benzimidazole portion of telmisartan may bind to a lipophilic pocket that includes Tyr113. On the other hand, the n-butyl group of irbesartan may bind to Tyr113. In conclusion, we confirmed that the slightly different structures of ARBs may be critical for binding to AT1 receptor and for the formation of unique modes of binding. PMID:24260317

The role of ryanodine receptor type 3 in a mouse model of Alzheimer disease

PubMed Central

Liu, Jie; Supnet, Charlene; Sun, Suya; Zhang, Hua; Good, Levi; Popugaeva, Elena; Bezprozvanny, Ilya

2014-01-01

Dysregulated endoplasmic reticulum (ER) calcium (Ca2+) signaling is reported to play an important role in Alzheimer disease (AD) pathogenesis. The role of ER Ca2+ release channels, the ryanodine receptors (RyanRs), has been extensively studied in AD models and RyanR expression and activity are upregulated in the brains of various familial AD (FAD) models. The objective of this study was to utilize a genetic approach to evaluate the importance of RyanR type 3 (RyanR3) in the context of AD pathology. PMID:24476841

Decreased CB receptor binding and cannabinoid signaling in three brain regions of a rat model of schizophrenia.

PubMed

Szűcs, Edina; Dvorácskó, Szabolcs; Tömböly, Csaba; Büki, Alexandra; Kékesi, Gabriella; Horváth, Gyöngyi; Benyhe, Sándor

2016-10-28

Schizophrenia is a serious mental health disorder characterized by several behavioral and biochemicel abnormalities. In a previous study we have shown that mu-opioid (MOP) receptor signaling is impaired in specific brain regions of our three-hit animal model of schizophrenia. Since the cannabinoid system is significantly influenced in schizophrenic patients, in the present work we investigated cannabinoid (CB) receptor binding and G-protein activation in cortical, subcortical and cerebellar regions of control and 'schizophrenic' rats. Cannabinoid agonist (WIN-55,212-2 mesylate) mediated G-protein activation was consistently decreased in all areas tested, and the difference was extremely significant in membranes prepared from the cerebellum. Interestingly, the cerebellar activity of WIN-55,212-2 stimulated G-proteins was substantially higher than those of cerebral cortex and subcortical region in control animals, indicating a primordial role of the cannabinoid system in the cerebellum. At the level of radioligand binding, the affinities of the CB receptors were also markedly decreased in the model animals. Capacity of the [ 3 H]WIN-55,212-2 binding was only higher in the cerebellum of 'schizophrenic' model rats. Taken together, in all three brain areas of model rats both cannabinoid receptor binding and cannabinoid agonist-mediated G-protein activation were regularly decreased. Our results revealed that besides the opioids, the endocannabinoid - cannabis receptor system also shows impairment in our rat model, increasing its face validity and translational utility. Copyright © 2016. Published by Elsevier Ireland Ltd.

Encompassing receptor flexibility in virtual screening using ensemble docking-based hybrid QSAR: discovery of novel phytochemicals for BACE1 inhibition.

PubMed

Chakraborty, Sandipan; Ramachandran, Balaji; Basu, Soumalee

2014-10-01

Mimicking receptor flexibility during receptor-ligand binding is a challenging task in computational drug design since it is associated with a large increase in the conformational search space. In the present study, we have devised an in silico design strategy incorporating receptor flexibility in virtual screening to identify potential lead compounds as inhibitors for flexible proteins. We have considered BACE1 (β-secretase), a key target protease from a therapeutic perspective for Alzheimer's disease, as the highly flexible receptor. The protein undergoes significant conformational transitions from open to closed form upon ligand binding, which makes it a difficult target for inhibitor design. We have designed a hybrid structure-activity model containing both ligand based descriptors and energetic descriptors obtained from molecular docking based on a dataset of structurally diverse BACE1 inhibitors. An ensemble of receptor conformations have been used in the docking study, further improving the prediction ability of the model. The designed model that shows significant prediction ability judged by several statistical parameters has been used to screen an in house developed 3-D structural library of 731 phytochemicals. 24 highly potent, novel BACE1 inhibitors with predicted activity (Ki) ≤ 50 nM have been identified. Detailed analysis reveals pharmacophoric features of these novel inhibitors required to inhibit BACE1.

Biochemical correlates in an animal model of depression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnson, J.O.

1986-01-01

A valid animal model of depression was used to explore specific adrenergic receptor differences between rats exhibiting aberrant behavior and control groups. Preliminary experiments revealed a distinct upregulation of hippocampal beta-receptors (as compared to other brain regions) in those animals acquiring a response deficit as a result of exposure to inescapable footshock. Concurrent studies using standard receptor binding techniques showed no large changes in the density of alpha-adrenergic, serotonergic, or dopaminergic receptor densities. This led to the hypothesis that the hippocampal beta-receptor in responses deficient animals could be correlated with the behavioral changes seen after exposure to the aversive stimulus.more » Normalization of the behavior through the administration of antidepressants could be expected to reverse the biochemical changes if these are related to the mechanism of action of antidepressant drugs. This study makes three important points: (1) there is a relevant biochemical change in the hippocampus of response deficient rats which occurs in parallel to a well-defined behavior, (2) the biochemical and behavioral changes are normalized by antidepressant treatments exhibiting both serotonergic and adrenergic mechanisms of action, and (3) the mode of action of antidepressants in this model is probably a combination of serotonergic and adrenergic influences modulating the hippocampal beta-receptor. These results are discussed in relation to anatomical and biochemical aspects of antidepressant action.« less

Smad Signaling Dynamics: Insights from a Parsimonious Model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wiley, H. S.; Shankaran, Harish

2008-09-09

The molecular mechanisms that transmit information from cell surface receptors to the nucleus are exceedingly complex; thus, much effort has been expended in developing computational models to understand these processes. A recent study on modeling the nuclear-cytoplasmic shuttling of Smad2-Smad4 complexes in response to transforming growth factor β (TGF-β) receptor activation has provided substantial insight into how this signaling network translates the degree of TGF-β receptor activation (input) into the amount of nuclear Smad2-Smad4 complexes (output). The study addressed this question by combining a simple, mechanistic model with targeted experiments, an approach that proved particularly powerful for exploring the fundamentalmore » properties of a complex signaling network. The mathematical model revealed that Smad nuclear-cytoplasmic dynamics enables a proportional, but time-delayed coupling between the input and the output. As a result, the output can faithfully track gradual changes in the input, while the rapid input fluctuations that constitute signaling noise are dampened out.« less

MK-801-Treated Oligodendrocytes as a Cellular Model to Study Schizophrenia.

PubMed

Brandão-Teles, Caroline; Martins-de-Souza, Daniel; Guest, Paul C; Cassoli, Juliana S

2017-01-01

Glutamate is the most important excitatory neurotransmitter in the brain. The N-methyl-D-aspartate (NMDA) subtype of glutamate receptor is found both in neurons and glial cells such as oligodendrocytes, which have been shown to be dysfunctional in schizophrenia. For this reasons, the oligodendrocyte MO3.13 cell line has been used to study glutamatergic dysfunction as a model of schizophrenia using the NMDA receptor antagonists such as MK-801 to block receptor function. Here, we describe a comprehensive protocol for culturing and carrying out proteomic analyses of MK-801-treated MO3.13 cells as a means of identifying potential new biomarkers and targets for drug discovery in schizophrenia research.

Nigral dopamine type-1 receptors are reduced in Huntington's disease: A postmortem autoradiographic study using ( sup 3 H)SCH 23390 and correlation with ( sup 3 H)forskolin binding

DOE Office of Scientific and Technical Information (OSTI.GOV)

Filloux, F.; Wagster, M.V.; Folstein, S.

1990-11-01

Intrastriatal injection of excitatory amino acids, particularly quinolinic acid, has been proposed as an animal model of Huntington's disease. Such neurotoxic lesions of caudate-putamen result in marked dopamine type-1 (D1) receptor losses in the injected nuclei as well as in the ipsilateral substantia nigra pars reticulata. Postmortem human substantia nigra from Huntington's disease brains and from control brains were examined using in vitro autoradiography. A marked reduction in ({sup 3}H)SCH 23390 binding (labeling D1 receptors) in the substantia nigra of postmortem brains of Huntington's patients was identified, thus paralleling the alterations seen in the animal models. A positive, statistically significantmore » correlation was also encountered between D1 receptor binding (labeled by ({sup 3}H)SCH 23390) and ({sup 3}H)forskolin binding (which identifies adenylate cyclase, a second messenger system linked to D1 receptor activation). The results suggest that in the human--as in lower vertebrates--D1 receptors are located on striatonigral terminals and that D1 receptor loss tends to be paralleled by a reduction in adenylate cyclase. Radioactive agents selective for the D1 receptor may prove useful in future studies of Huntington's disease using positron emission tomography scanning.« less

Tachykinins and Their Receptors: Contributions to Physiological Control and the Mechanisms of Disease

PubMed Central

Steinhoff, Martin S.; von Mentzer, Bengt; Geppetti, Pierangelo; Pothoulakis, Charalabos; Bunnett, Nigel W.

2014-01-01

The tachykinins, exemplified by substance P, are one of the most intensively studied neuropeptide families. They comprise a series of structurally related peptides that derive from alternate processing of three Tac genes and are expressed throughout the nervous and immune systems. Tachykinins interact with three neurokinin G protein-coupled receptors. The signaling, trafficking, and regulation of neurokinin receptors have also been topics of intense study. Tachykinins participate in important physiological processes in the nervous, immune, gastrointestinal, respiratory, urogenital, and dermal systems, including inflammation, nociception, smooth muscle contractility, epithelial secretion, and proliferation. They contribute to multiple diseases processes, including acute and chronic inflammation and pain, fibrosis, affective and addictive disorders, functional disorders of the intestine and urinary bladder, infection, and cancer. Neurokinin receptor antagonists are selective, potent, and show efficacy in models of disease. In clinical trials there is a singular success: neurokinin 1 receptor antagonists to treat nausea and vomiting. New information about the involvement of tachykinins in infection, fibrosis, and pruritus justifies further trials. A deeper understanding of disease mechanisms is required for the development of more predictive experimental models, and for the design and interpretation of clinical trials. Knowledge of neurokinin receptor structure, and the development of targeting strategies to disrupt disease-relevant subcellular signaling of neurokinin receptors, may refine the next generation of neurokinin receptor antagonists. PMID:24382888

Mathematical modelling of non-stationary fluctuation analysis for studying channel properties of synaptic AMPA receptors

PubMed Central

Benke, Timothy A; Lüthi, Andreas; Palmer, Mary J; Wikström, Martin A; Anderson, William W; Isaac, John T R; Collingridge, Graham L

2001-01-01

The molecular properties of synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors are an important factor determining excitatory synaptic transmission in the brain. Changes in the number (N) or single-channel conductance (γ) of functional AMPA receptors may underlie synaptic plasticity, such as long-term potentiation (LTP) and long-term depression (LTD). These parameters have been estimated using non-stationary fluctuation analysis (NSFA). The validity of NSFA for studying the channel properties of synaptic AMPA receptors was assessed using a cable model with dendritic spines and a microscopic kinetic description of AMPA receptors. Electrotonic, geometric and kinetic parameters were altered in order to determine their effects on estimates of the underlying γ. Estimates of γ were very sensitive to the access resistance of the recording (RA) and the mean open time of AMPA channels. Estimates of γ were less sensitive to the distance between the electrode and the synaptic site, the electrotonic properties of dendritic structures, recording electrode capacitance and background noise. Estimates of γ were insensitive to changes in spine morphology, synaptic glutamate concentration and the peak open probability (Po) of AMPA receptors. The results obtained using the model agree with biological data, obtained from 91 dendritic recordings from rat CA1 pyramidal cells. A correlation analysis showed that RA resulted in a slowing of the decay time constant of excitatory postsynaptic currents (EPSCs) by approximately 150 %, from an estimated value of 3.1 ms. RA also greatly attenuated the absolute estimate of γ by approximately 50-70 %. When other parameters remain constant, the model demonstrates that NSFA of dendritic recordings can readily discriminate between changes in γvs. changes in N or Po. Neither background noise nor asynchronous activation of multiple synapses prevented reliable discrimination between changes in γ and changes in either N or Po. The model (available online) can be used to predict how changes in the different properties of AMPA receptors may influence synaptic transmission and plasticity. PMID:11731574

Blocking beta 2-adrenergic receptor inhibits dendrite ramification in a mouse model of Alzheimer's disease.

PubMed

Wu, Qin; Sun, Jin-Xia; Song, Xiang-He; Wang, Jing; Xiong, Cun-Quan; Teng, Fei-Xiang; Gao, Cui-Xiang

2017-09-01

Dendrite ramification affects synaptic strength and plays a crucial role in memory. Previous studies revealed a correlation between beta 2-adrenergic receptor dysfunction and Alzheimer's disease (AD), although the mechanism involved is still poorly understood. The current study investigated the potential effect of the selective β 2 -adrenergic receptor antagonist, ICI 118551 (ICI), on Aβ deposits and AD-related cognitive impairment. Morris water maze test results demonstrated that the performance of AD-transgenic (TG) mice treated with ICI (AD-TG/ICI) was significantly poorer compared with NaCl-treated AD-TG mice (AD-TG/NaCl), suggesting that β 2 -adrenergic receptor blockage by ICI might reduce the learning and memory abilities of mice. Golgi staining and immunohistochemical staining revealed that blockage of the β 2 -adrenergic receptor by ICI treatment decreased the number of dendritic branches, and ICI treatment in AD-TG mice decreased the expression of hippocampal synaptophysin and synapsin 1. Western blot assay results showed that the blockage of β 2 -adrenergic receptor increased amyloid-β accumulation by downregulating hippocampal α-secretase activity and increasing the phosphorylation of amyloid precursor protein. These findings suggest that blocking the β 2 -adrenergic receptor inhibits dendrite ramification of hippocampal neurons in a mouse model of AD.

Blocking beta 2-adrenergic receptor inhibits dendrite ramification in a mouse model of Alzheimer's disease

PubMed Central

Wu, Qin; Sun, Jin-xia; Song, Xiang-he; Wang, Jing; Xiong, Cun-quan; Teng, Fei-xiang; Gao, Cui-xiang

2017-01-01

Dendrite ramification affects synaptic strength and plays a crucial role in memory. Previous studies revealed a correlation between beta 2-adrenergic receptor dysfunction and Alzheimer's disease (AD), although the mechanism involved is still poorly understood. The current study investigated the potential effect of the selective β2-adrenergic receptor antagonist, ICI 118551 (ICI), on Aβ deposits and AD-related cognitive impairment. Morris water maze test results demonstrated that the performance of AD-transgenic (TG) mice treated with ICI (AD-TG/ICI) was significantly poorer compared with NaCl-treated AD-TG mice (AD-TG/NaCl), suggesting that β2-adrenergic receptor blockage by ICI might reduce the learning and memory abilities of mice. Golgi staining and immunohistochemical staining revealed that blockage of the β2-adrenergic receptor by ICI treatment decreased the number of dendritic branches, and ICI treatment in AD-TG mice decreased the expression of hippocampal synaptophysin and synapsin 1. Western blot assay results showed that the blockage of β2-adrenergic receptor increased amyloid-β accumulation by downregulating hippocampal α-secretase activity and increasing the phosphorylation of amyloid precursor protein. These findings suggest that blocking the β2-adrenergic receptor inhibits dendrite ramification of hippocampal neurons in a mouse model of AD. PMID:29089997

Dimer-based model for heptaspanning membrane receptors.

PubMed

Franco, Rafael; Casadó, Vicent; Mallol, Josefa; Ferré, Sergi; Fuxe, Kjell; Cortés, Antonio; Ciruela, Francisco; Lluis, Carmen; Canela, Enric I

2005-07-01

The existence of intramembrane receptor-receptor interactions for heptaspanning membrane receptors is now fully accepted, but a model considering dimers as the basic unit that binds to two ligand molecules is lacking. Here, we propose a two-state-dimer model in which the ligand-induced conformational changes from one component of the dimer are communicated to the other. Our model predicts cooperativity in binding, which is relevant because the other current models fail to address this phenomenon satisfactorily. Our two-state-dimer model also predicts the variety of responses elicited by full or partial agonists, neutral antagonists and inverse agonists. This model can aid our understanding of the operation of heptaspanning receptors and receptor channels, and, potentially, be important for improving the treatment of cardiovascular, neurological and neuropsychyatric diseases.

Identification of endogenous surrogate ligands for human P2Y receptors through an in silico search.

PubMed

Hiramoto, Takeshi; Nonaka, Yosuke; Inoue, Kazuko; Yamamoto, Takefumi; Omatsu-Kanbe, Mariko; Matsuura, Hiroshi; Gohda, Keigo; Fujita, Norihisa

2004-05-01

G protein-coupled receptors (GPCRs) are distributed widely throughout the human body, and nearly 50% of current medicines act on a GPCR. GPCRs are considered to consist of seven transmembrane alpha-helices that form an alpha-helical bundle in which agonists and antagonists bind. A 3D structure of the target GPCR is indispensable for designing novel medicines acting on a GPCR. We have previously constructed the 3D structure of human P2Y(1) (hP2Y(1)) receptor, a GPCR, by homology modeling with the 3D structure of bovine rhodopsin as a template. In the present study, we have employed an in silico screening for compounds that could bind to the hP2Y(1)-receptor model using AutoDock 3.0. We selected 21 of the 30 top-ranked compounds, and by measuring intracellular Ca(2+) concentration, we identified 12 compounds that activated or blocked the hP2Y(1) receptor stably expressed in recombinant CHO cells. 5-Phosphoribosyl-1-pyrophosphate (PRPP) was found to activate the hP2Y(1) receptor with a low ED(50) value of 15 nM. The Ca(2+) assays showed it had no significant effect on P2Y(2), P2Y(6), or P2X(2) receptors, but acted as a weak agonist on the P2Y(12) receptor. This is the first study to rationally identify surrogate ligands for the P2Y-receptor family.

Exploring Molecular Mechanisms of Ligand Recognition by Opioid Receptors with Metadynamics†

PubMed Central

Provasi, Davide; Bortolato, Andrea; Filizola, Marta

2009-01-01

Opioid receptors are G protein-coupled receptors (GPCRs) of utmost significance in the development of potent analgesic drugs for the treatment of severe pain. An accurate evaluation at the molecular level of the ligand binding pathways into these receptors may play a key role in the design of new molecules with more desirable properties and reduced side effects. The recent characterization of high-resolution X-ray crystal structures of non-rhodopsin GPCRs for diffusible hormones and neurotransmitters presents an unprecedented opportunity to build improved homology models of opioid receptors, and to study in more detail their molecular mechanisms of ligand recognition. In this study, possible entry pathways of the non-selective antagonist naloxone (NLX) from the water environment into the well-accepted alkaloid binding pocket of a delta opioid receptor (DOR) molecular model based on the β2-adrenergic receptor crystal structure are explored using microsecond-scale well-tempered metadynamics simulations. Using as collective variables distances that account for the position of NLX and of the receptor extracellular loop 2 in relation to the DOR binding pocket, we were able to distinguish between the different states visited by the ligand (i.e., docked, undocked, and metastable bound intermediates), and to predict a free energy of binding close to experimental values after correcting for possible drawbacks of the sampling approach. The strategy employed herein holds promise for its application to the docking of diverse ligands to the opioid receptors as well as to other GPCRs. PMID:19785461

Exploring molecular mechanisms of ligand recognition by opioid receptors with metadynamics.

PubMed

Provasi, Davide; Bortolato, Andrea; Filizola, Marta

2009-10-27

Opioid receptors are G protein-coupled receptors (GPCRs) of utmost significance in the development of potent analgesic drugs for the treatment of severe pain. An accurate evaluation at the molecular level of the ligand binding pathways into these receptors may play a key role in the design of new molecules with more desirable properties and reduced side effects. The recent characterization of high-resolution X-ray crystal structures of non-rhodopsin GPCRs for diffusible hormones and neurotransmitters presents an unprecedented opportunity to build improved homology models of opioid receptors, and to study in more detail their molecular mechanisms of ligand recognition. In this study, possible pathways for entry of the nonselective antagonist naloxone (NLX) from the water environment into the well-accepted alkaloid binding pocket of a delta opioid receptor (DOR) molecular model based on the beta2-adrenergic receptor crystal structure are explored using microsecond-scale well-tempered metadynamics simulations. Using as collective variables distances that account for the position of NLX and of the receptor extracellular loop 2 in relation to the DOR binding pocket, we were able to distinguish between the different states visited by the ligand (i.e., docked, undocked, and metastable bound intermediates) and to predict a free energy of binding close to experimental values after correcting for possible drawbacks of the sampling approach. The strategy employed herein holds promise for its application to the docking of diverse ligands to the opioid receptors as well as to other GPCRs.

Effect of hypobaric hypoxia on the P2X receptors of pyramidal cells in the immature rat hippocampus CA1 sub-field.

PubMed

Zhao, Yan-Dong; Cheng, Sai-Yu; Ou, Shan; Xiao, Zhi; He, Wen-Juan; Jian-Cui; Ruan, Huai-Zhen

2012-01-01

This study was designed to evaluate the effect of hypobaric hypoxia (HH) on the function and expression of P2X receptors in rat hippocampus CA1 pyramidal cells. The functional changes of P2X receptors were investigated through the cell HH model and the expressional alterations of P2X receptors were observed through the animal HH model. P2X receptors mediated currents were recorded from the freshly dissociated CA1 pyramidal cells of 7-day-old SD rats by whole cell patch clamp recording. The expression and distribution of P2X receptors were observed through immunohistochemistry and western blot at HH 3-day and 7-day. In acute HH conditions, the amplitudes of ATP evoked peak currents were decreased compared to control. The immunohistochemistry and western blot results reflected there was no change in P2X receptors expression after 3 days HH injury, while P2X receptors expression was up-regulated in response to 7 days HH injury. These findings supported the possibility that the function of P2X receptors was sensitive to HH damage and long-term function decrease should result in the expression increase of P2X receptors.

Extinction of an instrumental response: a cognitive behavioral assay in Fmr1 knockout mice.

PubMed

Sidorov, M S; Krueger, D D; Taylor, M; Gisin, E; Osterweil, E K; Bear, M F

2014-06-01

Fragile X (FX) is the most common genetic cause of intellectual disability and autism. Previous studies have shown that partial inhibition of metabotropic glutamate receptor signaling is sufficient to correct behavioral phenotypes in a mouse model of FX, including audiogenic seizures, open-field hyperactivity and social behavior. These phenotypes model well the epilepsy (15%), hyperactivity (20%) and autism (30%) that are comorbid with FX in human patients. Identifying reliable and robust mouse phenotypes to model cognitive impairments is critical considering the 90% comorbidity of FX and intellectual disability. Recent work characterized a five-choice visuospatial discrimination assay testing cognitive flexibility, in which FX model mice show impairments associated with decreases in synaptic proteins in prefrontal cortex (PFC). In this study, we sought to determine whether instrumental extinction, another process requiring PFC, is altered in FX model mice, and whether downregulation of metabotropic glutamate receptor signaling pathways is sufficient to correct both visuospatial discrimination and extinction phenotypes. We report that instrumental extinction is consistently exaggerated in FX model mice. However, neither the extinction phenotype nor the visuospatial discrimination phenotype is corrected by approaches targeting metabotropic glutamate receptor signaling. This work describes a novel behavioral extinction assay to model impaired cognition in mouse models of neurodevelopmental disorders, provides evidence that extinction is exaggerated in the FX mouse model and suggests possible limitations of metabotropic glutamate receptor-based pharmacotherapy. © 2014 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Models for H₃ receptor antagonist activity of sulfonylurea derivatives.

PubMed

Khatri, Naveen; Madan, A K

2014-03-01

The histamine H₃ receptor has been perceived as an auspicious target for the treatment of various central and peripheral nervous system diseases. In present study, a wide variety of 60 2D and 3D molecular descriptors (MDs) were successfully utilized for the development of models for the prediction of antagonist activity of sulfonylurea derivatives for histamine H₃ receptors. Models were developed through decision tree (DT), random forest (RF) and moving average analysis (MAA). Dragon software version 6.0.28 was employed for calculation of values of diverse MDs of each analogue involved in the data set. The DT classified and correctly predicted the input data with an impressive non-error rate of 94% in the training set and 82.5% during cross validation. RF correctly classified the analogues into active and inactive with a non-error rate of 79.3%. The MAA based models predicted the antagonist histamine H₃ receptor activity with non-error rate up to 90%. Active ranges of the proposed MAA based models not only exhibited high potency but also showed improved safety as indicated by relatively high values of selectivity index. The statistical significance of the models was assessed through sensitivity, specificity, non-error rate, Matthew's correlation coefficient and intercorrelation analysis. Proposed models offer vast potential for providing lead structures for development of potent but safe H₃ receptor antagonist sulfonylurea derivatives. Copyright © 2013 Elsevier Inc. All rights reserved.

Active-State Model of a Dopamine D2 Receptor - Gαi Complex Stabilized by Aripiprazole-Type Partial Agonists

PubMed Central

Kling, Ralf C.; Tschammer, Nuska; Lanig, Harald; Clark, Timothy; Gmeiner, Peter

2014-01-01

Partial agonists exhibit a submaximal capacity to enhance the coupling of one receptor to an intracellular binding partner. Although a multitude of studies have reported different ligand-specific conformations for a given receptor, little is known about the mechanism by which different receptor conformations are connected to the capacity to activate the coupling to G-proteins. We have now performed molecular-dynamics simulations employing our recently described active-state homology model of the dopamine D2 receptor-Gαi protein-complex coupled to the partial agonists aripiprazole and FAUC350, in order to understand the structural determinants of partial agonism better. We have compared our findings with our model of the D2R-Gαi-complex in the presence of the full agonist dopamine. The two partial agonists are capable of inducing different conformations of important structural motifs, including the extracellular loop regions, the binding pocket and, in particular, intracellular G-protein-binding domains. As G-protein-coupling to certain intracellular epitopes of the receptor is considered the key step of allosterically triggered nucleotide-exchange, it is tempting to assume that impaired coupling between the receptor and the G-protein caused by distinct ligand-specific conformations is a major determinant of partial agonist efficacy. PMID:24932547

Effect of the explicit flexibility of the InhA enzyme from Mycobacterium tuberculosis in molecular docking simulations.

PubMed

Cohen, Elisangela M L; Machado, Karina S; Cohen, Marcelo; de Souza, Osmar Norberto

2011-12-22

Protein/receptor explicit flexibility has recently become an important feature of molecular docking simulations. Taking the flexibility into account brings the docking simulation closer to the receptors' real behaviour in its natural environment. Several approaches have been developed to address this problem. Among them, modelling the full flexibility as an ensemble of snapshots derived from a molecular dynamics simulation (MD) of the receptor has proved very promising. Despite its potential, however, only a few studies have employed this method to probe its effect in molecular docking simulations. We hereby use ensembles of snapshots obtained from three different MD simulations of the InhA enzyme from M. tuberculosis (Mtb), the wild-type (InhA_wt), InhA_I16T, and InhA_I21V mutants to model their explicit flexibility, and to systematically explore their effect in docking simulations with three different InhA inhibitors, namely, ethionamide (ETH), triclosan (TCL), and pentacyano(isoniazid)ferrate(II) (PIF). The use of fully-flexible receptor (FFR) models of InhA_wt, InhA_I16T, and InhA_I21V mutants in docking simulation with the inhibitors ETH, TCL, and PIF revealed significant differences in the way they interact as compared to the rigid, InhA crystal structure (PDB ID: 1ENY). In the latter, only up to five receptor residues interact with the three different ligands. Conversely, in the FFR models this number grows up to an astonishing 80 different residues. The comparison between the rigid crystal structure and the FFR models showed that the inclusion of explicit flexibility, despite the limitations of the FFR models employed in this study, accounts in a substantial manner to the induced fit expected when a protein/receptor and ligand approach each other to interact in the most favourable manner. Protein/receptor explicit flexibility, or FFR models, represented as an ensemble of MD simulation snapshots, can lead to a more realistic representation of the induced fit effect expected in the encounter and proper docking of receptors to ligands. The FFR models of InhA explicitly characterizes the overall movements of the amino acid residues in helices, strands, loops, and turns, allowing the ligand to properly accommodate itself in the receptor's binding site. Utilization of the intrinsic flexibility of Mtb's InhA enzyme and its mutants in virtual screening via molecular docking simulation may provide a novel platform to guide the rational or dynamical-structure-based drug design of novel inhibitors for Mtb's InhA. We have produced a short video sequence of each ligand (ETH, TCL and PIF) docked to the FFR models of InhA_wt. These videos are available at http://www.inf.pucrs.br/~osmarns/LABIO/Videos_Cohen_et_al_19_07_2011.htm.

Homology Models of Melatonin Receptors: Challenges and Recent Advances

PubMed Central

Pala, Daniele; Lodola, Alessio; Bedini, Annalida; Spadoni, Gilberto; Rivara, Silvia

2013-01-01

Melatonin exerts many of its actions through the activation of two G protein-coupled receptors (GPCRs), named MT1 and MT2. So far, a number of different MT1 and MT2 receptor homology models, built either from the prototypic structure of rhodopsin or from recently solved X-ray structures of druggable GPCRs, have been proposed. These receptor models differ in the binding modes hypothesized for melatonin and melatonergic ligands, with distinct patterns of ligand-receptor interactions and putative bioactive conformations of ligands. The receptor models will be described, and they will be discussed in light of the available information from mutagenesis experiments and ligand-based pharmacophore models. The ability of these ligand-receptor complexes to rationalize structure-activity relationships of known series of melatonergic compounds will be commented upon. PMID:23584026

Use of polyclonal and monoclonal antibodies to study hCG-receptor interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Milius, R.P.

1985-01-01

Although the glycoprotein hormones lutropin (LH), follitropin (FSH), and thyrotropin (TSH) bind to different receptors, each contains an identical alpha subunit. Specificity is somehow endowed by theta subunits which are distinct for each hormone. Human choriogonadotropin (hCG) is a natural LH analog that contains a beta subunit nearly identical to that of LH. The roles of these subunits in the recognition and high affinity binding of hCG to receptor was examined. Polyclonal and monoclonal antibodies specific for the individual subunits of hCG were used to probe the hormone-receptor interaction. Conformation-specific and sequence-specific antibodies were examined for their abilities to bindmore » Triton X-100-solubilized /sup 125/I-hCG-receptor complex and to inhibit hormone binding to crude rat ovarian membranes containing receptor. Even though the immunoreactive sites are not located on the receptor binding surface of the beta subunit, most, but not all, of these polyclonal and monoclonal antibodies were able to inhibit /sup 125/I-hCG binding to receptor. Although the inhibition of binding may be due to steric interference due to the size of the antibody molecules, a two-step model for hCG binding to receptor is presented that also explains these results. In this model, the beta subunit initially binds with the receptor with a highly specific but low affinity interaction. This activates a site for the high affinity binding of the alpha subunit and stabilization of the complex. This is an attractive model as it may be applied to other glycoprotein hormones sharing an alpha subunit.« less

Ligand-induced type II interleukin-4 receptor dimers are sustained by rapid re-association within plasma membrane microcompartments

NASA Astrophysics Data System (ADS)

Richter, David; Moraga, Ignacio; Winkelmann, Hauke; Birkholz, Oliver; Wilmes, Stephan; Schulte, Markos; Kraich, Michael; Kenneweg, Hella; Beutel, Oliver; Selenschik, Philipp; Paterok, Dirk; Gavutis, Martynas; Schmidt, Thomas; Garcia, K. Christopher; Müller, Thomas D.; Piehler, Jacob

2017-07-01

The spatiotemporal organization of cytokine receptors in the plasma membrane is still debated with models ranging from ligand-independent receptor pre-dimerization to ligand-induced receptor dimerization occurring only after receptor uptake into endosomes. Here, we explore the molecular and cellular determinants governing the assembly of the type II interleukin-4 receptor, taking advantage of various agonists binding the receptor subunits with different affinities and rate constants. Quantitative kinetic studies using artificial membranes confirm that receptor dimerization is governed by the two-dimensional ligand-receptor interactions and identify a critical role of the transmembrane domain in receptor dimerization. Single molecule localization microscopy at physiological cell surface expression levels, however, reveals efficient ligand-induced receptor dimerization by all ligands, largely independent of receptor binding affinities, in line with the similar STAT6 activation potencies observed for all IL-4 variants. Detailed spatiotemporal analyses suggest that kinetic trapping of receptor dimers in actin-dependent microcompartments sustains robust receptor dimerization and signalling.

Synergistically increased ILC2 and Th9 cells in lung tissue jointly promote the pathological process of asthma in mice.

PubMed

Ying, Xinyu; Su, Zhaoliang; Bie, Qingli; Zhang, Pan; Yang, Huijian; Wu, Yumin; Xu, Yunyun; Wu, Jing; Zhang, Mengying; Wang, Shengjun; Xu, Huaxi

2016-06-01

In recent years, T helper (Th) 9 cells have been demonstrated to be key mediators in immune responses in asthmatic lungs, and innate lymphoid cells 2 (ILC2s) have been described as a novel type of innate immunocyte with the ability to enhance immunoglobulin E (IgE) production. However, the interaction between ILC2s and Th9 cells in the pulmonary system of a mouse model of asthma remains to be elucidated. In the present study, the response state of lung tissue with regards to Th9 and ILC2s in a mouse model of asthma was investigated by detecting Th9‑ and ILC2‑associated cytokine receptors. The present study also investigated the association between the expression levels of the cytokine receptors in lung tissue samples and the IgE levels in sera samples from mouse models of asthma. Results from the present study demonstrated that the frequency of ILC2s and Th9 cells was significantly increased in the lung tissue samples, indicating that a Th2-type immune response had occurred. In addition, high mRNA expression levels of RAR‑related orphan receptor α, interleukin 1 receptor‑like 1, transcription factor PU.1 and interleukin (IL)‑9 were observed. Furthermore, IL‑5Rα, IL‑13Rα2 and high‑affinity IgE receptor were increased in mouse models of asthma, and a positive association was observed between the expression levels of ILC2‑ or Th9‑associated receptors in tissue samples and IgE levels in the sera. This indicated that ILC2s and Th9 were in a state of polarization and may promote each other in the lung tissue of mouse models of asthma, and that the lung tissue was responding to the two types of cells via increased expression of receptors.

MOLECULAR MODELING FOR PRIORITIZATION OF TOXICITY BIOASSAYS

EPA Science Inventory

The results of this study demonstrate the importance of receptor flexibility. However, when different receptors in the endocrine system are considered simultaneously most molecules, even weak environmental agents, are classified correctly. The ultimate goal of this research is ...

Docking model of the nicotinic acetylcholine receptor and nitromethylene neonicotinoid derivatives with a longer chiral substituent and their biological activities.

PubMed

Nagaoka, Hikaru; Nishiwaki, Hisashi; Kubo, Takuya; Akamatsu, Miki; Yamauchi, Satoshi; Shuto, Yoshihiro

2015-02-15

In the present study, nitromethylene neonicotinoid derivatives possessing substituents that contain a sulfur atom, oxygen atom or aromatic ring at position 5 on the imidazolidine ring were synthesized to evaluate their affinity for the nicotinic acetylcholine receptor (nAChR) and their insecticidal activity against adult female houseflies. Comparing the receptor affinity of the alkylated derivative with the receptor affinity of compounds possessing either ether or thioether groups revealed that conversion of the carbon atom to a sulfur atom did not influence the receptor affinity, whereas conversion to an oxygen atom was disadvantageous for the receptor affinity. The receptor affinity of compounds possessing a benzyl or phenyl group was lower than that of the unsubstituted compound. Analysis of the three-dimensional quantitative structure-activity relationship using comparative molecular field analysis demonstrated that steric hindrance of the receptor should exist around the C3 of an n-butyl group attached at position 5 on the imidazolidine ring. A docking study of the nAChR-ligand model suggested that the ligand-binding region expands as the length of the substituent increases by brushing against the amino acids that form the binding region. The insecticidal activity of the compounds was positively correlated with the receptor affinity by considering logP and the number of heteroatoms, including sulfur and oxygen atoms, in the substituents, suggesting that the insecticidal activity is influenced by the receptor affinity, hydrophobicity, and metabolic stability of the compounds. Copyright © 2015 Elsevier Ltd. All rights reserved.

An efficient model of human endometriosis by induced unopposed estrogenicity in baboons.

PubMed

Nair, Hareesh B; Baker, Robert; Owston, Michael A; Escalona, Renee; Dick, Edward J; VandeBerg, John L; Nickisch, Klaus J

2016-03-08

Endometriosis is a chronic estrogen-dependent disease that occurs in approximately 10% of reproductive age women. Baboons offer a clear benefit for studying the initiation and progression of endometriosis since baboon is very close to humans phylogenetically. Progestins are used in the treatment of endometriosis. The therapeutic window of progestins depends on the ratio of its affinity towards progesterone receptor agonism verses antagonism. The present study is to determine the role of pure antiprogestin in baboon endometriosis. We hypothesize that pure antiprogestin will induce unopposed estrogenicity and spontaneous endometriosis in baboons. The rate of endometrial invasion and attachment through modeled peritoneum in the presence and absence of progesterone and antiprogestin was evaluated in this study. A baboon model of endometriosis induced by unopposed estrogenicity using progesterone receptor antagonist (EC304) was used in this study. We observed EC304 has induced unopposed estrogenicity that deregulated proteins involved in attachment, invasion, cell growth, and steroid hormone receptors in this model. Our data suggest that depleting progesterone levels in the endometrium will increase estrogen hyper-responsiveness that leads to increased endometriotic lesion progression in the baboon (Papio anubis) model. This study reports a refined model of human endometriosis in baboons that could potentially be used to develop new diagnostic and therapeutic strategies for the benefit of women suffering from endometriosis.

GPR17: molecular modeling and dynamics studies of the 3-D structure and purinergic ligand binding features in comparison with P2Y receptors.

PubMed

Parravicini, Chiara; Ranghino, Graziella; Abbracchio, Maria P; Fantucci, Piercarlo

2008-06-04

GPR17 is a G-protein-coupled receptor located at intermediate phylogenetic position between two distinct receptor families: the P2Y and CysLT receptors for extracellular nucleotides and cysteinyl-LTs, respectively. We previously showed that GPR17 can indeed respond to both classes of endogenous ligands and to synthetic compounds active at the above receptor families, thus representing the first fully characterized non-peptide "hybrid" GPCR. In a rat brain focal ischemia model, the selective in vivo knock down of GPR17 by anti-sense technology or P2Y/CysLT antagonists reduced progression of ischemic damage, thus highlighting GPR17 as a novel therapeutic target for stroke. Elucidation of the structure of GPR17 and of ligand binding mechanisms are the necessary steps to obtain selective and potent drugs for this new potential target. On this basis, a 3-D molecular model of GPR17 embedded in a solvated phospholipid bilayer and refined by molecular dynamics simulations has been the first aim of this study. To explore the binding mode of the "purinergic" component of the receptor, the endogenous agonist UDP and two P2Y receptor antagonists demonstrated to be active on GPR17 (MRS2179 and cangrelor) were then modeled on the receptor. Molecular dynamics simulations suggest that GPR17 nucleotide binding pocket is similar to that described for the other P2Y receptors, although only one of the three basic residues that have been typically involved in ligand recognition is conserved (Arg255). The binding pocket is enclosed between the helical bundle and covered at the top by EL2. Driving interactions are H-bonds and salt bridges between the 6.55 and 6.52 residues and the phosphate moieties of the ligands. An "accessory" binding site in a region formed by the EL2, EL3 and the Nt was also found. Nucleotide binding to GPR17 occurs on the same receptor regions identified for already known P2Y receptors. Agonist/antagonist binding mode are similar, but not identical. An accessory external binding site could guide small ligands to the deeper principal binding site in a multi-step mechanism of activation. The nucleotide binding pocket appears to be unable to allocate the leukotrienic type ligands in the same effective way.

Receptor modeling for source apportionment of polycyclic aromatic hydrocarbons in urban atmosphere.

PubMed

Singh, Kunwar P; Malik, Amrita; Kumar, Ranjan; Saxena, Puneet; Sinha, Sarita

2008-01-01

This study reports source apportionment of polycyclic aromatic hydrocarbons (PAHs) in particulate depositions on vegetation foliages near highway in the urban environment of Lucknow city (India) using the principal components analysis/absolute principal components scores (PCA/APCS) receptor modeling approach. The multivariate method enables identification of major PAHs sources along with their quantitative contributions with respect to individual PAH. The PCA identified three major sources of PAHs viz. combustion, vehicular emissions, and diesel based activities. The PCA/APCS receptor modeling approach revealed that the combustion sources (natural gas, wood, coal/coke, biomass) contributed 19-97% of various PAHs, vehicular emissions 0-70%, diesel based sources 0-81% and other miscellaneous sources 0-20% of different PAHs. The contributions of major pyrolytic and petrogenic sources to the total PAHs were 56 and 42%, respectively. Further, the combustion related sources contribute major fraction of the carcinogenic PAHs in the study area. High correlation coefficient (R2 > 0.75 for most PAHs) between the measured and predicted concentrations of PAHs suggests for the applicability of the PCA/APCS receptor modeling approach for estimation of source contribution to the PAHs in particulates.

Isothermal titration calorimetry study of the interaction of sweeteners with fullerenols as an artificial sweet taste receptor model.

PubMed

Chen, Zhong-Xiu; Guo, Gang-Min; Deng, Shao-Ping

2009-04-08

A fullerenol-based synthetic sweetness receptor model, consisting of polyhydroxy groups for potential hydrogen bond donor along with a spherical hydrophobic center, was proposed according to the widely accepted sweetness hypothesis. An isothermal titration calorimetry (ITC) technique was used to study mimetic interaction of this sweet receptor model with a series of sweeteners having increasing sweetness intensity. The results showed that ITC is an effective method to provide thorough and precise characterization of the energies of molecular complex formation. Binding of all of the studied sweeteners with fullerenols was found through two sets of site models. More heat was released from sweeter synthetic compounds binding with fullerenols than from less sweet carbohydrates. The results imply that hydrogen bond formation is necessary for the sweeteners to bind to the fullerenol receptor in the first stage, whereas hydrophobic effect and conformation changes that lead to favorable entropy changes occur in most cases. The preliminary results of this study help to cover the lack of information about the thermodynamic basis of understanding of the initiation of the sweet sensation. It also adds complementary physicochemical measurements available for comparison with the sweetness hypothesis. On the other hand, a correlation between the thermodynamic parameters and sweetness intensity has been made as well, which exhibits potential as a useful tool in sensory analysis.

Simvastatin Prevents Dopaminergic Neurodegeneration in Experimental Parkinsonian Models: The Association with Anti-Inflammatory Responses

PubMed Central

Zhang, Limin; Wu, Aimin; Yang, Yu; Xiong, Zhaojun; Deng, Chao; Huang, Xu-Feng; Yenari, Midori A.; Yang, Yuan-Guo; Ying, Weihai; Wang, Qing

2011-01-01

Background In addition to their original applications to lowering cholesterol, statins display multiple neuroprotective effects. N-methyl-D-aspartate (NMDA) receptors interact closely with the dopaminergic system and are strongly implicated in therapeutic paradigms of Parkinson's disease (PD). This study aims to investigate how simvastatin impacts on experimental parkinsonian models via regulating NMDA receptors. Methodology/Principal Findings Regional changes in NMDA receptors in the rat brain and anxiolytic-like activity were examined after unilateral medial forebrain bundle lesion by 6-hydroxydopamine via a 3-week administration of simvastatin. NMDA receptor alterations in the post-mortem rat brain were detected by [3H]MK-801(Dizocilpine) binding autoradiography. 6-hydroxydopamine treated PC12 was applied to investigate the neuroprotection of simvastatin, the association with NMDA receptors, and the anti-inflammation. 6-hydroxydopamine induced anxiety and the downregulation of NMDA receptors in the hippocampus, CA1(Cornu Ammonis 1 Area), amygdala and caudate putamen was observed in 6-OHDA(6-hydroxydopamine) lesioned rats whereas simvastatin significantly ameliorated the anxiety-like activity and restored the expression of NMDA receptors in examined brain regions. Significant positive correlations were identified between anxiolytic-like activity and the restoration of expression of NMDA receptors in the hippocampus, amygdala and CA1 following simvastatin administration. Simvastatin exerted neuroprotection in 6-hydroxydopamine-lesioned rat brain and 6-hydroxydopamine treated PC12, partially by regulating NMDA receptors, MMP9 (matrix metalloproteinase-9), and TNF-a (tumour necrosis factor-alpha). Conclusions/Significance Our results provide strong evidence that NMDA receptor modulation after simvastatin treatment could partially explain its anxiolytic-like activity and anti-inflammatory mechanisms in experimental parkinsonian models. These findings contribute to a better understanding of the critical roles of simvastatin in treating PD via NMDA receptors. PMID:21731633

Binding Interactions of Dopamine and Apomorphine in D2High and D2Low States of Human Dopamine D2 Receptor Using Computational and Experimental Techniques.

PubMed

Durdagi, Serdar; Salmas, Ramin Ekhteiari; Stein, Matthias; Yurtsever, Mine; Seeman, Philip

2016-02-17

We have recently reported G-protein coupled receptor (GPCR) model structures for the active and inactive states of the human dopamine D2 receptor (D2R) using adrenergic crystal structures as templates. Since the therapeutic concentrations of dopamine agonists that suppress the release of prolactin are the same as those that act at the high-affinity state of the D2 receptor (D2High), D2High in the anterior pituitary gland is considered to be the functional state of the receptor. In addition, the therapeutic concentrations of anti-Parkinson drugs are also related to the dissociation constants in the D2High form of the receptor. The discrimination between the high- and low-affinity (D2Low) components of the D2R is not obvious and requires advanced computer-assisted structural biology investigations. Therefore, in this work, the derived D2High and D2Low receptor models (GPCR monomer and dimer three-dimensional structures) are used as drug-binding targets to investigate binding interactions of dopamine and apomorphine. The study reveals a match between the experimental dissociation constants of dopamine and apomorphine at their high- and low-affinity sites of the D2 receptor in monomer and dimer and their calculated dissociation constants. The allosteric receptor-receptor interaction for dopamine D2R dimer is associated with the accessibility of adjacent residues of transmembrane region 4. The measured negative cooperativity between agonist ligand at dopamine D2 receptor is also correctly predicted using the D2R homodimerization model.

Activation of spinal and supraspinal cannabinoid-1 receptors lead to antinociception in a rat model of neuropathic spinal cord injury pain

PubMed Central

Hama, Aldric; Sagen, Jacqueline

2011-01-01

Activation of CNS cannabinoid subtype-1 (CB1) receptors has been shown to mediate the antinociceptive and other effects of systemically administered CB receptor agonists. The endogenous peptide CB receptor ligand hemopressin (HE) has previously demonstrated an antinociceptive effect in rats with a hind paw inflammation, without exhibiting characteristic CB1 receptor-mediated side-effects. The current study evaluated the effect of intrathecal (i.t.) and intracerebroventricular (i.c.v.) injection of HE in a rat model of neuropathic spinal cord injury (SCI) pain. The non-subtype selective CB receptor agonist WIN 55,212-2 was also centrally administered in SCI rats as a comparator. Four weeks following an acute compression of the mid-thoracic spinal cord, rats displayed markedly decreased hind paw withdrawal thresholds, indicative of below-level neuropathic pain. Central administration of WIN 55,212-2 significantly increased withdrawal thresholds, whereas HE did not. Hemopressin has been reported to block CB1 receptors in vitro, similar to the CB1 receptor antagonist rimonabant. Pretreatment with rimonabant completely blocked the antinociceptive effect of centrally administered WIN 55,212-2, but pretreatment with HE did not. While the data confirm that activation of either supraspinal or spinal CB1 receptors leads to significant antinociception in SCI rats, the current data do not support an antinociceptive effect from an acute blockade of central CB1 receptors, HE’s putative antinociceptive mechanism, in neuropathic SCI rats. Although such a mechanism could be useful in other models of pain with a significant inflammatory component, the current data indicate that activation of CB1 receptors is needed to ameliorate neuropathic SCI pain. PMID:21813113

Simvastatin prevents dopaminergic neurodegeneration in experimental parkinsonian models: the association with anti-inflammatory responses.

PubMed

Yan, Junqiang; Xu, Yunqi; Zhu, Cansheng; Zhang, Limin; Wu, Aimin; Yang, Yu; Xiong, Zhaojun; Deng, Chao; Huang, Xu-Feng; Yenari, Midori A; Yang, Yuan-Guo; Ying, Weihai; Wang, Qing

2011-01-01

In addition to their original applications to lowering cholesterol, statins display multiple neuroprotective effects. N-methyl-D-aspartate (NMDA) receptors interact closely with the dopaminergic system and are strongly implicated in therapeutic paradigms of Parkinson's disease (PD). This study aims to investigate how simvastatin impacts on experimental parkinsonian models via regulating NMDA receptors. Regional changes in NMDA receptors in the rat brain and anxiolytic-like activity were examined after unilateral medial forebrain bundle lesion by 6-hydroxydopamine via a 3-week administration of simvastatin. NMDA receptor alterations in the post-mortem rat brain were detected by [³H]MK-801(Dizocilpine) binding autoradiography. 6-hydroxydopamine treated PC12 was applied to investigate the neuroprotection of simvastatin, the association with NMDA receptors, and the anti-inflammation. 6-hydroxydopamine induced anxiety and the downregulation of NMDA receptors in the hippocampus, CA1(Cornu Ammonis 1 Area), amygdala and caudate putamen was observed in 6-OHDA(6-hydroxydopamine) lesioned rats whereas simvastatin significantly ameliorated the anxiety-like activity and restored the expression of NMDA receptors in examined brain regions. Significant positive correlations were identified between anxiolytic-like activity and the restoration of expression of NMDA receptors in the hippocampus, amygdala and CA1 following simvastatin administration. Simvastatin exerted neuroprotection in 6-hydroxydopamine-lesioned rat brain and 6-hydroxydopamine treated PC12, partially by regulating NMDA receptors, MMP9 (matrix metalloproteinase-9), and TNF-a (tumour necrosis factor-alpha). Our results provide strong evidence that NMDA receptor modulation after simvastatin treatment could partially explain its anxiolytic-like activity and anti-inflammatory mechanisms in experimental parkinsonian models. These findings contribute to a better understanding of the critical roles of simvastatin in treating PD via NMDA receptors.

A cross-laboratory preclinical study on the effectiveness of interleukin-1 receptor antagonist in stroke

PubMed Central

Maysami, Samaneh; Wong, Raymond; Pradillo, Jesus M; Denes, Adam; Dhungana, Hiramani; Malm, Tarja; Koistinaho, Jari; Orset, Cyrille; Rahman, Mahbubur; Rubio, Marina; Schwaninger, Markus; Vivien, Denis; Bath, Philip M; Rothwell, Nancy J

2015-01-01

Stroke represents a global challenge and is a leading cause of permanent disability worldwide. Despite much effort, translation of research findings to clinical benefit has not yet been successful. Failure of neuroprotection trials is considered, in part, due to the low quality of preclinical studies, low level of reproducibility across different laboratories and that stroke co-morbidities have not been fully considered in experimental models. More rigorous testing of new drug candidates in different experimental models of stroke and initiation of preclinical cross-laboratory studies have been suggested as ways to improve translation. However, to our knowledge, no drugs currently in clinical stroke trials have been investigated in preclinical cross-laboratory studies. The cytokine interleukin 1 is a key mediator of neuronal injury, and the naturally occurring interleukin 1 receptor antagonist has been reported as beneficial in experimental studies of stroke. In the present paper, we report on a preclinical cross-laboratory stroke trial designed to investigate the efficacy of interleukin 1 receptor antagonist in different research laboratories across Europe. Our results strongly support the therapeutic potential of interleukin 1 receptor antagonist in experimental stroke and provide further evidence that interleukin 1 receptor antagonist should be evaluated in more extensive clinical stroke trials. PMID:26661169

Detection of angiotensin II binding to single adrenal zona glomerulosa cells by confocal Raman microspectroscopy

NASA Astrophysics Data System (ADS)

McCoy, Michael J.; Habermann, Timothy J.; Hanke, Craig J.; Adar, Fran; Campbell, William B.; Nithipatikom, Kasem

1999-04-01

We developed a confocal Raman microspectroscopic technique to study ligand-receptor bindings in single cells using Raman-labeled ligands and surface-enhanced Raman scattering (SERS). The adrenal zona glomerulosa (ZG) cells were used as a model in this study. ZG cells have a high density of angiotensin II (AII) receptors on the cellular membrane. There are two identified subtypes of AII receptors,namely AT1 and AT2 receptors. AII is a peptidic hormone, which upon binding to its receptors, stimulates the release of aldosterone from ZG cells. The cellular localization of these receptors subtypes was detected in single ZG cells by using immunocomplexation of receptors with specific antibodies and confocal Raman microspectroscopy. In the binding study, we used biotin-labeled AII to bind to its receptors in ZG cells. Then, avidin and Raman-labeled AII. The binding was measure directly on the single ZG cells. The results showed that the binding was displaced with unlabeled AII and specific AII antagonists. This is a rapid and sensitive technique for detection of cellular ligand bindings as well as antagonists screening in drug discovery.

Prediction of consensus binding mode geometries for related chemical series of positive allosteric modulators of adenosine and muscarinic acetylcholine receptors.

PubMed

Sakkal, Leon A; Rajkowski, Kyle Z; Armen, Roger S

2017-06-05

Following insights from recent crystal structures of the muscarinic acetylcholine receptor, binding modes of Positive Allosteric Modulators (PAMs) were predicted under the assumption that PAMs should bind to the extracellular surface of the active state. A series of well-characterized PAMs for adenosine (A 1 R, A 2A R, A 3 R) and muscarinic acetylcholine (M 1 R, M 5 R) receptors were modeled using both rigid and flexible receptor CHARMM-based molecular docking. Studies of adenosine receptors investigated the molecular basis of the probe-dependence of PAM activity by modeling in complex with specific agonist radioligands. Consensus binding modes map common pharmacophore features of several chemical series to specific binding interactions. These models provide a rationalization of how PAM binding slows agonist radioligand dissociation kinetics. M 1 R PAMs were predicted to bind in the analogous M 2 R PAM LY2119620 binding site. The M 5 R NAM (ML-375) was predicted to bind in the PAM (ML-380) binding site with a unique induced-fit receptor conformation. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Assessment and Challenges of Ligand Docking into Comparative Models of G-Protein Coupled Receptors

PubMed Central

Frimurer, Thomas M.; Meiler, Jens

2013-01-01

The rapidly increasing number of high-resolution X-ray structures of G-protein coupled receptors (GPCRs) creates a unique opportunity to employ comparative modeling and docking to provide valuable insight into the function and ligand binding determinants of novel receptors, to assist in virtual screening and to design and optimize drug candidates. However, low sequence identity between receptors, conformational flexibility, and chemical diversity of ligands present an enormous challenge to molecular modeling approaches. It is our hypothesis that rapid Monte-Carlo sampling of protein backbone and side-chain conformational space with Rosetta can be leveraged to meet this challenge. This study performs unbiased comparative modeling and docking methodologies using 14 distinct high-resolution GPCRs and proposes knowledge-based filtering methods for improvement of sampling performance and identification of correct ligand-receptor interactions. On average, top ranked receptor models built on template structures over 50% sequence identity are within 2.9 Å of the experimental structure, with an average root mean square deviation (RMSD) of 2.2 Å for the transmembrane region and 5 Å for the second extracellular loop. Furthermore, these models are consistently correlated with low Rosetta energy score. To predict their binding modes, ligand conformers of the 14 ligands co-crystalized with the GPCRs were docked against the top ranked comparative models. In contrast to the comparative models themselves, however, it remains difficult to unambiguously identify correct binding modes by score alone. On average, sampling performance was improved by 103 fold over random using knowledge-based and energy-based filters. In assessing the applicability of experimental constraints, we found that sampling performance is increased by one order of magnitude for every 10 residues known to contact the ligand. Additionally, in the case of DOR, knowledge of a single specific ligand-protein contact improved sampling efficiency 7 fold. These findings offer specific guidelines which may lead to increased success in determining receptor-ligand complexes. PMID:23844000

CoMSIA and Docking Study of Rhenium Based Estrogen Receptor Ligand Analogs

PubMed Central

Wolohan, Peter; Reichert, David E.

2007-01-01

OPLS all atom force field parameters were developed in order to model a diverse set of novel rhenium based estrogen receptor ligands whose relative binding affinities (RBA) to the estrogen receptor alpha isoform (ERα) with respect to 17β-Estradiol were available. The binding properties of these novel rhenium based organometallic complexes were studied with a combination of Comparative Molecular Similarity Indices Analysis (CoMSIA) and docking. A total of 29 estrogen receptor ligands consisting of 11 rhenium complexes and 18 organic ligands were docked inside the ligand-binding domain (LBD) of ERα utilizing the program Gold. The top ranked pose was used to construct CoMSIA models from a training set of 22 of the estrogen receptor ligands which were selected at random. In addition scoring functions from the docking runs and the polar volume (PV) were also studied to investigate their ability to predict RBA ERα. A partial least-squares analysis consisting of the CoMSIA steric, electrostatic and hydrophobic indices together with the polar volume proved sufficiently predictive having a correlation coefficient, r2, of 0.94 and a cross-validated correlation coefficient, q2, utilizing the leave one out method of 0.68. Analysis of the scoring functions from Gold showed particularly poor correlation to RBA ERα which did not improve when the rhenium complexes were extracted to leave the organic ligands. The combined CoMSIA and polar volume model ranked correctly the ligands in order of increasing RBA ERα, illustrating the utility of this method as a prescreening tool in the development of novel rhenium based estrogen receptor ligands. PMID:17280694

Modelling the interdependence between the stoichiometry of receptor oligomerization and ligand binding for a coexisting dimer/tetramer receptor system.

PubMed

Rovira, X; Vivó, M; Serra, J; Roche, D; Strange, P G; Giraldo, J

2009-01-01

Many G protein-coupled receptors have been shown to exist as oligomers, but the oligomerization state and the effects of this on receptor function are unclear. For some G protein-coupled receptors, in ligand binding assays, different radioligands provide different maximal binding capacities. Here we have developed mathematical models for co-expressed dimeric and tetrameric species of receptors. We have considered models where the dimers and tetramers are in equilibrium and where they do not interconvert and we have also considered the potential influence of the ligands on the degree of oligomerization. By analogy with agonist efficacy, we have considered ligands that promote, inhibit or have no effect on oligomerization. Cell surface receptor expression and the intrinsic capacity of receptors to oligomerize are quantitative parameters of the equations. The models can account for differences in the maximal binding capacities of radioligands in different preparations of receptors and provide a conceptual framework for simulation and data fitting in complex oligomeric receptor situations.

Modeling Interactions among Individual P2 Receptors to Explain Complex Response Patterns over a Wide Range of ATP Concentrations

PubMed Central

Xing, Shu; Grol, Matthew W.; Grutter, Peter H.; Dixon, S. Jeffrey; Komarova, Svetlana V.

2016-01-01

Extracellular ATP acts on the P2X family of ligand-gated ion channels and several members of the P2Y family of G protein-coupled receptors to mediate intercellular communication among many cell types including bone-forming osteoblasts. It is known that multiple P2 receptors are expressed on osteoblasts (P2X2,5,6,7 and P2Y1,2,4,6). In the current study, we investigated complex interactions within the P2 receptor network using mathematical modeling. To characterize individual P2 receptors, we extracted data from published studies of overexpressed human and rodent (rat and mouse) receptors and fit their dependencies on ATP concentration using the Hill equation. Next, we examined responses induced by an ensemble of endogenously expressed P2 receptors. Murine osteoblastic cells (MC3T3-E1 cells) were loaded with fluo-4 and stimulated with varying concentrations of extracellular ATP. Elevations in the concentration of cytosolic free calcium ([Ca2+]i) were monitored by confocal microscopy. Dependence of the calcium response on ATP concentration exhibited a complex pattern that was not explained by the simple addition of individual receptor responses. Fitting the experimental data with a combination of Hill equations from individual receptors revealed that P2Y1 and P2X7 mediated the rise in [Ca2+]i at very low and high ATP concentrations, respectively. Interestingly, to describe responses at intermediate ATP concentrations, we had to assume that a receptor with a K1∕2 in that range (e.g. P2Y4 or P2X5) exerts an inhibitory effect. This study provides new insights into the interactions among individual P2 receptors in producing an ensemble response to extracellular ATP. PMID:27468270

PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR (PPAR) AGONISTS AS PROMISING NEW MEDICATIONS FOR DRUG ADDICTION: PRECLINICAL EVIDENCE

PubMed Central

Foll, Bernard Le; Ciano, Patricia Di; Panlilio, Leigh V.; Goldberg, Steven R.; Ciccocioppo, Roberto

2013-01-01

This review examines the growing literature on the role of peroxisome proliferator-activated receptors (PPARs) in addiction. There are two subtypes of PPAR receptors that have been studied in addiction: PPAR-α and PPAR-γ. The role of each PPAR subtype in common models of addictive behavior, mainly pre-clinical models, is summarized. In particular, studies are reviewed that investigated the effects of PPAR-α agonists on relapse, sensitization, conditioned place preference, withdrawal and drug intake, and effects of PPAR-γ agonists on relapse, withdrawal and drug intake. Finally, studies that investigated the effects of PPAR agonists on neural pathways of addiction are reviewed. Taken together this preclinical data indicates that PPAR agonists are promising new medications for drug addiction treatment. PMID:23614675

An update on the role of the 5-hydroxytryptamine6 receptor in cognitive function.

PubMed

Fone, Kevin C F

2008-11-01

As the 5-hydroxytryptamine(6) (5-HT(6)) receptor is almost exclusively expressed in the CNS, particularly in areas associated with learning and memory, many studies have examined its role in cognitive function in the rodent, as reviewed herein. Most studies, in healthy adult rats, report that 5-HT(6) receptor antagonists enhance retention of spatial learning in the Morris water maze, improve consolidation in autoshaping tasks and reverse natural forgetting in object recognition. Antagonists appear to facilitate both cholinergic and glutamatergic neurotransmission, reversing scopolamine- and NMDA receptor antagonist-induced memory impairments. Recent reports show that the 5-HT(6) receptor antagonist, PRX-07034, restores the impairment of novel object recognition produced in rats reared in social isolation, a neurodevelopmental model producing behavioural changes similar to several core symptoms seen in schizophrenia. The 5-HT(6) receptor antagonist, Ro 04-6790, modestly improved reversal learning in isolation reared but not group-housed controls in the water maze. Ro 04-6790 also improved novel object discrimination both in adult rats that received chronic intermittent phencyclidine and drug-naïve 18-month-old rats. However, more information on their effect in animal models of schizophrenia and Alzheimer's disease is required. Several selective high-affinity 5-HT(6) receptor agonists developed recently also improve object discrimination and extra-dimensional set-shifting behaviour. Thus both 5-HT(6) receptor agonist and antagonist compounds show promise as pro-cognitive agents in pre-clinical studies but the explanation for their paradoxical analogous effect is currently unclear, and is discussed in this article.

Preliminary Results of the first European Source Apportionment intercomparison for Receptor and Chemical Transport Models

NASA Astrophysics Data System (ADS)

Belis, Claudio A.; Pernigotti, Denise; Pirovano, Guido

2017-04-01

Source Apportionment (SA) is the identification of ambient air pollution sources and the quantification of their contribution to pollution levels. This task can be accomplished using different approaches: chemical transport models and receptor models. Receptor models are derived from measurements and therefore are considered as a reference for primary sources urban background levels. Chemical transport model have better estimation of the secondary pollutants (inorganic) and are capable to provide gridded results with high time resolution. Assessing the performance of SA model results is essential to guarantee reliable information on source contributions to be used for the reporting to the Commission and in the development of pollution abatement strategies. This is the first intercomparison ever designed to test both receptor oriented models (or receptor models) and chemical transport models (or source oriented models) using a comprehensive method based on model quality indicators and pre-established criteria. The target pollutant of this exercise, organised in the frame of FAIRMODE WG 3, is PM10. Both receptor models and chemical transport models present good performances when evaluated against their respective references. Both types of models demonstrate quite satisfactory capabilities to estimate the yearly source contributions while the estimation of the source contributions at the daily level (time series) is more critical. Chemical transport models showed a tendency to underestimate the contribution of some single sources when compared to receptor models. For receptor models the most critical source category is industry. This is probably due to the variety of single sources with different characteristics that belong to this category. Dust is the most problematic source for Chemical Transport Models, likely due to the poor information about this kind of source in the emission inventories, particularly concerning road dust re-suspension, and consequently the little detail about the chemical components of this source used in the models. The sensitivity tests show that chemical transport models show better performances when displaying a detailed set of sources (14) than when using a simplified one (only 8). It was also observed that an enhanced vertical profiling can improve the estimation of specific sources, such as industry, under complex meteorological conditions and that an insufficient spatial resolution in urban areas can impact on the capabilities of models to estimate the contribution of diffuse primary sources (e.g. traffic). Both families of models identify traffic and biomass burning as the first and second most contributing categories, respectively, to elemental carbon. The results of this study demonstrate that the source apportionment assessment methodology developed by the JRC is applicable to any kind of SA model. The same methodology is implemented in the on-line DeltaSA tool to support source apportionment model evaluation (http://source-apportionment.jrc.ec.europa.eu/).

Structure-activity relationships for serotonin transporter and dopamine receptor selectivity.

PubMed

Agatonovic-Kustrin, Snezana; Davies, Paul; Turner, Joseph V

2009-05-01

Antipsychotic medications have a diverse pharmacology with affinity for serotonergic, dopaminergic, adrenergic, histaminergic and cholinergic receptors. Their clinical use now also includes the treatment of mood disorders, thought to be mediated by serotonergic receptor activity. The aim of our study was to characterise the molecular properties of antipsychotic agents, and to develop a model that would indicate molecular specificity for the dopamine (D(2)) receptor and the serotonin (5-HT) transporter. Back-propagation artificial neural networks (ANNs) were trained on a dataset of 47 ligands categorically assigned antidepressant or antipsychotic utility. The structure of each compound was encoded with 63 calculated molecular descriptors. ANN parameters including hidden neurons and input descriptors were optimised based on sensitivity analyses, with optimum models containing between four and 14 descriptors. Predicted binding preferences were in excellent agreement with clinical antipsychotic or antidepressant utility. Validated models were further tested by use of an external prediction set of five drugs with unknown mechanism of action. The SAR models developed revealed the importance of simple molecular characteristics for differential binding to the D(2) receptor and the 5-HT transporter. These included molecular size and shape, solubility parameters, hydrogen donating potential, electrostatic parameters, stereochemistry and presence of nitrogen. The developed models and techniques employed are expected to be useful in the rational design of future therapeutic agents.

Caffeinol at the receptor level; Anti-ischemic effect of NMDA receptor blockade is potentiated by caffeine

PubMed Central

Zhao, Xiurong; Strong, Roger; Piriyawat, Paisith; Palusinski, Robert; Grotta, James C.; Aronowski, Jaroslaw

2010-01-01

Background and Purpose Although caffeinol (combination of low dose of caffeine and ethanol) was shown to robustly reduce stroke damage in experimental models and is now in clinical evaluation for treatment of ischemic stroke, little is known about the potential mechanism of its action. Methods We have used an in vivo excitotoxicity model based on intracortical infusion of NMDA and model of reversible focal ischemia to demonstrate NMDA receptor inhibition as one potential mechanism of Caffeinol anti-ischemic activity. Results Caffeinol reduced the size of excitotoxic lesion and substitution of ethanol in Caffeinol with CNS-1102 and MK-801, but not with MgSO4, produced treatment with strong synergistic effect that was at least as robust in reducing ischemic damage as Caffeinol. This NMDA receptor antagonist and caffeine combination showed long window of opportunity, activity in spontaneously hypertensive rats, and unlike Caffeinol was fully effective in animals chronically pre-treated with ethanol. Conclusions Our study suggests that anti-excitotoxic properties may underlie some of the anti-ischemic effect of Caffeinol. This study provides strong evidence that the anti-ischemic effect of NMDA receptor blockers in general can be dramatically augmented by caffeine, thus opening a possibility for new utilization of NMDA-based pharmacology in the treatment of stroke. PMID:20044532

Painful purinergic receptors.

PubMed

Donnelly-Roberts, Diana; McGaraughty, Steve; Shieh, Char-Chang; Honore, Prisca; Jarvis, Michael F

2008-02-01

Multiple P2 receptor-mediated mechanisms exist by which ATP can alter nociceptive sensitivity following tissue injury. Evidence from a variety of experimental strategies, including genetic disruption studies and the development of selective antagonists, has indicated that the activation of P2X receptor subtypes, including P2X(3), P2X(2/3), P2X(4) and P2X(7), and P2Y (e.g., P2Y(2)) receptors, can modulate pain. For example, administration of a selective P2X(3) antagonist, A-317491, has been shown to effectively block both hyperalgesia and allodynia in different animal models of pathological pain. Intrathecally delivered antisense oligonucleotides targeting P2X(4) receptors decrease tactile allodynia following nerve injury. Selective antagonists for the P2X(7) receptor also reduce sensitization in animal models of inflammatory and neuropathic pain, providing evidence that purinergic glial-neural interactions are important modulators of noxious sensory neurotransmission. Furthermore, activation of P2Y(2) receptors leads to sensitization of polymodal transient receptor potential-1 receptors. Thus, ATP acting at multiple purinergic receptors, either directly on neurons (e.g., P2X(3), P2X(2/3), and P2Y receptors) or indirectly through neural-glial cell interactions (P2X(4) and P2X(7) receptors), alters nociceptive sensitivity. The development of selective antagonists for some of these P2 receptors has greatly aided investigations into the nociceptive role of ATP. This perspective highlights some of the recent advances to identify selective P2 receptor ligands, which has enhanced the investigation of ATP-related modulation of pain sensitivity.

Molecular dynamics simulations and docking studies on 3D models of the heterodimeric and homodimeric 5-HT(2A) receptor subtype.

PubMed

Bruno, Agostino; Beato, Claudia; Costantino, Gabriele

2011-04-01

G-protein coupled receptors may exist as functional homodimers, heterodimers and even as higher aggregates. In this work, we investigate the 5-HT(2A) receptor, which is a known target for antipsychotic drugs. Recently, 5-HT(2A) has been shown to form functional homodimers and heterodimers with the mGluR2 receptor. The objective of this study is to build up 3D models of the 5-HT(2A)/mGluR2 heterodimer and of the 5-HT(2A)-5-HT(2A) homodimer, and to evaluate the impact of the dimerization interface on the shape of the 5-HT(2A) binding pocket by using molecular dynamics simulations and docking studies. The heterodimer, homodimer and monomeric 5-HT(2A) receptors were simulated by molecular dynamics for 40 ns each. The trajectories were clustered and representative structures of six clusters for each system were generated. Inspection of the these representative structures clearly indicate an effect of the dimerization interface on the topology of the binding pocket. Docking studies allowed to generate receiver operating characteristic curves for a set of 5-HT(2A) ligands, indicating that different complexes prefer different classes of 5-HT(2A) ligands. This study clearly indicates that the presence of a dimerization interface must explicitly be considered when studying G-protein coupled receptors known to exist as dimers. Molecular dynamics simulation and cluster analysis are appropriate tools to study the phenomenon.

Factor VIII Interacts with the Endocytic Receptor Low-density Lipoprotein Receptor-related Protein 1 via an Extended Surface Comprising "Hot-Spot" Lysine Residues.

PubMed

van den Biggelaar, Maartje; Madsen, Jesper J; Faber, Johan H; Zuurveld, Marleen G; van der Zwaan, Carmen; Olsen, Ole H; Stennicke, Henning R; Mertens, Koen; Meijer, Alexander B

2015-07-03

Lysine residues are implicated in driving the ligand binding to the LDL receptor family. However, it has remained unclear how specificity is regulated. Using coagulation factor VIII as a model ligand, we now study the contribution of individual lysine residues in the interaction with the largest member of the LDL receptor family, low-density lipoprotein receptor-related protein (LRP1). Using hydrogen-deuterium exchange mass spectrometry (HDX-MS) and SPR interaction analysis on a library of lysine replacement variants as two independent approaches, we demonstrate that the interaction between factor VIII (FVIII) and LRP1 occurs over an extended surface containing multiple lysine residues. None of the individual lysine residues account completely for LRP1 binding, suggesting an additive binding model. Together with structural docking studies, our data suggest that FVIII interacts with LRP1 via an extended surface of multiple lysine residues that starts at the bottom of the C1 domain and winds around the FVIII molecule. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

The dopamine D2 receptor dimer and its interaction with homobivalent antagonists: homology modeling, docking and molecular dynamics.

PubMed

Kaczor, Agnieszka A; Jörg, Manuela; Capuano, Ben

2016-09-01

In order to apply structure-based drug design techniques to G protein-coupled receptor complexes, it is essential to model their 3D structure and to identify regions that are suitable for selective drug binding. For this purpose, we have developed and tested a multi-component protocol to model the inactive conformation of the dopamine D2 receptor dimer, suitable for interaction with homobivalent antagonists. Our approach was based on protein-protein docking, applying the Rosetta software to obtain populations of dimers as present in membranes with all the main possible interfaces. Consensus scoring based on the values and frequencies of best interfaces regarding four scoring parameters, Rosetta interface score, interface area, free energy of binding and energy of hydrogen bond interactions indicated that the best scored dimer model possesses a TM4-TM5-TM7-TM1 interface, which is in agreement with experimental data. This model was used to study interactions of the previously published dopamine D2 receptor homobivalent antagonists based on clozapine,1,4-disubstituted aromatic piperidines/piperazines and arylamidoalkyl substituted phenylpiperazine pharmacophores. It was found that the homobivalent antagonists stabilize the receptor-inactive conformation by maintaining the ionic lock interaction, and change the dimer interface by disrupting a set of hydrogen bonds and maintaining water- and ligand-mediated hydrogen bonds in the extracellular and intracellular part of the interface. Graphical Abstract Structure of the final model of the dopamine D2 receptor homodimer, indicating the distancebetween Tyr37 and Tyr 5.42 in the apo form (left) and in the complex with the ligand (right).

Quantitative Characterization of Shear-Induced Platelet Receptor Shedding: Glycoprotein Ibα, Glycoprotein VI, and Glycoprotein IIb/IIIa.

PubMed

Chen, Zengsheng; Koenig, Steven C; Slaughter, Mark S; Griffith, Bartley P; Wu, Zhongjun J

2017-11-07

The structural integrity of platelet receptors is essential for platelets to play the normal hemostatic function. The high non-physiologic shear stress (NPSS) commonly exists in blood-contacting medical devices and has been shown to cause platelet receptor shedding. The loss of platelet receptors may impair the normal hemostatic function of platelets. The aim of this study was to quantify NPSS-induced shedding of three key receptors on the platelet surface. Human blood was subjected to the matrix of well-defined shear stresses and exposure times, generated by using a custom-designed blood-shearing device. The expression of three key platelet receptors, glycoprotein (GP) Ibα, GPVI, and GPIIb/IIIa, in sheared blood was quantified using flow cytometry. The quantitative relationship between the loss of each of the three receptors on the platelet surface and shear condition (shear stress level and exposure time) was explored. It was found that these relationships followed well the power law functional form. The coefficients of the power law models for the shear-induced shedding of these platelet receptors were derived with coefficients of determination (R) of 0.77, 0.73, and 0.78, respectively. The power law models with these coefficients may be potentially used to predict the shear-induced platelet receptor shedding of human blood.

Ozanimod (RPC1063) is a potent sphingosine‐1‐phosphate receptor‐1 (S1P1) and receptor‐5 (S1P5) agonist with autoimmune disease‐modifying activity

PubMed Central

Clemons, B; Brooks, J; Brahmachary, E; Powell, R; Dedman, H; Desale, H G; Timony, G A; Martinborough, E; Rosen, H; Roberts, E; Boehm, M F; Peach, R J

2016-01-01

Background and Purpose Sphingosine1‐phosphate (S1P) receptors mediate multiple events including lymphocyte trafficking, cardiac function, and endothelial barrier integrity. Stimulation of S1P1 receptors sequesters lymphocyte subsets in peripheral lymphoid organs, preventing their trafficking to inflamed tissue sites, modulating immunity. Targeting S1P receptors for treating autoimmune disease has been established in clinical studies with the non‐selective S1P modulator, FTY720 (fingolimod, Gilenya™). The purpose of this study was to assess RPC1063 for its therapeutic utility in autoimmune diseases. Experimental Approach The specificity and potency of RPC1063 (ozanimod) was evaluated for all five S1P receptors, and its effect on cell surface S1P1 receptor expression, was characterized in vitro. The oral pharmacokinetic (PK) parameters and pharmacodynamic effects were established in rodents, and its activity in three models of autoimmune disease (experimental autoimmune encephalitis, 2,4,6‐trinitrobenzenesulfonic acid colitis and CD4+CD45RBhi T cell adoptive transfer colitis) was assessed. Key Results RPC1063 was specific for S1P1 and S1P5 receptors, induced S1P1 receptor internalization and induced a reversible reduction in circulating B and CCR7+ T lymphocytes in vivo. RPC1063 showed high oral bioavailability and volume of distribution, and a circulatory half‐life that supports once daily dosing. Oral RPC1063 reduced inflammation and disease parameters in all three autoimmune disease models. Conclusions and Implications S1P receptor selectivity, favourable PK properties and efficacy in three distinct disease models supports the clinical development of RPC1063 for the treatment of relapsing multiple sclerosis and inflammatory bowel disease, differentiates RPC1063 from other S1P receptor agonists, and could result in improved safety outcomes in the clinic. PMID:26990079

The Mouse Solitary Odorant Receptor Gene Promoters as Models for the Study of Odorant Receptor Gene Choice.

PubMed

Degl'Innocenti, Andrea; Parrilla, Marta; Harr, Bettina; Teschke, Meike

2016-01-01

In vertebrates, several anatomical regions located within the nasal cavity mediate olfaction. Among these, the main olfactory epithelium detects most conventional odorants. Olfactory sensory neurons, provided with cilia exposed to the air, detect volatile chemicals via an extremely large family of seven-transmembrane chemoreceptors named odorant receptors. Their genes are expressed in a monogenic and monoallelic fashion: a single allele of a single odorant receptor gene is transcribed in a given mature neuron, through a still uncharacterized molecular mechanism known as odorant receptor gene choice. Odorant receptor genes are typically arranged in genomic clusters, but a few are isolated (we call them solitary) from the others within a region broader than 1 Mb upstream and downstream with respect to their transcript's coordinates. The study of clustered genes is problematic, because of redundancy and ambiguities in their regulatory elements: we propose to use the solitary genes as simplified models to understand odorant receptor gene choice. Here we define number and identity of the solitary genes in the mouse genome (C57BL/6J), and assess the conservation of the solitary status in some mammalian orthologs. Furthermore, we locate their putative promoters, predict their homeodomain binding sites (commonly present in the promoters of odorant receptor genes) and compare candidate promoter sequences with those of wild-caught mice. We also provide expression data from histological sections. In the mouse genome there are eight intact solitary genes: Olfr19 (M12), Olfr49, Olfr266, Olfr267, Olfr370, Olfr371, Olfr466, Olfr1402; five are conserved as solitary in rat. These genes are all expressed in the main olfactory epithelium of three-day-old mice. The C57BL/6J candidate promoter of Olfr370 has considerably varied compared to its wild-type counterpart. Within the putative promoter for Olfr266 a homeodomain binding site is predicted. As a whole, our findings favor Olfr266 as a model gene to investigate odorant receptor gene choice.

To Live or to Die: Prosurvival Activity of PPARγ in Cancers

PubMed Central

Wang, Y. Lynn; Miao, Qi

2008-01-01

The role of PPARγ in tumorigenesis is controversial. In this article, we review and analyze literature from the past decade that highlights the potential proneoplastic activity of PPARγ. We discuss the following five aspects of the nuclear hormone receptor and its agonists: (1) relative expression of PPARγ in human tumor versus normal tissues; (2) receptor-dependent proneoplastic effects; (3) impact of PPARγ and its agonists on tumors in animal models; (4) clinical trials of thiazolidinediones (TZDs) in human malignancies; (5) TZDs as chemopreventive agents in epidemiology studies. The focus is placed on the most relevant in vivo animal models and human data. In vitro cell line studies are included only when the effects are shown to be dependent on the PPARγ receptor. PMID:18784849

Deficits in cognition and synaptic plasticity in a mouse model of Down syndrome ameliorated by GABAB receptor antagonists

PubMed Central

Kleschevnikov, A.M.; Belichenko, P.V.; Faizi, M.; Jacobs, L.F.; Htun, K.; Shamloo, M.; Mobley, W.C.

2012-01-01

Cognitive impairment in Down syndrome (DS) is characterized by deficient learning and memory. Mouse genetic models of DS exhibit impaired cognition in hippocampally mediated behavioral tasks and reduced synaptic plasticity of hippocampal pathways. Enhanced efficiency of GABAergic neurotransmission was implicated in those changes. We have recently shown that signaling through postsynaptic GABAB receptors is significantly increased in the dentate gyrus (DG) of Ts65Dn mice, a genetic model of DS. Here we examined a role for GABAB receptors in cognitive deficits in DS by defining the effect of selective GABAB receptor antagonists on behavior and synaptic plasticity of adult Ts65Dn mice. Treatment with the GABAB receptor antagonist CGP55845 restored memory of Ts65Dn mice in the novel place recognition, novel object recognition and contextual fear conditioning tasks, but did not affect locomotion and performance in T-maze. The treatment increased hippocampal levels of brain-derived neurotrophic factor (BDNF), equally in 2N and Ts65Dn mice. In hippocampal slices, treatment with the GABAB receptor antagonists CGP55845 or CGP52432 enhanced long-term potentiation (LTP) in the Ts65Dn DG. The enhancement of LTP was accompanied by an increase in the NMDA receptor-mediated component of the tetanus-evoked responses. These findings are evidence for a contribution of GABAB receptors to changes in hippocampal-based cognition in the Ts65Dn mouse. The ability to rescue cognitive performance through treatment with selective GABAB receptor antagonists motivates studies to further explore the therapeutic potential of these compounds in people with DS. PMID:22764230

Towards structural models of molecular recognition in olfactory receptors.

PubMed

Afshar, M; Hubbard, R E; Demaille, J

1998-02-01

The G protein coupled receptors (GPCR) are an important class of proteins that act as signal transducers through the cytoplasmic membrane. Understanding the structure and activation mechanism of these proteins is crucial for understanding many different aspects of cellular signalling. The olfactory receptors correspond to the largest family of GPCRs. Very little is known about how the structures of the receptors govern the specificity of interaction which enables identification of particular odorant molecules. In this paper, we review recent developments in two areas of molecular modelling: methods for modelling the configuration of trans-membrane helices and methods for automatic docking of ligands into receptor structures. We then show how a subset of these methods can be combined to construct a model of a rat odorant receptor interacting with lyral for which experimental data are available. This modelling can help us make progress towards elucidating the specificity of interactions between receptors and odorant molecules.

Bimodal Action of Protons on ATP Currents of Rat PC12 Cells

PubMed Central

Skorinkin, Andrei; Nistri, Andrea; Giniatullin, Rashid

2003-01-01

The mode of action of extracellular protons on ATP-gated P2X2 receptors remains controversial as either enhancement or depression of ATP-mediated currents has been reported. By investigating, at different pH, the electrophysiological effect of ATP on P2X2 receptors and complementing it with receptor modelling, the present study suggests a unified mechanism for both potentiation and inactivation of ATP receptors by protons. Our experiments on patch-clamped PC12 cells showed that, on the same cell, mild acidification potentiated currents induced by low ATP concentrations (<0.1 mM) and attenuated responses to high ATP concentrations (>1 mM) with emergence of current fading and rebound. To clarify the nature of the ATP/H+ interaction, we used the Ding and Sachs's “loop” receptor model which best describes the behavior of such receptors with two open states linked via one inactivated state. No effects by protons could be ascribed to H+-mediated open channel block. However, by assuming that protons facilitated binding of ATP to resting as well as open receptors, the model could closely replicate H+-induced potentiation of currents evoked by low ATP doses plus fading and rebound induced by high ATP doses. The latter phenomenon was due to receptor transition to the inactive state. The present data suggest that the high concentration of protons released with ATP (and catecholamines) from secretory vesicles may allow a dual action of H+ on P2X2 receptors. This condition might also occur on P2X2 receptors of central neurons exposed to low pH during ischemia. PMID:12810852

Exploring the stochastic dynamics of correlated movement of receptor proteins in plasma membranes in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Jung Y., E-mail: jyhuang@faculty.nctu.edu.tw; Lin, Chien Y.

Ligand-induced receptor dimerization plays a crucial role in the signaling process of living cells. In this study, we developed a theoretical model and performed single-molecule tracking to explore the correlated diffusion processes of liganded epidermal growth factor receptors prior to dimer formation. We disclosed that both an attractive potential between liganded receptor proteins in proximity and correlated fluctuations in the local environments of the proteins play an important role to produce the observed correlated movement of the receptors. This result can serve as the foundation to shed light on the way in which receptor functions are regulated in plasma membranesmore » in vivo.« less

Insights about Striatal Circuit Function and Schizophrenia from a Mouse Model of D2 Receptor Upregulation

PubMed Central

Simpson, Eleanor H.; Kellendonk, Christoph

2016-01-01

The dopamine hypothesis of schizophrenia is supported by a large number of imaging studies that have identified an increase in dopamine binding at the D2 receptor selectively in the striatum. Here we review a decade of work using a regionally restricted and temporally regulated transgenic mouse model to investigate the behavioral, molecular, electrophysiological, and anatomical consequences of selective D2 receptor upregulation in the striatum. These studies have identified new and potentially important biomarkers at the circuit and molecular level that can now be explored in patients with schizophrenia. They provide an example of how animal models and their detailed level of neurobiological analysis allow a deepening of our understanding of the relationship between neuronal circuit function and symptoms of schizophrenia, and as a consequence generate new hypotheses that are testable in patients. PMID:27720388

Synaptic transmission at functionally identified synapses in the enteric nervous system: roles for both ionotropic and metabotropic receptors.

PubMed

Gwynne, R M; Bornstein, J C

2007-03-01

Digestion and absorption of nutrients and the secretion and reabsorption of fluid in the gastrointestinal tract are regulated by neurons of the enteric nervous system (ENS), the extensive peripheral nerve network contained within the intestinal wall. The ENS is an important physiological model for the study of neural networks since it is both complex and accessible. At least 20 different neurochemically and functionally distinct classes of enteric neurons have been identified in the guinea pig ileum. These neurons express a wide range of ionotropic and metabotropic receptors. Synaptic potentials mediated by ionotropic receptors such as the nicotinic acetylcholine receptor, P2X purinoceptors and 5-HT(3) receptors are seen in many enteric neurons. However, prominent synaptic potentials mediated by metabotropic receptors, like the P2Y(1) receptor and the NK(1) receptor, are also seen in these neurons. Studies of synaptic transmission between the different neuron classes within the enteric neural pathways have shown that both ionotropic and metabotropic synaptic potentials play major roles at distinct synapses within simple reflex pathways. However, there are still functional synapses at which no known transmitter or receptor has been identified. This review describes the identified roles for both ionotropic and metabotropic neurotransmission at functionally defined synapses within the guinea pig ileum ENS. It is concluded that metabotropic synaptic potentials act as primary transmitters at some synapses. It is suggested identification of the interactions between different synaptic potentials in the production of complex behaviours will require the use of well validated computer models of the enteric neural circuitry.

Receptor Activity-modifying Protein-directed G Protein Signaling Specificity for the Calcitonin Gene-related Peptide Family of Receptors.

PubMed

Weston, Cathryn; Winfield, Ian; Harris, Matthew; Hodgson, Rose; Shah, Archna; Dowell, Simon J; Mobarec, Juan Carlos; Woodlock, David A; Reynolds, Christopher A; Poyner, David R; Watkins, Harriet A; Ladds, Graham

2016-10-14

The calcitonin gene-related peptide (CGRP) family of G protein-coupled receptors (GPCRs) is formed through the association of the calcitonin receptor-like receptor (CLR) and one of three receptor activity-modifying proteins (RAMPs). Binding of one of the three peptide ligands, CGRP, adrenomedullin (AM), and intermedin/adrenomedullin 2 (AM2), is well known to result in a Gα s -mediated increase in cAMP. Here we used modified yeast strains that couple receptor activation to cell growth, via chimeric yeast/Gα subunits, and HEK-293 cells to characterize the effect of different RAMP and ligand combinations on this pathway. We not only demonstrate functional couplings to both Gα s and Gα q but also identify a Gα i component to CLR signaling in both yeast and HEK-293 cells, which is absent in HEK-293S cells. We show that the CGRP family of receptors displays both ligand- and RAMP-dependent signaling bias among the Gα s , Gα i , and Gα q/11 pathways. The results are discussed in the context of RAMP interactions probed through molecular modeling and molecular dynamics simulations of the RAMP-GPCR-G protein complexes. This study further highlights the importance of RAMPs to CLR pharmacology and to bias in general, as well as identifying the importance of choosing an appropriate model system for the study of GPCR pharmacology. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

RECEPTOR MODEL DEVELOPMENT AND APPLICATION

EPA Science Inventory

Source apportionment (receptor) models are mathematical procedures for identifying and quantifying the sources of ambient air pollutants and their effects at a site (the receptor), primarily on the basis of species concentration measurements at the receptor, and generally without...

PM SOURCE APPORTIONMENT/RECEPTOR MODELING

EPA Science Inventory

Source apportionment (receptor) models are mathematical procedures for identifying and quantifying the sources of ambient air pollutants and their effects at a site (the receptor), primarily on the basis of species concentration measurements at the receptor, and generally without...

Phosphorylation of a Tyrosine Residue on Zap70 by Lck and Its Subsequent Binding via an SH2 Domain May Be a Key Gatekeeper of T Cell Receptor Signaling In Vivo.

PubMed

Thill, Peter A; Weiss, Arthur; Chakraborty, Arup K

2016-09-15

The initiation of signaling in T lymphocytes in response to the binding of the T cell receptor (TCR) to cognate ligands is a key step in the emergence of adaptive immune responses. Conventional models posit that TCR signaling is initiated by the phosphorylation of receptor-associated immune receptor activation motifs (ITAMs). The cytoplasmic tyrosine kinase Zap70 binds to phosphorylated ITAMs, is subsequently activated, and then propagates downstream signaling. While evidence for such models is provided by experiments with cell lines, in vivo, Zap70 is bound to phosphorylated ITAMs in resting T cells. However, Zap70 is activated only upon TCR binding to cognate ligand. We report the results of computational studies of a new model for the initiation of TCR signaling that incorporates these in vivo observations. Importantly, the new model is shown to allow better and faster TCR discrimination between self-ligands and foreign ligands. The new model is consistent with many past experimental observations, and experiments that could further test the model are proposed. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Analysis and modeling of heat-labile enterotoxins of Escherichia coli suggests a novel space with insights into receptor preference.

PubMed

Krishna Raja, M; Ghosh, Asit Ranjan; Vino, S; Sajitha Lulu, S

2015-01-01

Features of heat-labile enterotoxins of Escherichia coli which make them fit to use as novel receptors for antidiarrheals are not completely explored. Data-set of 14 different serovars of enterotoxigenic Escherichia coli producing heat-labile toxins were taken from NCBI Genbank database and used in the study. Sequence analysis showed mutations in different subunits and also at their interface residues. As these toxins lack crystallography structures, homology modeling using Modeller 9.11 led to the structural approximation for the E. coli producing heat-labile toxins. Interaction of modeled toxin subunits with proanthocyanidin, an antidiarrheal showed several strong hydrogen bonding interactions at the cost of minimized energy. The hits were subsequently characterized by molecular dynamics simulation studies to monitor their binding stabilities. This study looks into novel space where the ligand can choose the receptor preference not as a whole but as an individual subunit. Mutation at interface residues and interaction among subunits along with the binding of ligand to individual subunits would help to design a non-toxic labile toxin and also to improve the therapeutics.

Development of Source-Receptor matrix over South Korea in support of GAINS-Korea model

NASA Astrophysics Data System (ADS)

Choi, K. C.; Woo, J. H.; Kim, H. K.; Lee, Y. M.; Kim, Y.; Heyes, C.; Lee, J. B.; Song, C. K.; Han, J.

2014-12-01

A comprehensive and combined analysis of air pollution and climate change could reveal important synergies of emission control measures, which could be of high policy relevance. IIASA's GAINS model (The Greenhouse gas - Air pollution Interactions and Synergies) has been developed as a tool to identify emission control strategies that achieve given targets on air quality and greenhouse gas emissions at least costs. The GAINS-Korea Model, which is being jointly developed by Konkuk University and IIASA, should play an important role in understanding the impact of air quality improvements across the regions in Korea. Source-Receptor relationships (S-R) is an useful methodology in air pollution studies to determine the areas of origin of chemical compounds at receptor point, and thus be able to target actions to reduce pollutions. The GAINS model can assess the impact of emission reductions of sources on air quality in receptor regions based on S-R matrix, derived from chemical transport model. In order to develop S-R matrix for GAINS-Korea, the CAMx model with PSAT/OSAT tools was applied in this study. The coarse domain covers East Asia, and a nesting domain as main research area was used for Korea peninsula. To evaluate of S-R relationships, a modeling domain is divided into sixteen regions over South Korea with three outside of S. Korea countries (China, N. Korea and Japan) for estimating transboundary contributions. The results of our analysis will be presented at the conference.

Concepts in receptor optimization: targeting the RGD peptide.

PubMed

Chen, Wei; Chang, Chia-en; Gilson, Michael K

2006-04-12

Synthetic receptors have a wide range of potential applications, but it has been difficult to design low molecular weight receptors that bind ligands with high, "proteinlike" affinities. This study uses novel computational methods to understand why it is hard to design a high-affinity receptor and to explore the limits of affinity, with the bioactive peptide RGD as a model ligand. The M2 modeling method is found to yield excellent agreement with experiment for a known RGD receptor and then is used to analyze a series of receptors generated in silico with a de novo design algorithm. Forces driving binding are found to be systematically opposed by proportionate repulsions due to desolvation and entropy. In particular, strong correlations are found between Coulombic attractions and the electrostatic desolvation penalty and between the mean energy change on binding and the cost in configurational entropy. These correlations help explain why it is hard to achieve high affinity. The change in surface area upon binding is found to correlate poorly with affinity within this series. Measures of receptor efficiency are formulated that summarize how effectively a receptor uses surface area, total energy, and Coulombic energy to achieve affinity. Analysis of the computed efficiencies suggests that a low molecular weight receptor can achieve proteinlike affinity. It is also found that macrocyclization of a receptor can, unexpectedly, increase the entropy cost of binding because the macrocyclic structure further restricts ligand motion.

A receptor-based model for dopamine-induced fMRI signal

PubMed Central

Mandeville, Joseph. B.; Sander, Christin Y. M.; Jenkins, Bruce G.; Hooker, Jacob M.; Catana, Ciprian; Vanduffel, Wim; Alpert, Nathaniel M.; Rosen, Bruce R.; Normandin, Marc D.

2013-01-01

This report describes a multi-receptor physiological model of the fMRI temporal response and signal magnitude evoked by drugs that elevate synaptic dopamine in basal ganglia. The model is formulated as a summation of dopamine’s effects at D1-like and D2-like receptor families, which produce functional excitation and inhibition, respectively, as measured by molecular indicators like adenylate cyclase or neuroimaging techniques like fMRI. Functional effects within the model are described in terms of relative changes in receptor occupancies scaled by receptor densities and neuro-vascular coupling constants. Using literature parameters, the model reconciles many discrepant observations and interpretations of pre-clinical data. Additionally, we present data showing that amphetamine stimulation produces fMRI inhibition at low doses and a biphasic response at higher doses in the basal ganglia of non-human primates (NHP), in agreement with model predictions based upon the respective levels of evoked dopamine. Because information about dopamine release is required to inform the fMRI model, we simultaneously acquired PET 11C-raclopride data in several studies to evaluate the relationship between raclopride displacement and assumptions about dopamine release. At high levels of dopamine release, results suggest that refinements of the model will be required to consistently describe the PET and fMRI data. Overall, the remarkable success of the model in describing a wide range of preclinical fMRI data indicate that this approach will be useful for guiding the design and analysis of basic science and clinical investigations and for interpreting the functional consequences of dopaminergic stimulation in normal subjects and in populations with dopaminergic neuroadaptations. PMID:23466936

Preclinical Testing of Novel Oxytocin Receptor Activators in Models of Autism Phenotypes

DTIC Science & Technology

2014-09-01

AD_________________ Award Number: TITLE: Preclinical Testing of Novel Oxytocin Receptor Activators in Models of Autism ...AUG 2013-7 Aug 2014 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Preclinical Testing of Novel Oxytocin Receptor Activators in Models of Autism ...a genetic mouse model of autism -like phenotypes, the Grin1 knockdown mouse. The Grin1 gene encodes the NR1 subunit of the NMDA receptor . In the

Postdoctoral Fellow | Center for Cancer Research

Cancer.gov

The Neural Development Section (NDS) headed by Dr. Lino Tessarollo has an open postdoctoral fellow position. The candidate should have a background in neurobiology and basic expertise in molecular biology, cell biology, immunoistochemistry and biochemistry.  Experience in confocal analysis is desired. The NDS study the biology of neurotrophin and Trk receptors function by using both in vitro and in vivo approaches. Our group makes extensive use of engineered mouse models and cell culture models. The current research emphasis is on understanding the molecular mechanisms by which activated trk receptor function. Specifically, we are dissecting the molecular mechanism responsible for modulating Trk receptors activity, including their interaction with specific scaffold proteins and proteins leading to de-activation of Trk signaling. Moreover, we are attempting to identify new signaling pathways activated by truncated Trk receptors.

MULTIVARIATE RECEPTOR MODELS AND MODEL UNCERTAINTY. (R825173)

EPA Science Inventory

Abstract

Estimation of the number of major pollution sources, the source composition profiles, and the source contributions are the main interests in multivariate receptor modeling. Due to lack of identifiability of the receptor model, however, the estimation cannot be...

Molecular Imaging Provides Novel Insights on Estrogen Receptor Activity in Mouse Brain

PubMed Central

Stell, Alessia; Belcredito, Silvia; Ciana, Paolo; Maggi, Adriana

2009-01-01

Estrogen receptors have long been known to be expressed in several brain areas in addition to those directly involved in the control of reproductive functions. Investigations in humans and in animal models suggest a strong influence of estrogens on limbic and motor functions, yet the complexity and heterogeneity of neural tissue have limited our approaches to the full understanding of estrogen activity in the central nervous system. The aim of this study was to examine the transcriptional activity of estrogen receptors in the brain of male and female mice. Exploiting the ERE-Luc reporter mouse, we set up a novel, bioluminescence-based technique to study brain estrogen receptor transcriptional activity. Here we show, for the first time, that estrogen receptors are similarly active in male and female brains and that the estrous cycle affects estrogen receptor activity in regions of the central nervous system not known to be associated with reproductive functions. Because of its reproducibility and sensitivity, this novel bioluminescence application candidates as an innovative methodology for the study and development of drugs targeting brain estrogen receptors. PMID:19123998

Effects of neuronal nicotinic acetylcholine receptor allosteric modulators in animal behavior studies

PubMed Central

Pandya, Anshul. A.; Yakel, Jerrel L.

2013-01-01

Nicotinic acetylcholine receptors (nAChRs) are ligand-gated cation-conducting transmembrane channels from the cys-loop receptor superfamily. The neuronal subtypes of these receptors (e.g. the α7 and α4β2 subtypes) are involved in neurobehavioral processes such as anxiety, the central processing of pain, food intake, nicotine seeking behavior, and a number of cognitive functions like learning and memory. Neuronal nAChR dysfunction is involved in the pathophysiology of many neurological disorders, and behavioral studies in animals are useful models to assess the effects of compounds that act on these receptors. Allosteric modulators are ligands that bind to the receptors at sites other than the orthosteric site where acetylcholine, the endogenous agonist for the nAChRs, binds. While conventional ligands for the neuronal nAChRs have been studied for their behavioral effects in animals, allosteric modulators for these receptors have only recently gained attention, and research on their behavioral effects is growing rapidly. Here we will discuss the behavioral effects of allosteric modulators of the neuronal nAChRs. PMID:23732296

Molecular imaging provides novel insights on estrogen receptor activity in mouse brain.

PubMed

Stell, Alessia; Belcredito, Silvia; Ciana, Paolo; Maggi, Adriana

2008-01-01

Estrogen receptors have long been known to be expressed in several brain areas in addition to those directly involved in the control of reproductive functions. Investigations in humans and in animal models suggest a strong influence of estrogens on limbic and motor functions, yet the complexity and heterogeneity of neural tissue have limited our approaches to the full understanding of estrogen activity in the central nervous system. The aim of this study was to examine the transcriptional activity of estrogen receptors in the brain of male and female mice. Exploiting the ERE-Luc reporter mouse, we set up a novel, bioluminescence-based technique to study brain estrogen receptor transcriptional activity. Here we show, for the first time, that estrogen receptors are similarly active in male and female brains and that the estrous cycle affects estrogen receptor activity in regions of the central nervous system not known to be associated with reproductive functions. Because of its reproducibility and sensitivity, this novel bioluminescence application stands as a candidate as an innovative methodology for the study and development of drugs targeting brain estrogen receptors.

Beyond small molecule SAR – using the dopamine D3 receptor crystal structure to guide drug design

PubMed Central

Keck, Thomas M.; Burzynski, Caitlin; Shi, Lei; Newman, Amy Hauck

2016-01-01

The dopamine D3 receptor is a target of pharmacotherapeutic interest in a variety of neurological disorders including schizophrenia, restless leg syndrome, and drug addiction. The high protein sequence homology between the D3 and D2 receptors has posed a challenge to developing D3 receptor-selective ligands whose behavioral actions can be attributed to D3 receptor engagement, in vivo. However, through primarily small molecule structure-activity relationship (SAR) studies, a variety of chemical scaffolds have been discovered over the past two decades that have resulted in several D3 receptor-selective ligands with high affinity and in vivo activity. Nevertheless, viable clinical candidates remain limited. The recent determination of the high-resolution crystal structure of the D3 receptor has invigorated structure-based drug design, providing refinements to the molecular dynamic models and testable predictions about receptor-ligand interactions. This review will highlight recent preclinical and clinical studies demonstrating potential utility of D3 receptor-selective ligands in the treatment of addiction. In addition, new structure-based rational drug design strategies for D3 receptor-selective ligands that complement traditional small molecule SAR to improve the selectivity and directed efficacy profiles are examined. PMID:24484980

δ-Opioid receptor agonists inhibit migraine-related hyperalgesia, aversive state and cortical spreading depression in mice

PubMed Central

Pradhan, Amynah A; Smith, Monique L; Zyuzin, Jekaterina; Charles, Andrew

2014-01-01

Background and Purpose Migraine is an extraordinarily common brain disorder for which treatment options continue to be limited. Agonists that activate the δ-opioid receptor may be promising for the treatment of migraine as they are highly effective for the treatment of chronic rather than acute pain, do not induce hyperalgesia, have low abuse potential and have anxiolytic and antidepressant properties. The aim of this study was to investigate the therapeutic potential of δ-opioid receptor agonists for migraine by characterizing their effects in mouse migraine models. Experimental Approach Mechanical hypersensitivity was assessed in mice treated with acute and chronic doses of nitroglycerin (NTG), a known human migraine trigger. Conditioned place aversion to NTG was also measured as a model of migraine-associated negative affect. In addition, we assessed evoked cortical spreading depression (CSD), an established model of migraine aura, in a thinned skull preparation. Key Results NTG evoked acute and chronic mechanical and thermal hyperalgesia in mice, as well as conditioned place aversion. Three different δ-opioid receptor agonists, SNC80, ARM390 and JNJ20788560, significantly reduced NTG-evoked hyperalgesia. SNC80 also abolished NTG-induced conditioned place aversion, suggesting that δ-opioid receptor activation may also alleviate the negative emotional state associated with migraine. We also found that SNC80 significantly attenuated CSD, a model that is considered predictive of migraine preventive therapies. Conclusions and Implications These data show that δ-opioid receptor agonists modulate multiple basic mechanisms associated with migraine, indicating that δ-opioid receptors are a promising therapeutic target for this disorder. PMID:24467301

Ligand-guided optimization of CXCR4 homology models for virtual screening using a multiple chemotype approach

NASA Astrophysics Data System (ADS)

Neves, Marco A. C.; Simões, Sérgio; Sá e Melo, M. Luisa

2010-12-01

CXCR4 is a G-protein coupled receptor for CXCL12 that plays an important role in human immunodeficiency virus infection, cancer growth and metastasization, immune cell trafficking and WHIM syndrome. In the absence of an X-ray crystal structure, theoretical modeling of the CXCR4 receptor remains an important tool for structure-function analysis and to guide the discovery of new antagonists with potential clinical use. In this study, the combination of experimental data and molecular modeling approaches allowed the development of optimized ligand-receptor models useful for elucidation of the molecular determinants of small molecule binding and functional antagonism. The ligand-guided homology modeling approach used in this study explicitly re-shaped the CXCR4 binding pocket in order to improve discrimination between known CXCR4 antagonists and random decoys. Refinement based on multiple test-sets with small compounds from single chemotypes provided the best early enrichment performance. These results provide an important tool for structure-based drug design and virtual ligand screening of new CXCR4 antagonists.

Development and application of hybrid structure based method for efficient screening of ligands binding to G-protein coupled receptors

NASA Astrophysics Data System (ADS)

Kortagere, Sandhya; Welsh, William J.

2006-12-01

G-protein coupled receptors (GPCRs) comprise a large superfamily of proteins that are targets for nearly 50% of drugs in clinical use today. In the past, the use of structure-based drug design strategies to develop better drug candidates has been severely hampered due to the absence of the receptor's three-dimensional structure. However, with recent advances in molecular modeling techniques and better computing power, atomic level details of these receptors can be derived from computationally derived molecular models. Using information from these models coupled with experimental evidence, it has become feasible to build receptor pharmacophores. In this study, we demonstrate the use of the Hybrid Structure Based (HSB) method that can be used effectively to screen and identify prospective ligands that bind to GPCRs. Essentially; this multi-step method combines ligand-based methods for building enriched libraries of small molecules and structure-based methods for screening molecules against the GPCR target. The HSB method was validated to identify retinal and its analogues from a random dataset of ˜300,000 molecules. The results from this study showed that the 9 top-ranking molecules are indeed analogues of retinal. The method was also tested to identify analogues of dopamine binding to the dopamine D2 receptor. Six of the ten top-ranking molecules are known analogues of dopamine including a prodrug, while the other thirty-four molecules are currently being tested for their activity against all dopamine receptors. The results from both these test cases have proved that the HSB method provides a realistic solution to bridge the gap between the ever-increasing demand for new drugs to treat psychiatric disorders and the lack of efficient screening methods for GPCRs.

Assessing the role of metabotropic glutamate receptor 5 in multiple nociceptive modalities.

PubMed

Zhu, Chang Z; Wilson, Sonya G; Mikusa, Joseph P; Wismer, Carol T; Gauvin, Donna M; Lynch, James J; Wade, Carrie L; Decker, Michael W; Honore, Prisca

2004-12-15

Preclinical data, performed in a limited number of pain models, suggest that functional blockade of metabotropic glutamate (mGlu) receptors may be beneficial for pain management. In the present study, effects of 2-methyl-6-(phenylethynyl)-pyridine (MPEP), a potent, selective mGlu5 receptor antagonist, were examined in a wide variety of rodent nociceptive and hypersensitivity models in order to fully characterize the potential analgesic profile of mGlu5 receptor blockade. Effects of 3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pyridine (MTEP), as potent and selective as MPEP at mGlu5/mGlu1 receptors but more selective than MPEP at N-methyl-aspartate (NMDA) receptors, were also evaluated in selected nociceptive and side effect models. MPEP (3-30 mg/kg, i.p.) produced a dose-dependent reversal of thermal and mechanical hyperalgesia following complete Freund's adjuvant (CFA)-induced inflammatory hypersensitivity. Additionally, MPEP (3-30 mg/kg, i.p.) decreased thermal hyperalgesia observed in carrageenan-induced inflammatory hypersensitivity without affecting paw edema, abolished acetic acid-induced writhing activity in mice, and was shown to reduce mechanical allodynia and thermal hyperalgesia observed in a model of post-operative hypersensitivity and formalin-induced spontaneous pain. Furthermore, at 30 mg/kg, i.p., MPEP significantly attenuated mechanical allodynia observed in three neuropathic pain models, i.e. spinal nerve ligation, sciatic nerve constriction and vincristine-induced neuropathic pain. MTEP (3-30 mg/kg, i.p.) also potently reduced CFA-induced thermal hyperalgesia. However, at 100 mg/kg, i.p., MPEP and MTEP produced central nerve system (CNS) side effects as measured by rotarod performance and exploratory locomotor activity. These results suggest a role for mGlu5 receptors in multiple nociceptive modalities, though CNS side effects may be a limiting factor in developing mGlu5 receptor analgesic compounds.

Adenosine A2A receptor blockade attenuates spatial memory deficit and extent of demyelination areas in lyolecithin-induced demyelination model.

PubMed

Akbari, Atefeh; Khalili-Fomeshi, Mohsen; Ashrafpour, Manouchehr; Moghadamnia, Ali Akbar; Ghasemi-Kasman, Maryam

2018-05-03

In recent years, inactivation of A 2A adenosine receptors has been emerged as a novel strategy for treatment of several neurodegenerative diseases. Although numerous studies have shown the beneficial effects of A 2A receptors blockade on spatial memory, the impacts of selective adenosine A 2A receptors on memory performance has not yet been examined in the context of demyelination. In the present study, we evaluated the effect of A 2A receptor antagonist SCH58261 on spatial memory and myelination in an experimental model of focal demyelination in rat fimbria. Demyelination was induced by local injection of lysolecithin (LPC) 1% (2 μl) into the hippocampus fimbria. SCH58261 (20 μg/0.5 μl or 40 μg/0.5 μl) was daily injected intracerebroventricularly (i.c.v.) for 10 days post LPC injection. The Morris water maze test was used to assess the spatial learning and memory on day 6 post lesion. Myelin staining and immunostaining against astrocytes/microglia were carried out 10 days post LPC injection. The administration of adenosine A 2A receptor antagonist prevented the spatial memory impairment in LPC receiving animals. Myelin staining revealed that application of SCH58261 reduces the extent of demyelination areas in the fimbria. Furthermore, the level of astrocytes and microglia activation was attenuated following administration of A 2A receptor antagonist. Collectively, the results of this study suggest that A 2A receptor blockade can improve the spatial memory and protect myelin sheath, which might be considered as a novel therapeutic approach for multiple sclerosis disease. Copyright © 2017. Published by Elsevier Inc.

Human eosinophils - potential pharmacological model applied in human histamine H4 receptor research.

PubMed

Grosicki, Marek; Kieć-Kononowicz, Katarzyna

2015-01-01

Histamine and histamine receptors are well known for their immunomodulatory role in inflammation. In this review we describe the role of histamine and histamine H4 receptor on human eosinophils. In the first part of article we provide short summary of histamine and histamine receptors role in physiology and histamine related therapeutics used in clinics. We briefly describe the human histamine receptor H4 and its ligands, as well as human eosinophils. In the second part of the review we provide detailed description of known histamine effects on eosinophils including: intracellular calcium concentration flux, actin polymerization, cellular shape change, upregulation of adhesion proteins and cellular chemotaxis. We provide proofs that these effects are mainly connected with the activation of histamine H4 receptor. When examining experimental data we discuss the controversial results and limitations of the studies performed on isolated eosinophils. In conclusion we believe that studies on histamine H4 receptor on human eosinophils can provide interesting new biomarkers that can be used in clinical studies of histamine receptors, that in future might result in the development of new strategies in the treatment of chronic inflammatory conditions like asthma or allergy, in which eosinophils are involved.

Human Endometrial Adenocarcinoma Transplanted into Nude Mice: Growth Regulation by Estradiol

NASA Astrophysics Data System (ADS)

Satyaswaroop, P. G.; Zaino, R. J.; Mortel, R.

1983-01-01

A model for studying the growth of primary tumors of human endometrium and its regulation by 17β -estradiol has been developed in which ovariectomized nude mice are used as recipients. The receptors for sex steroids are maintained during serial transplantation of the tumor in this system. Although the rate of growth of receptor-negative endometrial tumors transplanted into ovariectomized nude mice is unaffected by the sustained presence or absence of estradiol, the growth of receptor-positive tumors is significantly increased by estradiol. Receptor-positive tumors treated with estradiol produced elevated concentrations of progesterone receptor. That the progesterone receptor is functional in this tumor is evident from the induction of estradiol 17β -dehydrogenase activity upon progestin administration. These findings are consistent with receptor-mediated regulation of growth of endometrial carcinoma.

Multiple receptor conformation docking, dock pose clustering and 3D QSAR studies on human poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors.

PubMed

Fatima, Sabiha; Jatavath, Mohan Babu; Bathini, Raju; Sivan, Sree Kanth; Manga, Vijjulatha

2014-10-01

Poly(ADP-ribose) polymerase-1 (PARP-1) functions as a DNA damage sensor and signaling molecule. It plays a vital role in the repair of DNA strand breaks induced by radiation and chemotherapeutic drugs; inhibitors of this enzyme have the potential to improve cancer chemotherapy or radiotherapy. Three-dimensional quantitative structure activity relationship (3D QSAR) models were developed using comparative molecular field analysis, comparative molecular similarity indices analysis and docking studies. A set of 88 molecules were docked into the active site of six X-ray crystal structures of poly(ADP-ribose)polymerase-1 (PARP-1), by a procedure called multiple receptor conformation docking (MRCD), in order to improve the 3D QSAR models through the analysis of binding conformations. The docked poses were clustered to obtain the best receptor binding conformation. These dock poses from clustering were used for 3D QSAR analysis. Based on MRCD and QSAR information, some key features have been identified that explain the observed variance in the activity. Two receptor-based QSAR models were generated; these models showed good internal and external statistical reliability that is evident from the [Formula: see text], [Formula: see text] and [Formula: see text]. The identified key features enabled us to design new PARP-1 inhibitors.

Regulation of Steroid Hormone Receptor Function By the 52-kDa FK506-Binding Protein (FKBP52)

PubMed Central

Sivils, Jeffrey C.; Storer, Cheryl L.; Galigniana, Mario D.; Cox, Marc B.

2011-01-01

The large FK506-binding protein FKBP52 has been characterized as an important positive regulator of androgen, glucocorticoid and progesterone receptor signaling pathways. FKBP52 associates with receptor-Hsp90 complexes and is proposed to have roles in both receptor hormone binding and receptor subcellular localization. Data from biochemical and cellular studies has been corroborated in whole animal models as fkbp52-deficient male and female mice display characteristics of androgen, glucocorticoid and/or progesterone insensitivity. FKBP52 receptor specificity and the specific phenotypes displayed by the fkbp52-deficient mice have firmly established FKBP52 as a promising target for the treatment of a variety of hormone-dependent diseases. Recent studies demonstrated that the FKBP52 FK1 domain and the proline-rich loop within this domain are functionally important for FKBP52 regulation of receptor function. Based on these data, efforts are currently underway to target the FKBP52 FK1 domain and the proline-rich loop with small molecule inhibitors. PMID:21511531

Molecular determinants of ligand binding modes in the histamine H(4) receptor: linking ligand-based three-dimensional quantitative structure-activity relationship (3D-QSAR) models to in silico guided receptor mutagenesis studies.

PubMed

Istyastono, Enade P; Nijmeijer, Saskia; Lim, Herman D; van de Stolpe, Andrea; Roumen, Luc; Kooistra, Albert J; Vischer, Henry F; de Esch, Iwan J P; Leurs, Rob; de Graaf, Chris

2011-12-08

The histamine H(4) receptor (H(4)R) is a G protein-coupled receptor (GPCR) that plays an important role in inflammation. Similar to the homologous histamine H(3) receptor (H(3)R), two acidic residues in the H(4)R binding pocket, D(3.32) and E(5.46), act as essential hydrogen bond acceptors of positively ionizable hydrogen bond donors in H(4)R ligands. Given the symmetric distribution of these complementary pharmacophore features in H(4)R and its ligands, different alternative ligand binding mode hypotheses have been proposed. The current study focuses on the elucidation of the molecular determinants of H(4)R-ligand binding modes by combining (3D) quantitative structure-activity relationship (QSAR), protein homology modeling, molecular dynamics simulations, and site-directed mutagenesis studies. We have designed and synthesized a series of clobenpropit (N-(4-chlorobenzyl)-S-[3-(4(5)-imidazolyl)propyl]isothiourea) derivatives to investigate H(4)R-ligand interactions and ligand binding orientations. Interestingly, our studies indicate that clobenpropit (2) itself can bind to H(4)R in two distinct binding modes, while the addition of a cyclohexyl group to the clobenpropit isothiourea moiety allows VUF5228 (5) to adopt only one specific binding mode in the H(4)R binding pocket. Our ligand-steered, experimentally supported protein modeling method gives new insights into ligand recognition by H(4)R and can be used as a general approach to elucidate the structure of protein-ligand complexes.

Amylin Enhances Amyloid-β Peptide Brain to Blood Efflux Across the Blood-Brain Barrier

PubMed Central

Mohamed, Loqman A.; Zhu, Haihao; Mousa, Youssef M.; Wang, Erming; Qiu, Wei Qiao; Kaddoumi, Amal

2017-01-01

Findings from Alzheimer’s disease (AD) mouse models showed that amylin treatment improved AD pathology and enhanced amyloid-β (Aβ) brain to blood clearance; however, the mechanism was not investigated. Using the Tg2576 AD mouse model, a single intraperitoneal injection of amylin significantly increased Aβ serum levels, and the effect was abolished by AC253, an amylin receptor antagonist, suggesting that amylin effect could be mediated by its receptor. Subsequent mechanistic studies showed amylin enhanced Aβ transport across a cell-based model of the blood-brain barrier (BBB), an effect that was abolished when the amylin receptor was inhibited by two amylin antagonists and by siRNA knockdown of amylin receptor Ramp3. To explain this finding, amylin effect on Aβ transport proteins expressed at the BBB was evaluated. Findings indicated that cells treated with amylin induced LRP1 expression, a major receptor involved in brain Aβ efflux, in plasma membrane fraction, suggesting intracellular translocation of LRP1 from the cytoplasmic pool. Increased LRP1 in membrane fraction could explain, at least in part, the enhanced uptake and transport of Aβ across the BBB. Collectively, our findings indicated that amylin induced Aβ brain to blood clearance through amylin receptor by inducing LRP1 subcellular translocation to the plasma membrane of the BBB endothelium. PMID:28059785

Activation and inhibition of mouse muscle and neuronal nicotinic acetylcholine receptors expressed in Xenopus oocytes.

PubMed

Papke, Roger L; Wecker, Lynn; Stitzel, Jerry A

2010-05-01

Transgenic mouse models with nicotinic acetylcholine receptor (nAChR) knockouts and knockins have provided important insights into the molecular substrates of addiction and disease. However, most studies of heterologously expressed neuronal nAChR have used clones obtained from other species, usually human or rat. In this work, we use mouse clones expressed in Xenopus oocytes to provide a relatively comprehensive characterization of the three primary classes of nAChR: muscle-type receptors, heteromeric neuronal receptors, and homomeric alpha7-type receptors. We evaluated the activation of these receptor subtypes with acetylcholine and cytisine-related compounds, including varenicline. We also characterized the activity of classic nAChR antagonists, confirming the utility of mecamylamine and dihydro-beta-erythroidine as selective antagonists in mouse models of alpha3beta4 and alpha4beta2 receptors, respectively. We also conducted an in-depth analysis of decamethonium and hexamethonium on muscle and neuronal receptor subtypes. Our data indicate that, as with receptors cloned from other species, pairwise expression of neuronal alpha and beta subunits in oocytes generates heterogeneous populations of receptors, most likely caused by variations in subunit stoichiometry. Coexpression of the mouse alpha5 subunit had varying effects, depending on the other subunits expressed. The properties of cytisine-related compounds are similar for mouse, rat, and human nAChR, except that varenicline produced greater residual inhibition of mouse alpha4beta2 receptors than with human receptors. We confirm that decamethonium is a partial agonist, selective for muscle-type receptors, but also note that it is a nondepolarizing antagonist for neuronal-type receptors. Hexamethonium was a relatively nonselective antagonist with mixed competitive and noncompetitive activity.

Activation and Inhibition of Mouse Muscle and Neuronal Nicotinic Acetylcholine Receptors Expressed in Xenopus Oocytes

PubMed Central

Wecker, Lynn; Stitzel, Jerry A.

2010-01-01

Transgenic mouse models with nicotinic acetylcholine receptor (nAChR) knockouts and knockins have provided important insights into the molecular substrates of addiction and disease. However, most studies of heterologously expressed neuronal nAChR have used clones obtained from other species, usually human or rat. In this work, we use mouse clones expressed in Xenopus oocytes to provide a relatively comprehensive characterization of the three primary classes of nAChR: muscle-type receptors, heteromeric neuronal receptors, and homomeric α7-type receptors. We evaluated the activation of these receptor subtypes with acetylcholine and cytisine-related compounds, including varenicline. We also characterized the activity of classic nAChR antagonists, confirming the utility of mecamylamine and dihydro-β-erythroidine as selective antagonists in mouse models of α3β4 and α4β2 receptors, respectively. We also conducted an in-depth analysis of decamethonium and hexamethonium on muscle and neuronal receptor subtypes. Our data indicate that, as with receptors cloned from other species, pairwise expression of neuronal α and β subunits in oocytes generates heterogeneous populations of receptors, most likely caused by variations in subunit stoichiometry. Coexpression of the mouse α5 subunit had varying effects, depending on the other subunits expressed. The properties of cytisine-related compounds are similar for mouse, rat, and human nAChR, except that varenicline produced greater residual inhibition of mouse α4β2 receptors than with human receptors. We confirm that decamethonium is a partial agonist, selective for muscle-type receptors, but also note that it is a nondepolarizing antagonist for neuronal-type receptors. Hexamethonium was a relatively nonselective antagonist with mixed competitive and noncompetitive activity. PMID:20100906

Impact of Measurement Uncertainties on Receptor Modeling of Speciated Atmospheric Mercury.

PubMed

Cheng, I; Zhang, L; Xu, X

2016-02-09

Gaseous oxidized mercury (GOM) and particle-bound mercury (PBM) measurement uncertainties could potentially affect the analysis and modeling of atmospheric mercury. This study investigated the impact of GOM measurement uncertainties on Principal Components Analysis (PCA), Absolute Principal Component Scores (APCS), and Concentration-Weighted Trajectory (CWT) receptor modeling results. The atmospheric mercury data input into these receptor models were modified by combining GOM and PBM into a single reactive mercury (RM) parameter and excluding low GOM measurements to improve the data quality. PCA and APCS results derived from RM or excluding low GOM measurements were similar to those in previous studies, except for a non-unique component and an additional component extracted from the RM dataset. The percent variance explained by the major components from a previous study differed slightly compared to RM and excluding low GOM measurements. CWT results were more sensitive to the input of RM than GOM excluding low measurements. Larger discrepancies were found between RM and GOM source regions than those between RM and PBM. Depending on the season, CWT source regions of RM differed by 40-61% compared to GOM from a previous study. No improvement in correlations between CWT results and anthropogenic mercury emissions were found.

Impact of Measurement Uncertainties on Receptor Modeling of Speciated Atmospheric Mercury

PubMed Central

Cheng, I.; Zhang, L.; Xu, X.

2016-01-01

Gaseous oxidized mercury (GOM) and particle-bound mercury (PBM) measurement uncertainties could potentially affect the analysis and modeling of atmospheric mercury. This study investigated the impact of GOM measurement uncertainties on Principal Components Analysis (PCA), Absolute Principal Component Scores (APCS), and Concentration-Weighted Trajectory (CWT) receptor modeling results. The atmospheric mercury data input into these receptor models were modified by combining GOM and PBM into a single reactive mercury (RM) parameter and excluding low GOM measurements to improve the data quality. PCA and APCS results derived from RM or excluding low GOM measurements were similar to those in previous studies, except for a non-unique component and an additional component extracted from the RM dataset. The percent variance explained by the major components from a previous study differed slightly compared to RM and excluding low GOM measurements. CWT results were more sensitive to the input of RM than GOM excluding low measurements. Larger discrepancies were found between RM and GOM source regions than those between RM and PBM. Depending on the season, CWT source regions of RM differed by 40–61% compared to GOM from a previous study. No improvement in correlations between CWT results and anthropogenic mercury emissions were found. PMID:26857835

Dose-dependent EEG effects of zolpidem provide evidence for GABA(A) receptor subtype selectivity in vivo.

PubMed

Visser, S A G; Wolters, F L C; van der Graaf, P H; Peletier, L A; Danhof, M

2003-03-01

Zolpidem is a nonbenzodiazepine GABA(A) receptor modulator that binds in vitro with high affinity to GABA(A) receptors expressing alpha(1) subunits but with relatively low affinity to receptors expressing alpha(2), alpha(3), and alpha(5) subunits. In the present study, it was investigated whether this subtype selectivity could be detected and quantified in vivo. Three doses (1.25, 5, and 25 mg) of zolpidem were administered to rats in an intravenous infusion over 5 min. The time course of the plasma concentrations was determined in conjunction with the change in the beta-frequency range of the EEG as pharmacodynamic endpoint. The concentration-effect relationship of the three doses showed a dose-dependent maximum effect and a dose-dependent potency. The data were analyzed for one- or two-site binding using two pharmacodynamic models based on 1) the descriptive model and 2) a novel mechanism-based pharmacokinetic/pharmacodynamic (PK/PD) model for GABA(A) receptor modulators that aims to separates drug- and system-specific properties, thereby allowing the estimation of in vivo affinity and efficacy. The application of two-site models significantly improved the fits compared with one-site models. Furthermore, in contrast to the descriptive model, the mechanism-based PK/PD model yielded dose-independent estimates for affinity (97 +/- 40 and 33,100 +/- 14,800 ng x ml(-1)). In conclusion, the mechanism-based PK/PD model is able to describe and explain the observed dose-dependent EEG effects of zolpidem and suggests the subtype selectivity of zolpidem in vivo.

Comparing the cardiovascular therapeutic indices of glycopyrronium and tiotropium in an integrated rat pharmacokinetic, pharmacodynamic and safety model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Trifilieff, Alexandre; Ethell, Brian T.; Sykes, David A.

Long acting inhaled muscarinic receptor antagonists, such as tiotropium, are widely used as bronchodilator therapy for chronic obstructive pulmonary disease (COPD). Although this class of compounds is generally considered to be safe and well tolerated in COPD patients the cardiovascular safety of tiotropium has recently been questioned. We describe a rat in vivo model that allows the concurrent assessment of muscarinic antagonist potency, bronchodilator efficacy and a potential for side effects, and we use this model to compare tiotropium with NVA237 (glycopyrronium bromide), a recently approved inhaled muscarinic antagonist for COPD. Anaesthetized Brown Norway rats were dosed intratracheally at 1more » or 6 h prior to receiving increasing doses of intravenous methacholine. Changes in airway resistance and cardiovascular function were recorded and therapeutic indices were calculated against the ED{sub 50} values for the inhibition of methacholine-induced bronchoconstriction. At both time points studied, greater therapeutic indices for hypotension and bradycardia were observed with glycopyrronium (19.5 and 28.5 fold at 1 h; > 200 fold at 6 h) than with tiotropium (1.5 and 4.2 fold at 1 h; 4.6 and 5.5 fold at 6 h). Pharmacokinetic, protein plasma binding and rat muscarinic receptor binding properties for both compounds were determined and used to generate an integrated model of systemic M{sub 2} muscarinic receptor occupancy, which predicted significantly higher M{sub 2} receptor blockade at ED{sub 50} doses with tiotropium than with glycopyrronium. In our preclinical model there was an improved safety profile for glycopyrronium when compared with tiotropium. - Highlights: • We use an in vivo rat model to study CV safety of inhaled muscarinic antagonists. • We integrate protein and receptor binding and PK of tiotropium and glycopyrrolate. • At ED{sub 50} doses for bronchoprotection we model systemic M{sub 2} receptor occupancy. • Glycopyrrolate demonstrates lower M{sub 2} occupancy at bronchoprotective doses. • Glycopyrrolate demonstrates an improved CV safety profile, versus tiotropium.« less

Multiple templates-based homology modeling enhances structure quality of AT1 receptor: validation by molecular dynamics and antagonist docking.

PubMed

Sokkar, Pandian; Mohandass, Shylajanaciyar; Ramachandran, Murugesan

2011-07-01

We present a comparative account on 3D-structures of human type-1 receptor (AT1) for angiotensin II (AngII), modeled using three different methodologies. AngII activates a wide spectrum of signaling responses via the AT1 receptor that mediates physiological control of blood pressure and diverse pathological actions in cardiovascular, renal, and other cell types. Availability of 3D-model of AT1 receptor would significantly enhance the development of new drugs for cardiovascular diseases. However, templates of AT1 receptor with low sequence similarity increase the complexity in straightforward homology modeling, and hence there is a need to evaluate different modeling methodologies in order to use the models for sensitive applications such as rational drug design. Three models were generated for AT1 receptor by, (1) homology modeling with bovine rhodopsin as template, (2) homology modeling with multiple templates and (3) threading using I-TASSER web server. Molecular dynamics (MD) simulation (15 ns) of models in explicit membrane-water system, Ramachandran plot analysis and molecular docking with antagonists led to the conclusion that multiple template-based homology modeling outweighs other methodologies for AT1 modeling.

An Analytical Model for Determining Two-Dimensional Receptor-Ligand Kinetics

PubMed Central

Cheung, Luthur Siu-Lun; Konstantopoulos, Konstantinos

2011-01-01

Cell-cell adhesive interactions play a pivotal role in major pathophysiological vascular processes, such as inflammation, infection, thrombosis, and cancer metastasis, and are regulated by hemodynamic forces generated by blood flow. Cell adhesion is mediated by the binding of receptors to ligands, which are both anchored on two-dimensional (2-D) membranes of apposing cells. Biophysical assays have been developed to determine the unstressed (no-force) 2-D affinity but fail to disclose its dependence on force. Here we develop an analytical model to estimate the 2-D kinetics of diverse receptor-ligand pairs as a function of force, including antibody-antigen, vascular selectin-ligand, and bacterial adhesin-ligand interactions. The model can account for multiple bond interactions necessary to mediate adhesion and resist detachment amid high hemodynamic forces. Using this model, we provide a generalized biophysical interpretation of the counterintuitive force-induced stabilization of cell rolling observed by a select subset of receptor-ligand pairs with specific intrinsic kinetic properties. This study enables us to understand how single-molecule and multibond biophysics modulate the macroscopic cell behavior in diverse pathophysiological processes. PMID:21575567

Secreted calmodulin-like skin protein inhibits neuronal death in cell-based Alzheimer's disease models via the heterotrimeric Humanin receptor

PubMed Central

Hashimoto, Y; Nawa, M; Kurita, M; Tokizawa, M; Iwamatsu, A; Matsuoka, M

2013-01-01

Humanin is a secreted bioactive peptide that is protective in a variety of death models, including cell-based neuronal death models related to Alzheimer's disease (AD). To mediate the protective effect in AD-related death models, Humanin signals via a cell-surface receptor that is generally composed of three subunits: ciliary neurotrophic factor receptor α, WSX-1 and gp130 (heterotrimeric Humanin receptor; htHNR). However, the protective effect of Humanin via the htHNR is weak (EC50=1–10 μℳ); therefore, it is possible that another physiological agonist for this receptor exists in vivo. In the current study, calmodulin-like skin protein (CLSP), a calmodulin relative with an undefined function, was shown to be secreted and inhibit neuronal death via the htHNR with an EC50 of 10–100 pℳ. CLSP was highly expressed in the skin, and the concentration in circulating normal human blood was ∼5 nℳ. When administered intraperitoneally in mice, recombinant CLSP was transported across the blood-cerebrospinal fluid (CSF)-barrier and its concentration in the CSF reaches 1/100 of its serum concentration at 1 h after injection. These findings suggest that CLSP is a physiological htHNR agonist. PMID:23519124

Modeling Canadian Quality Control Test Program for Steroid Hormone Receptors in Breast Cancer: Diagnostic Accuracy Study.

PubMed

Pérez, Teresa; Makrestsov, Nikita; Garatt, John; Torlakovic, Emina; Gilks, C Blake; Mallett, Susan

The Canadian Immunohistochemistry Quality Control program monitors clinical laboratory performance for estrogen receptor and progesterone receptor tests used in breast cancer treatment management in Canada. Current methods assess sensitivity and specificity at each time point, compared with a reference standard. We investigate alternative performance analysis methods to enhance the quality assessment. We used 3 methods of analysis: meta-analysis of sensitivity and specificity of each laboratory across all time points; sensitivity and specificity at each time point for each laboratory; and fitting models for repeated measurements to examine differences between laboratories adjusted by test and time point. Results show 88 laboratories participated in quality control at up to 13 time points using typically 37 to 54 histology samples. In meta-analysis across all time points no laboratories have sensitivity or specificity below 80%. Current methods, presenting sensitivity and specificity separately for each run, result in wide 95% confidence intervals, typically spanning 15% to 30%. Models of a single diagnostic outcome demonstrated that 82% to 100% of laboratories had no difference to reference standard for estrogen receptor and 75% to 100% for progesterone receptor, with the exception of 1 progesterone receptor run. Laboratories with significant differences to reference standard identified with Generalized Estimating Equation modeling also have reduced performance by meta-analysis across all time points. The Canadian Immunohistochemistry Quality Control program has a good design, and with this modeling approach has sufficient precision to measure performance at each time point and allow laboratories with a significantly lower performance to be targeted for advice.

Identifying Metabolically Active Chemicals Using a Consensus Quantitative Structure Activity Model for Estrogen Receptor Binding

EPA Science Inventory

Endocrine disrupting chemicals (EDCs) are abundant throughout the environment and can alter neurodevelopment, behavior, and reproductive success of humans and other species by perturbing signaling pathways related to the estrogen receptor (ER). A recent study compared results acr...

Molecular characterization and gene expression patterns of retinoid receptors, in normal and regenerating tissues of the sea cucumber, Holothuria glaberrima.

PubMed

Viera-Vera, Jorge; García-Arrarás, José E

2018-05-15

Retinoic acid receptors (RAR) and retinoid X receptors (RXR) are ligand-mediated transcription factors that synchronize intricate signaling networks in metazoans. Dimer formation between these two nuclear receptors mediates the recruitment of co-regulatory complexes coordinating the progression of signaling cascades during developmental and regenerative events. In the present study we identified and characterized the receptors for retinoic acid in the sea cucumber Holothuria glaberrima; a model system capable of regenerative organogenesis during adulthood. Molecular characterizations revealed the presence of three isoforms of RAR and two of RXR as a consequence of alternative splicing events. Various analyses including: primary structure sequencing, phylogenetic analysis, protein domain prediction, and multiple sequence alignment further confirmed their identity. Semiquantitative reverse transcription PCR analysis of each receptor isoform herein identified showed that the retinoid receptors are expressed in all tissues sampled: the mesenteries, respiratory trees, muscles, gonads, and the digestive tract. During regenerative organogenesis two of the receptors (RAR-L and RXR-T) showed differential expression in the posterior segment while RAR-S is differentially expressed in the anterior segment of the intestine. This work presents the first description of the components relaying the signaling for retinoic acid within this model system. Copyright © 2018 Elsevier B.V. All rights reserved.

Computational Analysis of the CB1 Carboxyl-terminus in the Receptor-G Protein Complex

PubMed Central

Shim, Joong-Youn; Khurana, Leepakshi; Kendall, Debra A.

2016-01-01

Despite the important role of the carboxyl-terminus (Ct) of the activated brain cannabinoid receptor one (CB1) in the regulation of G protein signaling, a structural understanding of interactions with G proteins is lacking. This is largely due to the highly flexible nature of the CB1 Ct that dynamically adapts its conformation to the presence of G proteins. In the present study, we explored how the CB1 Ct can interact with the G protein by building on our prior modeling of the CB1-Gi complex (Shim J-Y, Ahn KH, Kendall DA. The Journal of Biological Chemistry 2013;288:32449-32465) to incorporate a complete CB1 Ct (Glu416Ct–Leu472Ct). Based upon the structural constraints from NMR studies, we employed ROSETTA to predict tertiary folds, ZDOCK to predict docking orientation, and molecular dynamics (MD) simulations to obtain two distinct plausible models of CB1 Ct in the CB1-Gi complex. The resulting models were consistent with the NMR-determined helical structure (H9) in the middle region of the CB1 Ct. The CB1 Ct directly interacted with both Gα and Gβ and stabilized the receptor at the Gi interface. The results of site-directed mutagenesis studies of Glu416Ct, Asp423Ct, Asp428Ct, and Arg444Ct of CB1 Ct suggested that the CB1 Ct can influence receptor-G protein coupling by stabilizing the receptor at the Gi interface. This research provided, for the first time, models of the CB1 Ct in contact with the G protein. PMID:26994549

Towards Interpreting the Signal of CO2 Emissions from Megacities by Applying a Lagrangian Receptor-oriented Model to OCO-2 XCO2 data

NASA Astrophysics Data System (ADS)

Wu, D.; Lin, J. C.; Oda, T.; Ye, X.; Lauvaux, T.; Yang, E. G.; Kort, E. A.

2017-12-01

Urban regions are large emitters of CO2 whose emission inventories are still associated with large uncertainties. Therefore, a strong need exists to better quantify emissions from megacities using a top-down approach. Satellites — e.g., the Orbiting Carbon Observatory 2 (OCO-2), provide a platform for monitoring spatiotemporal column CO2 concentrations (XCO2). In this study, we present a Lagrangian receptor-oriented model framework and evaluate "model-retrieved" XCO2 by comparing against OCO-2-retrieved XCO2, for three megacities/regions (Riyadh, Cairo and Pearl River Delta). OCO-2 soundings indicate pronounced XCO2 enhancements (dXCO2) when crossing Riyadh, which are successfully captured by our model with a slight latitude shift. From this model framework, we can identify and compare the relative contributions of dXCO2 resulted from anthropogenic emission versus biospheric fluxes. In addition, to impose constraints on emissions for Riyadh through inversion methods, three uncertainties sources are addressed in this study, including 1) transport errors, 2) receptor and model setups in atmospheric models, and 3) urban emission uncertainties. For 1), we calculate transport errors by adding a wind error component to randomize particle distributions. For 2), a set of sensitivity tests using bootstrap method is performed to describe proper ways to setup receptors in Lagrangian models. For 3), both emission uncertainties from the Fossil Fuel Data Assimilation System (FFDAS) and the spread among three emission inventories are used to approximate an overall fractional uncertainty in modeled anthropogenic signal (dXCO2.anthro). Lastly, we investigate the definition of background (clean) XCO2 for megacities from retrieved XCO2 by means of statistical tools and our model framework.

Free energy force field (FEFF) 3D-QSAR analysis of a set of Plasmodium falciparum dihydrofolate reductase inhibitors

NASA Astrophysics Data System (ADS)

Santos-Filho, Osvaldo A.; Mishra, Rama K.; Hopfinger, A. J.

2001-09-01

Free energy force field (FEFF) 3D-QSAR analysis was used to construct ligand-receptor binding models for a set of 18 structurally diverse antifolates including pyrimethamine, cycloguanil, methotrexate, aminopterin and trimethoprim, and 13 pyrrolo[2,3-d]pyrimidines. The molecular target (`receptor') used was a 3D-homology model of a specific mutant type of Plasmodium falciparum (Pf) dihydrofolate reductase (DHFR). The dependent variable of the 3D-QSAR models is the IC50 inhibition constant for the specific mutant type of PfDHFR. The independent variables of the 3D-QSAR models (the descriptors) are scaled energy terms of a modified first-generation AMBER force field combined with a hydration shell aqueous solvation model and a collection of 2D-QSAR descriptors often used in QSAR studies. Multiple temperature molecular dynamics simulation (MDS) and the genetic function approximation (GFA) were employed using partial least square (PLS) and multidimensional linear regressions as the fitting functions to develop FEFF 3D-QSAR models for the binding process. The significant FEFF energy terms in the best 3D-QSAR models include energy contributions of the direct ligand-receptor interaction. Some changes in conformational energy terms of the ligand due to binding to the enzyme are also found to be important descriptors. The FEFF 3D-QSAR models indicate some structural features perhaps relevant to the mechanism of resistance of the PfDHFR to current antimalarials. The FEFF 3D-QSAR models are also compared to receptor-independent (RI) 4D-QSAR models developed in an earlier study and subsequently refined using recently developed generalized alignment rules.

Metabotropic Glutamate Receptors for Parkinson's Disease Therapy

PubMed Central

Gasparini, Fabrizio; Di Paolo, Thérèse; Gomez-Mancilla, Baltazar

2013-01-01

Excessive glutamatergic signalling within the basal ganglia is implicated in the progression of Parkinson's disease (PD) and inthe emergence of dyskinesia associated with long-term treatment with L-DOPA. There is considerable research focus on the discovery and development of compounds that modulate glutamatergic signalling via glutamate receptors, as treatments for PD and L-DOPA-induced dyskinesia (LID). Although initial preclinical studies with ionotropic glutamate receptor antagonists showed antiparkinsonian and antidyskinetic activity, their clinical use was limited due to psychiatric adverse effects, with the exception of amantadine, a weak N-methyl-d-aspartate (NMDA) antagonist, currently used to reduce dyskinesia in PD patients. Metabotropic receptor (mGlu receptor) modulators were considered to have a more favourable side-effect profile, and several agents have been studied in preclinical models of PD. The most promising results have been seen clinically with selective antagonists of mGlu5 receptor and preclinically with selective positive allosteric modulators of mGlu4 receptor. The growing understanding of glutamate receptor crosstalk also raises the possibility of more precise modulation of glutamatergic transmission, which may lead to the development of more effective agents for PD. PMID:23853735

A non-invasive test for receptor binding applied to nephrogenic diabetes insipidus.

PubMed Central

Britton, K. E.; Tedder, R. S.; Khokhar, A. M.; Brown, N. J.; Davison, A.; Slater, J. D.

1977-01-01

Studies in animals have determined the importance of specific receptors to the action of many hormones and drugs. In man, a non-invasive external counting technique has been used and absence of receptor function has been demonstrated in a patient with nephrogenic diabetes insipidus using radioactively labelled arginine vasopressin. This is in contrast to the findings in a patient with pituitary diabetes insipidus and a normal control. These results suggest a model for the study of hormone and drug kinetics in man avoiding multiple samplings of biological fluids. PMID:196275

Imaging Agonist-Induced D2/D3 Receptor Desensitization and Internalization In Vivo with PET/fMRI.

PubMed

Sander, Christin Y; Hooker, Jacob M; Catana, Ciprian; Rosen, Bruce R; Mandeville, Joseph B

2016-04-01

This study investigated the dynamics of dopamine receptor desensitization and internalization, thereby proposing a new technique for non-invasive, in vivo measurements of receptor adaptations. The D2/D3 agonist quinpirole, which induces receptor internalization in vitro, was administered at graded doses in non-human primates while imaging with simultaneous positron emission tomography (PET) and functional magnetic resonance imaging (fMRI). A pronounced temporal divergence between receptor occupancy and fMRI signal was observed: occupancy remained elevated while fMRI responded transiently. Analogous experiments with an antagonist (prochlorperazine) and a lower-affinity agonist (ropinirole) exhibited reduced temporal dissociation between occupancy and function, consistent with a mechanism of desensitization and internalization that depends upon drug efficacy and affinity. We postulated a model that incorporates internalization into a neurovascular-coupling relationship. This model yielded in vivo desensitization/internalization rates (0.2/min for quinpirole) consistent with published in vitro measurements. Overall, these results suggest that simultaneous PET/fMRI enables characterization of dynamic neuroreceptor adaptations in vivo, and may offer a first non-invasive method for assessing receptor desensitization and internalization.

Imaging Agonist-Induced D2/D3 Receptor Desensitization and Internalization In Vivo with PET/fMRI

PubMed Central

Sander, Christin Y; Hooker, Jacob M; Catana, Ciprian; Rosen, Bruce R; Mandeville, Joseph B

2016-01-01

This study investigated the dynamics of dopamine receptor desensitization and internalization, thereby proposing a new technique for non-invasive, in vivo measurements of receptor adaptations. The D2/D3 agonist quinpirole, which induces receptor internalization in vitro, was administered at graded doses in non-human primates while imaging with simultaneous positron emission tomography (PET) and functional magnetic resonance imaging (fMRI). A pronounced temporal divergence between receptor occupancy and fMRI signal was observed: occupancy remained elevated while fMRI responded transiently. Analogous experiments with an antagonist (prochlorperazine) and a lower-affinity agonist (ropinirole) exhibited reduced temporal dissociation between occupancy and function, consistent with a mechanism of desensitization and internalization that depends upon drug efficacy and affinity. We postulated a model that incorporates internalization into a neurovascular-coupling relationship. This model yielded in vivo desensitization/internalization rates (0.2/min for quinpirole) consistent with published in vitro measurements. Overall, these results suggest that simultaneous PET/fMRI enables characterization of dynamic neuroreceptor adaptations in vivo, and may offer a first non-invasive method for assessing receptor desensitization and internalization. PMID:26388148

Comparison of the Effects of the Selective Estrogen Receptor Modulators Ospemifene, Raloxifene, and Tamoxifen on Breast Tissue in Ex Vivo Culture.

PubMed

Eigeliene, Natalija; Erkkola, Risto; Härkönen, Pirkko

2016-01-01

Explant tissue culture provides a model for studying the direct effects of steroid hormones, their analogs, and novel hormonally active compounds on normal freshly isolated human breast tissues (HBTs). For this purpose, pre- and postmenopausal HBTs can be maintained in this culture system. The results demonstrate that the morphological integrity of HBT explants can be maintained in tissue culture up to 2 weeks and expression of differentiation markers, steroid hormone receptors, proliferation and apoptosis ratios can be evaluated as a response to hormonal stimulation. This chapter describes an ex vivo culture model that we have applied to study the effects of various hormonally active substances, including 17β-estradiol and selective estrogen receptor modulators (SERMs), on normal human breast tissues.

The role of the dopamine D1 receptor in social cognition: studies using a novel genetic rat model­

PubMed Central

VandenBroeke, Marie; Youn, Jiun; Ellenbroek, Arabella K.; Karel, Peter; Shan, Ling; van Boxtel, Ruben; Ooms, Sharon; Balemans, Monique; Langedijk, Jacqueline; Muller, Mareike; Vriend, Gert; Cools, Alexander R.; Cuppen, Edwin; Ellenbroek, Bart A.

2016-01-01

ABSTRACT Social cognition is an endophenotype that is impaired in schizophrenia and several other (comorbid) psychiatric disorders. One of the modulators of social cognition is dopamine, but its role is not clear. The effects of dopamine are mediated through dopamine receptors, including the dopamine D1 receptor (Drd1). Because current Drd1 receptor agonists are not Drd1 selective, pharmacological tools are not sufficient to delineate the role of the Drd1. Here, we describe a novel rat model with a genetic mutation in Drd1 in which we measured basic behavioural phenotypes and social cognition. The I116S mutation was predicted to render the receptor less stable. In line with this computational prediction, this Drd1 mutation led to a decreased transmembrane insertion of Drd1, whereas Drd1 expression, as measured by Drd1 mRNA levels, remained unaffected. Owing to decreased transmembrane Drd1 insertion, the mutant rats displayed normal basic motoric and neurological parameters, as well as locomotor activity and anxiety-like behaviour. However, measures of social cognition like social interaction, scent marking, pup ultrasonic vocalizations and sociability, were strongly reduced in the mutant rats. This profile of the Drd1 mutant rat offers the field of neuroscience a novel genetic rat model to study a series of psychiatric disorders including schizophrenia, autism, depression, bipolar disorder and drug addiction. PMID:27483345

Exploring Pharmacological Mechanisms of Lavender (Lavandula angustifolia) Essential Oil on Central Nervous System Targets

PubMed Central

López, Víctor; Nielsen, Birgitte; Solas, Maite; Ramírez, Maria J.; Jäger, Anna K.

2017-01-01

Lavender essential oil is traditionally used and approved by the European Medicines Agency (EMA) as herbal medicine to relieve stress and anxiety. Some animal and clinical studies reveal positive results in models of anxiety and depression although very little research has been done on molecular mechanisms. Our work consisted of evaluating the effects of lavender (Lavandula angustifolia) essential oil on central nervous system well-established targets, such as MAO-A, SERT, GABAAand NMDA receptors as well as in vitro models of neurotoxicity. The results showed that lavender essential oil and its main components exert affinity for the glutamate NMDA-receptor in a dose-dependent manner with an IC50 value of 0.04 μl/mL for lavender oil. In addition, lavender and linalool were also able to bind the serotonin transporter (SERT) whereas they did not show affinity for GABAA-benzodiazepine receptor. In three different models of neurotoxicity, lavender did not enhance the neurotoxic insult and improved viability of SH-SY5Y cells treated with hydrogen peroxide. According to our data, the anxiolytic and antidepressant-like effects attributed to lavender may be due to an antagonism on the NMDA-receptor and inhibition of SERT. This study suggests that lavender essential oil may exert pharmacological properties via modulating the NMDA receptor, the SERT as well as neurotoxicity induced by hydrogen peroxide. PMID:28579958

Physiological significance of ghrelin revealed by studies using genetically engineered mouse models with modifications in the ghrelin system.

PubMed

Ariyasu, Hiroyuki; Akamizu, Takashi

2015-01-01

Ghrelin, an endogenous ligand for the growth hormone (GH) secretagogue receptor (GHS-R or ghrelin receptor), is a 28-amino acid acylated peptide mainly produced in the stomach. The pharmacological administration of ghrelin is known to exert diverse effects, such as stimulating GH secretion, promoting food intake, and increasing adiposity. In recent years, genetically engineered mouse models have provided important insights into the physiology of various hormones. In this review, we discuss current knowledge regarding the physiological significance of ghrelin on the basis of studies using genetically engineered mouse models with modifications in the ghrelin system.

The fruit fly Drosophila melanogaster unfolds the secrets of innate immunity.

PubMed

Rämet, Mika

2012-09-01

In 2011, the Nobel Prize in Physiology and Medicine was rewarded, in part, for research on the Drosophila immune response. The research described the role of the Drosophila Toll receptor in antifungal resistance, and the subsequent characterization of Toll-like receptors in mammals reshaped our understanding of the immune system. This review summarizes the potential of the Drosophila model and describes the path that has lead Drosophila to become an important model to study immunity. Drosophila melanogaster has been one of the most fruitful models to study innate immunity. © 2012 The Author(s)/Acta Paediatrica © 2012 Foundation Acta Paediatrica.

Antidepressant-induced Dopamine Receptor Dysregulation: A Valid Animal Model of Manic-Depressive Illness

PubMed Central

Demontis, Francesca; Serra, Francesca; Serra, Gino

2017-01-01

Background: Mania seems to be associated with an increased dopamine (DA) transmission. Antidepressant treatments can induce mania in humans and potentiated DA transmission in animals, by sensitizing DA D2 receptors in the mesolimbic system. We have suggested that the sensitization of D2 receptors may be responsible of antidepressant-induced mania. This review aims to report the experimental evidence that led to the hypothesis that antidepressant-induced DA receptors dysregulation can be considered an animal model of bipolar disorder. Methods: We reviewed papers reporting preclinical and clinical studies on the role of DA in the mechanism of action of antidepressant treatments and in the patho-physiology of mood disorders. Results: A number of preclinical and clinical evidence suggests that mania could be associated with an increased DA activity, while a reduced function of this neurotransmission might underlie depression. Chronic treatment with imipramine induces a sensitization of DA D2 receptors in the mesolimbic system, followed, after drug discontinuation, by a reduced sensitivity associated with an increased immobility time in forced swimming test of depression (FST). Blockade of glutamate NMDA receptors by memantine administration prevents the imipramine effect on DA receptors sensitivity and on the FST. Conclusion: We suggest that chronic treatment with antidepressants induces a behavioural syndrome that mimics mania (the sensitization of DA receptors), followed by depression (desensitization of DA receptors and increased immobility time in the FST), i.e. an animal model of bipolar disorder. Moreover the observation that memantine prevents the “bipolar-like” behavior, suggests that the drug may have an antimanic and mood stabilizing effect. Preliminary clinical observations support this hypothesis. PMID:28503114

A desensitization-selective potentiator of AMPA-type glutamate receptors

PubMed Central

Sekiguchi, Masayuki; Nishikawa, Kaori; Aoki, Shunsuke; Wada, Keiji

2002-01-01

We examined the effects of PEPA, an allosteric potentiator of AMPA receptors, on AMPA receptor kinetics. PEPA did not affect the deactivation of glutamate responses but potently attenuated the extent of receptor desensitization without slowing the onset of desensitization in most of the recombinant AMPA receptors (GluR1-flip, GluR1-flop, GluR3-flip, GluR3-flip + GluR2-flip, and GluR3-flop + GluR2-flop) expressed in Xenopus oocytes. For the GluR3-flop subunit, PEPA attenuated the extent of desensitization and only weakly prolonged deactivation (1.3 fold). PEPA did not significantly affect recovery from desensitization in oocytes expressing GluR3-flip, GluR1-flop, and GluR1-flop, but weakly accelerated (2.6 fold) recovery from desensitization in oocytes expressing GluR3-flop. PEPA's effect on desensitization of GluR3-flop-containing receptors is unique in that onset is very slow. Simulation studies using simplified kinetic models for AMPA receptors are utilized to explore the differential effects of PEPA on GluR3-flip and -flop. It is possible to simulate the action on GluR3-flip by modulating two rate constants in a 12-state kinetic model. For simulation of the action on GluR3-flop, the 12-state kinetic model is not enough, and it is necessary to invoke a 13th state, a PEPA-bound receptor to which glutamate cannot bind. These results suggest that attenuation of extent of desensitization represents the principal mechanism underlying the potentiation of AMPA receptors by PEPA, and that PEPA exhibits different mechanisms with respect to GluR3-flip and GluR3-flop. PMID:12145103

A novel thromboxane receptor antagonist, nstpbp5185, inhibits platelet aggregation and thrombus formation in animal models.

PubMed

Huang, Shiu-Wen; Kuo, Heng-Lan; Hsu, Ming-Tsung; Tseng, Yufeng Jane; Lin, Shu-Wha; Kuo, Sheng-Chu; Peng, Hui-Chin; Lien, Jin-Cherng; Huang, Tur-Fu

2016-08-01

A novel benzimidazole derivative, nstpbp5185, was discovered through in vitro and in vivo evaluations for antiplatelet activity. Thromaboxane receptor (TP) is important in vascular physiology, haemostasis and pathophysiological thrombosis. Nstpbp5185 concentration-dependently inhibited human platelet aggregation caused by collagen, arachidonic acid and U46619. Nstpbp5185 caused a right-shift of the concentration-response curve of U46619 and competitively inhibited the binding of 3H-SQ-29548 to TP receptor expressed on HEK-293 cells, with an IC50 of 0.1 µM, indicating that nstpbp5185 is a TP antagonist. In murine thrombosis models, nstpbp5185 significantly prolonged the latent period in triggering platelet plug formation in mesenteric and FeCl3-induced thrombi formation, and increased the survival rate in pulmonary embolism model with less bleeding than aspirin. This study suggests nstpbp5185, an orally selective anti-thrombotic agent, acting through blockade of TXA2 receptor, may be efficacious for prevention or treatment of pathologic thrombosis.

Peripheral 5-HT3 Receptors Are Involved in the Antinociceptive Effect of Bunodosine 391.

PubMed

Ferreira Junior, Wilson Alves; Zaharenko, Andre Junqueira; Kazuma, Kohei; Picolo, Gisele; Gutierrez, Vanessa Pacciari; de Freitas, Jose Carlos; Konno, Katsuhiro; Cury, Yara

2017-12-27

Bunodosine 391 (BDS 391), a low molecular weight compound isolated from the sea anemone Bunodosoma cangicum , increases the nociceptive threshold and inhibits inflammatory hyperalgesia. Serotonin receptors are involved in those effects. In this study, we have expanded the characterization of the antinociceptive effect of BDS 391 demonstrating that, in rats: (a) the compound inhibits (1.2-12 ng/paw) overt pain, in the formalin test, and mechanical hyperalgesia (0.6-6.0 ng/paw) detected in a model of neuropathic pain; (b) intraplantar administration of ondansetron, a selective 5-HT3 receptor antagonist, blocks the effect of BDS 391, whereas ketanserin, a 5-HT2 receptor antagonist, partially reversed this effect, indicating the involvement of peripheral 5-HT2 and 5-HT3 receptors in BDS 391 antinociception; and (c) in binding assay studies, BDS 391 was not able to displace the selective 5-HT receptor antagonists, suggesting that this compound does not directly bind to these receptors. The effect of biguanide, a selective 5-HT3 receptor agonist, was also evaluated. The agonist inhibited the formalin's nociceptive response, supporting an antinociceptive role for 5-HT3 receptors. Our study is the first one to show that a non-peptidic low molecular weight compound obtained from a sea anemone is able to induce antinociception and that activation of peripheral 5-HT3 receptors contributes to this effect.

Peripheral 5-HT3 Receptors Are Involved in the Antinociceptive Effect of Bunodosine 391

PubMed Central

Ferreira Junior, Wilson Alves; Zaharenko, Andre Junqueira; Kazuma, Kohei; Picolo, Gisele; Gutierrez, Vanessa Pacciari; de Freitas, Jose Carlos; Konno, Katsuhiro

2017-01-01

Bunodosine 391 (BDS 391), a low molecular weight compound isolated from the sea anemone Bunodosoma cangicum, increases the nociceptive threshold and inhibits inflammatory hyperalgesia. Serotonin receptors are involved in those effects. In this study, we have expanded the characterization of the antinociceptive effect of BDS 391 demonstrating that, in rats: (a) the compound inhibits (1.2–12 ng/paw) overt pain, in the formalin test, and mechanical hyperalgesia (0.6–6.0 ng/paw) detected in a model of neuropathic pain; (b) intraplantar administration of ondansetron, a selective 5-HT3 receptor antagonist, blocks the effect of BDS 391, whereas ketanserin, a 5-HT2 receptor antagonist, partially reversed this effect, indicating the involvement of peripheral 5-HT2 and 5-HT3 receptors in BDS 391 antinociception; and (c) in binding assay studies, BDS 391 was not able to displace the selective 5-HT receptor antagonists, suggesting that this compound does not directly bind to these receptors. The effect of biguanide, a selective 5-HT3 receptor agonist, was also evaluated. The agonist inhibited the formalin’s nociceptive response, supporting an antinociceptive role for 5-HT3 receptors. Our study is the first one to show that a non-peptidic low molecular weight compound obtained from a sea anemone is able to induce antinociception and that activation of peripheral 5-HT3 receptors contributes to this effect. PMID:29280949

Molecular mechanism of action of pharmacoperone rescue of misrouted GPCR mutants: the GnRH receptor.

PubMed

Janovick, Jo Ann; Patny, Akshay; Mosley, Ralph; Goulet, Mark T; Altman, Michael D; Rush, Thomas S; Cornea, Anda; Conn, P Michael

2009-02-01

The human GnRH receptor (hGnRHR), a G protein-coupled receptor, is a useful model for studying pharmacological chaperones (pharmacoperones), drugs that rescue misfolded and misrouted protein mutants and restore them to function. This technique forms the basis of a therapeutic approach of rescuing mutants associated with human disease and restoring them to function. The present study relies on computational modeling, followed by site-directed mutagenesis, assessment of ligand binding, effector activation, and confocal microscopy. Our results show that two different chemical classes of pharmacoperones act to stabilize hGnRHR mutants by bridging residues D(98) and K(121). This ligand-mediated bridge serves as a surrogate for a naturally occurring and highly conserved salt bridge (E(90)-K(121)) that stabilizes the relation between transmembranes 2 and 3, which is required for passage of the receptor through the cellular quality control system and to the plasma membrane. Our model was used to reveal important pharmacophoric features, and then identify a novel chemical ligand, which was able to rescue a D(98) mutant of the hGnRHR that could not be rescued as effectively by previously known pharmacoperones.

Cannabidiol: Pharmacology and potential therapeutic role in epilepsy and other neuropsychiatric disorders

PubMed Central

Devinsky, Orrin; Cilio, Maria Roberta; Cross, Helen; Fernandez-Ruiz, Javier; French, Jacqueline; Hill, Charlotte; Katz, Russell; Di Marzo, Vincenzo; Jutras-Aswad, Didier; Notcutt, William George; Martinez-Orgado, Jose; Robson, Philip J.; Rohrback, Brian G.; Thiele, Elizabeth; Whalley, Benjamin; Friedman, Daniel

2015-01-01

Objective To present a summary of current scientific evidence about the cannabinoid, cannabidiol (CBD) with regards to their relevance to epilepsy and other selected neuropsychiatric disorders. Methods We summarize the presentations from a conference in which invited participants reviewed relevant aspects of the physiology, mechanisms of action, pharmacology and data from studies with animal models and human subjects. Results Cannabis has been used to treat disease since ancient times. Δ9-THC is the major psychoactive ingredient and cannabidiol (CBD) is the major non-psychoactive ingredient in cannabis. Cannabis and Δ9-THC are anticonvulsant in most animal models but can be proconvulsant in some healthy animals. Psychotropic effects of Δ9-THC limit tolerability. CBD is anticonvulsant in many acute animal models but there is limited data in chronic models. The antiepileptic mechanisms of CBD are not known, but may include effects on the equilibrative nucleoside transporter; the orphan G-protein-coupled receptor GPR55; the transient receptor potential of melastatin type 8 channel; the 5-HT1a receptor; the α3 and α1 glycine receptors; and the transient receptor potential of ankyrin type 1 channel. CBD has neuroprotective and anti-inflammatory effects. CBD appears to be well tolerated in humans but small and methodologically limited studies of CBD in human epilepsy have been inconclusive. More recent anecdotal reports of high-ratio CBD:Δ9-THC medical marijuana have claimed efficacy, but studies were not controlled. Significance CBD bears investigation in epilepsy and other neuropsychiatric disorders, including anxiety, schizophrenia, addiction and neonatal hypoxic-ischemic encephalopathy. However, we lack data from well-powered double-blind randomized, controlled studies on the efficacy of pure CBD for any disorder. Initial dose-tolerability and double-blind randomized, controlled studies focusing on target intractable epilepsy populations such as patients with Dravet and Lennox-Gastaut syndromes are being planned. Trials in other treatment-resistant epilepsies may also be warranted. PMID:24854329

Perampanel, an antagonist of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, for the treatment of epilepsy: studies in human epileptic brain and nonepileptic brain and in rodent models.

PubMed

Zwart, R; Sher, E; Ping, X; Jin, X; Sims, J R; Chappell, A S; Gleason, S D; Hahn, P J; Gardinier, K; Gernert, D L; Hobbs, J; Smith, J L; Valli, S N; Witkin, J M

2014-10-01

Perampanel [Fycompa, 2-(2-oxo-1-phenyl-5-pyridin-2-yl-1,2-dihydropyridin-3-yl)benzonitrile hydrate 4:3; Eisai Inc., Woodcliff Lake, NJ] is an AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor antagonist used as an adjunctive treatment of partial-onset seizures. We asked whether perampanel has AMPA receptor antagonist activity in both the cerebral cortex and hippocampus associated with antiepileptic efficacy and also in the cerebellum associated with motor side effects in rodent and human brains. We also asked whether epileptic or nonepileptic human cortex is similarly responsive to AMPA receptor antagonism by perampanel. In rodent models, perampanel decreased epileptic-like activity in multiple seizure models. However, doses of perampanel that had anticonvulsant effects were within the same range as those engendering motor side effects. Perampanel inhibited native rat and human AMPA receptors from the hippocampus as well as the cerebellum that were reconstituted into Xenopus oocytes. In addition, with the same technique, we found that perampanel inhibited AMPA receptors from hippocampal tissue that had been removed from a patient who underwent surgical resection for refractory epilepsy. Perampanel inhibited AMPA receptor-mediated ion currents from all the tissues investigated with similar potency (IC50 values ranging from 2.6 to 7.0 μM). Cortical slices from the left temporal lobe derived from the same patient were studied in a 60-microelectrode array. Large field potentials were evoked on at least 45 channels of the array, and 10 μM perampanel decreased their amplitude and firing rate. Perampanel also produced a 33% reduction in the branching parameter, demonstrating the effects of perampanel at the network level. These data suggest that perampanel blocks AMPA receptors globally across the brain to account for both its antiepileptic and side-effect profile in rodents and epileptic patients. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Comodulation of dopamine and serotonin on prefrontal cortical rhythms: a theoretical study

PubMed Central

Wang, Da-Hui; Wong-Lin, KongFatt

2013-01-01

The prefrontal cortex (PFC) is implicated to play an important role in cognitive control. Abnormal PFC activities and rhythms have been observed in some neurological and neuropsychiatric disorders, and evidences suggest influences from the neuromodulators dopamine (DA) and serotonin (5-HT). Despite the high level of interest in these brain systems, the combined effects of DA and 5-HT modulation on PFC dynamics remain unknown. In this work, we build a mathematical model that incorporates available experimental findings to systematically study the comodulation of DA and 5-HT on the network behavior, focusing on beta and gamma band oscillations. Single neuronal model shows pyramidal cells with 5-HT1A and 2A receptors can be non-monotonically modulated by 5-HT. Two-population excitatory-inhibitory type network consisting of pyramidal cells with D1 receptors can provide rich repertoires of oscillatory behavior. In particular, 5-HT and DA can modulate the amplitude and frequency of the oscillations, which can emerge or cease, depending on receptor types. Certain receptor combinations are conducive for the robustness of the oscillatory regime, or the existence of multiple discrete oscillatory regimes. In a multi-population heterogeneous model that takes into account possible combination of receptors, we demonstrate that robust network oscillations require high DA concentration. We also show that selective D1 receptor antagonists (agonists) tend to suppress (enhance) network oscillations, increase the frequency from beta toward gamma band, while selective 5-HT1A antagonists (agonists) act in opposite ways. Selective D2 or 5-HT2A receptor antagonists (agonists) can lead to decrease (increase) in oscillation amplitude, but only 5-HT2A antagonists (agonists) can increase (decrease) the frequency. These results are comparable to some pharmacological effects. Our work illustrates the complex mechanisms of DA and 5-HT when operating simultaneously through multiple receptors. PMID:23935568

Anti-inflammatory effects of PGE2 in the lung: role of the EP4 receptor subtype.

PubMed

Birrell, Mark A; Maher, Sarah A; Dekkak, Bilel; Jones, Victoria; Wong, Sissie; Brook, Peter; Belvisi, Maria G

2015-08-01

Asthma and chronic obstructive pulmonary disease (COPD) are chronic inflammatory diseases of the airway. Current treatment options (long acting β-adrenoceptor agonists and glucocorticosteroids) are not optimal as they are only effective in certain patient groups and safety concerns exist regarding both compound classes. Therefore, novel bronchodilator and anti-inflammatory strategies are being pursued. Prostaglandin E2 (PGE2) is an arachidonic acid-derived eicosanoid produced by the lung which acts on four different G-protein coupled receptors (EP1-4) to cause an array of beneficial and deleterious effects. The aim of this study was to identify the EP receptor mediating the anti-inflammatory actions of PGE2 in the lung using a range of cell-based assays and in vivo models. It was demonstrated in three distinct model systems (innate stimulus, lipopolysaccharide (LPS); allergic response, ovalbumin (OVA); inhaled pollutant, cigarette smoke) that mice missing functional EP4 (Ptger4(-/-)) receptors had higher levels of airway inflammation, suggesting that endogenous PGE2 was suppressing inflammation via EP4 receptor activation. Cell-based assay systems (murine and human monocytes/alveolar macrophages) demonstrated that PGE2 inhibited cytokine release from LPS-stimulated cells and that this was mimicked by an EP4 (but not EP1-3) receptor agonist and inhibited by an EP4 receptor antagonist. The anti-inflammatory effect occurred at the transcriptional level and was via the adenylyl cyclase/cAMP/ cAMP-dependent protein kinase (PKA) axis. This study demonstrates that EP4 receptor activation is responsible for the anti-inflammatory activity of PGE2 in a range of disease relevant models and, as such, could represent a novel therapeutic target for chronic airway inflammatory conditions. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

How to Choose the Suitable Template for Homology Modelling of GPCRs: 5-HT7 Receptor as a Test Case.

PubMed

Shahaf, Nir; Pappalardo, Matteo; Basile, Livia; Guccione, Salvatore; Rayan, Anwar

2016-09-01

G protein-coupled receptors (GPCRs) are a super-family of membrane proteins that attract great pharmaceutical interest due to their involvement in almost every physiological activity, including extracellular stimuli, neurotransmission, and hormone regulation. Currently, structural information on many GPCRs is mainly obtained by the techniques of computer modelling in general and by homology modelling in particular. Based on a quantitative analysis of eighteen antagonist-bound, resolved structures of rhodopsin family "A" receptors - also used as templates to build 153 homology models - it was concluded that a higher sequence identity between two receptors does not guarantee a lower RMSD between their structures, especially when their pair-wise sequence identity (within trans-membrane domain and/or in binding pocket) lies between 25 % and 40 %. This study suggests that we should consider all template receptors having a sequence identity ≤50 % with the query receptor. In fact, most of the GPCRs, compared to the currently available resolved structures of GPCRs, fall within this range and lack a correlation between structure and sequence. When testing suitability for structure-based drug design, it was found that choosing as a template the most similar resolved protein, based on sequence resemblance only, led to unsound results in many cases. Molecular docking analyses were carried out, and enrichment factors as well as attrition rates were utilized as criteria for assessing suitability for structure-based drug design. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Structural insights into transient receptor potential vanilloid type 1 (TRPV1) from homology modeling, flexible docking, and mutational studies.

PubMed

Lee, Jin Hee; Lee, Yoonji; Ryu, HyungChul; Kang, Dong Wook; Lee, Jeewoo; Lazar, Jozsef; Pearce, Larry V; Pavlyukovets, Vladimir A; Blumberg, Peter M; Choi, Sun

2011-04-01

The transient receptor potential vanilloid subtype 1 (TRPV1) is a non-selective cation channel composed of four monomers with six transmembrane helices (TM1-TM6). TRPV1 is found in the central and peripheral nervous system, and it is an important therapeutic target for pain relief. We describe here the construction of a tetrameric homology model of rat TRPV1 (rTRPV1). We experimentally evaluated by mutational analysis the contribution of residues of rTRPV1 contributing to ligand binding by the prototypical TRPV1 agonists, capsaicin and resiniferatoxin (RTX). We then performed docking analysis using our homology model. The docking results with capsaicin and RTX showed that our homology model was reliable, affording good agreement with our mutation data. Additionally, the binding mode of a simplified RTX (sRTX) ligand as predicted by the modeling agreed well with those of capsaicin and RTX, accounting for the high binding affinity of the sRTX ligand for TRPV1. Through the homology modeling, docking and mutational studies, we obtained important insights into the ligand-receptor interactions at the molecular level which should prove of value in the design of novel TRPV1 ligands.

Structural insights into transient receptor potential vanilloid type 1 (TRPV1) from homology modeling, flexible docking, and mutational studies

NASA Astrophysics Data System (ADS)

Lee, Jin Hee; Lee, Yoonji; Ryu, HyungChul; Kang, Dong Wook; Lee, Jeewoo; Lazar, Jozsef; Pearce, Larry V.; Pavlyukovets, Vladimir A.; Blumberg, Peter M.; Choi, Sun

2011-04-01

The transient receptor potential vanilloid subtype 1 (TRPV1) is a non-selective cation channel composed of four monomers with six transmembrane helices (TM1-TM6). TRPV1 is found in the central and peripheral nervous system, and it is an important therapeutic target for pain relief. We describe here the construction of a tetrameric homology model of rat TRPV1 (rTRPV1). We experimentally evaluated by mutational analysis the contribution of residues of rTRPV1 contributing to ligand binding by the prototypical TRPV1 agonists, capsaicin and resiniferatoxin (RTX). We then performed docking analysis using our homology model. The docking results with capsaicin and RTX showed that our homology model was reliable, affording good agreement with our mutation data. Additionally, the binding mode of a simplified RTX (sRTX) ligand as predicted by the modeling agreed well with those of capsaicin and RTX, accounting for the high binding affinity of the sRTX ligand for TRPV1. Through the homology modeling, docking and mutational studies, we obtained important insights into the ligand-receptor interactions at the molecular level which should prove of value in the design of novel TRPV1 ligands.

Endocytosis and membrane receptor internalization: implication of F-BAR protein Carom.

PubMed

Xu, Yanjie; Xia, Jixiang; Liu, Suxuan; Stein, Sam; Ramon, Cueto; Xi, Hang; Wang, Luqiao; Xiong, Xinyu; Zhang, Lixiao; He, Dingwen; Yang, William; Zhao, Xianxian; Cheng, Xiaoshu; Yang, Xiaofeng; Wang, Hong

2017-03-01

Endocytosis is a cellular process mostly responsible for membrane receptor internalization. Cell membrane receptors bind to their ligands and form a complex which can be internalized. We previously proposed that F-BAR protein initiates membrane curvature and mediates endocytosis via its binding partners. However, F-BAR protein partners involved in membrane receptor endocytosis and the regulatory mechanism remain unknown. In this study, we established database mining strategies to explore mechanisms underlying receptor-related endocytosis. We identified 34 endocytic membrane receptors and 10 regulating proteins in clathrin-dependent endocytosis (CDE), a major process of membrane receptor internalization. We found that F-BAR protein FCHSD2 (Carom) may facilitate endocytosis via 9 endocytic partners. Carom is highly expressed, along with highly expressed endocytic membrane receptors and partners, in endothelial cells and macrophages. We established 3 models of Carom-receptor complexes and their intracellular trafficking based on protein interaction and subcellular localization. We conclude that Carom may mediate receptor endocytosis and transport endocytic receptors to the cytoplasm for receptor signaling and lysosome/proteasome degradation, or to the nucleus for RNA processing, gene transcription and DNA repair.

An automated system for the analysis of G protein-coupled receptor transmembrane binding pockets: alignment, receptor-based pharmacophores, and their application.

PubMed

Kratochwil, Nicole A; Malherbe, Pari; Lindemann, Lothar; Ebeling, Martin; Hoener, Marius C; Mühlemann, Andreas; Porter, Richard H P; Stahl, Martin; Gerber, Paul R

2005-01-01

G protein-coupled receptors (GPCRs) share a common architecture consisting of seven transmembrane (TM) domains. Various lines of evidence suggest that this fold provides a generic binding pocket within the TM region for hosting agonists, antagonists, and allosteric modulators. Here, a comprehensive and automated method allowing fast analysis and comparison of these putative binding pockets across the entire GPCR family is presented. The method relies on a robust alignment algorithm based on conservation indices, focusing on pharmacophore-like relationships between amino acids. Analysis of conservation patterns across the GPCR family and alignment to the rhodopsin X-ray structure allows the extraction of the amino acids lining the TM binding pocket in a so-called ligand binding pocket vector (LPV). In a second step, LPVs are translated to simple 3D receptor pharmacophore models, where each amino acid is represented by a single spherical pharmacophore feature and all atomic detail is omitted. Applications of the method include the assessment of selectivity issues, support of mutagenesis studies, and the derivation of rules for focused screening to identify chemical starting points in early drug discovery projects. Because of the coarseness of this 3D receptor pharmacophore model, however, meaningful scoring and ranking procedures of large sets of molecules are not justified. The LPV analysis of the trace amine-associated receptor family and its experimental validation is discussed as an example. The value of the 3D receptor model is demonstrated for a class C GPCR family, the metabotropic glutamate receptors.

Quantitative pharmacological analysis of antagonist binding kinetics at CRF1 receptors in vitro and in vivo

PubMed Central

Ramsey, Simeon J; Attkins, Neil J; Fish, Rebecca; van der Graaf, Piet H

2011-01-01

BACKGROUND AND PURPOSE A series of novel non-peptide corticotropin releasing factor type-1 receptor (CRF1) antagonists were found to display varying degrees of insurmountable and non-competitive behaviour in functional in vitro assays. We describe how we attempted to relate this behaviour to ligand receptor-binding kinetics in a quantitative manner and how this resulted in the development and implementation of an efficient pharmacological screening method based on principles described by Motulsky and Mahan. EXPERIMENTAL APPROACH A non-equilibrium binding kinetic assay was developed to determine the receptor binding kinetics of non-peptide CRF1 antagonists. Nonlinear, mixed-effects modelling was used to obtain estimates of the compounds association and dissociation rates. We present an integrated pharmacokinetic–pharmacodynamic (PKPD) approach, whereby the time course of in vivo CRF1 receptor binding of novel compounds can be predicted on the basis of in vitro assays. KEY RESULTS The non-competitive antagonist behaviour appeared to be correlated to the CRF1 receptor off-rate kinetics. The integrated PKPD model suggested that, at least in a qualitative manner, the in vitro assay can be used to triage and select compounds for further in vivo investigations. CONCLUSIONS AND IMPLICATIONS This study provides evidence for a link between ligand offset kinetics and insurmountable/non-competitive antagonism at the CRF1 receptor. The exact molecular pharmacological nature of this association remains to be determined. In addition, we have developed a quantitative framework to study and integrate in vitro and in vivo receptor binding kinetic behaviour of CRF1 receptor antagonists in an efficient manner in a drug discovery setting. PMID:21449919

Novel targeted approaches to treating biliary tract cancer: the dual epidermal growth factor receptor and ErbB-2 tyrosine kinase inhibitor NVP-AEE788 is more efficient than the epidermal growth factor receptor inhibitors gefitinib and erlotinib.

PubMed

Wiedmann, Marcus; Feisthammel, Jürgen; Blüthner, Thilo; Tannapfel, Andrea; Kamenz, Thomas; Kluge, Annett; Mössner, Joachim; Caca, Karel

2006-08-01

Aberrant activation of the epidermal growth factor receptor is frequently observed in neoplasia, notably in tumors of epithelial origin. Attempts to treat such tumors with epidermal growth factor receptor antagonists resulted in remarkable success in recent studies. Little is known, however, about the efficacy of this therapy in biliary tract cancer. Protein expression of epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 was assessed in seven human biliary tract cancer cell lines by immunoblotting. In addition, histological sections from 19 patients with extrahepatic cholangiocarcinoma were analyzed for epidermal growth factor receptor, ErbB-2 and vascular endothelial growth factor receptor-2 expression by immunohistochemistry. Moreover, we sequenced the cDNA products representing the entire epidermal growth factor receptor coding region of the seven cell lines, and searched for genomic epidermal growth factor receptor amplifications and polysomy by fluorescence in-situ hybridization. Cell growth inhibition by gefitinib erlotinib and NVP-AEE788 was studied in vitro by automated cell counting. In addition, the anti-tumoral effect of erlotinib and NVP-AEE788 was studied in a chimeric mouse model. The anti-tumoral drug mechanism in this model was assessed by MIB-1 antibody staining, terminal deoxynucleotidyl transfer-mediated dUTP nick end-labelling assay, von Willebrand factor staining, and immunoblotting for p-p42/44 (p-Erk1/2, p-MAPK) and p-AKT. Immunoblotting revealed expression of epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 in all biliary tract cancer cell lines. EGFR was detectable in six of 19 (32%) extrahepatic human cholangiocarcinoma tissue samples, ErbB-2 in 16 of 19 (84%), and vascular endothelial growth factor receptor-2 in nine of 19 (47%). Neither epidermal growth factor receptor mutations nor amplifications or polysomy were found in the seven biliary tract cancer cell lines. Gefitinib, erlotinib and NVP-AEE788 caused a significant growth inhibition in vitro; however, there was a significant difference in efficacy (NVP-AEE788>erlotinib>gefitinib). After 14 days of in-vivo treatment, using the chimeric mouse model, tumors had a significantly reduced volume and mass after NVP-AEE788, but not after erlotinib treatment, as compared with placebo. Reduction of proliferation (signalling via the mitogen-activated protein kinase pathway), induction of apoptosis and inhibition of angiogenesis were the main mechanisms of drug action. No significant reduction of anti-apoptotic AKT phosphorylation, however, occurred, which may be a possible counter mechanism of the tumor. Epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 expression was detectable in biliary tract cancer, and receptor inhibition exerts marked effects on tumor growth in vitro and in vivo, which was strongest for the dual EGFR/ErbB-2 inhibitor NVP-AEE788. Therefore, further clinical evaluation of this new drug for the treatment of biliary tract cancer is recommended.

Type I interferon signals in macrophages and dendritic cells control dengue virus infection: implications for a new mouse model to test dengue vaccines.

PubMed

Züst, Roland; Toh, Ying-Xiu; Valdés, Iris; Cerny, Daniela; Heinrich, Julia; Hermida, Lisset; Marcos, Ernesto; Guillén, Gerardo; Kalinke, Ulrich; Shi, Pei-Yong; Fink, Katja

2014-07-01

Dengue virus (DENV) infects an estimated 400 million people every year, causing prolonged morbidity and sometimes mortality. Development of an effective vaccine has been hampered by the lack of appropriate small animal models; mice are naturally not susceptible to DENV and only become infected if highly immunocompromised. Mouse models lacking both type I and type II interferon (IFN) receptors (AG129 mice) or the type I IFN receptor (IFNAR(-/-) mice) are susceptible to infection with mouse-adapted DENV strains but are severely impaired in mounting functional immune responses to the virus and thus are of limited use for study. Here we used conditional deletion of the type I IFN receptor (IFNAR) on individual immune cell subtypes to generate a minimally manipulated mouse model that is susceptible to DENV while retaining global immune competence. Mice lacking IFNAR expression on CD11c(+) dendritic cells and LysM(+) macrophages succumbed completely to DENV infection, while mice deficient in the receptor on either CD11c(+) or LysM(+) cells were susceptible to infection but often resolved viremia and recovered fully from infection. Conditional IFNAR mice responded with a swift and strong CD8(+) T-cell response to viral infection, compared to a weak response in IFNAR(-/-) mice. Furthermore, mice lacking IFNAR on either CD11c(+) or LysM(+) cells were also sufficiently immunocompetent to raise a protective immune response to a candidate subunit vaccine against DENV-2. These data demonstrate that mice with conditional deficiencies in expression of the IFNAR represent improved models for the study of DENV immunology and screening of vaccine candidates. Dengue virus infects 400 million people every year worldwide, causing 100 million clinically apparent infections, which can be fatal if untreated. Despite many years of research, there are no effective vaccine and no antiviral treatment available for dengue. Development of vaccines has been hampered in particular by the lack of a suitable small animal model. Mouse models used to test dengue vaccine are deficient in interferon (IFN) type I signaling and severely immunocompromised and therefore likely not ideal for the testing of vaccines. In this study, we explored alternative models lacking the IFN receptor only on certain cell types. We show that mice lacking the IFN receptor on either CD11c- or LysM-expressing cells (conditional IFNAR mice) are susceptible to dengue virus infection. Importantly, we demonstrate that conditional IFN receptor knockout mice generate a better immune response to live virus and a candidate dengue vaccine compared to IFNAR mice and are resistant to subsequent challenge. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Design, synthesis, pharmacological evaluation and in silico ADMET prediction of novel substituted benzimidazole derivatives as angiotensin II-AT1 receptor antagonists based on predictive 3D QSAR models.

PubMed

Vyas, V K; Gupta, N; Ghate, M; Patel, S

2014-01-01

In this study we designed novel substituted benzimidazole derivatives and predicted their absorption, distribution, metabolism, excretion and toxicity (ADMET) properties, based on a predictive 3D QSAR study on 132 substituted benzimidazoles as AngII-AT1 receptor antagonists. The two best predicted compounds were synthesized and evaluated for AngII-AT1 receptor antagonism. Three different alignment tools for comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were used. The best 3D QSAR models were obtained using the rigid body (Distill) alignment method. CoMFA and CoMSIA models were found to be statistically significant with leave-one-out correlation coefficients (q(2)) of 0.630 and 0.623, respectively, cross-validated coefficients (r(2)cv) of 0.651 and 0.630, respectively, and conventional coefficients of determination (r(2)) of 0.848 and 0.843, respectively. 3D QSAR models were validated using a test set of 24 compounds, giving satisfactory predicted results (r(2)pred) of 0.727 and 0.689 for the CoMFA and CoMSIA models, respectively. We have identified some key features in substituted benzimidazole derivatives, such as lipophilicity and H-bonding at the 2- and 5-positions of the benzimidazole nucleus, respectively, for AT1 receptor antagonistic activity. We designed 20 novel substituted benzimidazole derivatives and predicted their activity. In silico ADMET properties were also predicted for these designed molecules. Finally, the compounds with best predicted activity were synthesized and evaluated for in vitro angiotensin II-AT1 receptor antagonism.

Differentiation in the angiotensin II receptor 1 blocker class on autonomic function.

PubMed

Krum, H

2001-09-01

Autonomic function is disordered in cardiovascular disease states such as chronic heart failure (CHF) and hypertension. Interactions between the renin-angiotensin-aldosterone system (RAAS) and the sympathetic nervous system (SNS) may potentially occur at a number of sites. These include central sites (eg, rostral ventrolateral medulla), at the level of baroreflex control, and at the sympathetic prejunctional angiotensin II receptor 1 (AT(1)) receptor, which is facilitatory for norepinephrine release from the sympathetic nerve terminal. Therefore, drugs that block the RAAS may be expected to improve autonomic dysfunction in cardiovascular disease states. In order to test the hypothesis that RAAS inhibition directly reduces SNS activity, a pithed rat model of sympathetic stimulation has been established. In this model, an increase in frequency of stimulation results in a pressor response that is sympathetically mediated and highly reproducible. This pressor response is enhanced in the presence of angiotensin II and is reduced in the presence of nonselective AIIRAs that block both AT(1) and AT(2) receptor subtypes (eg, saralasin). AT(1)-selective antagonists have also been studied in this model, at pharmacologically relevant doses. In one such study, only the AT(1) blocker eprosartan reduced sympathetically stimulated increases in blood pressure, whereas comparable doses of losartan, valsartan, and irbesartan did not. The reason(s) for the differences between eprosartan and other agents of this class on sympathetic modulation are not clear, but may relate to the chemical structure of the drug (a non- biphenyl tetrazole structure that is chemically distinct from the structure of other AIIRAs), receptor binding characteristics (competitive), or unique effects on presynaptic AT(1) receptors.

Development of a population pharmacokinetic model to predict brain distribution and dopamine D2 receptor occupancy of raclopride in non-anesthetized rat.

PubMed

Wong, Yin Cheong; Ilkova, Trayana; van Wijk, Rob C; Hartman, Robin; de Lange, Elizabeth C M

2018-01-01

Raclopride is a selective antagonist of the dopamine D2 receptor. It is one of the most frequently used in vivo D2 tracers (at low doses) for assessing drug-induced receptor occupancy (RO) in animals and humans. It is also commonly used as a pharmacological blocker (at high doses) to occupy the available D2 receptors and antagonize the action of dopamine or drugs on D2 in preclinical studies. The aims of this study were to comprehensively evaluate its pharmacokinetic (PK) profiles in different brain compartments and to establish a PK-RO model that could predict the brain distribution and RO of raclopride in the freely moving rat using a LC-MS based approach. Rats (n=24) received a 10-min IV infusion of non-radiolabeled raclopride (1.61μmol/kg, i.e. 0.56mg/kg). Plasma and the brain tissues of striatum (with high density of D2 receptors) and cerebellum (with negligible amount of D2 receptors) were collected. Additional microdialysis experiments were performed in some rats (n=7) to measure the free drug concentration in the extracellular fluid of the striatum and cerebellum. Raclopride concentrations in all samples were analyzed by LC-MS. A population PK-RO model was constructed in NONMEM to describe the concentration-time profiles in the unbound plasma, brain extracellular fluid and brain tissue compartments and to estimate the RO based on raclopride-D2 receptor binding kinetics. In plasma raclopride showed a rapid distribution phase followed by a slower elimination phase. The striatum tissue concentrations were consistently higher than that of cerebellum tissue throughout the whole experimental period (10-h) due to higher non-specific tissue binding and D2 receptor binding in the striatum. Model-based simulations accurately predicted the literature data on rat plasma PK, brain tissue PK and D2 RO at different time points after intravenous or subcutaneous administration of raclopride at tracer dose (RO <10%), sub-pharmacological dose (RO 10%-30%) and pharmacological dose (RO >30%). For the first time a predictive model that could describe the quantitative in vivo relationship between dose, PK and D2 RO of raclopride in non-anesthetized rat was established. The PK-RO model could facilitate the selection of optimal dose and dosing time when raclopride is used as tracer or as pharmacological blocker in various rat studies. The LC-MS based approach, which doses and quantifies a non-radiolabeled tracer, could be useful in evaluating the systemic disposition and brain kinetics of tracers. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

The cubic ternary complex receptor-occupancy model. III. resurrecting efficacy.

PubMed

Weiss, J M; Morgan, P H; Lutz, M W; Kenakin, T P

1996-08-21

Early work in pharmacology characterized the interaction of receptors and ligands in terms of two parameters, affinity and efficacy, an approach we term the bipartite view. A precise formulation of efficacy only exists for very simple pharmacological models. Here we extend the notion of efficacy to models that incorporate receptor activation and G-protein coupling. Using the cubic ternary complex model, we show that efficacy is not purely a property of the ligand-receptor interaction; it also depends upon the distributional details of the receptor species in the native receptor ensemble. This suggests a distinction between what we call potential efficacy (a vector) and realized efficacy (a scalar). To each receptor species in the native receptor ensemble we assign a part-worth utility; taken together these utilities comprise the potential efficacy vector. Realized efficacy is the expectation of these part-worth utilities with respect to the frequency distribution of receptor species in the native receptor ensemble. In the parlance of statistical decision theory, the binding of a ligand to a receptor ensemble is a random prospect and realized efficacy is the utility of this prospect. We explore the implications that our definition of efficacy has for understanding agonism and in assessing the legitimacy of the bipartite view in pharmacology.

Comparison of receptor models for source apportionment of the PM10 in Zaragoza (Spain).

PubMed

Callén, M S; de la Cruz, M T; López, J M; Navarro, M V; Mastral, A M

2009-08-01

Receptor models are useful to understand the chemical and physical characteristics of air pollutants by identifying their sources and by estimating contributions of each source to receptor concentrations. In this work, three receptor models based on principal component analysis with absolute principal component scores (PCA-APCS), Unmix and positive matrix factorization (PMF) were applied to study for the first time the apportionment of the airborne particulate matter less or equal than 10microm (PM10) in Zaragoza, Spain, during 1year sampling campaign (2003-2004). The PM10 samples were characterized regarding their concentrations in inorganic components: trace elements and ions and also organic components: polycyclic aromatic hydrocarbons (PAH) not only in the solid phase but also in the gas phase. A comparison of the three receptor models was carried out in order to do a more robust characterization of the PM10. The three models predicted that the major sources of PM10 in Zaragoza were related to natural sources (60%, 75% and 47%, respectively, for PCA-APCS, Unmix and PMF) although anthropogenic sources also contributed to PM10 (28%, 25% and 39%). With regard to the anthropogenic sources, while PCA and PMF allowed high discrimination in the sources identification associated with different combustion sources such as traffic and industry, fossil fuel, biomass and fuel-oil combustion, heavy traffic and evaporative emissions, the Unmix model only allowed the identification of industry and traffic emissions, evaporative emissions and heavy-duty vehicles. The three models provided good correlations between the experimental and modelled PM10 concentrations with major precision and the closest agreement between the PMF and PCA models.

Methods to Protect from Skeletal Cardiovascular Insufficiency: Improving Soldier Performance in Adverse Environments

DTIC Science & Technology

2006-11-01

A specific objective here is to determine the role and subtype of adenosine receptors that mediate skeletal muscle protection using a quantitative... using a mouse hindlimb model and it defined adenosine A3 receptors as one of the skeletal muscle protective adenosine receptors. The study also...from the US Army Medical Research and Materiel Command Human Subjects Research Review Board on 9/9/2005. Twenty two subjects were consented but

Caffeine and Selective Adenosine Receptor Antagonists as New Therapeutic Tools for the Motivational Symptoms of Depression

PubMed Central

López-Cruz, Laura; Salamone, John D.; Correa, Mercè

2018-01-01

Major depressive disorder is one of the most common and debilitating psychiatric disorders. Some of the motivational symptoms of depression, such anergia (lack of self-reported energy) and fatigue are relatively resistant to traditional treatments such as serotonin uptake inhibitors. Thus, new pharmacological targets are being investigated. Epidemiological data suggest that caffeine consumption can have an impact on aspects of depressive symptomatology. Caffeine is a non-selective adenosine antagonist for A1/A2A receptors, and has been demonstrated to modulate behavior in classical animal models of depression. Moreover, selective adenosine receptor antagonists are being assessed for their antidepressant effects in animal studies. This review focuses on how caffeine and selective adenosine antagonists can improve different aspects of depression in humans, as well as in animal models. The effects on motivational symptoms of depression such as anergia, fatigue, and psychomotor slowing receive particular attention. Thus, the ability of adenosine receptor antagonists to reverse the anergia induced by dopamine antagonism or depletion is of special interest. In conclusion, although further studies are needed, it appears that caffeine and selective adenosine receptor antagonists could be therapeutic agents for the treatment of motivational dysfunction in depression. PMID:29910727

Molecular interactions of agonist and inverse agonist ligands at serotonin 5-HT2C G protein-coupled receptors: computational ligand docking and molecular dynamics studies validated by experimental mutagenesis results

NASA Astrophysics Data System (ADS)

Córdova-Sintjago, Tania C.; Liu, Yue; Booth, Raymond G.

2015-02-01

To understand molecular determinants for ligand activation of the serotonin 5-HT2C G protein-coupled receptor (GPCR), a drug target for obesity and neuropsychiatric disorders, a 5-HT2C homology model was built according to an adrenergic β2 GPCR (β2AR) structure and validated using a 5-HT2B GPCR crystal structure. The models were equilibrated in a simulated phosphatidyl choline membrane for ligand docking and molecular dynamics studies. Ligands included (2S, 4R)-(-)-trans-4-(3'-bromo- and trifluoro-phenyl)-N,N-dimethyl-1,2,3,4-tetrahydronaphthalene-2-amine (3'-Br-PAT and 3'-CF3-PAT), a 5-HT2C agonist and inverse agonist, respectively. Distinct interactions of 3'-Br-PAT and 3'-CF3-PAT at the wild-type (WT) 5-HT2C receptor model were observed and experimental 5-HT2C receptor mutagenesis studies were undertaken to validate the modelling results. For example, the inverse agonist 3'-CF3-PAT docked deeper in the WT 5-HT2C binding pocket and altered the orientation of transmembrane helices (TM) 6 in comparison to the agonist 3'-Br-PAT, suggesting that changes in TM orientation that result from ligand binding impact function. For both PATs, mutation of 5-HT2C residues S3.36, T3.37, and F5.47 to alanine resulted in significantly decreased affinity, as predicted from modelling results. It was concluded that upon PAT binding, 5-HT2C residues T3.37 and F5.47 in TMs 3 and 5, respectively, engage in inter-helical interactions with TMs 4 and 6, respectively. The movement of TMs 5 and 6 upon agonist and inverse agonist ligand binding observed in the 5-HT2C receptor modelling studies was similar to movements reported for the activation and deactivation of the β2AR, suggesting common mechanisms among aminergic neurotransmitter GPCRs.

Customizing G Protein-coupled receptor models for structure-based virtual screening.

PubMed

de Graaf, Chris; Rognan, Didier

2009-01-01

This review will focus on the construction, refinement, and validation of G Protein-coupled receptor models for the purpose of structure-based virtual screening. Practical tips and tricks derived from concrete modeling and virtual screening exercises to overcome the problems and pitfalls associated with the different steps of the receptor modeling workflow will be presented. These examples will not only include rhodopsin-like (class A), but also secretine-like (class B), and glutamate-like (class C) receptors. In addition, the review will present a careful comparative analysis of current crystal structures and their implication on homology modeling. The following themes will be discussed: i) the use of experimental anchors in guiding the modeling procedure; ii) amino acid sequence alignments; iii) ligand binding mode accommodation and binding cavity expansion; iv) proline-induced kinks in transmembrane helices; v) binding mode prediction and virtual screening by receptor-ligand interaction fingerprint scoring; vi) extracellular loop modeling; vii) virtual filtering schemes. Finally, an overview of several successful structure-based screening shows that receptor models, despite structural inaccuracies, can be efficiently used to find novel ligands.

μ Opioid receptor: novel antagonists and structural modeling

NASA Astrophysics Data System (ADS)

Kaserer, Teresa; Lantero, Aquilino; Schmidhammer, Helmut; Spetea, Mariana; Schuster, Daniela

2016-02-01

The μ opioid receptor (MOR) is a prominent member of the G protein-coupled receptor family and the molecular target of morphine and other opioid drugs. Despite the long tradition of MOR-targeting drugs, still little is known about the ligand-receptor interactions and structure-function relationships underlying the distinct biological effects upon receptor activation or inhibition. With the resolved crystal structure of the β-funaltrexamine-MOR complex, we aimed at the discovery of novel agonists and antagonists using virtual screening tools, i.e. docking, pharmacophore- and shape-based modeling. We suggest important molecular interactions, which active molecules share and distinguish agonists and antagonists. These results allowed for the generation of theoretically validated in silico workflows that were employed for prospective virtual screening. Out of 18 virtual hits evaluated in in vitro pharmacological assays, three displayed antagonist activity and the most active compound significantly inhibited morphine-induced antinociception. The new identified chemotypes hold promise for further development into neurochemical tools for studying the MOR or as potential therapeutic lead candidates.

Thymic involution in the suspended rat model for weightlessness - Decreased glucocorticoid receptor concentration

NASA Technical Reports Server (NTRS)

Steffen, J. M.; Musacchia, X. J.

1984-01-01

Hindlimb muscle atrophy, thymic involution and adrenal hypertrophy in rats during spaceflight can be simulated using suspension models. Skeletal muscle and thymus are sensitive to gluco-corticoids (GC), and previous studies have demonstrated that muscle atrophy in suspended rats is associated with increased GC receptor concentration. The objectives were to confirm thymic involution during suspension, and determine if involution correlated with increased GC receptor concentration. Seven days of antiorthostatic (AO) suspension of rats produced a significant (P less than 0.001) reduction in thymic wet weight not associated with an alteration of percent water content. GC receptor concentration (pmol/mg protein) decreased 20 percent (P less than 0.025) in thymus glands from 7 day AO suspended rats. Suspension, therefore, is associated with involution of the thymus, but this is not dependent upon AO positioning. Thymus GC receptor concentrations were depressed in 7-day suspended rats, in contrast with previous observations on skeletal muscle, suggesting that different mechanisms may underlie these responses.

Exploration of N-arylpiperazine Binding Sites of D2 Dopaminergic Receptor.

PubMed

Soskic, Vukic; Sukalovic, Vladimir; Kostic-Rajacic, Sladjana

2015-01-01

The crystal structures of the D3 dopamine receptor and several other G-protein coupled receptors (GPCRs) were published in recent times. Those 3D structures are used by us and other scientists as a template for the homology modeling and ligand docking analysis of related GPCRs. Our main scientific interest lies in the field of pharmacologically active N-arylpiperazines that exhibit antipsychotic and/or antidepressant properties, and as such are dopaminergic and serotonergic receptor ligands. In this short review article we are presenting synthesis and biological data on the new N-arylpipereazine as well our results on molecular modeling of the interactions of those N-arylpiperazines with the model of D2 dopamine receptors. To obtain that model the crystal structure of the D3 dopamine receptor was used. Our results show that the N-arylpiperazines binding site consists of two pockets: one is the orthosteric binding site where the N-arylpiperazine part of the ligand is docked and the second is a non-canonical accessory binding site for N-arylpipereazine that is formed by a second extracellular loop (ecl2) of the receptor. Until now, the structure of this receptor region was unresolved in crystal structure analyses of the D3 dopamine receptor. To get a more complete picture of the ligand - receptor interaction, DFT quantum mechanical calculations on N-arylpiperazine were performed and the obtained models were used to examine those interactions.

Expression analysis in a rat psychosis model identifies novel candidate genes validated in a large case–control sample of schizophrenia

PubMed Central

Ingason, A; Giegling, I; Hartmann, A M; Genius, J; Konte, B; Friedl, M; Ripke, S; Sullivan, P F; St. Clair, D; Collier, D A; O'Donovan, M C; Mirnics, K; Rujescu, D

2015-01-01

Antagonists of the N-methyl-D-aspartate (NMDA)-type glutamate receptor induce psychosis in healthy individuals and exacerbate schizophrenia symptoms in patients. In this study we have produced an animal model of NMDA receptor hypofunction by chronically treating rats with low doses of the NMDA receptor antagonist MK-801. Subsequently, we performed an expression study and identified 20 genes showing altered expression in the brain of these rats compared with untreated animals. We then explored whether the human orthologs of these genes are associated with schizophrenia in the largest schizophrenia genome-wide association study published to date, and found evidence for association for 4 out of the 20 genes: SF3B1, FOXP1, DLG2 and VGLL4. Interestingly, three of these genes, FOXP1, SF3B1 and DLG2, have previously been implicated in neurodevelopmental disorders. PMID:26460480

Expression analysis in a rat psychosis model identifies novel candidate genes validated in a large case-control sample of schizophrenia.

PubMed

Ingason, A; Giegling, I; Hartmann, A M; Genius, J; Konte, B; Friedl, M; Ripke, S; Sullivan, P F; St Clair, D; Collier, D A; O'Donovan, M C; Mirnics, K; Rujescu, D

2015-10-13

Antagonists of the N-methyl-D-aspartate (NMDA)-type glutamate receptor induce psychosis in healthy individuals and exacerbate schizophrenia symptoms in patients. In this study we have produced an animal model of NMDA receptor hypofunction by chronically treating rats with low doses of the NMDA receptor antagonist MK-801. Subsequently, we performed an expression study and identified 20 genes showing altered expression in the brain of these rats compared with untreated animals. We then explored whether the human orthologs of these genes are associated with schizophrenia in the largest schizophrenia genome-wide association study published to date, and found evidence for association for 4 out of the 20 genes: SF3B1, FOXP1, DLG2 and VGLL4. Interestingly, three of these genes, FOXP1, SF3B1 and DLG2, have previously been implicated in neurodevelopmental disorders.

Dimensionality of Motion and Binding Valency Govern Receptor-Ligand Kinetics As Revealed by Agent-Based Modeling.

PubMed

Lehnert, Teresa; Figge, Marc Thilo

2017-01-01

Mathematical modeling and computer simulations have become an integral part of modern biological research. The strength of theoretical approaches is in the simplification of complex biological systems. We here consider the general problem of receptor-ligand binding in the context of antibody-antigen binding. On the one hand, we establish a quantitative mapping between macroscopic binding rates of a deterministic differential equation model and their microscopic equivalents as obtained from simulating the spatiotemporal binding kinetics by stochastic agent-based models. On the other hand, we investigate the impact of various properties of B cell-derived receptors-such as their dimensionality of motion, morphology, and binding valency-on the receptor-ligand binding kinetics. To this end, we implemented an algorithm that simulates antigen binding by B cell-derived receptors with a Y-shaped morphology that can move in different dimensionalities, i.e., either as membrane-anchored receptors or as soluble receptors. The mapping of the macroscopic and microscopic binding rates allowed us to quantitatively compare different agent-based model variants for the different types of B cell-derived receptors. Our results indicate that the dimensionality of motion governs the binding kinetics and that this predominant impact is quantitatively compensated by the bivalency of these receptors.

Modeling G Protein-Coupled Receptors: a Concrete Possibility

PubMed Central

Costanzi, Stefano

2010-01-01

G protein-coupled receptors (GPCRs) are a large superfamily of membrane bound signaling proteins that are involved in the regulation of a wide range of physiological functions and constitute the most common target for therapeutic intervention. Due to the paucity of crystal structures, homology modeling has become a widespread technique for the construction of GPCR models, which have been applied to the study of their structure-function relationships and to the identification of lead ligands through virtual screening. Rhodopsin has been for years the only available template. However, recent breakthroughs in GPCR crystallography have led to the solution of the structures of a few additional receptors. In light of these newly elucidated crystal structures, we have been able to produce a substantial amount of data to demonstrate that accurate models of GPCRs in complex with their ligands can be constructed through homology modeling followed by fully flexible molecular docking. These results have been confirmed by our success in the first blind assessment of GPCR modeling and docking, organized in coordination with the solution of the X-ray structure of the adenosine A2A receptor. Taken together, these data indicate that: a) the transmembrane helical bundle can be modeled with considerable accuracy; b) predicting the binding mode of a ligand, although doable, is challenging; c) modeling of the extracellular and intracellular loops is still problematic. PMID:21253444

Purinergic receptor X7 is a key modulator of metabolic oxidative stress-mediated autophagy and inflammation in experimental nonalcoholic steatohepatitis

PubMed Central

Das, Suvarthi; Seth, Ratanesh Kumar; Kumar, Ashutosh; Kadiiska, Maria B.; Michelotti, Gregory; Diehl, Anna Mae

2013-01-01

Recent studies indicate that metabolic oxidative stress, autophagy, and inflammation are hallmarks of nonalcoholic steatohepatitis (NASH) progression. However, the molecular mechanisms that link these important events in NASH remain unclear. In this study, we investigated the mechanistic role of purinergic receptor X7 (P2X7) in modulating autophagy and resultant inflammation in NASH in response to metabolic oxidative stress. The study uses two rodent models of NASH. In one of them, a CYP2E1 substrate bromodichloromethane is used to induce metabolic oxidative stress and NASH. Methyl choline-deficient diet feeding is used for the other NASH model. CYP2E1 and P2X7 receptor gene-deleted mice are used to establish their roles in regulating metabolic oxidative stress and autophagy. Autophagy gene expression, protein levels, confocal microscopy based-immunolocalization of lysosome-associated membrane protein (LAMP)2A and histopathological analysis were performed. CYP2E1-dependent metabolic oxidative stress induced increases in P2X7 receptor expression and chaperone-mediated autophagy markers LAMP2A and heat shock cognate 70 but caused depletion of light chain 3 isoform B (LC3B) protein levels. P2X7 receptor gene deletion significantly decreased LAMP2A and inflammatory indicators while significantly increasing LC3B protein levels compared with wild-type mice treated with bromodichloromethane. P2X7 receptor-deleted mice were also protected from NASH pathology as evidenced by decreased inflammation and fibrosis. Our studies establish that P2X7 receptor is a key regulator of autophagy induced by metabolic oxidative stress in NASH, thereby modulating hepatic inflammation. Furthermore, our findings presented here form a basis for P2X7 receptor as a potential therapeutic target in the treatment for NASH. PMID:24157968

Integrin receptor involvement in actin cable formation in an in vitro model of events associated with wound contraction.

PubMed

Stephens, P; Genever, P G; Wood, E J; Raxworthy, M J

1997-01-01

Actin cables have been reported to act in vivo as contractile 'purse strings' capable of closing embryonic wounds through generation of circumferential tension. Furthermore, their involvement in wounds within in vitro model systems suggests that actin cable contraction may be an important mechanism involved in the process of wound closure. The aim of this study therefore, was to investigate the appearance of actin cables in a contracting fibroblast populated collagen lattice, an in vitro model of events associated with wound contraction. Utilising this in vitro model, the time-course of actin cable production was investigated and the involvement of integrin receptors analysed using immunofluorescent labelling techniques. Over a period of hours distinct cellular cable-like structures developed at the edges of collagen lattices coinciding with the onset of contraction. Cellular organisation within the cable was evident as was polymerisation of actin microfilaments into elongated stress fibres forming a continuous cell-cell 'actin cable' around the circumference of the lattice. Immunolocalisation demonstrated that integrin receptor subunits beta 1 and alpha 2 but not alpha 5 were involved in apparent intimate cell-cell contact between juxtaposed fibroblasts within this actin cable. This study demonstrates the involvement of integrin receptors in actin cable formation within collagen lattice systems undergoing reorganisation. Such integrin involvement may enable participating cells to respond to the tensional status of their surrounding environment and via cell-cell communication, to permit a co-ordinated contraction of the cable. It is concluded that integrin receptor involvement in active actin cable contraction may be involved in the process of wound contraction.

Comparison of source apportionment of PM2.5 using receptor models in the main hub port city of East Asia: Busan

NASA Astrophysics Data System (ADS)

Jeong, Ju-Hee; Shon, Zang-Ho; Kang, Minsung; Song, Sang-Keun; Kim, Yoo-Keun; Park, Jinsoo; Kim, Hyunjae

2017-01-01

The contributions of various PM2.5 emission sources to ambient PM2.5 levels during 2013 in the main hub port city (Busan, South Korea) of East Asia was quantified using several receptor modeling techniques. Three receptor models of principal component analysis/absolute principal component score (PCA/APCS), positive matrix factorization (PMF), and chemical mass balance (CMB) were used to apportion the source of PM2.5 obtained from the target city. The results of the receptor models indicated that the secondary formation of PM2.5 was the dominant (45-60%) contributor to PM2.5 levels in the port city of Busan. The PMF and PCA/APCS suggested that ship emission was a non-negligible contributor of PM2.5 (up to about 10%) in the study area, whereas it was a negligible contributor based on CMB. The magnitude of source contribution estimates to PM2.5 levels differed significantly among these three models due to their limitations (e.g., PM2.5 emission source profiles and restrictions of the models). Potential source contribution function and concentration-weighted trajectory analyses indicated that long-range transport from sources in the eastern China and Yellow Sea contributed significantly to the level of PM2.5 in Busan.

A1 adenosine receptor attenuates intracerebral hemorrhage-induced secondary brain injury in rats by activating the P38-MAPKAP2-Hsp27 pathway.

PubMed

Zhai, Weiwei; Chen, Dongdong; Shen, Haitao; Chen, Zhouqing; Li, Haiying; Yu, Zhengquan; Chen, Gang

2016-06-14

This study was designed to determine the role of the A1 adenosine receptors in intracerebral hemorrhage (ICH)-induced secondary brain injury and the underlying mechanisms. A collagenase-induced ICH model was established in Sprague-Dawley rats, and cultured primary rat cortical neurons were exposed to oxyhemoglobin at a concentration of 10 μM to mimic ICH in vitro. The A1 adenosine receptor agonist N(6)-cyclohexyladenosine and antagonist 8-phenyl-1,3-dipropylxanthine were used to study the role of A1 adenosine receptor in ICH-induced secondary brain injury, and antagonists of P38 and Hsp27 were used to study the underlying mechanisms of A1 adenosine receptor actions. The protein level of A1 adenosine receptor was significantly increased by ICH, while there was no significant change in protein levels of the other 3 adenosine receptors. In addition, the A1 adenosine receptor expression could be increased by N(6)-cyclohexyladenosine and decreased by 8-phenyl-1,3-dipropylxanthine under ICH conditions. Activation of the A1 adenosine receptor attenuated neuronal apoptosis in the subcortex, which was associated with increased phosphorylation of P38, MAPK, MAPKAP2, and Hsp27. Inhibition of the A1 adenosine receptor resulted in opposite effects. Finally, the neuroprotective effect of the A1 adenosine receptor agonist N(6)-cyclohexyladenosine was inhibited by antagonists of P38 and Hsp27. This study demonstrates that activation of the A1 adenosine receptor by N(6)-cyclohexyladenosine could prevent ICH-induced secondary brain injury via the P38-MAPKAP2-Hsp27 pathway.

Quantitative measurement of cell membrane receptor internalization by the nanoluciferase reporter: Using the G protein-coupled receptor RXFP3 as a model.

PubMed

Liu, Yu; Song, Ge; Shao, Xiao-Xia; Liu, Ya-Li; Guo, Zhan-Yun

2015-02-01

Nanoluciferase (NanoLuc) is a newly developed small luciferase reporter with the brightest bioluminescence to date. In the present work, we developed NanoLuc as a sensitive bioluminescent reporter to measure quantitatively the internalization of cell membrane receptors, based on the pH dependence of the reporter activity. The G protein-coupled receptor RXFP3, the cognate receptor of relaxin-3/INSL7, was used as a model receptor. We first generated stable HEK293T cells that inducibly coexpressed a C-terminally NanoLuc-tagged human RXFP3 and a C-terminally enhanced green fluorescent protein (EGFP)-tagged human RXFP3. The C-terminal EGFP-tag and NanoLuc-tag had no detrimental effects on the ligand-binding potency and intracellular trafficking of RXFP3. Based on the fluorescence of the tagged EGFP reporter, the ligand-induced RXFP3 internalization was visualized directly under a fluorescence microscope. Based on the bioluminescence of the tagged NanoLuc reporter, the ligand-induced RXFP3 internalization was measured quantitatively by a convenient bioluminescent assay. Coexpression of an EGFP-tagged inactive [E141R]RXFP3 had no detrimental effect on the ligand-binding potency and ligand-induced internalization of the NanoLuc-tagged wild-type RXFP3, suggesting that the mutant RXFP3 and wild-type RXFP3 worked independently. The present bioluminescent internalization assay could be extended to other G protein-coupled receptors and other cell membrane receptors to study ligand-receptor and receptor-receptor interactions. Copyright © 2014 Elsevier B.V. All rights reserved.

Restoration of Tumor Immune Surveillance via Targeting of IL-13Receptor-α2

PubMed Central

Fichtner-Feigl, Stefan; Terabe, Masaki; Kitani, Atsushi; Young, Cheryl A.; Fuss, Ivan; Geissler, Edward K.; Schlitt, Hans-Jürgen; Berzofsky, Jay A.; Strober, Warren

2009-01-01

In previous studies we described a “counter-immunosurveillance” mechanism initiated by tumor-activated, IL-13-producing NKT cells that signal Gr-1+ cells to produce TGF-β1, a cytokine that suppresses the activity of tumor-inhibiting cytolytic CD8+ T cells. Here we show that in two tumor models (the CT-26 metastatic colon cancer and the 15-12RM fibrosarcoma regressor models) this counter-surveillance mechanism requires the expression of a novel IL-13 receptor, IL-13Rα2, on Gr-1intermediate cells, since down-regulation of IL-13Rα2 expression or the AP-1 signal generated by the receptor via in vivo administration of specific siRNA or decoy oligonucleotides leads to loss of TGF-β1 production. Furthermore, acting on prior studies showing that IL-13Rα2 expression is induced (in part) by TNF-α, we show that receptor expression and TGF-β1 production is inhibited by administration of a TNF-α neutralizing substance, TNF-αR-Fc (etanercept). Taking advantage of this latter fact, we then demonstrate in the CT-26 model that counter-immunosurveillance could be inhibited, anti-CT-26-specific CD8+ cytolytic activity restored, and CT-26 metastatic tumor nodules greatly decreased by administration of TNF-αR-Fc. Corroborative data was obtained using the 15-12RM fibrosarcoma model. These studies point to the prevention of metastatic cancer with an available agent with already known clinically acceptable adverse effects and toxicity. PMID:18451175

43.1 GENETIC INSIGHTS LEAD TO DISCOVERY OF SELECTIVE ACTIVATORS OF MGLU1 AND MGLU3 METABOTROPIC GLUTAMATE RECEPTORS AS POTENTIAL TREATMENTS FOR SCHIZOPHRENIA

PubMed Central

Conn, P Jeffrey; Yohn, Samantha; Stansley, Branden; Foster, Dan; Plumley, Hyekyung; Lindsley, Craig

2018-01-01

Abstract Background A large number of clinical and preclinical studies suggest that dysfunction at synapses for the excitatory neurotransmitter glutamate may play a critical role in the pathophysiological changes that underlie each of the major symptom clusters observed in schizophrenia patients. Interestingly, recent genetic studies identified multiple nonsynonymous single nucleotide polymorphisms (SNPs) in the human genes encoding two specific subtypes of metabotropic glutamate (mGlu) receptors that are associated with schizophrenia. These include GRM1 and GRM3, the genes encoding for the mGlu1 and mGlu3 receptor subtypes respectively. Furthermore, postmortem studies suggest that expression of these mGlu receptor subtypes is altered brains of schizophrenia patients compared to controls. Mutations in GRM1 were identified a range of schizophrenia patients, whereas SNPs in the human gene encoding mGlu3 (GRM3) are selectively associated with poor performance on cognitive tests that are dependent on function of the prefrontal cortex (PFC) and hippocampus. These studies raise the possibility that disrupted signaling of mGlu1 and/or mGlu3 could contribute to the symptoms of schizophrenia and that selective modulators of these receptors could provide a novel approach to treatment of this disorder. Methods Wild-type and mutant forms of mGlu receptors were expressed in cell lines and used for discovery and optimization of highly selective positive allosteric modulators (PAMs) of mGlu1 and mGlu3. Optimized mGlu1 and mGlu3 PAMs were then used along with mouse genetic studies to evaluate the roles of these receptors in specific basal ganglia and forebrain circuits that have been implicated in schizophrenia. Finally, these compounds were used in animal models to assess potential efficacy in rodent models that are relevant for reducing positive, negative, and cognitive symptoms that are observed in schizophrenia patients. Results GRM1 mutations associated with schizophrenia were found to reduce mGlu1 signaling, suggesting that loss of function of this receptor could contribute to symptoms associated with schizophrenia. Furthermore, we found that highly selective mGlu1 PAMs reverse deficits in mGlu1 signaling observed in these mutant receptors, induced a profound reduction in dopamine release in striatal areas implicated in schizophrenia, and have robust antipsychotic-like effects that are mediated by localized inhibition of dopamine release in striatum. In contrast to existing antipsychotic medications, selective mGlu1 PAMs also improve motivation and reduce anhedonia in animal models. Interestingly, selective mGlu3 PAMs have multiple effects in the prefrontal cortex and hippocampus that would be expected to improve cognitive function. Consistent with this, highly selective mGlu3 PAMs have robust cognition-enhancing effects in rodent models that are relevant for the cognitive deficits observed in schizophrenia patients. Discussion These studies provide exciting new evidence that highly selective activators of two glutamate receptors identified in human genetic studies have potential utility in treatment of positive (mGLu1), negative (mGlu1), and cognitive (mGlu3) symptoms of schizophrenia patients. Furthermore, the novel mGlu1 and mGlu3 PAMs discovered in these studies provide excellent drug leads for further optimization and ultimate clinical testing.

New techniques for positron emission tomography in the study of human neurological disorders

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kuhl, D.E.

1992-07-01

The general goals of the physics and kinetic modeling projects are to: (1) improve the quantitative information extractable from PET images, and (2) develop, implement and optimize tracer kinetic models for new PET neurotransmitter/receptor ligands aided by computer simulations. Work towards improving PET quantification has included projects evaluating: (1) iterative reconstruction algorithms using supplemental boundary information, (2) automated registration of dynamic PET emission and transmission data using sinogram edge detection, and (3) automated registration of multiple subjects to a common coordinate system, including the use of non-linear warping methods. Simulation routines have been developed providing more accurate representation of datamore » generated from neurotransmitter/receptor studies. Routines consider data generated from complex compartmental models, high or low specific activity administrations, non-specific binding, pre- or post-injection of cold or competing ligands, temporal resolution of the data, and radiolabeled metabolites. Computer simulations and human PET studies have been performed to optimize kinetic models for four new neurotransmitter/receptor ligands, [{sup 11}C]TRB (muscarinic), [{sup 11}C]flumazenil (benzodiazepine), [{sup 18}F]GBR12909, (dopamine), and [{sup 11}C]NMPB (muscarinic).« less

New techniques for positron emission tomography in the study of human neurological disorders

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kuhl, D.E.

1992-01-01

The general goals of the physics and kinetic modeling projects are to: (1) improve the quantitative information extractable from PET images, and (2) develop, implement and optimize tracer kinetic models for new PET neurotransmitter/receptor ligands aided by computer simulations. Work towards improving PET quantification has included projects evaluating: (1) iterative reconstruction algorithms using supplemental boundary information, (2) automated registration of dynamic PET emission and transmission data using sinogram edge detection, and (3) automated registration of multiple subjects to a common coordinate system, including the use of non-linear warping methods. Simulation routines have been developed providing more accurate representation of datamore » generated from neurotransmitter/receptor studies. Routines consider data generated from complex compartmental models, high or low specific activity administrations, non-specific binding, pre- or post-injection of cold or competing ligands, temporal resolution of the data, and radiolabeled metabolites. Computer simulations and human PET studies have been performed to optimize kinetic models for four new neurotransmitter/receptor ligands, ({sup 11}C)TRB (muscarinic), ({sup 11}C)flumazenil (benzodiazepine), ({sup 18}F)GBR12909, (dopamine), and ({sup 11}C)NMPB (muscarinic).« less

The Mouse Solitary Odorant Receptor Gene Promoters as Models for the Study of Odorant Receptor Gene Choice

PubMed Central

Degl'Innocenti, Andrea

2016-01-01

Background In vertebrates, several anatomical regions located within the nasal cavity mediate olfaction. Among these, the main olfactory epithelium detects most conventional odorants. Olfactory sensory neurons, provided with cilia exposed to the air, detect volatile chemicals via an extremely large family of seven-transmembrane chemoreceptors named odorant receptors. Their genes are expressed in a monogenic and monoallelic fashion: a single allele of a single odorant receptor gene is transcribed in a given mature neuron, through a still uncharacterized molecular mechanism known as odorant receptor gene choice. Aim Odorant receptor genes are typically arranged in genomic clusters, but a few are isolated (we call them solitary) from the others within a region broader than 1 Mb upstream and downstream with respect to their transcript's coordinates. The study of clustered genes is problematic, because of redundancy and ambiguities in their regulatory elements: we propose to use the solitary genes as simplified models to understand odorant receptor gene choice. Procedures Here we define number and identity of the solitary genes in the mouse genome (C57BL/6J), and assess the conservation of the solitary status in some mammalian orthologs. Furthermore, we locate their putative promoters, predict their homeodomain binding sites (commonly present in the promoters of odorant receptor genes) and compare candidate promoter sequences with those of wild-caught mice. We also provide expression data from histological sections. Results In the mouse genome there are eight intact solitary genes: Olfr19 (M12), Olfr49, Olfr266, Olfr267, Olfr370, Olfr371, Olfr466, Olfr1402; five are conserved as solitary in rat. These genes are all expressed in the main olfactory epithelium of three-day-old mice. The C57BL/6J candidate promoter of Olfr370 has considerably varied compared to its wild-type counterpart. Within the putative promoter for Olfr266 a homeodomain binding site is predicted. As a whole, our findings favor Olfr266 as a model gene to investigate odorant receptor gene choice. PMID:26794459

Association between p75 neurotrophin receptor gene expression and cell apoptosis in tissues surrounding hematomas in rat models of intracerebral hemorrhage.

PubMed

He, Baixiang; Bao, Gang; Guo, Shiwen; Xu, Gaofeng; Li, Qi; Wang, Ning

2012-03-15

Animal models of intracerebral hemorrhage were established by injection of autologous blood into the caudate nucleus in rats. Cell apoptosis was measured by flow cytometry and immunohistochemical staining of the p75 neurotrophin receptor. p75 neurotrophin receptor protein was detected by immunohistochemistry. p75 neurotrophin receptor mRNA was examined by quantitative real-time polymerase chain reactions. At 24 hours after modeling, cellular apoptosis occured around hematoma with upregulation of p75 neurotrophin receptor protein and mRNA was observed, which directly correlated to apoptosis. This observation indicated that p75 neurotrophin receptor upregulation was associated with cell apoptosis around hematomas after intracerebral hemorrhage.

Principles of antibody-mediated TNF receptor activation

PubMed Central

Wajant, H

2015-01-01

From the beginning of research on receptors of the tumor necrosis factor (TNF) receptor superfamily (TNFRSF), agonistic antibodies have been used to stimulate TNFRSF receptors in vitro and in vivo. Indeed, CD95, one of the first cloned TNFRSF receptors, was solely identified as the target of cell death-inducing antibodies. Early on, it became evident from in vitro studies that valency and Fcγ receptor (FcγR) binding of antibodies targeting TNFRSF receptors can be of crucial relevance for agonistic activity. TNFRSF receptor-specific antibodies of the IgM subclass and secondary cross-linked or aggregation prone dimeric antibodies typically display superior agonistic activity compared with dimeric antibodies. Likewise, anchoring of antibodies to cell surface-expressed FcγRs potentiate their ability to trigger TNFRSF receptor signaling. However, only recently has the relevance of oligomerization and FcγR binding for the in vivo activity of antibody-induced TNFRSF receptor activation been straightforwardly demonstrated in vivo. This review discusses the crucial role of oligomerization and/or FcγR binding for antibody-mediated TNFRSF receptor stimulation in light of current models of TNFRSF receptor activation and especially the overwhelming relevance of these issues for the rational development of therapeutic TNFRSF receptor-targeting antibodies. PMID:26292758

Modulation of opioid analgesia by agmatine.

PubMed

Kolesnikov, Y; Jain, S; Pasternak, G W

1996-01-18

Administered alone, agmatine at doses of 0.1 or 10 mg/kg is without effect in the mouse tailflick assay. However, agmatine enhances morphine analgesia in a dose-dependent manner, shifting morphine's ED50 over 5-fold. A far greater effect is observed when morphine is given intrathecally (9-fold shift) than after intracerebroventricular administration (2-fold). In contrast to the potentiation of morphine analgesia, agmatine (10 mg/kg) has no effect on morphine's inhibition of gastrointestinal transit. delta-Opioid receptor-mediated analgesia also is potentiated by agmatine, but kappa1-receptor-mediated (U50,488H; trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl] benzeneacetemide) and kappa3-opioid receptor-mediated (naloxone benzoylhydrazone) analgesia is not significantly enhanced by any dose of agmatine tested in this acute model. In chronic studies, agmatine at a low dose (0.1 mg/kg) which does not affect morphine analgesia acutely prevents tolerance following chronic morphine dosing for 10 days. A higher agmatine dose (10 mg/kg) has a similar effect. Agmatine also blocks tolerance to the delta-opioid receptor ligand [D-Pen2,D-Pen5]enkephalin given intrathecally, but not to the kappa3-opioid receptor agonist naloxone benzoylhydrazone. Despite its inactivity on kappa1-opioid analgesia in the acute model, agmatine prevents kappa1-opioid receptor-mediated tolerance. These studies demonstrate the dramatic interactions between agmatine and opioid analgesia and tolerance.

A novel antagonist of the prostaglandin E(2) EP(4) receptor inhibits Th1 differentiation and Th17 expansion and is orally active in arthritis models.

PubMed

Chen, Q; Muramoto, K; Masaaki, N; Ding, Y; Yang, H; Mackey, M; Li, W; Inoue, Y; Ackermann, K; Shirota, H; Matsumoto, I; Spyvee, M; Schiller, S; Sumida, T; Gusovsky, F; Lamphier, M

2010-05-01

Rheumatoid arthritis (RA) is an autoimmune disorder involving subsets of activated T cells, in particular T helper (Th) 1 and Th17 cells, which infiltrate and damage tissues and induce inflammation. Prostaglandin E(2) (PGE(2)) enhances the Th17 response, exacerbates collagen-induced arthritis (CIA) and promotes inflammatory pain. The current study investigated whether selective antagonism of the PGE(2) EP(4) receptor would suppress Th1/Th17 cell development and inflammatory arthritis in animal models of RA. Effects of PGE(2) and a novel EP(4) receptor antagonist ER-819762 on Th1 differentiation, interleukin-23 (IL-23) production by dendritic cells (DCs), and Th17 development were assessed in vitro. The effect of ER-819762 was evaluated in CIA and glucose-6-phosphate isomerase (GPI)-induced arthritis models. In addition, the effects of ER-819762 on pain were evaluated in a model of chronic inflammatory pain induced by complete Freund's adjuvant (CFA) in the rat. Stimulation of the EP(4) receptor enhanced Th1 differentiation via phosphatidylinositol 3 kinase signalling, selectively promoted Th17 cell expansion, and induced IL-23 secretion by activated DCs, effects suppressed by ER-819762 or anti-PGE(2) antibody. Oral administration of ER-19762 suppressed Th1 and Th17 cytokine production, suppressed disease in collagen- and GPI-induced arthritis in mice, and suppressed CFA-induced inflammatory pain in rats. PGE(2) stimulates EP(4) receptors to promote Th1 differentiation and Th17 expansion and is critically involved in development of arthritis in two animal models. Selective suppression of EP(4) receptor signalling may have therapeutic value in RA both by modifying inflammatory arthritis and by relieving pain.

Melanocortin 1 receptor and skin pathophysiology: beyond colour, much more than meets the eye.

PubMed

García-Borrón, José Carlos; Olivares, Concepción

2014-06-01

The melanocortin 1 receptor (MC1R), a G protein-coupled receptor preferentially expressed in melanocytes, mediates the pigmentary effects of α melanocyte-stimulating hormone (αMSH). MC1R is also expressed in other cutaneous cell types, particularly keratinocytes and dermal fibroblasts, suggesting non-pigmentary actions of the αMSH/MC1R system. Böhm and Stegemann now report a dramatic effect of mouse Mc1r functional status on susceptibility to skin fibrosis and collagen types I and III metabolism, in a study combining the powerful mouse model provided by the natural Mc1r(e/e) knockout and an established model of skin fibrosis. The study underscores the antifibrotic role for the skin αMSH/MC1R system. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Structural complementarity of Toll/interleukin-1 receptor domains in Toll-like receptors and the adaptors Mal and MyD88.

PubMed

Dunne, Aisling; Ejdeback, Mikael; Ludidi, Phumzile L; O'Neill, Luke A J; Gay, Nicholas J

2003-10-17

The Toll/interleukin 1 receptor (TIR) domain is a region found in the cytoplasmic tails of members of the Toll-like receptor/interleukin-1 receptor superfamily. The domain is essential for signaling and is also found in the adaptor proteins Mal (MyD88 adaptor-like) and MyD88, which function to couple activation of the receptor to downstream signaling components. Experimental structures of two Toll/interleukin 1 receptor domains reveal a alpha-beta-fold similar to that of the bacterial chemotaxis protein CheY, and other evidence suggests that the adaptors can make heterotypic interactions with both the receptors and themselves. Here we show that the purified TIR domains of Mal and MyD88 can form stable heterodimers and also that Mal homodimers and oligomers are dissociated in the presence of ATP. To identify structural features that may contribute to the formation of signaling complexes, we produced models of the TIR domains from human Toll-like receptor 4 (TLR4), Mal, and MyD88. We found that although the overall fold is conserved the electrostatic surface potentials are quite distinct. Docking studies of the models suggest that Mal and MyD88 bind to different regions in TLRs 2 and 4, a finding consistent with a cooperative role of the two adaptors in signaling. Mal and MyD88 are predicted to interact at a third non-overlapping site, suggesting that the receptor and adaptors may form heterotetrameric complexes. The theoretical model of the interactions is supported by experimental data from glutathione S-transferase pull-downs and co-immunoprecipitations. Neither theoretical nor experimental data suggest a direct role for the conserved proline in the BB-loop in the association of TLR4, Mal, and MyD88. Finally we show a sequence relationship between the Drosophila protein Tube and Mal that may indicate a functional equivalence of these two adaptors in the Drosophila and vertebrate Toll pathways.

5HT-1A receptors and anxiety-like behaviours: studies in rats with constitutionally upregulated/downregulated serotonin transporter.

PubMed

Bordukalo-Niksic, Tatjana; Mokrovic, Gordana; Stefulj, Jasminka; Zivin, Marko; Jernej, Branimir; Cicin-Sain, Lipa

2010-12-01

Altered activity of brain serotonergic (5HT) system has been implicated in a wide range of behaviours and behavioural disorders, including anxiety. Functioning of 5HT-1A receptor has been suggested as a modulator of emotional balance in both, normal and pathological forms of anxiety. Here, we studied serotonergic modulation of anxiety-like behaviour using a genetic rat model with constitutional differences in 5HT homeostasis, named Wistar-Zagreb 5HT (WZ-5HT) rats. The model, consisting of high-5HT and low-5HT sublines, was developed by selective breeding of animals for extreme activities of peripheral (platelet) 5HT transporter, but selection process had affected also central 5HT homeostasis, as evidenced from neurochemical and behavioural studies. Anxiety-like behaviour in WZ-5HT rats was evaluated by two commonly used paradigms: open field and elevated-plus maze. The involvement of 5HT-1A receptors in behavioural response was assessed by measuring mRNA expression in cell bodies (raphe nuclei) and projection regions (frontal cortex, hippocampus) by use of RT-PCR and in situ hybridization, and by measuring functionality of cortical 5HT-1A receptors by use of [(3)H]8-OH-DPAT radioligand binding. Animals from the high-5HT subline exhibit increased anxiety-like behaviour and decreased exploratory activity when exposed to novel environment. No measurable differences in constitutional (baseline) functionality or expression of 5HT-1A receptors between sublines were found. The results support contribution of increased serotonergic functioning to the anxiety-like behaviour. They also validate the high-5HT subline of WZ-5HT rats as a potential model to study mechanisms of anxiety, especially of its nonpathological form, while the low-5HT subline may be useful to model sensation seeking phenotype. Copyright (c) 2010 Elsevier B.V. All rights reserved.

A new biochromatography model based on DNA origami assembled PPARγ: construction and evaluation.

PubMed

Zhou, Jie; Meng, Lingchang; Sun, Chong; Chen, Shanshan; Sun, Fang; Luo, Pei; Zhao, Yongxing

2017-05-01

As drug targets, receptors have potential to screen drugs. Silica is an attractive support to immobilize receptors; however, the lack of biocompatibility makes it easier for receptors to lose bioactivity, which remains an obstacle to its widespread use. With the advantage of biocompatibility, DNA origami can be used as a biological carrier to improve the biocompatibility of silica and assemble receptors. In this study, a new biochromatography model based on DNA origami was constructed. A large quantity of M13ssDNA was used as a scaffold, leading to significant costs, so M13ssDNA was self-produced from the bacteriophage particles. This approach is demonstrated using the ligand binding domain of gamma isoform peroxisome proliferator-activated receptor (PPARγ-LBD) as a research object. PPARγ-LBD was assembled on DNA origami carrier and then coupled on the surface of silica. The products were packed into the column as stationary phase to construct the biochromatography with the ability to recognize drugs. Affinity and specificity of the biochromatography model were evaluated by HPLC. The final results showed that the biochromatography could recognize rosiglitazone specifically, which further proved that the model could screen chemical compositions interacted with PPARγ. It was the first time to take advantage of DNA origami to assemble PPARγ to construct biochromatography. The new biochromatography model has the advantages of being efficient, convenient, and high-throughput. This method affords a new way to rapidly and conveniently screen active ingredients from complex sample plant extracts and natural product-like libraries.

Continuous delivery of naltrexone and nalmefene leads to tolerance in reducing alcohol drinking and to supersensitivity of brain opioid receptors.

PubMed

Korpi, Esa R; Linden, Anni-Maija; Hytönen, Heidi R; Paasikoski, Nelli; Vashchinkina, Elena; Dudek, Mateusz; Herr, Deron R; Hyytiä, Petri

2017-07-01

Opioid antagonist treatments reduce alcohol drinking in rodent models and in alcohol-dependent patients, with variable efficacy across different studies. These treatments may suffer from the development of tolerance and opioid receptor supersensitivity, as suggested by preclinical models showing activation of these processes during and after subchronic high-dose administration of the short-acting opioid antagonist naloxone. In the present study, we compared equipotent low and moderate daily doses of naltrexone and nalmefene, two opioid antagonists in the clinical practice for treatment of alcoholism. The antagonists were given here subcutaneously for 7 days either as daily injections or continuous osmotic minipump-driven infusions to alcohol-preferring AA rats having trained to drink 10% alcohol in a limited access protocol. One day after stopping the antagonist treatment, [ 35 S]GTPγS autoradiography on brain cryostat sections was carried out to examine the coupling of receptors to G protein activation. The results prove the efficacy of repeated injections over infused opioid antagonists in reducing alcohol drinking. Tolerance to the reducing effect on alcohol drinking and to the enhancement of G protein coupling to μ-opioid receptors in various brain regions were consistently detected only after infused antagonists. Supersensitivity of κ-opioid receptors was seen in the ventral and dorsal striatal regions especially by infused nalmefene. Nalmefene showed no clear agonistic activity in rat brain sections or at human recombinant κ-opioid receptors. The findings support the as-needed dosing practice, rather than the standard continual dosing, in the treatment of alcoholism with opioid receptor antagonists. © 2016 Society for the Study of Addiction.

Atmospheric aerosol source identification and estimates of source contributions to air pollution in Dundee, UK

NASA Astrophysics Data System (ADS)

Qin, Y.; Oduyemi, K.

Anthropogenic aerosol (PM 10) emission sources sampled at an air quality monitoring station in Dundee have been analysed. However, the information on local natural aerosol emission sources was unavailable. A method that combines receptor model and atmospheric dispersion model was used to identify aerosol sources and estimate source contributions to air pollution. The receptor model identified five sources. These are aged marine aerosol source with some chlorine replaced by sulphate, secondary aerosol source of ammonium sulphate, secondary aerosol source of ammonium nitrate, soil and construction dust source, and incinerator and fuel oil burning emission source. For the vehicle emission source, which has been comprehensively described in the atmospheric emission inventory but cannot be identified by the receptor model, an atmospheric dispersion model was used to estimate its contributions. In Dundee, a significant percentage, 67.5%, of the aerosol mass sampled at the study station could be attributed to the six sources named above.

Effects of GABA receptor antagonists on thresholds of P23H rat retinal ganglion cells to electrical stimulation of the retina

NASA Astrophysics Data System (ADS)

Jensen, Ralph J.; Rizzo, Joseph F., III

2011-06-01

An electronic retinal prosthesis may provide useful vision for patients suffering from retinitis pigmentosa (RP). In animal models of RP, the amount of current needed to activate retinal ganglion cells (RGCs) is higher than in normal, healthy retinas. In this study, we sought to reduce the stimulation thresholds of RGCs in a degenerate rat model (P23H-line 1) by blocking GABA receptor mediated inhibition in the retina. We examined the effects of TPMPA, a GABAC receptor antagonist, and SR95531, a GABAA receptor antagonist, on the electrically evoked responses of RGCs to biphasic current pulses delivered to the subretinal surface through a 400 µm diameter electrode. Both TPMPA and SR95531 reduced the stimulation thresholds of ON-center RGCs on average by 15% and 20% respectively. Co-application of the two GABA receptor antagonists had the greatest effect, on average reducing stimulation thresholds by 32%. In addition, co-application of the two GABA receptor antagonists increased the magnitude of the electrically evoked responses on average three-fold. Neither TPMPA nor SR95531, applied alone or in combination, had consistent effects on the stimulation thresholds of OFF-center RGCs. We suggest that the effects of the GABA receptor antagonists on ON-center RGCs may be attributable to blockage of GABA receptors on the axon terminals of ON bipolar cells.

Effects of L-histidine depletion and L-tyrosine/L-phenylalanine depletion on sensory and motor processes in healthy volunteers

PubMed Central

van Ruitenbeek, P; Sambeth, A; Vermeeren, A; Young, SN; Riedel, WJ

2009-01-01

Background and purpose: Animal studies show that histamine plays a role in cognitive functioning and that histamine H3-receptor antagonists, which increase histaminergic function through presynaptic receptors, improve cognitive performance in models of clinical cognitive deficits. In order to test such new drugs in humans, a model for cognitive impairments induced by low histaminergic functions would be useful. Studies with histamine H1-receptor antagonists have shown limitations as a model. Here we evaluated whether depletion of L-histidine, the precursor of histamine, was effective in altering measures associated with histamine in humans and the behavioural and electrophysiological (event-related-potentials) effects. Experimental approach: Seventeen healthy volunteers completed a three-way, double-blind, crossover study with L-histidine depletion, L-tyrosine/L-phenylalanine depletion (active control) and placebo as treatments. Interactions with task manipulations in a choice reaction time task were studied. Task demands were increased using visual stimulus degradation and increased response complexity. In addition, subjective and objective measures of sedation and critical tracking task performance were assessed. Key results: Measures of sedation and critical tracking task performance were not affected by treatment. L-histidine depletion was effective and enlarged the effect of response complexity as measured with the response-locked lateralized readiness potential onset latency. Conclusions and implications: L-histidine depletion affected response- but not stimulus-related processes, in contrast to the effects of H1-receptor antagonists which were previously found to affect primarily stimulus-related processes. L-histidine depletion is promising as a model for histamine-based cognitive impairment. However, these effects need to be confirmed by further studies. PMID:19413574

Can human allergy drug fexofenadine, an antagonist of histamine (H1) receptor, be used to treat dog and cat? Homology modeling, docking and molecular dynamic Simulation of three H1 receptors in complex with fexofenadine.

PubMed

Sader, Safaa; Cai, Jun; Muller, Anna C G; Wu, Chun

2017-08-01

Fexofenadine, a potent antagonist to human histamine 1 (H 1 ) receptor, is a non-sedative third generation antihistamine that is widely used to treat various human allergic conditions such as allergic rhinitis, conjunctivitis and atopic dermatitis. Encouragingly, it's been successfully used to treat canine atopic dermatitis, this supports the notion that it might have a great potential for treating other canine allergic conditions and other mammal pets such as dog. Regrettably, while there is a myriad of studies conducted on the interactions of antihistamines with human H 1 receptor, the similar studies on non-human pet H 1 are considerably scarce. The published studies using the first and second generation antihistamines drugs have shown that the antihistamine response is varied and unpredictable. Thus, to probe its efficacy on pet, the homology models of dog and cat H 1 receptors were built based on the crystal structure of human H 1 receptor bound to antagonist doxepin (PDB 3RZE) and fexofenadine was subsequently docked to human, dog and cat H 1 receptors. The docked complexes are then subjected to 1000ns molecular dynamics (MD) simulations with explicit membrane. Our calculated MM/GBSA binding energies indicated that fexofenadine binds comparably to the three receptors; and our MD data also showed the binding poses, structural and dynamic features among three receptors are very similar. Therefore, our data supported the application of fexofenadine to the H 1 related allergic conditions of dog and cat. Nonetheless, subtle systemic differences among human, dog and cat H 1 receptors were also identified. Clearly, there is still a space to develop a more selective, potent and safe antihistamine alternatives such as Fexofenadine for dog or cat based on these differences. Our computation approach might provide a fast and economic way to predict if human antihistamine drugs can also be safely and efficaciously administered to animals. Copyright © 2017 Elsevier Inc. All rights reserved.

Serotonergic and dopaminergic distinctions in the behavioral pharmacology of (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) and lysergic acid diethylamide (LSD).

PubMed

Schindler, Emmanuelle A D; Dave, Kuldip D; Smolock, Elaine M; Aloyo, Vincent J; Harvey, John A

2012-03-01

After decades of social stigma, hallucinogens have reappeared in the clinical literature demonstrating unique benefits in medicine. The precise behavioral pharmacology of these compounds remains unclear, however. Two commonly studied hallucinogens, (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) and lysergic acid diethylamide (LSD), were investigated both in vivo and in vitro to determine the pharmacology of their behavioral effects in an animal model. Rabbits were administered DOI or LSD and observed for head bob behavior after chronic drug treatment or after pretreatment with antagonist ligands. The receptor binding characteristics of DOI and LSD were studied in vitro in frontocortical homogenates from naïve rabbits or ex vivo in animals receiving an acute drug injection. Both DOI- and LSD-elicited head bobs required serotonin(2A) (5-HT(2A)) and dopamine(1) (D(1)) receptor activation. Serotonin(2B/2C) receptors were not implicated in these behaviors. In vitro studies demonstrated that LSD and the 5-HT(2A/2C) receptor antagonist, ritanserin, bound frontocortical 5-HT(2A) receptors in a pseudo-irreversible manner. In contrast, DOI and the 5-HT(2A/2C) receptor antagonist, ketanserin, bound reversibly. These binding properties were reflected in ex vivo binding studies. The two hallucinogens also differed in that LSD showed modest D(1) receptor binding affinity whereas DOI had negligible binding affinity at this receptor. Although DOI and LSD differed in their receptor binding properties, activation of 5-HT(2A) and D(1) receptors was a common mechanism for eliciting head bob behavior. These findings implicate these two receptors in the mechanism of action of hallucinogens. Copyright © 2011 Elsevier Inc. All rights reserved.

Application of receptor models on water quality data in source apportionment in Kuantan River Basin

PubMed Central

2012-01-01

Recent techniques in the management of surface river water have been expanding the demand on the method that can provide more representative of multivariate data set. A proper technique of the architecture of artificial neural network (ANN) model and multiple linear regression (MLR) provides an advance tool for surface water modeling and forecasting. The development of receptor model was applied in order to determine the major sources of pollutants at Kuantan River Basin, Malaysia. Thirteen water quality parameters were used in principal component analysis (PCA) and new variables of fertilizer waste, surface runoff, anthropogenic input, chemical and mineral changes and erosion are successfully developed for modeling purposes. Two models were compared in terms of efficiency and goodness-of-fit for water quality index (WQI) prediction. The results show that APCS-ANN model gives better performance with high R2 value (0.9680) and small root mean square error (RMSE) value (2.6409) compared to APCS-MLR model. Meanwhile from the sensitivity analysis, fertilizer waste acts as the dominant pollutant contributor (59.82%) to the basin studied followed by anthropogenic input (22.48%), surface runoff (13.42%), erosion (2.33%) and lastly chemical and mineral changes (1.95%). Thus, this study concluded that receptor modeling of APCS-ANN can be used to solve various constraints in environmental problem that exist between water distribution variables toward appropriate water quality management. PMID:23369363

In silico Exploration of the Conformational Universe of GPCRs.

PubMed

Rodríguez-Espigares, Ismael; Kaczor, Agnieszka A; Selent, Jana

2016-07-01

The structural plasticity of G protein coupled receptors (GPCRs) leads to a conformational universe going from inactive to active receptor states with several intermediate states. Many of them have not been captured yet and their role for GPCR activation is not well understood. The study of this conformational space and the transition dynamics between different receptor populations is a major challenge in molecular biophysics. The rational design of effector molecules that target such receptor populations allows fine-tuning receptor signalling with higher specificity to produce drugs with safer therapeutic profiles. In this minireview, we outline highly conserved receptor regions which are considered determinant for the establishment of distinct receptor states. We then discuss in-silico approaches such as dimensionality reduction methods and Markov State Models to explore the GPCR conformational universe and exploit the obtained conformations through structure-based drug design. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Feng; Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030; Yang, Yong, E-mail: yyang@houstonmethodist.org

Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocationmore » of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers.« less

The effects of BMY-14802 against L-DOPA- and dopamine agonist-induced dyskinesia in the hemiparkinsonian rat

PubMed Central

Bhide, Nirmal; Lindenbach, David; Surrena, Margaret A.; Goldenberg, Adam A.; Bishop, Christopher; Berger, S. Paul; Paquette, Melanie A.

2013-01-01

Rationale L-DOPA continues to be the primary treatment for patients with Parkinson’s disease; however, the benefits of long-term treatment are often accompanied by debilitating side effects known as dyskinesias. In recent years, several 5-HT1A receptor agonists have been found to reduce dyskinesia in clinical and experimental models of PD. The purported sigma-1 antagonist, BMY-14802 has been previously demonstrated to reduce L-DOPA induced dyskinesia in a 5-HT1A receptor dependent manner. Objective In the present study, we extend these findings by examining the anti-dyskinetic potential of BMY-14802 against L-DOPA, the D1 receptor agonist SKF81297 and the D2 receptor agonist, Quinpirole, in the hemi-parkinsonian rat model. In addition, the receptor specificity of BMY-14802’s effects was evaluated using WAY-100635, a 5-HT1A receptor antagonist. Results Results confirmed the dose-dependent (20>10>5 mg/kg) anti-dyskinetic effects of BMY-14802 against L-DOPA with preservation of antiparkinsonian efficacy at 10 mg/kg. BMY-14802 at 10 and 20 mg/kg also reduced dyskinesia induced by both D1 and D2 receptor agonists. Additionally, BMY-14802’s anti-dyskinetic effects against L-DOPA, but not SKF81297 or Quinpirole, were reversed by WAY-100635 (0.5 mg/kg). Conclusion Collectively, these findings demonstrate that BMY-14802 provides anti-dyskinetic relief against L-DOPA and direct DA agonist in a preclinical model of PD, acting via multiple receptor systems and supports the utility of such compounds for the improved treatment of PD. PMID:23389756

5HT2A receptor blockade in dorsomedial striatum reduces repetitive behaviors in BTBR mice.

PubMed

Amodeo, D A; Rivera, E; Cook, E H; Sweeney, J A; Ragozzino, M E

2017-03-01

Restricted and repetitive behaviors are a defining feature of autism, which can be expressed as a cognitive flexibility deficit or stereotyped, motor behaviors. There is limited knowledge about the underlying neuropathophysiology contributing to these behaviors. Previous findings suggest that central 5HT 2A receptor activity is altered in autism, while recent work indicates that systemic 5HT 2A receptor antagonist treatment reduces repetitive behaviors in an idiopathic model of autism. 5HT 2A receptors are expressed in the orbitofrontal cortex and striatum. These two regions have been shown to be altered in autism. The present study investigated whether 5HT 2A receptor blockade in the dorsomedial striatum or orbitofrontal cortex in the BTBR mouse strain, an idiopathic model of autism, affects the phenotype related to restricted and repetitive behaviors. Microinfusion of the 5HT 2A receptor antagonist, M100907 into the dorsomedial striatum alleviated a reversal learning impairment and attenuated grooming behavior. M100907 infusion into the orbitofrontal cortex increased perseveration during reversal learning and potentiated grooming. These findings suggest that increased 5HT 2A receptor activity in the dorsomedial striatum may contribute to behavioral inflexibility and stereotyped behaviors in the BTBR mouse. 5HT 2A receptor signaling in the orbitofrontal cortex may be critical for inhibiting a previously learned response during reversal learning and expression of stereotyped behavior. The present results suggest which brain areas exhibit abnormalities underlying repetitive behaviors in an idiopathic mouse model of autism, as well as which brain areas systemic treatment with M100907 may principally act on in BTBR mice to attenuate repetitive behaviors. © 2016 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

5-HT2C receptor involvement in the control of persistence in the reinforced spatial alternation animal model of obsessive-compulsive disorder.

PubMed

Papakosta, Vassiliki-Maria; Kalogerakou, Stamatina; Kontis, Dimitris; Anyfandi, Eleni; Theochari, Eirini; Boulougouris, Vasileios; Papadopoulos, Sokrates; Panagis, George; Tsaltas, Eleftheria

2013-04-15

The serotonergic system is implicated in the pathophysiology of obsessive-compulsive disorder (OCD). However, the distinct role of serotonin (5-HT) receptor subtypes remains unclear. This study investigates the contribution of 5-HT2A and 5-HT2C receptors in the modulation of persistence in the reinforced spatial alternation model of OCD. Male Wistar rats were assessed for spontaneous and pharmacologically induced (by m-chlorophenylpiperazine: mCPP) directional persistence in the reinforced alternation OCD model. Systemic administration of mCPP (non-specific 5-HT agonist, 2.5mg/kg), M100907 (selective 5-HT2A receptor antagonist, 0.08 mg/kg), SB242084 (selective 5-HT2C receptor antagonist, 0.5 mg/kg) and vehicle was used. Experiment 1 investigated M100907 and SB242084 effects in animals spontaneously exhibiting high and low persistence during the early stages of alternation training. Experiment 2 investigated M100900 and SB242084 effects on mCPP-induced persistence. Under the regime used in Experiment 1, 5-HT2A or 5-HT2C receptor antagonism did not affect spontaneous directional persistence in either high or low persistence groups. In Experiment 2, 5-HT2C but not 5-HT2A receptor antagonism significantly reduced, but did not abolish, mCPP-induced directional persistence. These findings suggest that 5-HT2C but not 5-HT2A receptors contribute to the modulation of mCPP-induced persistent behaviour, raising the possibility that the use of 5-HT2C antagonists may have a therapeutic value in OCD. Copyright © 2013 Elsevier B.V. All rights reserved.

Design of ET(B) receptor agonists: NMR spectroscopic and conformational studies of ET7-21[Leu7, Aib11, Cys(Acm)15].

PubMed

Hewage, Chandralal M; Jiang, Lu; Parkinson, John A; Ramage, Robert; Sadler, Ian H

2002-03-01

In a previous report we have shown that the endothelin-B receptor-selective linear endothelin peptide, ET-1[Cys (Acm)1,15, Ala3, Leu7, Aib11], folds into an alpha-helical conformation in a methanol-d3/water co-solvent [Hewage et al. (1998) FEBS Lett., 425, 234-238]. To study the requirements for the structure-activity relationships, truncated analogues of this peptide were subjected to further studies. Here we report the solution conformation of ET7-21[Leu7, Aib11, Cys(Acm)15], in a methanol-d3/water co-solvent at pH 3.6, by NMR spectroscopic and molecular modelling studies. Further truncation of this short peptide results in it displaying poor agonist activity. The modelled structure shows that the peptide folds into an alpha-helical conformation between residues Lys9-His16, whereas the C-terminus prefers no fixed conformation. This truncated linear endothelin analogue is pivotal for designing endothelin-B receptor agonists.

Targeting neurotransmitter receptors with nanoparticles in vivo allows single-molecule tracking in acute brain slices

NASA Astrophysics Data System (ADS)

Varela, Juan A.; Dupuis, Julien P.; Etchepare, Laetitia; Espana, Agnès; Cognet, Laurent; Groc, Laurent

2016-03-01

Single-molecule imaging has changed the way we understand many biological mechanisms, particularly in neurobiology, by shedding light on intricate molecular events down to the nanoscale. However, current single-molecule studies in neuroscience have been limited to cultured neurons or organotypic slices, leaving as an open question the existence of fast receptor diffusion in intact brain tissue. Here, for the first time, we targeted dopamine receptors in vivo with functionalized quantum dots and were able to perform single-molecule tracking in acute rat brain slices. We propose a novel delocalized and non-inflammatory way of delivering nanoparticles (NPs) in vivo to the brain, which allowed us to label and track genetically engineered surface dopamine receptors in neocortical neurons, revealing inherent behaviour and receptor activity regulations. We thus propose a NP-based platform for single-molecule studies in the living brain, opening new avenues of research in physiological and pathological animal models.

Is the isolated ligand binding domain a good model of the domain in the native receptor?

PubMed

Deming, Dustin; Cheng, Qing; Jayaraman, Vasanthi

2003-05-16

Numerous studies have used the atomic level structure of the isolated ligand binding domain of the glutamate receptor to elucidate the agonist-induced activation and desensitization processes in this group of proteins. However, no study has demonstrated the structural equivalence of the isolated ligand binding fragments and the protein in the native receptor. In this report, using visible absorption spectroscopy we show that the electronic environment of the antagonist 6-cyano-7-nitro-2,3-dihydroxyquinoxaline is identical for the isolated protein and the native glutamate receptors expressed in cells. Our results hence establish that the local structure of the ligand binding site is the same in the two proteins and validate the detailed structure-function relationships that have been developed based on a comparison of the structure of the isolated ligand binding domain and electrophysiological consequences in the native receptor.

The role of the dopamine D1 receptor in social cognition: studies using a novel genetic rat model.

PubMed

Homberg, Judith R; Olivier, Jocelien D A; VandenBroeke, Marie; Youn, Jiun; Ellenbroek, Arabella K; Karel, Peter; Shan, Ling; van Boxtel, Ruben; Ooms, Sharon; Balemans, Monique; Langedijk, Jacqueline; Muller, Mareike; Vriend, Gert; Cools, Alexander R; Cuppen, Edwin; Ellenbroek, Bart A

2016-10-01

Social cognition is an endophenotype that is impaired in schizophrenia and several other (comorbid) psychiatric disorders. One of the modulators of social cognition is dopamine, but its role is not clear. The effects of dopamine are mediated through dopamine receptors, including the dopamine D1 receptor (Drd1). Because current Drd1 receptor agonists are not Drd1 selective, pharmacological tools are not sufficient to delineate the role of the Drd1. Here, we describe a novel rat model with a genetic mutation in Drd1 in which we measured basic behavioural phenotypes and social cognition. The I116S mutation was predicted to render the receptor less stable. In line with this computational prediction, this Drd1 mutation led to a decreased transmembrane insertion of Drd1, whereas Drd1 expression, as measured by Drd1 mRNA levels, remained unaffected. Owing to decreased transmembrane Drd1 insertion, the mutant rats displayed normal basic motoric and neurological parameters, as well as locomotor activity and anxiety-like behaviour. However, measures of social cognition like social interaction, scent marking, pup ultrasonic vocalizations and sociability, were strongly reduced in the mutant rats. This profile of the Drd1 mutant rat offers the field of neuroscience a novel genetic rat model to study a series of psychiatric disorders including schizophrenia, autism, depression, bipolar disorder and drug addiction. © 2016. Published by The Company of Biologists Ltd.

Exploring the Interaction between TSC2, PTEN, and the NMDA Receptor in Animal Models of Tuberous Sclerosis

DTIC Science & Technology

2014-09-01

Sunnen CN, Crowell B, Lee GH, Anderson AE, and D’Arcangelo G. Examination of the Role of Pten in Ionotropic Glutamate Receptor Expression. National...PTEN, and the NMDA Receptor in Animal Models of Tuberous Sclerosis PRINCIPAL INVESTIGATOR: D’Arcangelo, Gabriella CONTRACTING...June 2014 4. TITLE AND SUBTITLE Exploring the Interaction between TSC2, PTEN, and the NMDA Receptor in Animal Models of Tuberous Sclerosis 5a

Odor Preference Learning and Memory Modify GluA1 Phosphorylation and GluA1 Distribution in the Neonate Rat Olfactory Bulb: Testing the AMPA Receptor Hypothesis in an Appetitive Learning Model

ERIC Educational Resources Information Center

Cui, Wen; Darby-King, Andrea; Grimes, Matthew T.; Howland, John G.; Wang, Yu Tian; McLean, John H.; Harley, Carolyn W.

2011-01-01

An increase in synaptic AMPA receptors is hypothesized to mediate learning and memory. AMPA receptor increases have been reported in aversive learning models, although it is not clear if they are seen with memory maintenance. Here we examine AMPA receptor changes in a cAMP/PKA/CREB-dependent appetitive learning model: odor preference learning in…

Decreased levels of NMDA but not AMPA receptors in the lipid-raft fraction of 3xTg-AD model of Alzheimer's disease: Relation to Arc/Arg3.1 protein expression.

PubMed

Morin, Jean-Pascal; Díaz-Cintra, Sofía; Bermúdez-Rattoni, Federico; Delint-Ramírez, Ilse

2016-11-01

It was recently suggested that alteration in lipid raft composition in Alzheimer's disease may lead to perturbations in neurons signalosome, which may help explain the deficits observed in synaptic plasticity mechanisms and long-term memory impairments in AD models. As a first effort to address this issue, we evaluated lipid-raft contents of distinct NMDA and AMPA receptor subunits in the hippocampus of the 3xTg-AD model of Alzheimer's disease. Our results show that compared to controls, 10 months-old 3xTg-AD mice have diminished levels of NMDA receptors in rafts but not in post-synaptic density or total fractions. Additionally, the levels of GluR1 were unaltered in all the analyzed fractions. Finally, we went on to show that the diminished levels of NMDA receptors in rafts correlated with diminished global levels of Arc/Arg3.1, a synaptic protein with a central role in long-term memory formation. This study adds to our current understanding of the signaling pathways disruptions observed in current Alzheimer's disease models. Copyright © 2016 Elsevier Ltd. All rights reserved.

Arrestin Scaffolds NHERF1 to the P2Y12 Receptor to Regulate Receptor Internalization*

PubMed Central

Nisar, Shaista P.; Cunningham, Margaret; Saxena, Kunal; Pope, Robert J.; Kelly, Eamonn; Mundell, Stuart J.

2012-01-01

We have recently shown in a patient with mild bleeding that the PDZ-binding motif of the platelet G protein-coupled P2Y12 receptor (P2Y12R) is required for effective receptor traffic in human platelets. In this study we show for the first time that the PDZ motif-binding protein NHERF1 exerts a major role in potentiating G protein-coupled receptor (GPCR) internalization. NHERF1 interacts with the C-tail of the P2Y12R and unlike many other GPCRs, NHERF1 interaction is required for effective P2Y12R internalization. In vitro and prior to agonist stimulation P2Y12R/NHERF1 interaction requires the intact PDZ binding motif of this receptor. Interestingly on receptor stimulation NHERF1 no longer interacts directly with the receptor but instead binds to the receptor via the endocytic scaffolding protein arrestin. These findings suggest a novel model by which arrestin can serve as an adaptor to promote NHERF1 interaction with a GPCR to facilitate effective NHERF1-dependent receptor internalization. PMID:22610101

Arrestin scaffolds NHERF1 to the P2Y12 receptor to regulate receptor internalization.

PubMed

Nisar, Shaista P; Cunningham, Margaret; Saxena, Kunal; Pope, Robert J; Kelly, Eamonn; Mundell, Stuart J

2012-07-13

We have recently shown in a patient with mild bleeding that the PDZ-binding motif of the platelet G protein-coupled P2Y(12) receptor (P2Y(12)R) is required for effective receptor traffic in human platelets. In this study we show for the first time that the PDZ motif-binding protein NHERF1 exerts a major role in potentiating G protein-coupled receptor (GPCR) internalization. NHERF1 interacts with the C-tail of the P2Y(12)R and unlike many other GPCRs, NHERF1 interaction is required for effective P2Y(12)R internalization. In vitro and prior to agonist stimulation P2Y(12)R/NHERF1 interaction requires the intact PDZ binding motif of this receptor. Interestingly on receptor stimulation NHERF1 no longer interacts directly with the receptor but instead binds to the receptor via the endocytic scaffolding protein arrestin. These findings suggest a novel model by which arrestin can serve as an adaptor to promote NHERF1 interaction with a GPCR to facilitate effective NHERF1-dependent receptor internalization.

Current perspectives on selective dopamine D3 receptor antagonists as pharmacotherapeutics for addictions and related disorders

PubMed Central

Heidbreder, Christian A.; Newman, Amy H.

2011-01-01

Repeated exposure to drugs of abuse produces long-term molecular and neurochemical changes that may explain the core features of addiction, such as the compulsive seeking and taking of the drug, as well as the risk of relapse. A growing number of new molecular and cellular targets of addictive drugs have been identified, and rapid advances are being made in relating those targets to specific behavioral phenotypes in animal models of addiction. In this context, the pattern of expression of the dopamine (DA) D3 receptor in the rodent and human brain and changes in this pattern in response to drugs of abuse have contributed primarily to direct research efforts toward the development of selective DA D3 receptor antagonists. Growing preclinical evidence indicates that these compounds may actually regulate the motivation to self-administer drugs and disrupt drug-associated cue-induced craving. This report will be divided into three parts. First, preclinical evidence in support of the efficacy of selective DA D3 receptor antagonists in animal models of drug addiction will be reviewed. The effects of mixed DA D2/D3 receptor antagonists will not be discussed here because most of these compounds have low selectivity at the D3 versus D2 receptor, and their efficacy profile is related primarily to functional antagonism at D2 receptors and possibly interactions with other neurotransmitter systems. Second, major advances in medicinal chemistry for the identification and optimization of selective DA D3 receptor antagonists and partial agonists will be analyzed. Third, translational research from preclinical efficacy studies to so-called proof-of-concept studies for drug addiction indications will be discussed. PMID:20201845

Stereochemical studies of the monocyclic agouti-related protein (103-122) Arg-Phe-Phe residues: conversion of a melanocortin-4 receptor antagonist into an agonist and results in the discovery of a potent and selective melanocortin-1 agonist.

PubMed

Joseph, Christine G; Wang, Xiang S; Scott, Joseph W; Bauzo, Rayna M; Xiang, Zhimin; Richards, Nigel G; Haskell-Luevano, Carrie

2004-12-30

The agouti-related protein (AGRP) is an endogenous antagonist of the centrally expressed melanocortin receptors. The melanocortin-4 receptor (MC4R) is involved in energy homeostasis, food intake, sexual function, and obesity. The endogenous hAGRP protein is 132 amino acids in length, possesses five disulfide bridges at the C-terminus of the molecule, and is expressed in the hypothalamus of the brain. We have previously reported that a monocyclic hAGRP(103-122) peptide is an antagonist at the melanocortin receptors expressed in the brain. Stereochemical inversion from the endogenous l- to d-isomers of single or multiple amino acid modifications in this monocyclic truncated hAGRP sequence resulted in molecules that are converted from melanocortin receptor antagonists into melanocortin receptor agonists. The Asp-Pro-Ala-Ala-Thr-Ala-Tyr-cyclo[Cys-Arg-DPhe-DPhe-Asn-Ala-Phe-Cys]-Tyr-Ala-Arg-Lys-Leu peptide resulted in a 60 nM melanocortin-1 receptor agonist that is 100-fold selective versus the mMC4R, 1000-fold selective versus the mMC3R, and ca. 180-fold selective versus the mMC5R. In attempts to identify putative ligand-receptor interactions that may be participating in the agonist induced stimulation of the MC4R, selected ligands were docked into a homology molecular model of the mMC4R. These modeling studies have putatively identified hAGRP ligand DArg111-mMC4RAsn115 (TM3) and the hAGRP DPhe113-mMC4RPhe176 (TM4) interactions as important for agonist activity.

Current perspectives on selective dopamine D(3) receptor antagonists as pharmacotherapeutics for addictions and related disorders.

PubMed

Heidbreder, Christian A; Newman, Amy H

2010-02-01

Repeated exposure to drugs of abuse produces long-term molecular and neurochemical changes that may explain the core features of addiction, such as the compulsive seeking and taking of the drug, as well as the risk of relapse. A growing number of new molecular and cellular targets of addictive drugs have been identified, and rapid advances are being made in relating those targets to specific behavioral phenotypes in animal models of addiction. In this context, the pattern of expression of the dopamine (DA) D(3) receptor in the rodent and human brain and changes in this pattern in response to drugs of abuse have contributed primarily to direct research efforts toward the development of selective DA D(3) receptor antagonists. Growing preclinical evidence indicates that these compounds may actually regulate the motivation to self-administer drugs and disrupt drug-associated cue-induced craving. This report will be divided into three parts. First, preclinical evidence in support of the efficacy of selective DA D(3) receptor antagonists in animal models of drug addiction will be reviewed. The effects of mixed DA D(2)/D(3) receptor antagonists will not be discussed here because most of these compounds have low selectivity at the D(3) versus D(2) receptor, and their efficacy profile is related primarily to functional antagonism at D(2) receptors and possibly interactions with other neurotransmitter systems. Second, major advances in medicinal chemistry for the identification and optimization of selective DA D(3) receptor antagonists and partial agonists will be analyzed. Third, translational research from preclinical efficacy studies to so-called proof-of-concept studies for drug addiction indications will be discussed.

Endocytosis and membrane receptor internalization: implication of F-BAR protein Carom

PubMed Central

Xu, Yanjie; Liu, Suxuan; Xia, Jixiang; Stein, Sam; Ramon, Cueto; Xi, Hang; Wang, Luqiao; Xiong, Xinyu; Zhang, Lixiao; He, Dingwen; Yang, William; Zhao, Xianxian; Cheng, Xiaoshu; Yang, Xiaofeng; Wang, Hong

2016-01-01

Endocytosis is a cellular process mostly responsible for membrane receptor internalization. Cell membrane receptors bind to their ligands and form a complex which can be internalized. We previously proposed that F-BAR protein initiates membrane curvature and mediates endocytosis via their binding partners. However, F-BAR protein partners involved in membrane receptor endocytosis and the regulatory mechanism remain unknown. In this study, we established a group of database mining strategies to explore mechanisms underlying receptor-related endocytosis. We identified 34 endocytic membrane receptors and 10 regulating proteins for vesicle formation in clathrin-dependent endocytosis (CDE), a major process of membrane receptor internalization. We found that F-BAR protein FCHSD2 (Carom) may facilitate endocytosis via 9 endocytic partners. Carom is highly expressed, along with highly expressed endocytic membrane receptors and partners, in endothelial cells and macrophages. We established 3 models of Carom-receptor complex and their intracellular trafficking based on protein-protein interaction and subcellular localization. We conclude that Carom may mediate receptor endocytosis and transport endocytic receptors to the cytoplasm for receptor signaling and lysosome/proteasome degradation, or to the nucleus for RNA processing, gene transcription and DNA repair. PMID:28199211

Modeling Conformational Transitions and Energetics of Ligand Binding with the Glutamate Receptor Ligand Binding Domain

NASA Astrophysics Data System (ADS)

Kurnikova, Maria

2009-03-01

Understanding of protein motion and energetics of conformational transitions is crucial to understanding protein function. The glutamate receptor ligand binding domain (GluR2 S1S2) is a two lobe protein, which binds ligand at the interface of two lobes and undergoes conformational transition. The cleft closure conformational transition of S1S2 has been implicated in gating of the ion channel formed by the transmembrane domain of the receptor. In this study we present a composite multi-faceted theoretical analysis of the detailed mechanism of this conformational transition based on rigid cluster decomposition of the protein structure [1] and identifying hydrogen bonds that are responsible for stabilizing the closed conformation [2]. Free energy of the protein reorganization upon ligand binding was calculated using combined Thermodynamic Integration (TI) and Umbrella Sampling (US) simulations [3]. Ligand -- protein interactions in the binding cleft were analyzed using Molecular Dynamics, continuum electrostatics and QM/MM models [4]. All model calculations compare well with corresponding experimental measurements. [4pt] [1] Protein Flexibility using Constraints from Molecular Dynamics Simulations T. Mamonova, B. Hespenheide, R. Straub, M. F. Thorpe, M. G. Kurnikova , Phys. Biol., 2, S137 (2005)[0pt] [2] Theoretical Study of the Glutamate Receptor Ligand Binding Domain Flexibility and Conformational Reorganization T. Mamonova, K. Speranskiy, and M. Kurnikova , Prot.: Struct., Func., Bioinf., 73,656 (2008)[0pt] [3] Energetics of the cleft closing transition and glutamate binding in the Glutamate Receptor ligand Binding Domain T. Mamonova, M. Yonkunas, and M. Kurnikova Biochemistry 47, 11077 (2008)[0pt] [4] On the Binding Determinants of the Glutamate Agonist with the Glutamate Receptor Ligand Binding Domain K. Speranskiy and M. Kurnikova Biochemistry 44, 11208 (2005)

Effects of Vanadium-Containing Compounds on Membrane Lipids and on Microdomains Used in Receptor-Mediated Signaling

PubMed Central

Roess, Deborah A.; Smith, Steven M. L.; Winter, Peter; Zhou, Jun; Dou, Ping; Baruah, Bharat; Trujillo, Alejandro M.; Levinger, Nancy E.; Yang, Xioda; Barisas, B. George; Crans, Debbie C.

2011-01-01

There is increasing evidence for the involvement of plasma membrane microdomains in insulin receptor function. Moreover, disruption of these structures, which are typically enriched in sphingomyelin and cholesterol, results in insulin resistance. Treatment strategies for insulin resistance include the use of vanadium compounds which have been shown in animal models to enhance insulin responsiveness. One possible mechanism for insulin-enhancing effects might involve direct effects of vanadium compounds on membrane lipid organization. These changes in lipid organization promote the partitioning of insulin receptors and other receptors into membrane microdomains where receptors are optimally functional. To explore this possibility, we have used several strategies involving vanadium complexes such as [VO2dipic]− (pyridin-2,6-dicarboxylatodioxovanadium(V)), decavanadate (V10O286−, V10), BMOV (bis(maltolato)oxovanadium(IV)) and [VO(saltris)]2 (2-salicylideniminato-2-(hydroxymethyl)-1,3-dihydroxypropane-oxovanadium(V)). Our strategies include an evaluation of interactions between vanadium-containing compounds and model lipid systems, an evaluation of the effects of vanadium compounds on lipid fluidity in erythrocyte membranes, and studies of the effects of vanadium-containing compounds on signaling events initiated by receptors known to use membrane microdomains as signaling platforms. PMID:18729092

Anesthetic sites and allosteric mechanisms of action on Cys-loop ligand-gated ion channels.

PubMed

Forman, Stuart A; Miller, Keith W

2011-02-01

The Cys-loop ligand-gated ion channel superfamily is a major group of neurotransmitter-activated receptors in the central and peripheral nervous system. The superfamily includes inhibitory receptors stimulated by γ-aminobutyric acid (GABA) and glycine and excitatory receptors stimulated by acetylcholine and serotonin. The first part of this review presents current evidence on the location of the anesthetic binding sites on these channels and the mechanism by which binding to these sites alters their function. The second part of the review addresses the basis for this selectivity, and the third part describes the predictive power of a quantitative allosteric model showing the actions of etomidate on γ-aminobutyric acid type A receptors (GABA(A)Rs). General anesthetics at clinical concentrations inhibit the excitatory receptors and enhance the inhibitory receptors. The location of general anesthetic binding sites on these receptors is being defined by photoactivable analogues of general anesthetics. The receptor studied most extensively is the muscle-type nicotinic acetylcholine receptor (nAChR), and progress is now being made with GABA(A)Rs. There are three categories of sites that are all in the transmembrane domain: 1) within a single subunit's four-helix bundle (intrasubunit site; halothane and etomidate on the δ subunit of AChRs); 2) between five subunits in the transmembrane conduction pore (channel lumen sites; etomidate and alcohols on nAChR); and 3) between two subunits (subunit interface sites; etomidate between the α1 and β2/3 subunits of the GABA(A)R). These binding sites function allosterically. Certain conformations of a receptor bind the anesthetic with greater affinity than others. Time-resolved photolabelling of some sites occurs within milliseconds of channel opening on the nAChR but not before. In GABA(A)Rs, electrophysiological data fit an allosteric model in which etomidate binds to and stabilizes the open state, increasing both the fraction of open channels and their lifetime. As predicted by the model, the channel-stabilizing action of etomidate is so strong that higher concentrations open the channel in the absence of agonist. The formal functional paradigm presented for etomidate may apply to other potent general anesthetic drugs. Combining photolabelling with structure-function mutational studies in the context of allosteric mechanisms should lead us to a more detailed understanding of how and where these important drugs act.

Chronic MK-801 Application in Adolescence and Early Adulthood: A Spatial Working Memory Deficit in Adult Long-Evans Rats But No Changes in the Hippocampal NMDA Receptor Subunits

PubMed Central

Uttl, Libor; Petrasek, Tomas; Sengul, Hilal; Svojanovska, Marketa; Lobellova, Veronika; Vales, Karel; Radostova, Dominika; Tsenov, Grygoriy; Kubova, Hana; Mikulecka, Anna; Svoboda, Jan; Stuchlik, Ales

2018-01-01

The role of NMDA receptors in learning, memory and hippocampal function has long been recognized. Post-mortem studies have indicated that the expression or subunit composition of the NMDA glutamate receptor subtype might be related to the impaired cognitive functions found in schizophrenia patients. NMDA receptor antagonists have been used to develop animal models of this disorder. There is accumulating evidence showing that not only the acute but also the chronic application of NMDA receptor antagonists may induce schizophrenia-like alterations in behavior and brain functions. However, limited evidence is available regarding the consequences of NMDA receptor blockage during periods of adolescence and early adulthood. This study tested the hypothesis that a 2-week treatment of male Long-Evans and Wistar rats with dizocilpine (MK-801; 0.5 mg/kg daily) starting at postnatal days (PD) 30 and 60 would cause a long-term cognitive deficit and changes in the levels of NMDA receptor subunits. The working memory version of the Morris water maze (MWM) and active place avoidance with reversal on a rotating arena (Carousel) requiring cognitive coordination and flexibility probed cognitive functions and an elevated-plus maze (EPM) was used to measure anxiety-like behavior. The western blot method was used to determine changes in NMDA receptor subunit levels in the hippocampus. Our results showed no significant changes in behaviors in Wistar rats. Slightly elevated anxiety-like behavior was observed in the EPM in Long-Evans rats with the onset of treatment on PD 30. Furthermore, Long-Evans rats treated from PD 60 displayed impaired working memory in the MWM. There were; however, no significant changes in the levels of NMDA receptor subunits because of MK-801 administration. These findings suggest that a 2-week treatment starting on PD 60 in Long-Evans rats leads to long-term changes in working memory, but this deficit is not paralleled by changes in NMDA receptor subunits. These results support the face validity, but not construct validity of this model. We suggest that chronic treatment of adolescent and adult rats does not constitute a plausible animal model of schizophrenia. PMID:29487522

Chronic MK-801 Application in Adolescence and Early Adulthood: A Spatial Working Memory Deficit in Adult Long-Evans Rats But No Changes in the Hippocampal NMDA Receptor Subunits.

PubMed

Uttl, Libor; Petrasek, Tomas; Sengul, Hilal; Svojanovska, Marketa; Lobellova, Veronika; Vales, Karel; Radostova, Dominika; Tsenov, Grygoriy; Kubova, Hana; Mikulecka, Anna; Svoboda, Jan; Stuchlik, Ales

2018-01-01

The role of NMDA receptors in learning, memory and hippocampal function has long been recognized. Post-mortem studies have indicated that the expression or subunit composition of the NMDA glutamate receptor subtype might be related to the impaired cognitive functions found in schizophrenia patients. NMDA receptor antagonists have been used to develop animal models of this disorder. There is accumulating evidence showing that not only the acute but also the chronic application of NMDA receptor antagonists may induce schizophrenia-like alterations in behavior and brain functions. However, limited evidence is available regarding the consequences of NMDA receptor blockage during periods of adolescence and early adulthood. This study tested the hypothesis that a 2-week treatment of male Long-Evans and Wistar rats with dizocilpine (MK-801; 0.5 mg/kg daily) starting at postnatal days (PD) 30 and 60 would cause a long-term cognitive deficit and changes in the levels of NMDA receptor subunits. The working memory version of the Morris water maze (MWM) and active place avoidance with reversal on a rotating arena (Carousel) requiring cognitive coordination and flexibility probed cognitive functions and an elevated-plus maze (EPM) was used to measure anxiety-like behavior. The western blot method was used to determine changes in NMDA receptor subunit levels in the hippocampus. Our results showed no significant changes in behaviors in Wistar rats. Slightly elevated anxiety-like behavior was observed in the EPM in Long-Evans rats with the onset of treatment on PD 30. Furthermore, Long-Evans rats treated from PD 60 displayed impaired working memory in the MWM. There were; however, no significant changes in the levels of NMDA receptor subunits because of MK-801 administration. These findings suggest that a 2-week treatment starting on PD 60 in Long-Evans rats leads to long-term changes in working memory, but this deficit is not paralleled by changes in NMDA receptor subunits. These results support the face validity, but not construct validity of this model. We suggest that chronic treatment of adolescent and adult rats does not constitute a plausible animal model of schizophrenia.

Prolonged exposure to WIN55,212-2 causes down-regulation of the CB1 receptor and the development of tolerance to its anticonvulsant effects in the hippocampal neuronal culture model of acquired epilepsy

PubMed Central

Blair, Robert E.; Deshpande, Laxmikant S.; Sombati, Sompong; Elphick, Maurice R.; Martin, Billy R.; DeLorenzo, Robert J.

2009-01-01

Summary Cannabinoids have been shown to cause CB1-receptor dependent anticonvulsant activity in both in vivo and in vitro models of status epilepticus (SE) and acquired epilepsy (AE). It has been further demonstrated in these models that the endocannabinoid system functions in a tonic manner to suppress seizure discharges through a CB1-receptor dependent pathway. Although acute cannabinoid treatment has anticonvulsant activity, little is known concerning the effects of prolonged exposure to CB1 agonists and development of tolerance on the epileptic phenotype. This study was carried out to evaluate the effects of prolonged exposure to the CB1 agonist WIN55,212-2 on seizure activity in a hippocampal neuronal culture model of low-Mg2+ induced spontaneous recurrent epileptiform discharges (SREDs). Following low-Mg2+ induced SREDs, cultures were returned to maintenance media containing 10, 100 or 1000 nM WIN55,212-2 from 4 to 24 hours. Whole-cell current-clamp analysis of WIN55,212-2 treated cultures revealed a concentration-dependent increase in SRED frequency. Immunocytochemical staining revealed that WIN55,212-2 treatment induced a concentration-dependent down-regulation of the CB1 receptor in neuronal processes and at both glutamatergic and GABAergic presynaptic terminals. Prolonged exposure to the inactive enantiomer WIN55,212-3 in low-Mg2+ treated cultures had no effect on the frequency of SREDs or CB1 receptor staining. The results from this study further substantiate a role for a tonic CB1 receptor-dependent endocannabinoid regulation of seizure discharge and suggest that prolonged exposure to cannabinoids results in the development of tolerance to the anticonvulsant effects of cannabinoids and an exacerbation of seizure activity in the epileptic phenotype. PMID:19540252

Inverse agonist and neutral antagonist actions of synthetic compounds at an insect 5-HT1 receptor.

PubMed

Troppmann, B; Balfanz, S; Baumann, A; Blenau, W

2010-04-01

5-Hydroxytryptamine (5-HT) has been shown to control and modulate many physiological and behavioural functions in insects. In this study, we report the cloning and pharmacological properties of a 5-HT(1) receptor of an insect model for neurobiology, physiology and pharmacology. A cDNA encoding for the Periplaneta americana 5-HT(1) receptor was amplified from brain cDNA. The receptor was stably expressed in HEK 293 cells, and the functional and pharmacological properties were determined in cAMP assays. Receptor distribution was investigated by RT-PCR and by immunocytochemistry using an affinity-purified polyclonal antiserum. The P. americana 5-HT(1) receptor (Pea5-HT(1)) shares pronounced sequence and functional similarity with mammalian 5-HT(1) receptors. Activation with 5-HT reduced adenylyl cyclase activity in a dose-dependent manner. Pea5-HT(1) was expressed as a constitutively active receptor with methiothepin acting as a neutral antagonist, and WAY 100635 as an inverse agonist. Receptor mRNA was present in various tissues including brain, salivary glands and midgut. Receptor-specific antibodies showed that the native protein was expressed in a glycosylated form in membrane samples of brain and salivary glands. This study marks the first pharmacological identification of an inverse agonist and a neutral antagonist at an insect 5-HT(1) receptor. The results presented here should facilitate further analyses of 5-HT(1) receptors in mediating central and peripheral effects of 5-HT in insects.

5-HT7 receptors as modulators of neuronal excitability, synaptic transmission and plasticity: physiological role and possible implications in autism spectrum disorders

PubMed Central

Ciranna, Lucia; Catania, Maria Vincenza

2014-01-01

Serotonin type 7 receptors (5-HT7) are expressed in several brain areas, regulate brain development, synaptic transmission and plasticity, and therefore are involved in various brain functions such as learning and memory. A number of studies suggest that 5-HT7 receptors could be potential pharmacotherapeutic target for cognitive disorders. Several abnormalities of serotonergic system have been described in patients with autism spectrum disorder (ASD), including abnormal activity of 5-HT transporter, altered blood and brain 5-HT levels, reduced 5-HT synthesis and altered expression of 5-HT receptors in the brain. A specific role for 5-HT7 receptors in ASD has not yet been demonstrated but some evidence implicates their possible involvement. We have recently shown that 5-HT7 receptor activation rescues hippocampal synaptic plasticity in a mouse model of Fragile X Syndrome, a monogenic cause of autism. Several other studies have shown that 5-HT7 receptors modulate behavioral flexibility, exploratory behavior, mood disorders and epilepsy, which include core and co-morbid symptoms of ASD. These findings further suggest an involvement of 5-HT7 receptors in ASD. Here, we review the physiological roles of 5-HT7 receptors and their implications in Fragile X Syndrome and other ASD. PMID:25221471

Dissociating anxiolytic and sedative effects of GABAAergic drugs using temperature and locomotor responses to acute stress

PubMed Central

Klanker, Marianne; Groenink, Lucianne; Korte, S. Mechiel; Cook, James M.; Van Linn, Michael L.; Hopkins, Seth C.; Olivier, Berend

2009-01-01

Rationale The stress-induced hyperthermia (SIH) model is an anxiety model that uses the transient rise in body temperature in response to acute stress. Benzodiazepines produce anxiolytic as well as sedative side effects through nonselective binding to GABAA receptor subunits. The GABAA receptor α1 subunit is associated with sedation, whereas the GABAA receptor α2 and α3 subunits are involved in anxiolytic effects. Objectives We therefore examined the effects of (non) subunit-selective GABAA receptor agonists on temperature and locomotor responses to novel cage stress. Results Using telemetric monitoring of temperature and locomotor activity, we found that nonsubunit-selective GABAA receptor agonist diazepam as well as the α3 subunit-selective receptor agonist TP003 dose-dependently attenuated SIH and locomotor responses. Administration of GABAA receptor α1-selective agonist zolpidem resulted in profound hypothermia and locomotor sedation. The GABAA receptor α1-selective antagonist βCCt antagonized the hypothermia, but did not reverse the SIH response attenuation caused by diazepam and zolpidem. These results suggest an important regulating role for the α1 subunit in thermoregulation and sedation. Ligands of extrasynaptic GABAA receptors such as alcohol and nonbenzodiazepine THIP attenuated the SIH response only at high doses. Conclusions The present study confirms a putative role for the GABAA receptor α1 subunit in hypothermia and sedation and supports a role for α2/3 subunit GABAA receptor agonists in anxiety processes. In conclusion, we show that home cage temperature and locomotor responses to novel home cage stress provide an excellent tool to assess both anxiolytic and sedative effects of various (subunit-selective) GABAAergic compounds. PMID:19169673

Operational models of pharmacological agonism.

PubMed

Black, J W; Leff, P

1983-12-22

The traditional receptor-stimulus model of agonism began with a description of drug action based on the law of mass action and has developed by a series of modifications, each accounting for new experimental evidence. By contrast, in this paper an approach to modelling agonism is taken that begins with the observation that experimental agonist-concentration effect, E/[A], curves are commonly hyperbolic and develops using the deduction that the relation between occupancy and effect must be hyperbolic if the law of mass action applies at the agonist-receptor level. The result is a general model that explicitly describes agonism by three parameters: an agonist-receptor dissociation constant, KA; the total receptor concentration, [R0]; and a parameter, KE, defining the transduction of agonist-receptor complex, AR, into pharmacological effect. The ratio, [R0]/KE, described here as the 'transducer ratio', tau, is a logical definition for the efficacy of an agonist in a system. The model may be extended to account for non-hyperbolic E/[A] curves with no loss of meaning. Analysis shows that an explicit formulation of the traditional receptor-stimulus model is one particular form of the general model but that it is not the simplest. An alternative model is proposed, representing the cognitive and transducer functions of a receptor, that describes agonist action with one fewer parameter than the traditional model. In addition, this model provides a chemical definition of intrinsic efficacy making this parameter experimentally accessible in principle. The alternative models are compared and contrasted with regard to their practical and conceptual utilities in experimental pharmacology.

Activation and desensitization of ionotropic glutamate receptors by selectively triggering pre-existing motions.

PubMed

Krieger, James; Lee, Ji Young; Greger, Ingo H; Bahar, Ivet

2018-02-23

Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that are key players in synaptic transmission and plasticity. They are composed of four subunits, each containing four functional domains, the quaternary packing and collective structural dynamics of which are important determinants of their molecular mechanism of function. With the explosion of structural studies on different members of the family, including the structures of activated open channels, the mechanisms of action of these central signaling machines are now being elucidated. We review the current state of computational studies on two major members of the family, AMPA and NMDA receptors, with focus on molecular simulations and elastic network model analyses that have provided insights into the coupled movements of extracellular and transmembrane domains. We describe the newly emerging mechanisms of activation, allosteric signaling and desensitization, as mainly a selective triggering of pre-existing soft motions, as deduced from computational models and analyses that leverage structural data on intact AMPA and NMDA receptors in different states. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Dynamic dual-tracer MRI-guided fluorescence tomography to quantify receptor density in vivo

PubMed Central

Davis, Scott C.; Samkoe, Kimberley S.; Tichauer, Kenneth M.; Sexton, Kristian J.; Gunn, Jason R.; Deharvengt, Sophie J.; Hasan, Tayyaba; Pogue, Brian W.

2013-01-01

The up-regulation of cell surface receptors has become a central focus in personalized cancer treatment; however, because of the complex nature of contrast agent pharmacokinetics in tumor tissue, methods to quantify receptor binding in vivo remain elusive. Here, we present a dual-tracer optical technique for noninvasive estimation of specific receptor binding in cancer. A multispectral MRI-coupled fluorescence molecular tomography system was used to image the uptake kinetics of two fluorescent tracers injected simultaneously, one tracer targeted to the receptor of interest and the other tracer a nontargeted reference. These dynamic tracer data were then fit to a dual-tracer compartmental model to estimate the density of receptors available for binding in the tissue. Applying this approach to mice with deep-seated gliomas that overexpress the EGF receptor produced an estimate of available receptor density of 2.3 ± 0.5 nM (n = 5), consistent with values estimated in comparative invasive imaging and ex vivo studies. PMID:23671066

STRESS RESPONSE STUDIES USING ANIMAL MODELS

EPA Science Inventory

This presentation will provide the evidence that ozone exposure in animal models induce neuroendocrine stress response and this stress response modulates lung injury and inflammation through adrenergic and glucocorticoid receptors.

PGE2 maintains the tone of the guinea pig trachea through a balance between activation of contractile EP1 receptors and relaxant EP2 receptors

PubMed Central

Säfholm, J; Dahlén, S-E; Delin, I; Maxey, K; Stark, K; Cardell, L-O; Adner, M

2013-01-01

Background and Purpose The guinea pig trachea (GPT) is commonly used in airway pharmacology. The aim of this study was to define the expression and function of EP receptors for PGE2 in GPT as there has been ambiguity concerning their role. Experimental Approach Expression of mRNA for EP receptors and key enzymes in the PGE2 pathway were assessed by real-time PCR using species-specific primers. Functional studies of GPT were performed in tissue organ baths. Key Results Expression of mRNA for the four EP receptors was found in airway smooth muscle. PGE2 displayed a bell-shaped concentration–response curve, where the initial contraction was inhibited by the EP1 receptor antagonist ONO-8130 and the subsequent relaxation by the EP2 receptor antagonist PF-04418948. Neither EP3 (ONO-AE5-599) nor EP4 (ONO-AE3-208) selective receptor antagonists affected the response to PGE2. Expression of COX-2 was greater than COX-1 in GPT, and the spontaneous tone was most effectively abolished by selective COX-2 inhibitors. Furthermore, ONO-8130 and a specific PGE2 antibody eliminated the spontaneous tone, whereas the EP2 antagonist PF-04418948 increased it. Antagonists of other prostanoid receptors had no effect on basal tension. The relaxant EP2 response to PGE2 was maintained after long-term culture, whereas the contractile EP1 response showed homologous desensitization to PGE2, which was prevented by COX-inhibitors. Conclusions and Implications Endogenous PGE2, synthesized predominantly by COX-2, maintains the spontaneous tone of GPT by a balance between contractile EP1 receptors and relaxant EP2 receptors. The model may be used to study interactions between EP receptors. PMID:22934927

Discovery of indolylpiperazinylpyrimidines with dual-target profiles at adenosine A2A and dopamine D2 receptors for Parkinson's disease treatment

PubMed Central

Paira, Priyankar; Tan, Aaron; Herr, Deron Raymond; Lim, Kah Leong; Ng, Chee Hoe; Venkatesan, Gopalakrishnan; Klotz, Karl-Norbert; Federico, Stephanie; Spalluto, Giampiero; Cheong, Siew Lee; Chen, Yu Zong

2018-01-01

Parkinson’s disease (PD) is a neurodegenerative disorder characterized by progressive loss of dopaminergic neurons in the substantia nigra of the human brain, leading to depletion of dopamine production. Dopamine replacement therapy remains the mainstay for attenuation of PD symptoms. Nonetheless, the potential benefit of current pharmacotherapies is mostly limited by adverse side effects, such as drug-induced dyskinesia, motor fluctuations and psychosis. Non-dopaminergic receptors, such as human A2A adenosine receptors, have emerged as important therapeutic targets in potentiating therapeutic effects and reducing the unwanted side effects. In this study, new chemical entities targeting both human A2A adenosine receptor and dopamine D2 receptor were designed and evaluated. Two computational methods, namely support vector machine (SVM) models and Tanimoto similarity-based clustering analysis, were integrated for the identification of compounds containing indole-piperazine-pyrimidine (IPP) scaffold. Subsequent synthesis and testing resulted in compounds 5 and 6, which acted as human A2A adenosine receptor binders in the radioligand competition assay (Ki = 8.7–11.2 μM) as well as human dopamine D2 receptor binders in the artificial cell membrane assay (EC50 = 22.5–40.2 μM). Moreover, compound 5 showed improvement in movement and mitigation of the loss of dopaminergic neurons in Drosophila models of PD. Furthermore, in vitro toxicity studies on compounds 5 and 6 did not reveal any mutagenicity (up to 100 μM), hepatotoxicity (up to 30 μM) or cardiotoxicity (up to 30 μM). PMID:29304113

An anti-nicotinic cognitive challenge model using mecamylamine in comparison with the anti-muscarinic cognitive challenge using scopolamine.

PubMed

Baakman, Anne Catrien; Alvarez-Jimenez, Ricardo; Rissmann, Robert; Klaassen, Erica S; Stevens, Jasper; Goulooze, Sebastiaan C; den Burger, Jeroen C G; Swart, Eleonora L; van Gerven, Joop M A; Groeneveld, Geert Jan

2017-08-01

The muscarinic acetylcholine receptor antagonist scopolamine is often used for proof-of-pharmacology studies with pro-cognitive compounds. From a pharmacological point of view, it would seem more rational to use a nicotinic rather than a muscarinic anticholinergic challenge to prove pharmacology of a nicotinic acetylcholine receptor agonist. This study aims to characterize a nicotinic anticholinergic challenge model using mecamylamine and to compare it to the scopolamine model. In this double-blind, placebo-controlled, four-way cross-over trial, 12 healthy male subjects received oral mecamylamine 10 and 20 mg, intravenous scopolamine 0.5 mg and placebo. Pharmacokinetics were analysed using non-compartmental analysis. Pharmacodynamic effects were measured with a multidimensional test battery that includes neurophysiological, subjective, (visuo)motor and cognitive measurements. All treatments were safe and well tolerated. Mecamylamine had a t max of 2.5 h and a C max of 64.5 ng ml -1 for the 20 mg dose. Mecamylamine had a dose-dependent effect decreasing the adaptive tracking performance and VAS alertness, and increasing the finger tapping and visual verbal learning task performance time and errors. Scopolamine significantly affected almost all pharmacodynamic tests. This study demonstrated that mecamylamine causes nicotinic receptor specific temporary decline in cognitive functioning. Compared with the scopolamine model, pharmacodynamic effects were less pronounced at the dose levels tested; however, mecamylamine caused less sedation. The cognitive effects of scopolamine might at least partly be caused by sedation. Whether the mecamylamine model can be used for proof-of-pharmacology of nicotinic acetylcholine receptor agonists remains to be established. © 2017 The British Pharmacological Society.

Dynamic evolution of the GnRH receptor gene family in vertebrates.

PubMed

Williams, Barry L; Akazome, Yasuhisa; Oka, Yoshitaka; Eisthen, Heather L

2014-10-25

Elucidating the mechanisms underlying coevolution of ligands and receptors is an important challenge in molecular evolutionary biology. Peptide hormones and their receptors are excellent models for such efforts, given the relative ease of examining evolutionary changes in genes encoding for both molecules. Most vertebrates possess multiple genes for both the decapeptide gonadotropin releasing hormone (GnRH) and for the GnRH receptor. The evolutionary history of the receptor family, including ancestral copy number and timing of duplications and deletions, has been the subject of controversy. We report here for the first time sequences of three distinct GnRH receptor genes in salamanders (axolotls, Ambystoma mexicanum), which are orthologous to three GnRH receptors from ranid frogs. To understand the origin of these genes within the larger evolutionary context of the gene family, we performed phylogenetic analyses and probabilistic protein homology searches of GnRH receptor genes in vertebrates and their near relatives. Our analyses revealed four points that alter previous views about the evolution of the GnRH receptor gene family. First, the "mammalian" pituitary type GnRH receptor, which is the sole GnRH receptor in humans and previously presumed to be highly derived because it lacks the cytoplasmic C-terminal domain typical of most G-protein coupled receptors, is actually an ancient gene that originated in the common ancestor of jawed vertebrates (Gnathostomata). Second, unlike previous studies, we classify vertebrate GnRH receptors into five subfamilies. Third, the order of subfamily origins is the inverse of previous proposed models. Fourth, the number of GnRH receptor genes has been dynamic in vertebrates and their ancestors, with multiple duplications and losses. Our results provide a novel evolutionary framework for generating hypotheses concerning the functional importance of structural characteristics of vertebrate GnRH receptors. We show that five subfamilies of vertebrate GnRH receptors evolved early in the vertebrate phylogeny, followed by several independent instances of gene loss. Chief among cases of gene loss are humans, best described as degenerate with respect to GnRH receptors because we retain only a single, ancient gene.

Cannabidivarin-rich cannabis extracts are anticonvulsant in mouse and rat via a CB1 receptor-independent mechanism

PubMed Central

Hill, T D M; Cascio, M-G; Romano, B; Duncan, M; Pertwee, R G; Williams, C M; Whalley, B J; Hill, A J

2013-01-01

BACKGROUND AND PURPOSE Epilepsy is the most prevalent neurological disease and is characterized by recurrent seizures. Here, we investigate (i) the anticonvulsant profiles of cannabis-derived botanical drug substances (BDSs) rich in cannabidivarin (CBDV) and containing cannabidiol (CBD) in acute in vivo seizure models and (ii) the binding of CBDV BDSs and their components at cannabinoid CB1 receptors. EXPERIMENTAL APPROACH The anticonvulsant profiles of two CBDV BDSs (50–422 mg·kg−1) were evaluated in three animal models of acute seizure. Purified CBDV and CBD were also evaluated in an isobolographic study to evaluate potential pharmacological interactions. CBDV BDS effects on motor function were also investigated using static beam and grip strength assays. Binding of CBDV BDSs to cannabinoid CB1 receptors was evaluated using displacement binding assays. KEY RESULTS CBDV BDSs exerted significant anticonvulsant effects in the pentylenetetrazole (≥100 mg·kg−1) and audiogenic seizure models (≥87 mg·kg−1), and suppressed pilocarpine-induced convulsions (≥100 mg·kg−1). The isobolographic study revealed that the anticonvulsant effects of purified CBDV and CBD were linearly additive when co-administered. Some motor effects of CBDV BDSs were observed on static beam performance; no effects on grip strength were found. The Δ9-tetrahydrocannabinol and Δ9-tetrahydrocannabivarin content of CBDV BDS accounted for its greater affinity for CB1 cannabinoid receptors than purified CBDV. CONCLUSIONS AND IMPLICATIONS CBDV BDSs exerted significant anticonvulsant effects in three models of seizure that were not mediated by the CB1 cannabinoid receptor and were of comparable efficacy with purified CBDV. These findings strongly support the further clinical development of CBDV BDSs for the treatment of epilepsy. PMID:23902406

Assessing and predicting drug-induced anticholinergic risks: an integrated computational approach.

PubMed

Xu, Dong; Anderson, Heather D; Tao, Aoxiang; Hannah, Katia L; Linnebur, Sunny A; Valuck, Robert J; Culbertson, Vaughn L

2017-11-01

Anticholinergic (AC) adverse drug events (ADEs) are caused by inhibition of muscarinic receptors as a result of designated or off-target drug-receptor interactions. In practice, AC toxicity is assessed primarily based on clinician experience. The goal of this study was to evaluate a novel concept of integrating big pharmacological and healthcare data to assess clinical AC toxicity risks. AC toxicity scores (ATSs) were computed using drug-receptor inhibitions identified through pharmacological data screening. A longitudinal retrospective cohort study using medical claims data was performed to quantify AC clinical risks. ATS was compared with two previously reported toxicity measures. A quantitative structure-activity relationship (QSAR) model was established for rapid assessment and prediction of AC clinical risks. A total of 25 common medications, and 575,228 exposed and unexposed patients were analyzed. Our data indicated that ATS is more consistent with the trend of AC outcomes than other toxicity methods. Incorporating drug pharmacokinetic parameters to ATS yielded a QSAR model with excellent correlation to AC incident rate ( R 2 = 0.83) and predictive performance (cross validation Q 2 = 0.64). Good correlation and predictive performance ( R 2 = 0.68/ Q 2 = 0.29) were also obtained for an M2 receptor-specific QSAR model and tachycardia, an M2 receptor-specific ADE. Albeit using a small medication sample size, our pilot data demonstrated the potential and feasibility of a new computational AC toxicity scoring approach driven by underlying pharmacology and big data analytics. Follow-up work is under way to further develop the ATS scoring approach and clinical toxicity predictive model using a large number of medications and clinical parameters.

Docking and free energy simulations to predict conformational domains involved in hCG-LH receptor interactions using recombinant antibodies.

PubMed

Majumdar, Ritankar; Railkar, Reema; Dighe, Rajan R

2011-11-01

Single chain fragment variables (ScFvs) have been extensively employed in studying the protein-protein interactions. ScFvs derived from phage display libraries have an additional advantage of being generated against a native antigen, circumventing loss of information on conformational epitopes. In the present study, an attempt has been made to elucidate human chorionic gonadotropin (hCG)-luteinizing hormone (LH) receptor interactions by using a neutral and two inhibitory ScFvs against hCG. The objective was to dock a computationally derived model of these ScFvs onto the crystal structure of hCG and understand the differential roles of the mapped epitopes in hCG-LH receptor interactions. An anti-hCG ScFv, whose epitope was mapped previously using biochemical tools, served as the positive control for assessing the quality of docking analysis. To evaluate the role of specific side chains at the hCG-ScFv interface, binding free energy as well as residue interaction energies of complexes in solution were calculated using molecular mechanics Poisson-Boltzmann/surface area method after performing the molecular dynamic simulations on the selected hCG-ScFv models and validated using biochemical and SPR analysis. The robustness of these calculations was demonstrated by comparing the theoretically determined binding energies with the experimentally obtained kinetic parameters for hCG-ScFv complexes. Superimposition of hCG-ScFv model onto a model of hCG complexed with the 51-266 residues of LH receptor revealed importance of the residues previously thought to be unimportant for hormone binding and response. This analysis provides an alternate tool for understanding the structure-function analysis of ligand-receptor interactions. Copyright © 2011 Wiley-Liss, Inc.

A Novel Approach for Determining Source–Receptor Relationships in Model Simulations: A Case Study of Black Carbon Transport in Northern Hemisphere Winter

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Po-Lun; Gattiker, J. R.; Liu, Xiaohong

2013-06-27

A Gaussian process (GP) emulator is applied to quantify the contribution of local and remote emissions of black carbon (BC) on the BC concentrations in different regions using a Latin Hypercube sampling strategy for emission perturbations in the offline version of the Community Atmosphere Model Version 5.1 (CAM5) simulations. The source-receptor relationships are computed based on simulations constrained by a standard free-running CAM5 simulation and the ERA-Interim reanalysis product. The analysis demonstrates that the emulator is capable of retrieving the source-receptor relationships based on a small number of CAM5 simulations. Most regions are found susceptible to their local emissions. Themore » emulator also finds that the source-receptor relationships retrieved from the model-driven and the reanalysis-driven simulations are very similar, suggesting that the simulated circulation in CAM5 resembles the assimilated meteorology in ERA-Interim. The robustness of the results provides confidence for applying the emulator to detect dose-response signals in the climate system.« less

Progesterone Signaling Inhibits Cervical Carcinogenesis in Mice

PubMed Central

Yoo, Young A; Son, Jieun; Mehta, Fabiola F.; DeMayo, Francesco J.; Lydon, John P.; Chung, Sang-Hyuk

2014-01-01

Human papillomavirus is the main cause of cervical cancer, yet other nonviral cofactors are also required for the disease. The uterine cervix is a hormone-responsive tissue, and female hormones have been implicated in cervical carcinogenesis. A transgenic mouse model expressing human papillomavirus oncogenes E6 and/or E7 has proven useful to study a mechanism of hormone actions in the context of this common malignancy. Estrogen and estrogen receptor α are required for the development of cervical cancer in this mouse model. Estrogen receptor α is known to up-regulate expression of the progesterone receptor, which, on activation by its ligands, either promotes or inhibits carcinogenesis, depending on the tissue context. Here, we report that progesterone receptor inhibits cervical and vaginal epithelial cell proliferation in a ligand-dependent manner. We also report that synthetic progestin medroxyprogesterone acetate promotes regression of cancers and precancerous lesions in the female lower reproductive tracts (ie, cervix and vagina) in the human papillomavirus transgenic mouse model. Our results provide the first experimental evidence that supports the hypothesis that progesterone signaling is inhibitory for cervical carcinogenesis in vivo. PMID:24012679

A Kinetic Model for Calcium Dynamics in RAW 264.7 Cells: 2. Knockdown Response and Long-Term Response

PubMed Central

Maurya, Mano Ram; Subramaniam, Shankar

2007-01-01

This article addresses how quantitative models such as the one proposed in the companion article can be used to study cellular network perturbations such as knockdowns and pharmacological perturbations in a predictive manner. Using the kinetic model for cytosolic calcium dynamics in RAW 264.7 cells developed in the companion article, the calcium response to complement 5a (C5a) for the knockdown of seven proteins (C5a receptor; G-β-2; G-α,i-2,3; regulator of G-protein signaling-10; G-protein coupled receptor kinase-2; phospholipase C β-3; arrestin) is predicted and validated against the data from the Alliance for Cellular Signaling. The knockdown responses provide insights into how altered expressions of important proteins in disease states result in intermediate measurable phenotypes. Long-term response and long-term dose response have also been predicted, providing insights into how the receptor desensitization, internalization, and recycle result in tolerance. Sensitivity analysis of long-term response shows that the mechanisms and parameters in the receptor recycle path are important for long-term calcium dynamics. PMID:17483189

D-cycloserine in Schizophrenia: New Strategies for Improving Clinical Outcomes by Enhancing Plasticity

PubMed Central

Goff, Donald C.

2017-01-01

Background Dysregulation of N-methyl D-aspartate (NMDA) receptor signaling is strongly implicated in schizophrenia. Based on the ketamine model of NMDA receptor hypoactivity, therapeutic approaches designed to maintain a sustained increase in agonist activity at the glycine site of the NMDA receptor have produced promising, although inconsistent, efficacy for negative symptoms. Methods A review of the published literature on D-cycloserine (DCS) pharmacology in animal models and in clinical studies was performed. Findings relevant to DCS effects on memory and plasticity and their potential clinical application to schizophrenia were summarized. Results Studies in animals and clinical trials in patients with anxiety disorders have demonstrated that single or intermittent dosing with DCS enhances memory consolidation. Preliminary trials in patients with schizophrenia suggest that intermittent dosing with DCS may produce persistent improvement of negative symptoms and enhance learning when combined with cognitive behavioral therapy for delusions or with cognitive remediation. The pharmacology of DCS is complex, since it acts as a “super agonist” at NMDA receptors containing GluN2C subunits and, under certain conditions, it may act as an antagonist at NMDA receptors containing GluN2B subunits. Conclusions There are preliminary findings that support a role for D-cycloserine in schizophrenia as a strategy to enhance neuroplasticity and memory. However, additional studies with DCS are needed to confirm these findings. In addition, clinical trials with positive and negative allosteric modulators with greater specificity for NMDA receptor subtypes are needed to identify the optimal strategy for enhancing neuroplasticity in schizophrenia. PMID:26915421

Characterization of SV-40 Tag rats as a model to study prostate cancer

PubMed Central

2009-01-01

Background Prostate cancer is the second most frequently diagnosed cancer in men. Animal models that closely mimic clinical disease in humans are invaluable tools in the fight against prostate cancer. Recently, a Simian Virus-40 T-antigen (SV-40 Tag) targeted probasin promoter rat model was developed. This model, however, has not been extensively characterized; hence we have investigated the ontogeny of prostate cancer and determined the role of sex steroid receptor and insulin-like growth factor-1 (IGF-1) signaling proteins in the novel SV-40 Tag rat. Methods The SV-40 Tag rat was histopathologically characterized for time to tumor development, incidence and multiplicity and in the ventral, dorsal, lateral and anterior lobes of the prostate. Immunoassay techniques were employed to measure cell proliferation, apoptosis, and sex steroid receptor and growth factor signaling-related proteins. Steroid hormone concentrations were measured via coated well enzyme linked immunosorbent assay (ELISA) kits. Results Prostatic intraepithelial neoplasia (PIN) and well-differentiated prostate cancer developed as early as 2 and 10 weeks of age, respectively in the ventral prostate (VP) followed by in the dorsolateral (DLP). At 8 weeks of age, testosterone and dihydrotestosterone (DHT) concentrations in SV-40 Tag rats were increased when compared to non-transgenic rats. High cell proliferation and apoptotic indices were found in VP and DLP of transgenic rats. Furthermore, we observed increased protein expression of androgen receptor, IGF-1, IGF-1 receptor, and extracellular signal-regulated kinases in the prostates of SV-40 Tag rats. Conclusion The rapid development of PIN and prostate cancer in conjunction with the large prostate size makes the SV-40 Tag rat a useful model for studying prostate cancer. This study provides evidence of the role of sex steroid and growth factor proteins in prostate cancer development and defines appropriate windows of opportunity for preclinical trials and aids in the rational design of chemoprevention, intervention, regression, and therapeutic studies using prostate cancer rodent models. PMID:19171036

Characterization of air manganese exposure estimates for residents in two Ohio towns

PubMed Central

Colledge, Michelle A.; Julian, Jaime R.; Gocheva, Vihra V.; Beseler, Cheryl L.; Roels, Harry A.; Lobdell, Danelle T.; Bowler, Rosemarie M.

2016-01-01

This study was conducted to derive receptor-specific outdoor exposure concentrations of total suspended particulate (TSP) and respirable (dae ≤ 10 μm) air manganese (air-Mn) for East Liverpool and Marietta (Ohio) in the absence of facility emissions data, but where long-term air measurements were available. Our “site-surface area emissions method” used U.S. Environmental Protection Agency’s (EPA) AERMOD (AMS/EPA Regulatory Model) dispersion model and air measurement data to estimate concentrations for residential receptor sites in the two communities. Modeled concentrations were used to create ratios between receptor points and calibrated using measured data from local air monitoring stations. Estimated outdoor air-Mn concentrations were derived for individual study subjects in both towns. The mean estimated long-term air-Mn exposure levels for total suspended particulate were 0.35 μg/m3 (geometric mean [GM]) and 0.88 μg/m3 (arithmetic mean [AM]) in East Liverpool (range: 0.014–6.32 μg/m3) and 0.17 μg/m3 (GM) and 0.21 μg/m3 (AM) in Marietta (range: 0.03–1.61 μg/m3). Modeled results compared well with averaged ambient air measurements from local air monitoring stations. Exposure to respirable Mn particulate matter (PM10; PM <10 μm) was higher in Marietta residents. Implications Few available studies evaluate long-term health outcomes from inhalational manganese (Mn) exposure in residential populations, due in part to challenges in measuring individual exposures. Local long-term air measurements provide the means to calibrate models used in estimating long-term exposures. Furthermore, this combination of modeling and ambient air sampling can be used to derive receptor-specific exposure estimates even in the absence of source emissions data for use in human health outcome studies. PMID:26211636

Functional reconstitution of Drosophila melanogaster NMJ glutamate receptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Tae Hee; Dharkar, Poorva; Mayer, Mark L.

The Drosophila larval neuromuscular junction (NMJ), at which glutamate acts as the excitatory neurotransmitter, is a widely used model for genetic analysis of synapse function and development. Despite decades of study, the inability to reconstitute NMJ glutamate receptor function using heterologous expression systems has complicated the analysis of receptor function, such that it is difficult to resolve the molecular basis for compound phenotypes observed in mutant flies. In this paper, we find that Drosophila Neto functions as an essential component required for the function of NMJ glutamate receptors, permitting analysis of glutamate receptor responses in Xenopus oocytes. Finally, in combinationmore » with a crystallographic analysis of the GluRIIB ligand binding domain, we use this system to characterize the subunit dependence of assembly, channel block, and ligand selectivity for Drosophila NMJ glutamate receptors.« less

Functional reconstitution of Drosophila melanogaster NMJ glutamate receptors

DOE PAGES

Han, Tae Hee; Dharkar, Poorva; Mayer, Mark L.; ...

2015-04-27

The Drosophila larval neuromuscular junction (NMJ), at which glutamate acts as the excitatory neurotransmitter, is a widely used model for genetic analysis of synapse function and development. Despite decades of study, the inability to reconstitute NMJ glutamate receptor function using heterologous expression systems has complicated the analysis of receptor function, such that it is difficult to resolve the molecular basis for compound phenotypes observed in mutant flies. In this paper, we find that Drosophila Neto functions as an essential component required for the function of NMJ glutamate receptors, permitting analysis of glutamate receptor responses in Xenopus oocytes. Finally, in combinationmore » with a crystallographic analysis of the GluRIIB ligand binding domain, we use this system to characterize the subunit dependence of assembly, channel block, and ligand selectivity for Drosophila NMJ glutamate receptors.« less

On the intrinsic flexibility of the opioid receptor through multiscale modeling approaches

NASA Astrophysics Data System (ADS)

Vercauteren, Daniel; FosséPré, Mathieu; Leherte, Laurence; Laaksonen, Aatto

Numerous releases of G protein-coupled receptors crystalline structures created the opportunity for computational methods to widely explore their dynamics. Here, we study the biological implication of the intrinsic flexibility properties of opioid receptor OR. First, one performed classical all-atom (AA) Molecular Dynamics (MD) simulations of OR in its apo-form. We highlighted that the various degrees of bendability of the α-helices present important consequences on the plasticity of the binding site. Hence, this latter adopts a wide diversity of shape and volume, explaining why OR interacts with very diverse ligands. Then, one introduces a new strategy for parameterizing purely mechanical but precise coarse-grained (CG) elastic network models (ENMs). The CG ENMs reproduced in a high accurate way the flexibility properties of OR versus the AA simulations. At last, one uses network modularization to design multi-grained (MG) models. They represent a novel type of low resolution models, different in nature versus CG models as being true multi-resolution models, i . e ., each MG grouping a different number of residues. The three parts constitute hierarchical and multiscale approach for tackling the flexibility of OR.

Reading Meta-Television: A New Model for Reader-Response Criticism.

ERIC Educational Resources Information Center

Olson, Scott R.

Many current models of television viewing regard viewers either as passive receptors, active participants, or addled dupes. A study proposed a more flexible model for television viewing research. The study used the television program "St. Elsewhere," an example of "meta-television" (television programming which contains hidden…

Surveying GPCR solubilisation conditions using surface plasmon resonance.

PubMed

Navratilova, Iva Hopkins; Aristotelous, Tonia; Bird, Louise E; Hopkins, Andrew L

2018-06-15

Biophysical screening techniques, such as surface plasmon resonance, enable detailed kinetic analysis of ligands binding to solubilised G-protein coupled receptors. The activity of a receptor solubilised out of the membrane is crucially dependent on the environment in which it is suspended. Finding the right conditions is challenging due to the number of variables to investigate in order to determine the optimum solubilisation buffer for any given receptor. In this study we used surface plasmon resonance technology to screen a variety of solubilisation conditions including buffers and detergents for two model receptors: CXCR4 and CCR5. We tested 950 different combinations of solubilisation conditions for both receptors. The activity of both receptors was monitored by using conformation dependent monoclonal antibodies and the binding of small molecule ligands. Despite both receptors belonging to the chemokine receptor family they show some differences in their preference for solubilisation conditions that provide the highest level of binding for both the conformation dependent antibodies and small molecules. The study described here is focused not only on finding the best solubilisation conditions for each receptor, but also on factors that determine the sensitivity of the assay for each receptor. We also suggest how these data about different buffers and detergents can be used as a guide for selecting solubilisation conditions for other membrane proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

Spring-Loaded Model Revisited: Paramyxovirus Fusion Requires Engagement of a Receptor Binding Protein beyond Initial Triggering of the Fusion Protein▿

PubMed Central

Porotto, Matteo; DeVito, Ilaria; Palmer, Samantha G.; Jurgens, Eric M.; Yee, Jia L.; Yokoyama, Christine C.; Pessi, Antonello; Moscona, Anne

2011-01-01

During paramyxovirus entry into a host cell, receptor engagement by a specialized binding protein triggers conformational changes in the adjacent fusion protein (F), leading to fusion between the viral and cell membranes. According to the existing paradigm of paramyxovirus membrane fusion, the initial activation of F by the receptor binding protein sets off a spring-loaded mechanism whereby the F protein progresses independently through the subsequent steps in the fusion process, ending in membrane merger. For human parainfluenza virus type 3 (HPIV3), the receptor binding protein (hemagglutinin-neuraminidase [HN]) has three functions: receptor binding, receptor cleaving, and activating F. We report that continuous receptor engagement by HN activates F to advance through the series of structural rearrangements required for fusion. In contrast to the prevailing model, the role of HN-receptor engagement in the fusion process is required beyond an initiating step, i.e., it is still required even after the insertion of the fusion peptide into the target cell membrane, enabling F to mediate membrane merger. We also report that for Nipah virus, whose receptor binding protein has no receptor-cleaving activity, the continuous stimulation of the F protein by a receptor-engaged binding protein is key for fusion. We suggest a general model for paramyxovirus fusion activation in which receptor engagement plays an active role in F activation, and the continued engagement of the receptor binding protein is essential to F protein function until the onset of membrane merger. This model has broad implications for the mechanism of paramyxovirus fusion and for strategies to prevent viral entry. PMID:21976650

Spring-loaded model revisited: paramyxovirus fusion requires engagement of a receptor binding protein beyond initial triggering of the fusion protein.

PubMed

Porotto, Matteo; Devito, Ilaria; Palmer, Samantha G; Jurgens, Eric M; Yee, Jia L; Yokoyama, Christine C; Pessi, Antonello; Moscona, Anne

2011-12-01

During paramyxovirus entry into a host cell, receptor engagement by a specialized binding protein triggers conformational changes in the adjacent fusion protein (F), leading to fusion between the viral and cell membranes. According to the existing paradigm of paramyxovirus membrane fusion, the initial activation of F by the receptor binding protein sets off a spring-loaded mechanism whereby the F protein progresses independently through the subsequent steps in the fusion process, ending in membrane merger. For human parainfluenza virus type 3 (HPIV3), the receptor binding protein (hemagglutinin-neuraminidase [HN]) has three functions: receptor binding, receptor cleaving, and activating F. We report that continuous receptor engagement by HN activates F to advance through the series of structural rearrangements required for fusion. In contrast to the prevailing model, the role of HN-receptor engagement in the fusion process is required beyond an initiating step, i.e., it is still required even after the insertion of the fusion peptide into the target cell membrane, enabling F to mediate membrane merger. We also report that for Nipah virus, whose receptor binding protein has no receptor-cleaving activity, the continuous stimulation of the F protein by a receptor-engaged binding protein is key for fusion. We suggest a general model for paramyxovirus fusion activation in which receptor engagement plays an active role in F activation, and the continued engagement of the receptor binding protein is essential to F protein function until the onset of membrane merger. This model has broad implications for the mechanism of paramyxovirus fusion and for strategies to prevent viral entry.

A combined ligand-based and target-based drug design approach for G-protein coupled receptors: application to salvinorin A, a selective kappa opioid receptor agonist

NASA Astrophysics Data System (ADS)

Singh, Nidhi; Chevé, Gwénaël; Ferguson, David M.; McCurdy, Christopher R.

2006-08-01

Combined ligand-based and target-based drug design approaches provide a synergistic advantage over either method individually. Therefore, we set out to develop a powerful virtual screening model to identify novel molecular scaffolds as potential leads for the human KOP (hKOP) receptor employing a combined approach. Utilizing a set of recently reported derivatives of salvinorin A, a structurally unique KOP receptor agonist, a pharmacophore model was developed that consisted of two hydrogen bond acceptor and three hydrophobic features. The model was cross-validated by randomizing the data using the CatScramble technique. Further validation was carried out using a test set that performed well in classifying active and inactive molecules correctly. Simultaneously, a bovine rhodopsin based "agonist-bound" hKOP receptor model was also generated. The model provided more accurate information about the putative binding site of salvinorin A based ligands. Several protein structure-checking programs were used to validate the model. In addition, this model was in agreement with the mutation experiments carried out on KOP receptor. The predictive ability of the model was evaluated by docking a set of known KOP receptor agonists into the active site of this model. The docked scores correlated reasonably well with experimental p K i values. It is hypothesized that the integration of these two independently generated models would enable a swift and reliable identification of new lead compounds that could reduce time and cost of hit finding within the drug discovery and development process, particularly in the case of GPCRs.

Designing of a fluoride selective receptor through molecular orbital engineering

NASA Astrophysics Data System (ADS)

Mishra, Rakesh K.; Kumar, Virendra; Diwan, Uzra; Upadhyay, K. K.; Roy Chowdhury, P. K.

2012-11-01

The stepwise substitution of appropriate groups over the 3-[(2,4-dinitro-phenyl)-hydrazono]-butyric acid ethyl ester (R3) lead receptor R1 which showed selectivity towards fluoride in DMSO. The UV-vis and 1H NMR titration studies revealed the details of the binding between receptor R1 and fluoride. The receptor R1 also recognized fluoride in a toothpaste solution to as low as 50 ppm. The theoretical simulations of recognition event at Density Functional Theory (DFT) level using B3LYP/6-31G** basis set and polarizable continuum model (PCM) approach lead a semi-quantitative match with the experimental results.

Regulation and Functional Implications of Opioid Receptor Splicing in Opioid Pharmacology and HIV Pathogenesis

PubMed Central

Regan, Patrick M.; Langford, T. Dianne; Khalili, Kamel

2015-01-01

Despite the identification and characterization of four opioid receptor subtypes and the genes from which they are encoded, pharmacological data does not conform to the predications of a four opioid receptor model. Instead, current studies of opioid pharmacology suggest the existence of additional receptor subtypes; however, no additional opioid receptor subtype has been identified to date. It is now understood that this discrepancy is due to the generation of multiple isoforms of opioid receptor subtypes. While several mechanisms are utilized to generate these isoforms, the primary mechanism involves alternative splicing of the pre-mRNA transcript. Extensive alternative splicing patterns for opioid receptors have since been identified and discrepancies in opioid pharmacology are now partially attributed to variable expression of these isoforms. Recent studies have been successful in characterizing the localization of these isoforms as well as their specificity in ligand binding; however, the regulation of opioid receptor splicing specificity is poorly characterized. Furthermore, the functional significance of individual receptor isoforms and the extent to which opioid- and/or HIV-mediated changes in the opioid receptor isoform profile contributes to altered opioid pharmacology or the well-known physiological role of opioids in the exacerbation of HIV neurocognitive dysfunction is unknown. As such, the current review details constitutive splicing mechanisms as well as the specific architecture of opioid receptor genes, transcripts, and receptors in order to highlight the current understanding of opioid receptor isoforms, potential mechanisms of their regulation and signaling, and their functional significance in both opioid pharmacology and HIV-associated neuropathology. PMID:26529364

Synthesis, molecular modeling studies and biological evaluation of fluorine substituted analogs of GW 501516.

PubMed

Ciocoiu, Calin C; Ravna, Aina W; Sylte, Ingebrigt; Rustan, Arild C; Hansen, Trond Vidar

2011-12-01

(±)-2-Fluoro-2-(2-methyl-4-(((4-methyl-2-(4-(trifluoromethyl)phenyl)thiazol-5-yl)methyl)thio)phenoxy)acetic acid (2a) has been prepared and subjected to biological testing against all three subtypes of the PPARs. This compound exhibited agonist effects with EC(50) values of 560 and 55 nM against PPARα and PPARδ, respectively, in a luciferase assay. Moreover, compound (±)-2a also exhibited potent ability to induce oleic acid oxidation in a human myotube cell assay with EC(50)=3.7 nM. Compound (±)-2a can be classified as a dual PPARα/δ agonist with a 10-fold higher potency against the PPARδ receptor than against the PPARα receptor. Molecular modeling studies revealed that both enantiomers of 2a bind to the PPARδ receptor with similar binding energies. Copyright © 2011 Elsevier Ltd. All rights reserved.

Evidence for spinal N-methyl-d-aspartate receptor involvement in prolonged chemical nociception in the rat.

PubMed

Haley, Jane E; Dickenson, Anthony H

2016-08-15

We used in vivo electrophysiology and a model of more persistent nociceptive inputs to monitor spinal cord neuronal activity in anaesthetised rats to reveal the pharmacology of enhanced pain signalling. The study showed that all responses were blocked by non-selective antagonism of glutamate receptors but a selective and preferential role of the N-methyl-d-aspartate (NMDA) receptor in the prolonged plastic responses was clearly seen. The work lead to many publications, initially preclinical but increasingly from patient studies, showing the importance of the NMDA receptor in central sensitisation within the spinal cord and how this could relate to persistent pain states. This article is part of a Special Issue entitled SI:50th Anniversary Issue. Copyright © 2016 Elsevier B.V. All rights reserved.

Development and Validation of Decision Forest Model for Estrogen Receptor Binding Prediction of Chemicals Using Large Data Sets.

PubMed

Ng, Hui Wen; Doughty, Stephen W; Luo, Heng; Ye, Hao; Ge, Weigong; Tong, Weida; Hong, Huixiao

2015-12-21

Some chemicals in the environment possess the potential to interact with the endocrine system in the human body. Multiple receptors are involved in the endocrine system; estrogen receptor α (ERα) plays very important roles in endocrine activity and is the most studied receptor. Understanding and predicting estrogenic activity of chemicals facilitates the evaluation of their endocrine activity. Hence, we have developed a decision forest classification model to predict chemical binding to ERα using a large training data set of 3308 chemicals obtained from the U.S. Food and Drug Administration's Estrogenic Activity Database. We tested the model using cross validations and external data sets of 1641 chemicals obtained from the U.S. Environmental Protection Agency's ToxCast project. The model showed good performance in both internal (92% accuracy) and external validations (∼ 70-89% relative balanced accuracies), where the latter involved the validations of the model across different ER pathway-related assays in ToxCast. The important features that contribute to the prediction ability of the model were identified through informative descriptor analysis and were related to current knowledge of ER binding. Prediction confidence analysis revealed that the model had both high prediction confidence and accuracy for most predicted chemicals. The results demonstrated that the model constructed based on the large training data set is more accurate and robust for predicting ER binding of chemicals than the published models that have been developed using much smaller data sets. The model could be useful for the evaluation of ERα-mediated endocrine activity potential of environmental chemicals.

Human tumor xenografts in mouse as a model for evaluating therapeutic efficacy of monoclonal antibodies or antibody-drug conjugate targeting receptor tyrosine kinases.

PubMed

Feng, Liang; Wang, Wei; Yao, Hang-Ping; Zhou, Jianwei; Zhang, Ruiwen; Wang, Ming-Hai

2015-01-01

Targeting receptor tyrosine kinases by therapeutic monoclonal antibodies and antibody-drug conjugates has met with tremendous success in clinical oncology. Currently, numerous therapeutic monoclonal antibodies are under preclinical development. The potential for moving candidate antibodies into clinical trials relies heavily on therapeutic efficacy validated by human tumor xenografts in mice. Here we describe methods used to determine therapeutic efficacy of monoclonal antibodies or antibody-drug conjugates specific to human receptor tyrosine kinase using human tumor xenografts in mice as the model. The end point of the study is to determine whether treatment of tumor-bearing mice with a monoclonal antibody or antibody-drug conjugates results in significant delay of tumor growth.

The antimalarial drugs quinine, chloroquine and mefloquine are antagonists at 5-HT3 receptors

PubMed Central

Thompson, A J; Lochner, M; Lummis, S C R

2007-01-01

Background and Purpose: The antimalarial compounds quinine, chloroquine and mefloquine affect the electrophysiological properties of Cys-loop receptors and have structural similarities to 5-HT3 receptor antagonists. They may therefore act at 5-HT3 receptors. Experimental Approach: The effects of quinine, chloroquine and mefloquine on electrophysiological and ligand binding properties of 5-HT3A receptors expressed in HEK 293 cells and Xenopus oocytes were examined. The compounds were also docked into models of the binding site. Key Results: 5-HT3 responses were blocked with IC 50 values of 13.4 μM, 11.8 μM and 9.36 μM for quinine, chloroquine and mefloquine. Schild plots indicated quinine and chloroquine behaved competitively with pA 2 values of 4.92 (K B=12.0 μM) and 4.97 (K B=16.4 μM). Mefloquine displayed weakly voltage-dependent, non-competitive inhibition consistent with channel block. On and off rates for quinine and chloroquine indicated a simple bimolecular reaction scheme. Quinine, chloroquine and mefloquine displaced [3H]granisetron with K i values of 15.0, 24.2 and 35.7 μ M. Docking of quinine into a homology model of the 5-HT3 receptor binding site located the tertiary ammonium between W183 and Y234, and the quinoline ring towards the membrane, stabilised by a hydrogen bond with E129. For chloroquine, the quinoline ring was positioned between W183 and Y234 and the tertiary ammonium stabilised by interactions with F226. Conclusions and Implications: This study shows that quinine and chloroquine competitively inhibit 5-HT3 receptors, while mefloquine inhibits predominantly non-competitively. Both quinine and chloroquine can be docked into a receptor binding site model, consistent with their structural homology to 5-HT3 receptor antagonists. PMID:17502851

Upregulation of the endothelin A (ETA) receptor and its association with neurodegeneration in a rodent model of glaucoma.

PubMed

McGrady, Nolan R; Minton, Alena Z; Stankowska, Dorota L; He, Shaoqing; Jefferies, Hayden B; Krishnamoorthy, Raghu R

2017-03-01

Primary open angle glaucoma is a heterogeneous group of optic neuropathies that results in optic nerve degeneration and a loss of retinal ganglion cells (RGCs) ultimately causing blindness if allowed to progress. Elevation of intraocular pressure (IOP) is the most attributable risk factor for developing glaucoma and lowering of IOP is currently the only available therapy. However, despite lowering IOP, neurodegenerative effects persist in some patients. Hence, it would be beneficial to develop approaches to promote neuroprotection of RGCs in addition to IOP lowering therapies. The endothelin system is a key target for intervention against glaucomatous neurodegeneration. The endothelin family of peptides and receptors, particularly endothelin-1 (ET-1) and endothelin B (ET B ) receptor, has been shown to have neurodegenerative roles in glaucoma. The purpose of this study was to examine changes in endothelin A (ET A ) receptor protein expression in the retinas of adult male Brown Norway rats following IOP elevation by the Morrison's model of ocular hypertension and the impact of ET A receptor overexpression on RGC viability in vitro. IOP elevation was carried out in one eye of Brown Norway rats by injection of hypertonic saline through episcleral veins. After 2 weeks of IOP elevation, immunohistochemical analysis of retinal sections from rat eyes showed an increasing trend in immunostaining for ET A receptors in multiple retinal layers including the inner plexiform layer, ganglion cell layer and outer plexiform layer. Following 4 weeks of IOP elevation, a significant increase in immunostaining for ET A receptor expression was found in the retina, primarily in the inner plexiform layer and ganglion cells. A modest increase in staining for ET A receptors was also found in the outer plexiform layer in the retina of rats with IOP elevation. Cell culture studies showed that overexpression of ET A receptors in 661W cells as well as primary RGCs decreases cell viability, compared to empty vector transfected cells. Adeno-associated virus mediated overexpression of the ET A receptor produced an increase in the ET B receptor in primary RGCs. Elevated IOP results in an appreciable change in ET A receptor expression in the retina. Overexpression of the ET A receptor results in an overall decrease in cell viability, accompanied by an increase in ET B receptor levels, suggesting the involvement of both ET A and ET B receptors in mediating cell death. These findings raise possibilities for the development of ET A /ET B dual receptor antagonists as neuroprotective treatments for glaucomatous neuropathy.

CoMFA analyses of C-2 position salvinorin A analogs at the kappa-opioid receptor provides insights into epimer selectivity.

PubMed

McGovern, Donna L; Mosier, Philip D; Roth, Bryan L; Westkaemper, Richard B

2010-04-01

The highly potent and kappa-opioid (KOP) receptor-selective hallucinogen Salvinorin A and selected analogs have been analyzed using the 3D quantitative structure-affinity relationship technique Comparative Molecular Field Analysis (CoMFA) in an effort to derive a statistically significant and predictive model of salvinorin affinity at the KOP receptor and to provide additional statistical support for the validity of previously proposed structure-based interaction models. Two CoMFA models of Salvinorin A analogs substituted at the C-2 position are presented. Separate models were developed based on the radioligand used in the kappa-opioid binding assay, [(3)H]diprenorphine or [(125)I]6 beta-iodo-3,14-dihydroxy-17-cyclopropylmethyl-4,5 alpha-epoxymorphinan ([(125)I]IOXY). For each dataset, three methods of alignment were employed: a receptor-docked alignment derived from the structure-based docking algorithm GOLD, another from the ligand-based alignment algorithm FlexS, and a rigid realignment of the poses from the receptor-docked alignment. The receptor-docked alignment produced statistically superior results compared to either the FlexS alignment or the realignment in both datasets. The [(125)I]IOXY set (Model 1) and [(3)H]diprenorphine set (Model 2) gave q(2) values of 0.592 and 0.620, respectively, using the receptor-docked alignment, and both models produced similar CoMFA contour maps that reflected the stereoelectronic features of the receptor model from which they were derived. Each model gave significantly predictive CoMFA statistics (Model 1 PSET r(2)=0.833; Model 2 PSET r(2)=0.813). Based on the CoMFA contour maps, a binding mode was proposed for amine-containing Salvinorin A analogs that provides a rationale for the observation that the beta-epimers (R-configuration) of protonated amines at the C-2 position have a higher affinity than the corresponding alpha-epimers (S-configuration). (c) 2010. Published by Elsevier Inc.

Glial cells isolated from dorsal root ganglia express prostaglandin E(2) (EP(4)) and prostacyclin (IP) receptors.

PubMed

Ng, Kai Yu; Wong, Yung Hou; Wise, Helen

2011-07-01

Isolated cells from adult rat dorsal root ganglia (DRG) are frequently used as a model system to study responses of primary sensory neurons to nociceptor sensitizing agents such as prostaglandin E(2) and prostacyclin, which are presumed to act only on the neurons in typical mixed cell cultures. In the present study, we evaluated the expression of prostaglandin E(2) (EP(4)) and prostacyclin (IP) receptors in cultures of mixed DRG cells and in purified DRG glia. We show here that EP(4) and IP receptor agonists stimulated adenylyl cyclase activity in both mixed DRG cells and in purified DRG glia, and that these responses were specifically inhibited by EP(4) and IP receptor antagonists, respectively. The presence of EP(4) and IP receptors in DRG glia was further confirmed by the expression of EP(4) and IP receptor immunoreactivity and mRNA. With the increasing awareness of neuron-glial interactions within intact DRG and the use of isolated DRG cells in the study of mechanisms underlying nociception, it will be essential to consider the role played by EP(4) and IP receptor-expressing glial cells when evaluating prostanoid-induced sensitization of DRG neurons. Copyright © 2011 Elsevier B.V. All rights reserved.

A combined computational and structural model of the full-length human prolactin receptor

PubMed Central

Bugge, Katrine; Papaleo, Elena; Haxholm, Gitte W.; Hopper, Jonathan T. S.; Robinson, Carol V.; Olsen, Johan G.; Lindorff-Larsen, Kresten; Kragelund, Birthe B.

2016-01-01

The prolactin receptor is an archetype member of the class I cytokine receptor family, comprising receptors with fundamental functions in biology as well as key drug targets. Structurally, each of these receptors represent an intriguing diversity, providing an exceptionally challenging target for structural biology. Here, we access the molecular architecture of the monomeric human prolactin receptor by combining experimental and computational efforts. We solve the NMR structure of its transmembrane domain in micelles and collect structural data on overlapping fragments of the receptor with small-angle X-ray scattering, native mass spectrometry and NMR spectroscopy. Along with previously published data, these are integrated by molecular modelling to generate a full receptor structure. The result provides the first full view of a class I cytokine receptor, exemplifying the architecture of more than 40 different receptor chains, and reveals that the extracellular domain is merely the tip of a molecular iceberg. PMID:27174498

Mechanisms of inverse agonist action at D2 dopamine receptors

PubMed Central

Roberts, David J; Strange, Philip G

2005-01-01

Mechanisms of inverse agonist action at the D2(short) dopamine receptor have been examined. Discrimination of G-protein-coupled and -uncoupled forms of the receptor by inverse agonists was examined in competition ligand-binding studies versus the agonist [3H]NPA at a concentration labelling both G-protein-coupled and -uncoupled receptors. Competition of inverse agonists versus [3H]NPA gave data that were fitted best by a two-binding site model in the absence of GTP but by a one-binding site model in the presence of GTP. Ki values were derived from the competition data for binding of the inverse agonists to G-protein-uncoupled and -coupled receptors. Kcoupled and Kuncoupled were statistically different for the set of compounds tested (ANOVA) but the individual values were different in a post hoc test only for (+)-butaclamol. These observations were supported by simulations of these competition experiments according to the extended ternary complex model. Inverse agonist efficacy of the ligands was assessed from their ability to reduce agonist-independent [35S]GTPγS binding to varying degrees in concentration–response curves. Inverse agonism by (+)-butaclamol and spiperone occurred at higher potency when GDP was added to assays, whereas the potency of (−)-sulpiride was unaffected. These data show that some inverse agonists ((+)-butaclamol, spiperone) achieve inverse agonism by stabilising the uncoupled form of the receptor at the expense of the coupled form. For other compounds tested, we were unable to define the mechanism. PMID:15735658

Neuropeptide S attenuates neuropathological, neurochemical and behavioral changes induced by the NMDA receptor antagonist MK-801

PubMed Central

Okamura, Naoe; Reinscheid, Rainer K.; Ohgake, Shintaro; Iyo, Masaomi; Hashimoto, Kenji

2009-01-01

Neuropeptide S (NPS) and its cognate receptor were reported to mediate anxiolytic-like and arousal effects. NPS receptors are predominantly expressed in the brain, especially in limbic structures, including amygdala, olfactory nucleus, subiculum and retrosplenial cortex. In contrast, the NPS precursor is expressed in only a few brainstem nuclei where it is co-expressed with various excitatory transmitters, including glutamate. The current study investigates interactions of the NPS system with glutamatergic neurotransmission. It has been suggested that dysfunctions in glutamatergic neurotransmission via N-methyl-D-aspartate (NMDA) receptors might be involved in the pathophysiology of schizophrenia since NMDA receptor antagonists, such as MK-801, have been shown to induce psychotic-like behavior in humans and animal models. Also, MK-801 is known to produce histological changes such as cytoplasmic vacuoles in retrosplenial cortex neurons where NPS receptors are highly expressed. In this study we show that NPS is able to alleviate neuropathological, neurochemical and behavioral changes produced by NMDA receptor antagonists. NPS treatment attenuated MK-801-induced vacuolization in the rat retrosplenial cortex in a dose dependent manner that can be blocked by an NPS receptor-selective antagonist. NPS also suppressed MK-801-induced increases of extracellular acetylcholine levels in the retrosplenial cortex. In the prepulse inhibition (PPI) assay, animals pretreated with NPS recovered significantly from MK-801-induced disruption of PPI. Our study suggests that NPS may have protective effects against the neurotoxic and behavioral changes produced by NMDA receptor antagonists and that NPS receptor agonists may elicit antipsychotic effects. PMID:19576911

Stochastic lattice model of synaptic membrane protein domains.

PubMed

Li, Yiwei; Kahraman, Osman; Haselwandter, Christoph A

2017-05-01

Neurotransmitter receptor molecules, concentrated in synaptic membrane domains along with scaffolds and other kinds of proteins, are crucial for signal transmission across chemical synapses. In common with other membrane protein domains, synaptic domains are characterized by low protein copy numbers and protein crowding, with rapid stochastic turnover of individual molecules. We study here in detail a stochastic lattice model of the receptor-scaffold reaction-diffusion dynamics at synaptic domains that was found previously to capture, at the mean-field level, the self-assembly, stability, and characteristic size of synaptic domains observed in experiments. We show that our stochastic lattice model yields quantitative agreement with mean-field models of nonlinear diffusion in crowded membranes. Through a combination of analytic and numerical solutions of the master equation governing the reaction dynamics at synaptic domains, together with kinetic Monte Carlo simulations, we find substantial discrepancies between mean-field and stochastic models for the reaction dynamics at synaptic domains. Based on the reaction and diffusion properties of synaptic receptors and scaffolds suggested by previous experiments and mean-field calculations, we show that the stochastic reaction-diffusion dynamics of synaptic receptors and scaffolds provide a simple physical mechanism for collective fluctuations in synaptic domains, the molecular turnover observed at synaptic domains, key features of the observed single-molecule trajectories, and spatial heterogeneity in the effective rates at which receptors and scaffolds are recycled at the cell membrane. Our work sheds light on the physical mechanisms and principles linking the collective properties of membrane protein domains to the stochastic dynamics that rule their molecular components.

Polygonatum sibiricum rhizome promotes sleep by regulating non-rapid eye movement and GABAergic/serotonergic receptors in rodent models.

PubMed

Jo, Kyungae; Suh, Hyung Joo; Choi, Hyeon-Son

2018-05-29

The aim of this study is to investigate the sleep-promoting effect of a water extract of the Polygonatum sibiricum rhizome (PSE) in rodent models. PSE contained oleamide (0.10 mg/g extract) and glyceryl monolinoleate (0.17 mg/g extract), which are recognized as sleep-promoting substances. In pentobarbital-induced sleep model at hypnotic level, PSE (160 mg/kg) administration significantly decreased sleep latency time by 29% (2.7 min) and increased sleep duration time by 70% (68.4 min) compared with the normal control (3.8 min and 40.7 min, respectively). In the electroencephalography (EEG) analysis of rats, PSE-mediated sleep promotion accompanied the change of sleep architecture including increase of non-rapid eye movement (NREM) and decrease of REM. This sleep promoting effect was more obvious in caffeine-induced awakening model; total sleep time was increased by 40% along with increased NREM by PSE treatment at 160 mg/kg. In addition, PSE significantly increased the protein and mRNA levels of GABA A -R2 and 5-HT1A receptor, the major sleep-related neurotransmitter receptors. Furthermore, glyceryl monolinoleate and oleamide effectively bound to GABA A receptor in a competitive binding assay. These results indicate that PSE-mediated sleep-promoting effect is associated with the extension of NREM and upregulation of GABA A -R2 and 5-HT1A, and is mediated by binding to the GABA A receptor in vertebrate models. Copyright © 2018. Published by Elsevier Masson SAS.

Rosetta FlexPepDock ab-initio: simultaneous folding, docking and refinement of peptides onto their receptors.

PubMed

Raveh, Barak; London, Nir; Zimmerman, Lior; Schueler-Furman, Ora

2011-04-29

Flexible peptides that fold upon binding to another protein molecule mediate a large number of regulatory interactions in the living cell and may provide highly specific recognition modules. We present Rosetta FlexPepDock ab-initio, a protocol for simultaneous docking and de-novo folding of peptides, starting from an approximate specification of the peptide binding site. Using the Rosetta fragments library and a coarse-grained structural representation of the peptide and the receptor, FlexPepDock ab-initio samples efficiently and simultaneously the space of possible peptide backbone conformations and rigid-body orientations over the receptor surface of a given binding site. The subsequent all-atom refinement of the coarse-grained models includes full side-chain modeling of both the receptor and the peptide, resulting in high-resolution models in which key side-chain interactions are recapitulated. The protocol was applied to a benchmark in which peptides were modeled over receptors in either their bound backbone conformations or in their free, unbound form. Near-native peptide conformations were identified in 18/26 of the bound cases and 7/14 of the unbound cases. The protocol performs well on peptides from various classes of secondary structures, including coiled peptides with unusual turns and kinks. The results presented here significantly extend the scope of state-of-the-art methods for high-resolution peptide modeling, which can now be applied to a wide variety of peptide-protein interactions where no prior information about the peptide backbone conformation is available, enabling detailed structure-based studies and manipulation of those interactions. © 2011 Raveh et al.

Rosetta FlexPepDock ab-initio: Simultaneous Folding, Docking and Refinement of Peptides onto Their Receptors

PubMed Central

Raveh, Barak; London, Nir; Zimmerman, Lior; Schueler-Furman, Ora

2011-01-01

Flexible peptides that fold upon binding to another protein molecule mediate a large number of regulatory interactions in the living cell and may provide highly specific recognition modules. We present Rosetta FlexPepDock ab-initio, a protocol for simultaneous docking and de-novo folding of peptides, starting from an approximate specification of the peptide binding site. Using the Rosetta fragments library and a coarse-grained structural representation of the peptide and the receptor, FlexPepDock ab-initio samples efficiently and simultaneously the space of possible peptide backbone conformations and rigid-body orientations over the receptor surface of a given binding site. The subsequent all-atom refinement of the coarse-grained models includes full side-chain modeling of both the receptor and the peptide, resulting in high-resolution models in which key side-chain interactions are recapitulated. The protocol was applied to a benchmark in which peptides were modeled over receptors in either their bound backbone conformations or in their free, unbound form. Near-native peptide conformations were identified in 18/26 of the bound cases and 7/14 of the unbound cases. The protocol performs well on peptides from various classes of secondary structures, including coiled peptides with unusual turns and kinks. The results presented here significantly extend the scope of state-of-the-art methods for high-resolution peptide modeling, which can now be applied to a wide variety of peptide-protein interactions where no prior information about the peptide backbone conformation is available, enabling detailed structure-based studies and manipulation of those interactions. PMID:21572516

Different amounts of ejaculatory activity, a natural rewarding behavior, induce differential mu and delta opioid receptor internalization in the rat's ventral tegmental area.

PubMed

Garduño-Gutiérrez, René; León-Olea, Martha; Rodríguez-Manzo, Gabriela

2013-12-06

Opioid receptors internalize upon specific agonist stimulation. The in vivo significance of receptor internalization is not well established, partly due to the limited in vivo models used to study this phenomenon. Ejaculation promotes endogenous opioid release which activates opioid receptors at the brain, including the mesolimbic system and medial preoptic area. The objective of the present work was to analyze if there was a correlation between the degree of in vivo mu (MOR) and delta opioid receptor (DOR) internalization in the ventral tegmental area and the execution of different amounts of ejaculatory behavior of male rats. To this aim, we analyzed the brains of rats that ejaculated once or six successive times and of sexually exhausted rats with an established sexual inhibition, using immunofluorescence and confocal microscopy. Results showed that MOR and DOR internalization increased as a consequence of ejaculation. There was a relationship between the amount of sexual activity executed and the degree of internalization for MOR, but not for DOR. MOR internalization was larger in rats that ejaculated repeatedly than in animals ejaculating only once. Significant DOR internalization was found only in animals ejaculating once. Changes in MOR, DOR and beta arrestin2 detection, associated to sexual activity, were also found. It is suggested that copulation to satiety might be useful as a model system to study the biological significance of receptor internalization. © 2013 Published by Elsevier B.V.

Centrally mediated antinociceptive effects of cannabinoid receptor ligands in rat models of nociception

PubMed Central

Hama, Aldric; Sagen, Jacqueline

2011-01-01

The endogenous nonapeptide hemopressin (HE) demonstrates potent block of the cannabinoid subtype-1 (CB1) receptor in vitro and robust antinociception in vivo. The current study evaluated the effects of centrally administered HE in mechanistically distinct pre-clinical rat models of pain—the hot plate test and the hind paw formalin test. The non-subtype selective CB receptor agonist WIN 55,212-2 was tested concurrently as a positive control. In the hot plate test, neither intrathecal (i.t.) HE nor WIN 55,212-2 significantly altered the latency to respond to noxious heat. By contrast, i.t. HE and WIN 55,212-2 significantly reduced pain-related behaviors in the formalin test. Possible HE functionality as a CB1 receptor antagonist at the spinal level was evaluated in the formalin test. Intrathecal pretreatment with HE did not attenuate the antinociceptive effect of i.t. WIN 55,212-2. However, pretreatment with the CB1 receptor antagonist rimonabant did; i.t. rimonabant pretreatment was not antinociceptive. Potential supraspinal antinociceptive activity of HE was also evaluated. Whereas intracerebroventricular (i.c.v.) injection of WIN 55,212-2 reduced pain-related behaviors in the formalin test, interestingly, i.c.v. HE increased behaviors. In the current study, an antinociceptive effect with the CB receptor ligand HE was obtained under the specific condition of tissue injury and not in the uninjured state. Thus, HE could be a useful analgesic peptide with a novel spinal mechanism of action. PMID:21958947

Pharmacological identification of cholinergic receptor subtypes on Drosophila melanogaster larval heart.

PubMed

Malloy, Cole A; Ritter, Kyle; Robinson, Jonathan; English, Connor; Cooper, Robin L

2016-01-01

The Drosophila melanogaster heart is a popular model in which to study cardiac physiology and development. Progress has been made in understanding the role of endogenous compounds in regulating cardiac function in this model. It is well characterized that common neurotransmitters act on many peripheral and non-neuronal tissues as they flow through the hemolymph of insects. Many of these neuromodulators, including acetylcholine (ACh), have been shown to act directly on the D. melanogaster larval heart. ACh is a primary neurotransmitter in the central nervous system (CNS) of vertebrates and at the neuromuscular junctions on skeletal and cardiac tissue. In insects, ACh is the primary excitatory neurotransmitter of sensory neurons and is also prominent in the CNS. A full understanding regarding the regulation of the Drosophila cardiac physiology by the cholinergic system remains poorly understood. Here we use semi-intact D. melanogaster larvae to study the pharmacological profile of cholinergic receptor subtypes, nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors (mAChRs), in modulating heart rate (HR). Cholinergic receptor agonists, nicotine and muscarine both increase HR, while nAChR agonist clothianidin exhibits no significant effect when exposed to an open preparation at concentrations as low as 100 nM. In addition, both nAChR and mAChR antagonists increase HR as well but also display capabilities of blocking agonist actions. These results provide evidence that both of these receptor subtypes display functional significance in regulating the larval heart's pacemaker activity.

Adenosine A2A Receptor Blockade or Deletion Diminishes Fibrocyte Accumulation in the Skin in a Murine Model of Scleroderma, Bleomycin-induced Fibrosis

PubMed Central

Katebi, Majid; Fernandez, Patricia; Chan, Edwin S. L.; Cronstein, Bruce N.

2015-01-01

Peripheral blood fibrocytes are a newly identified circulating leukocyte subpopulation that migrates into injured tissue where it may display fibroblast-like properties and participate in wound healing and fibrosis of skin and other organs. Previous studies in our lab demonstrated that A2A receptor-deficient and A2A antagonist-treated mice were protected from developing bleomycin-induced dermal fibrosis, thus the aim of this study was to determine whether the adenosine A2A receptor regulates recruitment of fibrocytes to the dermis in this bleomycin-induced model of dermal fibrosis. Sections of skin from normal mice and bleomycin-treated wild type, A2A knockout and A2A antagonist-treated mice were stained for Procollagen α2 Type I and CD34 and the double stained cells, fibrocytes, were counted in the tissue sections. There were more fibrocytes in the dermis of bleomycin-treated mice than normal mice and the increase was abrogated by deletion or blockade of adenosine A2A receptors. Because fibrocytes play a central role in tissue fibrosis these results suggest that diminished adenosine A2A receptor-mediated recruitment of fibrocytes into tissue may play a role in the pathogenesis of fibrosing diseases of the skin. Moreover, these results provide further evidence that adenosine A2A receptors may represent a new target for the treatment of such fibrosing diseases as scleroderma or nephrogenic fibrosing dermopathy. PMID:18709547

5-Amino-6-chloro-N-[(1-isobutylpiperidin-4-yl)methyl]-2-methylimidazo[1,2-alpha]pyridine-8-carboxamide (CJ-033,466), a novel and selective 5-hydroxytryptamine4 receptor partial agonist: pharmacological profile in vitro and gastroprokinetic effect in conscious dogs.

PubMed

Mikami, Tadayoshi; Ochi, Yasuo; Suzuki, Keiko; Saito, Toshiyuki; Sugie, Yutaka; Sakakibara, Minoru

2008-04-01

5-Hydroxytryptamine (5-HT) receptors and dopamine(2) (D(2)) receptor modulate gastrointestinal motility. Gastroprokinetic agents that act on several 5-HT receptor subtypes and/or D(2) receptors are used clinically. Although the 5-HT(4) receptor is known to mediate the gastroprokinetic effects of these agents, the absence of highly selective 5-HT(4) receptor agonists has made it difficult to confirm the physiological consequences of selective 5-HT(4) receptor stimulation. In this study, we report the in vitro pharmacological profiles and the in vivo gastroprokinetic effects of 5-amino-6-chloro-N-[(1-isobutylpiperidin-4-yl)methyl]-2-methylimidazo[1,2-alpha]pyridine-8-carboxamide (CJ-033,466), a novel, potent, and selective 5-HT(4) partial agonist. Compared with preceding 5-HT(4) agonists such as cisapride, mosapride, and tegaserod, CJ-033,466 had a superior in vitro profile, with nanomolar agonistic activities for the 5-HT(4) receptor and 1000-fold greater selectivity for the 5-HT(4) receptor over other 5-HT and D(2) receptors. In vivo studies in conscious dogs showed that CJ-033,466 dose-dependently stimulated gastric antral motility in both the fasted and postprandial states at the same dose range and that it was 30 times more potent than cisapride. Furthermore, CJ-033,466 accelerated the gastric emptying rate in a gastroparesis dog model at the minimally effective dose established in the gastric motility study. In conclusion, CJ-033,466 is a potent and highly selective 5-HT(4) agonist that stimulates physiologically coordinated gastric motility, and it has no activity on other 5-HT receptor subtypes and D(2) receptors. Therefore, CJ-033,466 could be used to treat gastroparesis, providing better gastroprokinetics and reduced side effects mediated by the other receptors.

All-Atom Structural Models of the Transmembrane Domains of Insulin and Type 1 Insulin-Like Growth Factor Receptors

PubMed Central

Mohammadiarani, Hossein; Vashisth, Harish

2016-01-01

The receptor tyrosine kinase superfamily comprises many cell-surface receptors including the insulin receptor (IR) and type 1 insulin-like growth factor receptor (IGF1R) that are constitutively homodimeric transmembrane glycoproteins. Therefore, these receptors require ligand-triggered domain rearrangements rather than receptor dimerization for activation. Specifically, binding of peptide ligands to receptor ectodomains transduces signals across the transmembrane domains for trans-autophosphorylation in cytoplasmic kinase domains. The molecular details of these processes are poorly understood in part due to the absence of structures of full-length receptors. Using MD simulations and enhanced conformational sampling algorithms, we present all-atom structural models of peptides containing 51 residues from the transmembrane and juxtamembrane regions of IR and IGF1R. In our models, the transmembrane regions of both receptors adopt helical conformations with kinks at Pro961 (IR) and Pro941 (IGF1R), but the C-terminal residues corresponding to the juxtamembrane region of each receptor adopt unfolded and flexible conformations in IR as opposed to a helix in IGF1R. We also observe that the N-terminal residues in IR form a kinked-helix sitting at the membrane–solvent interface, while homologous residues in IGF1R are unfolded and flexible. These conformational differences result in a larger tilt-angle of the membrane-embedded helix in IGF1R in comparison to IR to compensate for interactions with water molecules at the membrane–solvent interfaces. Our metastable/stable states for the transmembrane domain of IR, observed in a lipid bilayer, are consistent with a known NMR structure of this domain determined in detergent micelles, and similar states in IGF1R are consistent with a previously reported model of the dimerized transmembrane domains of IGF1R. Our all-atom structural models suggest potentially unique structural organization of kinase domains in each receptor. PMID:27379020

Angiotensin II AT1 receptor blocker candesartan prevents the fast up-regulation of cerebrocortical benzodiazepine-1 receptors induced by acute inflammatory and restraint stress

PubMed Central

Sánchez-Lemus, Enrique; Honda, Masaru; Saavedra, Juan M.

2012-01-01

Centrally acting Angiotensin II AT1 receptor blockers (ARBs) protect from stress-induced disorders and decrease anxiety in a model of inflammatory stress, the systemic injection of bacterial endotoxin lipopolysaccharide (LPS). In order to better understand the anxiolytic effect of ARBs, we treated rats with LPS (50 µg/kg) with or without three days of pretreatment with the ARB candesartan (1 mg/kg/day), and studied cortical benzodiazepine (BZ) and corticotrophin-releasing factor (CRF) receptors. We compared the cortical BZ and CRF receptors expression pattern induced by LPS with that produced in restraint stress. Inflammation stress produced a generalized increase in cortical BZ1 receptors and reduced mRNA expression of the GABAA receptor γ2 subunit in cingulate cortex; changes were prevented by candesartan pretreatment. Moreover, restraint stress produced similar increases in cortical BZ1 receptor binding, and candesartan prevented these changes. Treatment with candesartan alone increased cortical BZ1 binding, and decreased γ2 subunit mRNA expression in the cingulate cortex. Conversely, we did not find changes in CRF1 receptor expression in any of the cortical areas studied, either after inflammation or restraint stress. Cortical CRF2 receptor binding was undetectable, but CRF2 mRNA expression was decreased by inflammation stress, a change prevented by candesartan. We conclude that stress promotes rapid and widespread changes in cortical BZ1 receptor expression; and that the stress-induced BZ1 receptor expression is under the control of AT1 receptor activity. The results suggest that the anti-anxiety effect of ARBs may be associated with their capacity to regulate stress-induced alterations in cortical BZ1 receptors. PMID:22503782

ATMOSPHERIC MERCURY DEPOSITION TO LAKE MICHIGAN DURING THE LAKE MICHIGAN MASS BALANCE STUDY

EPA Science Inventory

Wet and dry mercury (Hg) deposition were calculated to Lake Michigan using a hybrid receptor modeling framework. The model utilized mercury monitoring data collected during the Lake Michigan Mass Balance Study and the Atmospheric Exchange Over Lakes and Oceans Study together w...

Imidazole derivatives as angiotensin II AT1 receptor blockers: Benchmarks, drug-like calculations and quantitative structure-activity relationships modeling

NASA Astrophysics Data System (ADS)

Alloui, Mebarka; Belaidi, Salah; Othmani, Hasna; Jaidane, Nejm-Eddine; Hochlaf, Majdi

2018-03-01

We performed benchmark studies on the molecular geometry, electron properties and vibrational analysis of imidazole using semi-empirical, density functional theory and post Hartree-Fock methods. These studies validated the use of AM1 for the treatment of larger systems. Then, we treated the structural, physical and chemical relationships for a series of imidazole derivatives acting as angiotensin II AT1 receptor blockers using AM1. QSAR studies were done for these imidazole derivatives using a combination of various physicochemical descriptors. A multiple linear regression procedure was used to design the relationships between molecular descriptor and the activity of imidazole derivatives. Results validate the derived QSAR model.

Allosteric modulation of sigma-1 receptors elicits anti-seizure activities.

PubMed

Guo, Lin; Chen, Yanke; Zhao, Rui; Wang, Guanghui; Friedman, Eitan; Zhang, Ao; Zhen, Xuechu

2015-08-01

Application of orthosteric sigma-1 receptor agonists as anti-seizure drugs has been hindered by questionable efficacy and potential adverse effects. Here, we have investigated the anti-seizure effects of the novel and potent allosteric modulator of sigma-1 receptors, SKF83959 and its derivative SOMCL-668 (3-methyl-phenyl-2,3,4,5-tetrahydro-1H-benzo[d]azepin-7-ol). The anti-seizure effects of SKF83959 were investigated in three mouse models, maximal electroshock seizures, pentylenetetrazole-induced convulsions and kainic acid-induced 'status epilepticus'. Also, in rats, the cortical epileptiform activity induced by topical application of picrotoxin was recorded in electrocorticograms. In rat hippocampal brain slices, effects of the drugs on the high potassium-evoked epileptiform local field potentials were studied. Anti-seizure activities of SOMCL-668, a newly developed sigma-1 receptor selective allosteric modulator, were also investigated. SKF83959 (20, 40 mg·kg(-1) ) exhibited anti -seizure actitity in the three mouse models and reduced the cortical epileptiform activity without alteration of spontaneous motor activity and motor coordination. These effects were blocked by the sigma-1 receptor antagonist BD1047, but not the dopamine D1 receptor antagonist SCH23390. SKF83959 alone did not directly inhibit the epileptiform firing of CA3 neurons induced by high potassium in hippocampal slices, but did potentiate inhibition by the orthosteric sigma-1 receptor agonist SKF10047. Lastly, a selective sigma-1 receptor allosteric modulator SOMCL-668, which does not bind to dopamine receptors, exerted similar anti-seizure activities. SKF83959 and SOMCL-668 displayed anti-seizure activities, indicating that allosteric modulation of sigma-1 receptors may provide a novel approach for discovering new anti-seizure drugs. © 2015 The British Pharmacological Society.

Acetaminophen Metabolite N-Acylphenolamine Induces Analgesia via Transient Receptor Potential Vanilloid 1 Receptors Expressed on the Primary Afferent Terminals of C-fibers in the Spinal Dorsal Horn.

PubMed

Ohashi, Nobuko; Uta, Daisuke; Sasaki, Mika; Ohashi, Masayuki; Kamiya, Yoshinori; Kohno, Tatsuro

2017-08-01

The widely used analgesic acetaminophen is metabolized to N-acylphenolamine, which induces analgesia by acting directly on transient receptor potential vanilloid 1 or cannabinoid 1 receptors in the brain. Although these receptors are also abundant in the spinal cord, no previous studies have reported analgesic effects of acetaminophen or N-acylphenolamine mediated by the spinal cord dorsal horn. We hypothesized that clinical doses of acetaminophen induce analgesia via these spinal mechanisms. We assessed our hypothesis in a rat model using behavioral measures. We also used in vivo and in vitro whole cell patch-clamp recordings of dorsal horn neurons to assess excitatory synaptic transmission. Intravenous acetaminophen decreased peripheral pinch-induced excitatory responses in the dorsal horn (53.1 ± 20.7% of control; n = 10; P < 0.01), while direct application of acetaminophen to the dorsal horn did not reduce these responses. Direct application of N-acylphenolamine decreased the amplitudes of monosynaptic excitatory postsynaptic currents evoked by C-fiber stimulation (control, 462.5 ± 197.5 pA; N-acylphenolamine, 272.5 ± 134.5 pA; n = 10; P = 0.022) but not those evoked by stimulation of Aδ-fibers. These phenomena were mediated by transient receptor potential vanilloid 1 receptors, but not cannabinoid 1 receptors. The analgesic effects of acetaminophen and N-acylphenolamine were stronger in rats experiencing an inflammatory pain model compared to naïve rats. Our results suggest that the acetaminophen metabolite N-acylphenolamine induces analgesia directly via transient receptor potential vanilloid 1 receptors expressed on central terminals of C-fibers in the spinal dorsal horn and leads to conduction block, shunt currents, and desensitization of these fibers.

High occupancy of sigma-1 receptors in the human brain after single oral administration of fluvoxamine: a positron emission tomography study using [11C]SA4503.

PubMed

Ishikawa, Masatomo; Ishiwata, Kiichi; Ishii, Kenji; Kimura, Yuichi; Sakata, Muneyuki; Naganawa, Mika; Oda, Keiichi; Miyatake, Ryousuke; Fujisaki, Mihisa; Shimizu, Eiji; Shirayama, Yukihiko; Iyo, Masaomi; Hashimoto, Kenji

2007-10-15

Sigma-1 receptors might be implicated in the pathophysiology of psychiatric diseases, as well as in the mechanisms of action of some selective serotonin reuptake inhibitors (SSRIs). Among the several SSRIs, fluvoxamine has the highest affinity for sigma-1 receptors (Ki = 36 nM), whereas paroxetine shows low affinity (Ki = 1893 nM). The present study was undertaken to examine whether fluvoxamine binds to sigma-1 receptors in living human brain. A dynamic positron emission tomography (PET) data acquisition using the selective sigma-1 receptor ligand [(11)C]SA4503 was performed with arterial blood sampling to evaluate quantitatively the binding of [(11)C]SA4503 to sigma-1 receptors in 15 healthy male volunteers. Each subject had two PET scans before and after randomly receiving a single dose of either fluvoxamine (50, 100, 150, or 200 mg) or paroxetine (20 mg). The binding potential of [(11)C]SA4503 in 9 regions of the brain was calculated by a 2-tissue 3-compartment model. In addition, we examined the effects of functional polymorphisms of the sigma-1 receptor (SIGMAR1) gene on the binding potential of [(11)C]SA4503. Fluvoxamine bound to sigma-1 receptors in all brain regions in a dose-dependent manner, whereas paroxetine did not bind to sigma-1 receptors. However, there was no association between the SIGMAR1 gene polymorphism GC-241-240TT and binding potential. The study demonstrated that fluvoxamine bound to sigma-1 receptors in living human brain at therapeutic doses. These findings suggest that sigma-1 receptors may play an important role in the mechanism of action of fluvoxamine.

Allosteric modulation model of the mu opioid receptor by herkinorin, a potent not alkaloidal agonist

NASA Astrophysics Data System (ADS)

Marmolejo-Valencia, A. F.; Martínez-Mayorga, K.

2017-05-01

Modulation of opioid receptors is the primary choice for pain management and structural information studies have gained new horizons with the recently available X-ray crystal structures. Herkinorin is one of the most remarkable salvinorin A derivative with high affinity for the mu opioid receptor, moderate selectivity and lack of nitrogen atoms on its structure. Surprisingly, binding models for herkinorin are lacking. In this work, we explore binding models of herkinorin using automated docking, molecular dynamics simulations, free energy calculations and available experimental information. Our herkinorin D-ICM-1 binding model predicted a binding free energy of -11.52 ± 1.14 kcal mol-1 by alchemical free energy estimations, which is close to the experimental values -10.91 ± 0.2 and -10.80 ± 0.05 kcal mol-1 and is in agreement with experimental structural information. Specifically, D-ICM-1 molecular dynamics simulations showed a water-mediated interaction between D-ICM-1 and the amino acid H2976.52, this interaction coincides with the co-crystallized ligands. Another relevant interaction, with N1272.63, allowed to rationalize herkinorin's selectivity to mu over delta opioid receptors. Our suggested binding model for herkinorin is in agreement with this and additional experimental data. The most remarkable observation derived from our D-ICM-1 model is that herkinorin reaches an allosteric sodium ion binding site near N1503.35. Key interactions in that region appear relevant for the lack of β-arrestin recruitment by herkinorin. This interaction is key for downstream signaling pathways involved in the development of side effects, such as tolerance. Future SAR studies and medicinal chemistry efforts will benefit from the structural information presented in this work.

Comprehensive Reactive Receiver Modeling for Diffusive Molecular Communication Systems: Reversible Binding, Molecule Degradation, and Finite Number of Receptors.

PubMed

Ahmadzadeh, Arman; Arjmandi, Hamidreza; Burkovski, Andreas; Schober, Robert

2016-10-01

This paper studies the problem of receiver modeling in molecular communication systems. We consider the diffusive molecular communication channel between a transmitter nano-machine and a receiver nano-machine in a fluid environment. The information molecules released by the transmitter nano-machine into the environment can degrade in the channel via a first-order degradation reaction and those that reach the receiver nano-machine can participate in a reversible bimolecular reaction with receiver receptor proteins. Thereby, we distinguish between two scenarios. In the first scenario, we assume that the entire surface of the receiver is covered by receptor molecules. We derive a closed-form analytical expression for the expected received signal at the receiver, i.e., the expected number of activated receptors on the surface of the receiver. Then, in the second scenario, we consider the case where the number of receptor molecules is finite and the uniformly distributed receptor molecules cover the receiver surface only partially. We show that the expected received signal for this scenario can be accurately approximated by the expected received signal for the first scenario after appropriately modifying the forward reaction rate constant. The accuracy of the derived analytical results is verified by Brownian motion particle-based simulations of the considered environment, where we also show the impact of the effect of receptor occupancy on the derived analytical results.

Receptor dimer stabilization by hierarchical plasma membrane microcompartments regulates cytokine signaling

PubMed Central

You, Changjiang; Marquez-Lago, Tatiana T.; Richter, Christian Paolo; Wilmes, Stephan; Moraga, Ignacio; Garcia, K. Christopher; Leier, André; Piehler, Jacob

2016-01-01

The interaction dynamics of signaling complexes is emerging as a key determinant that regulates the specificity of cellular responses. We present a combined experimental and computational study that quantifies the consequences of plasma membrane microcompartmentalization for the dynamics of type I interferon receptor complexes. By using long-term dual-color quantum dot (QD) tracking, we found that the lifetime of individual ligand-induced receptor heterodimers depends on the integrity of the membrane skeleton (MSK), which also proved important for efficient downstream signaling. By pair correlation tracking and localization microscopy as well as by fast QD tracking, we identified a secondary confinement within ~300-nm-sized zones. A quantitative spatial stochastic diffusion-reaction model, entirely parameterized on the basis of experimental data, predicts that transient receptor confinement by the MSK meshwork allows for rapid reassociation of dissociated receptor dimers. Moreover, the experimentally observed apparent stabilization of receptor dimers in the plasma membrane was reproduced by simulations of a refined, hierarchical compartment model. Our simulations further revealed that the two-dimensional association rate constant is a key parameter for controlling the extent of MSK-mediated stabilization of protein complexes, thus ensuring the specificity of this effect. Together, experimental evidence and simulations support the hypothesis that passive receptor confinement by MSK-based microcompartmentalization promotes maintenance of signaling complexes in the plasma membrane. PMID:27957535

A spatial focusing model for G protein signals. Regulator of G protein signaling (RGS) protien-mediated kinetic scaffolding.

PubMed

Zhong, Huailing; Wade, Susan M; Woolf, Peter J; Linderman, Jennifer J; Traynor, John R; Neubig, Richard R

2003-02-28

Regulators of G protein signaling (RGS) are GTPase-accelerating proteins (GAPs), which can inhibit heterotrimeric G protein pathways. In this study, we provide experimental and theoretical evidence that high concentrations of receptors (as at a synapse) can lead to saturation of GDP-GTP exchange making GTP hydrolysis rate-limiting. This results in local depletion of inactive heterotrimeric G-GDP, which is reversed by RGS GAP activity. Thus, RGS enhances receptor-mediated G protein activation even as it deactivates the G protein. Evidence supporting this model includes a GTP-dependent enhancement of guanosine 5'-3-O-(thio)triphosphate (GTPgammaS) binding to G(i) by RGS. The RGS domain of RGS4 is sufficient for this, not requiring the NH(2)- or COOH-terminal extensions. Furthermore, a kinetic model including only the GAP activity of RGS replicates the GTP-dependent enhancement of GTPgammaS binding observed experimentally. Finally in a Monte Carlo model, this mechanism results in a dramatic "spatial focusing" of active G protein. Near the receptor, G protein activity is maintained even with RGS due to the ability of RGS to reduce depletion of local Galpha-GDP levels permitting rapid recoupling to receptor and maintained G protein activation near the receptor. In contrast, distant signals are suppressed by the RGS, since Galpha-GDP is not depleted there. Thus, a novel RGS-mediated "kinetic scaffolding" mechanism is proposed which narrows the spatial range of active G protein around a cluster of receptors limiting the spill-over of G protein signals to more distant effector molecules, thus enhancing the specificity of G(i) protein signals.

Decreasing the Expression of GABAA α5 Subunit-Containing Receptors Partially Improves Cognitive, Electrophysiological, and Morphological Hippocampal Defects in the Ts65Dn Model of Down Syndrome.

PubMed

Vidal, Verónica; García-Cerro, Susana; Martínez, Paula; Corrales, Andrea; Lantigua, Sara; Vidal, Rebeca; Rueda, Noemí; Ozmen, Laurence; Hernández, Maria-Clemencia; Martínez-Cué, Carmen

2018-06-01

Trisomy 21 or Down syndrome (DS) is the most common cause of intellectual disability of a genetic origin. The Ts65Dn (TS) mouse, which is the most commonly used and best-characterized mouse model of DS, displays many of the cognitive, neuromorphological, and biochemical anomalies that are found in the human condition. One of the mechanisms that have been proposed to be responsible for the cognitive deficits in this mouse model is impaired GABA-mediated inhibition. Because of the well-known modulatory role of GABA A α5 subunit-containing receptors in cognitive processes, these receptors are considered to be potential targets for improving the intellectual disability in DS. The chronic administration of GABA A α5-negative allosteric modulators has been shown to be procognitive without anxiogenic or proconvulsant side effects. In the present study, we use a genetic approach to evaluate the contribution of GABA A α5 subunit-containing receptors to the cognitive, electrophysiological, and neuromorphological deficits in TS mice. We show that reducing the expression of GABA A α5 receptors by deleting one or two copies of the Gabra5 gene in TS mice partially ameliorated the cognitive impairments, improved long-term potentiation, enhanced neural differentiation and maturation, and normalized the density of the GABAergic synapse markers. Reducing the gene dosage of Gabra5 in TS mice did not induce motor alterations and anxiety or affect the viability of the mice. Our results provide further evidence of the role of GABA A α5 receptor-mediated inhibition in cognitive impairment in the TS mouse model of DS.

PM2.5 pollution from household solid fuel burning practices in Central India: 2. Application of receptor models for source apportionment.

PubMed

Matawle, Jeevan Lal; Pervez, Shamsh; Deb, Manas Kanti; Shrivastava, Anjali; Tiwari, Suresh

2018-02-01

USEPA's UNMIX, positive matrix factorization (PMF) and effective variance-chemical mass balance (EV-CMB) receptor models were applied to chemically speciated profiles of 125 indoor PM 2.5 measurements, sampled longitudinally during 2012-2013 in low-income group households of Central India which uses solid fuels for cooking practices. Three step source apportionment studies were carried out to generate more confident source characterization. Firstly, UNMIX6.0 extracted initial number of source factors, which were used to execute PMF5.0 to extract source-factor profiles in second step. Finally, factor analog locally derived source profiles were supplemented to EV-CMB8.2 with indoor receptor PM 2.5 chemical profile to evaluate source contribution estimates (SCEs). The results of combined use of three receptor models clearly describe that UNMIX and PMF are useful tool to extract types of source categories within small receptor dataset and EV-CMB can pick those locally derived source profiles for source apportionment which are analog to PMF-extracted source categories. The source apportionment results have also shown three fold higher relative contribution of solid fuel burning emissions to indoor PM 2.5 compared to those measurements reported for normal households with LPG stoves. The previously reported influential source marker species were found to be comparatively similar to those extracted from PMF fingerprint plots. The comparison between PMF and CMB SCEs results were also found to be qualitatively similar. The performance fit measures of all three receptor models were cross-verified and validated and support each other to gain confidence in source apportionment results.

Inverse agonist and neutral antagonist actions of synthetic compounds at an insect 5-HT1 receptor

PubMed Central

Troppmann, B; Balfanz, S; Baumann, A; Blenau, W

2010-01-01

Background and purpose: 5-Hydroxytryptamine (5-HT) has been shown to control and modulate many physiological and behavioural functions in insects. In this study, we report the cloning and pharmacological properties of a 5-HT1 receptor of an insect model for neurobiology, physiology and pharmacology. Experimental approach: A cDNA encoding for the Periplaneta americana 5-HT1 receptor was amplified from brain cDNA. The receptor was stably expressed in HEK 293 cells, and the functional and pharmacological properties were determined in cAMP assays. Receptor distribution was investigated by RT-PCR and by immunocytochemistry using an affinity-purified polyclonal antiserum. Key results: The P. americana 5-HT1 receptor (Pea5-HT1) shares pronounced sequence and functional similarity with mammalian 5-HT1 receptors. Activation with 5-HT reduced adenylyl cyclase activity in a dose-dependent manner. Pea5-HT1 was expressed as a constitutively active receptor with methiothepin acting as a neutral antagonist, and WAY 100635 as an inverse agonist. Receptor mRNA was present in various tissues including brain, salivary glands and midgut. Receptor-specific antibodies showed that the native protein was expressed in a glycosylated form in membrane samples of brain and salivary glands. Conclusions and implications: This study marks the first pharmacological identification of an inverse agonist and a neutral antagonist at an insect 5-HT1 receptor. The results presented here should facilitate further analyses of 5-HT1 receptors in mediating central and peripheral effects of 5-HT in insects. PMID:20233210

Modelling and simulation of biased agonism dynamics at a G protein-coupled receptor.

PubMed

Bridge, L J; Mead, J; Frattini, E; Winfield, I; Ladds, G

2018-04-07

Theoretical models of G protein-coupled receptor (GPCR) concentration-response relationships often assume an agonist producing a single functional response via a single active state of the receptor. These models have largely been analysed assuming steady-state conditions. There is now much experimental evidence to suggest that many GPCRs can exist in multiple receptor conformations and elicit numerous functional responses, with ligands having the potential to activate different signalling pathways to varying extents-a concept referred to as biased agonism, functional selectivity or pluri-dimensional efficacy. Moreover, recent experimental results indicate a clear possibility for time-dependent bias, whereby an agonist's bias with respect to different pathways may vary dynamically. Efforts towards understanding the implications of temporal bias by characterising and quantifying ligand effects on multiple pathways will clearly be aided by extending current equilibrium binding and biased activation models to include G protein activation dynamics. Here, we present a new model of time-dependent biased agonism, based on ordinary differential equations for multiple cubic ternary complex activation models with G protein cycle dynamics. This model allows simulation and analysis of multi-pathway activation bias dynamics at a single receptor for the first time, at the level of active G protein (α GTP ), towards the analysis of dynamic functional responses. The model is generally applicable to systems with N G G proteins and N* active receptor states. Numerical simulations for N G =N * =2 reveal new insights into the effects of system parameters (including cooperativities, and ligand and receptor concentrations) on bias dynamics, highlighting new phenomena including the dynamic inter-conversion of bias direction. Further, we fit this model to 'wet' experimental data for two competing G proteins (G i and G s ) that become activated upon stimulation of the adenosine A 1 receptor with adenosine derivative compounds. Finally, we show that our model can qualitatively describe the temporal dynamics of this competing G protein activation. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

An Allosteric Coagonist Model for Propofol Effects on α1β2γ2L γ-Aminobutyric Acid Type A Receptors

PubMed Central

Ruesch, Dirk; Neumann, Elena; Wulf, Hinnerk; Forman, Stuart A.

2011-01-01

Background Propofol produces its major actions via γ-aminobutyric acid type A (GABAA) receptors. At low concentrations, propofol enhances agonist-stimulated GABAA receptor activity, and high propofol concentrations directly activate receptors. Etomidate produces similar effects, and there is convincing evidence that a single class of etomidate sites mediate both agonist modulation and direct GABAA receptor activation. It is unknown if the propofol binding site(s) on GABAA receptors that modulate agonist-induced activity also mediate direct activation. Methods GABAA α1β2γ2L receptors were heterologously expressed in Xenopus oocytes and activity was quantified using voltage clamp electrophysiology. We tested whether propofol and etomidate display the same linkage between agonist modulation and direct activation of GABAA receptors by identifying equi-efficacious drug solutions for direct activation. We then determined whether these drug solutions produce equal modulation of GABA-induced receptor activity. We also measured propofol-dependent direct activation and modulation of low GABA responses. Allosteric coagonist models similar to that established for etomidate, but with variable numbers of propofol sites, were fitted to combined data. Results Solutions of 19 μM propofol and 10 μM etomidate were found to equally activate GABAA receptors. These two drug solutions also produced indistinguishable modulation of GABA-induced receptor activity. Combined electrophysiological data behaved in a manner consistent with allosteric co-agonist models with more than one propofol site. The best fit was observed when the model assumed three equivalent propofol sites. Conclusions Our results support the hypothesis that propofol, like etomidate, acts at GABAA receptor sites mediating both GABA modulation and direct activation. PMID:22104494

Glutamate metabotropic receptors as targets for drug therapy in epilepsy.

PubMed

Moldrich, Randal X; Chapman, Astrid G; De Sarro, Giovambattista; Meldrum, Brian S

2003-08-22

Metabotropic glutamate (mGlu) receptors have multiple actions on neuronal excitability through G-protein-linked modifications of enzymes and ion channels. They act presynaptically to modify glutamatergic and gamma-aminobutyric acid (GABA)-ergic transmission and can contribute to long-term changes in synaptic function. The recent identification of subtype-selective agonists and antagonists has permitted evaluation of mGlu receptors as potential targets in the treatment of epilepsy. Agonists acting on group I mGlu receptors (mGlu1 and mGlu5) are convulsant. Antagonists acting on mGlu1 or mGlu5 receptors are anticonvulsant against 3,5-dihydroxyphenylglycine (DHPG)-induced seizures and in mouse models of generalized motor seizures and absence seizures. The competitive, phenylglycine mGlu1/5 receptor antagonists generally require intracerebroventricular administration for potent anticonvulsant efficacy but noncompetitive antagonists, e.g., (3aS,6aS)-6a-naphthalen-2-ylmethyl-5-methyliden-hexahydrocyclopenta[c]furan-1-on (BAY36-7620), 2-methyl-6-(phenylethynyl)pyridine hydrochloride (MPEP), and 2-methyl-6-(2-phenylethenyl)pyridine (SIB-1893) block generalized seizures with systemic administration. Agonists acting on group II mGlu receptors (mGlu2, mGlu3) to reduce glutamate release are anticonvulsant, e.g., 2R,4R-aminopyrrolidine-2,4-dicarboxylate [(2R,4R)-APDC], (+)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (LY354740), and (-)-2-oxa-4-aminobicyclo[3.1.0]hexane-4,6-dicarboxylate (LY379268). The classical agonists acting on group III mGlu receptors such as L-(+)-2-amino-4-phosphonobutyric acid, and L-serine-O-phosphate are acutely proconvulsant with some anticonvulsant activity. The more recently identified agonists (R,S)-4-phosphonophenylglycine [(R,S)-PPG] and (S)-3,4-dicarboxyphenylglycine [(S)-3,4-DCPG] and (1S,3R,4S)-1-aminocyclopentane-1,2,4-tricarboxylic acid [ACPT-1] are all anticonvulsant without proconvulsant effects. Studies in animal models of kindling reveal some efficacy of mGlu receptor ligands against fully kindled limbic seizures. In genetic mouse models, mGlu1/5 antagonists and mGlu2/3 agonists are effective against absence seizures. Thus, antagonists at group I mGlu receptors and agonists at groups II and III mGlu receptors are potential antiepileptic agents, but their clinical usefulness will depend on their acute and chronic side effects. Potential also exists for combining mGlu receptor ligands with other glutamatergic and non-glutamatergic agents to produce an enhanced anticonvulsant effect. This review also discusses what is known about mGlu receptor expression and function in rodent epilepsy models and human epileptic conditions.

A Role for D1 Dopamine Receptors in Striatal Methamphetamine-Induced Neurotoxicity

PubMed Central

Friend, Danielle M.; Keefe, Kristen A.

2015-01-01

Methamphetamine (METH) exposure results in long-term damage to the dopamine system in both human METH abusers and animal models. One factor that has been heavily implicated in this METH-induced damage to the dopaminergic system is the activation of D1 Dopamine (DA) receptors. However, a significant caveat to the studies investigating the role of the receptor in such toxicity is that genetic and pharmacological manipulations of the D1 DA receptor also mitigate METH-induced hyperthermia. Importantly, METH-induced hyperthermia is tightly associated with the neurotoxicity, such that simply cooling animals during METH exposure protects against the neurotoxicity. Therefore, it is difficult to determine whether D1 DA receptors per se play an important role in METH-induced neurotoxicity or whether the protection observed simply resulted from a mitigation of METH-induced hyperthermia. To answer this important question, the current study infused a D1 DA receptor antagonist into striatum during METH exposure while controlling for METH-induced hyperthermia. Here we found that even when METH-induced hyperthermia is maintained, the coadministration of a D1 DA receptor antagonist protects against METH-induced neurotoxicity, strongly suggesting that D1 DA receptors play an important role in METH-induced neurotoxicity apart from the mitigation of METH-induced hyperthermia. PMID:23994061

A role for D1 dopamine receptors in striatal methamphetamine-induced neurotoxicity.

PubMed

Friend, Danielle M; Keefe, Kristen A

2013-10-25

Methamphetamine (METH) exposure results in long-term damage to the dopamine system in both human METH abusers and animal models. One factor that has been heavily implicated in this METH-induced damage to the dopaminergic system is the activation of D1 dopamine (DA) receptors. However, a significant caveat to the studies investigating the role of the receptor in such toxicity is that genetic and pharmacological manipulations of the D1 DA receptor also mitigate METH-induced hyperthermia. Importantly, METH-induced hyperthermia is tightly associated with the neurotoxicity, such that simply cooling animals during METH exposure protects against the neurotoxicity. Therefore, it is difficult to determine whether D1 DA receptors per se play an important role in METH-induced neurotoxicity or whether the protection observed simply resulted from a mitigation of METH-induced hyperthermia. To answer this important question, the current study infused a D1 DA receptor antagonist into striatum during METH exposure while controlling for METH-induced hyperthermia. Here we found that even when METH-induced hyperthermia is maintained, the coadministration of a D1 DA receptor antagonist protects against METH-induced neurotoxicity, strongly suggesting that D1 DA receptors play an important role in METH-induced neurotoxicity apart from the mitigation of METH-induced hyperthermia. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Improving database enrichment through ensemble docking

NASA Astrophysics Data System (ADS)

Rao, Shashidhar; Sanschagrin, Paul C.; Greenwood, Jeremy R.; Repasky, Matthew P.; Sherman, Woody; Farid, Ramy

2008-09-01

While it may seem intuitive that using an ensemble of multiple conformations of a receptor in structure-based virtual screening experiments would necessarily yield improved enrichment of actives relative to using just a single receptor, it turns out that at least in the p38 MAP kinase model system studied here, a very large majority of all possible ensembles do not yield improved enrichment of actives. However, there are combinations of receptor structures that do lead to improved enrichment results. We present here a method to select the ensembles that produce the best enrichments that does not rely on knowledge of active compounds or sophisticated analyses of the 3D receptor structures. In the system studied here, the small fraction of ensembles of up to 3 receptors that do yield good enrichments of actives were identified by selecting ensembles that have the best mean GlideScore for the top 1% of the docked ligands in a database screen of actives and drug-like "decoy" ligands. Ensembles of two receptors identified using this mean GlideScore metric generally outperform single receptors, while ensembles of three receptors identified using this metric consistently give optimal enrichment factors in which, for example, 40% of the known actives outrank all the other ligands in the database.

Endogenous opioid mechanisms partially mediate P2X3/P2X2/3-related antinociception in rat models of inflammatory and chemogenic pain but not neuropathic pain.

PubMed

McGaraughty, Steve; Honore, Prisca; Wismer, Carol T; Mikusa, Joseph; Zhu, Chang Z; McDonald, Heath A; Bianchi, Bruce; Faltynek, Connie R; Jarvis, Michael F

2005-09-01

P2X3/P2X2/3 receptors have emerged as important components of nociception. However, there is limited information regarding the neurochemical systems that are affected by antagonism of the P2X3/P2X2/3 receptor and that ultimately contribute to the ensuing antinociception. In order to determine if the endogenous opioid system is involved in this antinociception, naloxone was administered just prior to the injection of a selective P2X3/P2X2/3 receptor antagonist, A-317491, in rat models of neuropathic, chemogenic, and inflammatory pain. Naloxone (1-10 mg kg(-1), i.p.), dose-dependently reduced the antinociceptive effects of A-317491 (1-300 micromol kg(-1), s.c.) in the CFA model of thermal hyperalgesia and the formalin model of chemogenic pain (2nd phase), but not in the L5-L6 spinal nerve ligation model of neuropathic allodynia. In comparison experiments, the same doses of naloxone blocked or attenuated the actions of morphine (2 or 8 mg kg(-1), s.c.) in each of these behavioral models. Injection of a peripheral opioid antagonist, naloxone methiodide (10 mg kg(-1), i.p.), did not affect A-317491-induced antinociception in the CFA and formalin assays, suggesting that the opioid component of this antinociception occurred within the CNS. Furthermore, this utilization of the central opioid system could be initiated by antagonism of spinal P2X3/P2X2/3 receptors since the antinociceptive actions of intrathecally delivered A-317491 (30 nmol) in the formalin model were reduced by both intrathecally (10-50 nmol) and systemically (10 mg kg(-1), i.p.) administered naloxone. This utilization of the opioid system was not specific to A-317491 since suramin-, a nonselective P2X receptor antagonist, induced antinociception was also attenuated by naloxone. In in vitro studies, A-317491 (3-100 microM) did not produce any agonist response at delta opioid receptors expressed in NG108-15 cells. A-317491 had been previously shown to be inactive at the kappa and mu opioid receptors. Furthermore, naloxone, at concentrations up to 1 mM, did not compete for [3H] A-317491 binding in 1321N1 cells expressing human P2X3 receptors. Taken together, these results indicate that antagonism of spinal P2X3/P2X2/3 receptors results in an indirect activation of the opioid system to alleviate inflammatory hyperalgesia and chemogenic nociception.

Comparing pharmacophore models derived from crystallography and NMR ensembles

NASA Astrophysics Data System (ADS)

Ghanakota, Phani; Carlson, Heather A.

2017-11-01

NMR and X-ray crystallography are the two most widely used methods for determining protein structures. Our previous study examining NMR versus X-Ray sources of protein conformations showed improved performance with NMR structures when used in our Multiple Protein Structures (MPS) method for receptor-based pharmacophores (Damm, Carlson, J Am Chem Soc 129:8225-8235, 2007). However, that work was based on a single test case, HIV-1 protease, because of the rich data available for that system. New data for more systems are available now, which calls for further examination of the effect of different sources of protein conformations. The MPS technique was applied to Growth factor receptor bound protein 2 (Grb2), Src SH2 homology domain (Src-SH2), FK506-binding protein 1A (FKBP12), and Peroxisome proliferator-activated receptor-γ (PPAR-γ). Pharmacophore models from both crystal and NMR ensembles were able to discriminate between high-affinity, low-affinity, and decoy molecules. As we found in our original study, NMR models showed optimal performance when all elements were used. The crystal models had more pharmacophore elements compared to their NMR counterparts. The crystal-based models exhibited optimum performance only when pharmacophore elements were dropped. This supports our assertion that the higher flexibility in NMR ensembles helps focus the models on the most essential interactions with the protein. Our studies suggest that the "extra" pharmacophore elements seen at the periphery in X-ray models arise as a result of decreased protein flexibility and make very little contribution to model performance.

Free fatty acid (FFA) and hydroxy carboxylic acid (HCA) receptors.

PubMed

Offermanns, Stefan

2014-01-01

Saturated and unsaturated free fatty acids (FFAs), as well as hydroxy carboxylic acids (HCAs) such as lactate and ketone bodies, are carriers of metabolic energy, precursors of biological mediators, and components of biological structures. However, they are also able to exert cellular effects through G protein-coupled receptors named FFA1-FFA4 and HCA1-HCA3. Work during the past decade has shown that these receptors are widely expressed in the human body and regulate the metabolic, endocrine, immune and other systems to maintain homeostasis under changing dietary conditions. The development of genetic mouse models and the generation of synthetic ligands of individual FFA and HCA receptors have been instrumental in identifying cellular and biological functions of these receptors. These studies have produced strong evidence that several FFA and HCA receptors can be targets for the prevention and treatment of various diseases, including type 2 diabetes mellitus, obesity, and inflammation.

Dantrolene, a treatment for Alzheimer disease?

PubMed

Liang, Li; Wei, Huafeng

2015-01-01

Alzheimer disease (AD) is a fatal progressive disease and the most common form of dementia without effective treatments. Previous studies support that the disruption of endoplasmic reticulum Ca through overactivation of ryanodine receptors plays an important role in the pathogenesis of AD. Normalization of intracellular Ca homeostasis could be an effective strategy for AD therapies. Dantrolene, an antagonist of ryanodine receptors and an FDA-approved drug for clinical treatment of malignant hyperthermia and muscle spasms, exhibits neuroprotective effects in multiple models of neurodegenerative disorders. Recent preclinical studies consistently support the therapeutic effects of dantrolene in various types of AD animal models and were summarized in the current review.

Homology modeling study toward identifying structural properties in the HA2 B-loop that would influence the HA1 receptor-binding site.

PubMed

Cueno, Marni E; Imai, Kenichi; Shimizu, Kazufumi; Ochiai, Kuniyasu

2013-07-01

Influenza hemagglutinin (HA) consists of a fibrous globular stem (HA2) inserted into the viral membrane supporting a globular head (HA1). HA1 receptor-binding has been hypothesized to be structurally correlated to the HA2 B-loop, however, this was never fully understood. Here, we elucidated the structural relationship between the HA2 B-loop and the HA1 receptor-binding site (RBS). Throughout this study, we analyzed 2486 H1N1 HA homology models obtained from human, swine and avian strains during 1976-2012. Quality of all homology models were verified before further analyses. We established that amino acid residue 882 is putatively strain-conserved and differs in the human (K882), swine (H882) and avian (N882) strains. Moreover, we observed that the amino acid at residue 882 and, similarly, its orientation has the potential to influence the HA1 RBS diameter measurements which we hypothesize may consequentially affect influenza H1N1 viral infectivity, immune escape, transmissibility, and evolution. Copyright © 2013 Elsevier Inc. All rights reserved.

Mesenchymal stem cells cannot affect mRNA expression of toll-like receptors in different tissues during sepsis.

PubMed

Pedrazza, Leonardo; Pereira, Talita Carneiro Brandão; Abujamra, Ana Lucia; Nunes, Fernanda Bordignon; Bogo, Maurício Reis; de Oliveira, Jarbas Rodrigues

2017-07-01

Experimental animal models and human clinical studies support a crucial role for TLRs in infectious diseases. The aim of this study was to test the ability of MSCs, which have immunomodulatory effects, of altering the mRNA expression of toll-like receptors during a experimental model of sepsis in different tissues. Three experimental groups (male C57BL/6 mice) were formed for the test: control group, untreated septic group and septic group treated with MSCs (1 × 10 6 cells/animal). Lungs, cortex, kidney, liver and colon tissue were dissected after 12 h of sepsis induction and TLR2/3/4/9 mRNA were evaluated by RT-qPCR. We observed a decrease of TLR2 and 9 mRNA expression in the liver of the sepsis group, while TLR3 was decreased in the lung and liver. No change was found between the sepsis group and the sepsis + MSC group. In this model of experimental sepsis the MSCs were unable to modify the mRNA expression of the different toll-like receptors evaluated.

Exploration of optimal dosing regimens of haloperidol, a D2 Antagonist, via modeling and simulation analysis in a D2 receptor occupancy study.

PubMed

Lim, Hyeong-Seok; Kim, Su Jin; Noh, Yook-Hwan; Lee, Byung Chul; Jin, Seok-Joon; Park, Hyun Soo; Kim, Soohyeon; Jang, In-Jin; Kim, Sang Eun

2013-03-01

To evaluate the potential usage of D(2) receptor occupancy (D2RO) measured by positron emission tomography (PET) in antipsychotic development. In this randomized, parallel group study, eight healthy male volunteers received oral doses of 0.5 (n = 3), 1 (n = 2), or 3 mg (n = 3) of haloperidol once daily for 7 days. PET's were scanned before haloperidol, and on days 8, 12, with serial pharmacokinetic sampling on day 7. Pharmacokinetics and binding potential to D(2) receptor in putamen and caudate nucleus over time were analyzed using NONMEM, and simulations for the profiles of D2RO over time on various regimens of haloperidol were conducted to find the optimal dosing regimens. One compartment model with a saturable binding compartment, and inhibitory E(max) model in the effect compartment best described the data. Plasma haloperidol concentrations at half-maximal inhibition were 0.791 and 0.650 ng/ml, in putamen and caudate nucleus. Simulation suggested haloperidol 2 mg every 12 h is near the optimal dose. This study showed that sparse D2RO measurements in steady state pharmacodynamic design after multiple dosing could reveal the possibility of treatment effect of D(2) antagonist, and could identify the potential optimal doses for later clinical studies by modeling and simulation.

The neuropsychopharmacology of phencyclidine: from NMDA receptor hypofunction to the dopamine hypothesis of schizophrenia.

PubMed

Jentsch, J D; Roth, R H

1999-03-01

Administration of noncompetitive NMDA/glutamate receptor antagonists, such as phencyclidine (PCP) and ketamine, to humans induces a broad range of schizophrenic-like symptomatology, findings that have contributed to a hypoglutamatergic hypothesis of schizophrenia. Moreover, a history of experimental investigations of the effects of these drugs in animals suggests that NMDA receptor antagonists may model some behavioral symptoms of schizophrenia in nonhuman subjects. In this review, the usefulness of PCP administration as a potential animal model of schizophrenia is considered. To support the contention that NMDA receptor antagonist administration represents a viable model of schizophrenia, the behavioral and neurobiological effects of these drugs are discussed, especially with regard to differing profiles following single-dose and long-term exposure. The neurochemical effects of NMDA receptor antagonist administration are argued to support a neurobiological hypothesis of schizophrenia, which includes pathophysiology within several neurotransmitter systems, manifested in behavioral pathology. Future directions for the application of NMDA receptor antagonist models of schizophrenia to preclinical and pathophysiological research are offered.

Immune Receptors and Co-receptors in Antiviral Innate Immunity in Plants.

PubMed

Gouveia, Bianca C; Calil, Iara P; Machado, João Paulo B; Santos, Anésia A; Fontes, Elizabeth P B

2016-01-01

Plants respond to pathogens using an innate immune system that is broadly divided into PTI (pathogen-associated molecular pattern- or PAMP-triggered immunity) and ETI (effector-triggered immunity). PTI is activated upon perception of PAMPs, conserved motifs derived from pathogens, by surface membrane-anchored pattern recognition receptors (PRRs). To overcome this first line of defense, pathogens release into plant cells effectors that inhibit PTI and activate effector-triggered susceptibility (ETS). Counteracting this virulence strategy, plant cells synthesize intracellular resistance (R) proteins, which specifically recognize pathogen effectors or avirulence (Avr) factors and activate ETI. These coevolving pathogen virulence strategies and plant resistance mechanisms illustrate evolutionary arms race between pathogen and host, which is integrated into the zigzag model of plant innate immunity. Although antiviral immune concepts have been initially excluded from the zigzag model, recent studies have provided several lines of evidence substantiating the notion that plants deploy the innate immune system to fight viruses in a manner similar to that used for non-viral pathogens. First, most R proteins against viruses so far characterized share structural similarity with antibacterial and antifungal R gene products and elicit typical ETI-based immune responses. Second, virus-derived PAMPs may activate PTI-like responses through immune co-receptors of plant PTI. Finally, and even more compelling, a viral Avr factor that triggers ETI in resistant genotypes has recently been shown to act as a suppressor of PTI, integrating plant viruses into the co-evolutionary model of host-pathogen interactions, the zigzag model. In this review, we summarize these important progresses, focusing on the potential significance of antiviral immune receptors and co-receptors in plant antiviral innate immunity. In light of the innate immune system, we also discuss a newly uncovered layer of antiviral defense that is specific to plant DNA viruses and relies on transmembrane receptor-mediated translational suppression for defense.

A multiscale computational approach to dissect early events in the Erb family receptor mediated activation, differential signaling, and relevance to oncogenic transformations.

PubMed

Liu, Yingting; Purvis, Jeremy; Shih, Andrew; Weinstein, Joshua; Agrawal, Neeraj; Radhakrishnan, Ravi

2007-06-01

We describe a hierarchical multiscale computational approach based on molecular dynamics simulations, free energy-based molecular docking simulations, deterministic network-based kinetic modeling, and hybrid discrete/continuum stochastic dynamics protocols to study the dimer-mediated receptor activation characteristics of the Erb family receptors, specifically the epidermal growth factor receptor (EGFR). Through these modeling approaches, we are able to extend the prior modeling of EGF-mediated signal transduction by considering specific EGFR tyrosine kinase (EGFRTK) docking interactions mediated by differential binding and phosphorylation of different C-terminal peptide tyrosines on the RTK tail. By modeling signal flows through branching pathways of the EGFRTK resolved on a molecular basis, we are able to transcribe the effects of molecular alterations in the receptor (e.g., mutant forms of the receptor) to differing kinetic behavior and downstream signaling response. Our molecular dynamics simulations show that the drug sensitizing mutation (L834R) of EGFR stabilizes the active conformation to make the system constitutively active. Docking simulations show preferential characteristics (for wildtype vs. mutant receptors) in inhibitor binding as well as preferential enhancement of phosphorylation of particular substrate tyrosines over others. We find that in comparison to the wildtype system, the L834R mutant RTK preferentially binds the inhibitor erlotinib, as well as preferentially phosphorylates the substrate tyrosine Y1068 but not Y1173. We predict that these molecular level changes result in preferential activation of the Akt signaling pathway in comparison to the Erk signaling pathway for cells with normal EGFR expression. For cells with EGFR over expression, the mutant over activates both Erk and Akt pathways, in comparison to wildtype. These results are consistent with qualitative experimental measurements reported in the literature. We discuss these consequences in light of how the network topology and signaling characteristics of altered (mutant) cell lines are shaped differently in relationship to native cell lines.

Combined 3D-QSAR, molecular docking and molecular dynamics study on thyroid hormone activity of hydroxylated polybrominated diphenyl ethers to thyroid receptors β

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Xiaolin; Ye, Li; Wang, Xiaoxiang

2012-12-15

Several recent reports suggested that hydroxylated polybrominated diphenyl ethers (HO-PBDEs) may disturb thyroid hormone homeostasis. To illuminate the structural features for thyroid hormone activity of HO-PBDEs and the binding mode between HO-PBDEs and thyroid hormone receptor (TR), the hormone activity of a series of HO-PBDEs to thyroid receptors β was studied based on the combination of 3D-QSAR, molecular docking, and molecular dynamics (MD) methods. The ligand- and receptor-based 3D-QSAR models were obtained using Comparative Molecular Similarity Index Analysis (CoMSIA) method. The optimum CoMSIA model with region focusing yielded satisfactory statistical results: leave-one-out cross-validation correlation coefficient (q{sup 2}) was 0.571 andmore » non-cross-validation correlation coefficient (r{sup 2}) was 0.951. Furthermore, the results of internal validation such as bootstrapping, leave-many-out cross-validation, and progressive scrambling as well as external validation indicated the rationality and good predictive ability of the best model. In addition, molecular docking elucidated the conformations of compounds and key amino acid residues at the docking pocket, MD simulation further determined the binding process and validated the rationality of docking results. -- Highlights: ► The thyroid hormone activities of HO-PBDEs were studied by 3D-QSAR. ► The binding modes between HO-PBDEs and TRβ were explored. ► 3D-QSAR, molecular docking, and molecular dynamics (MD) methods were performed.« less

F104S c-Mpl responds to a transmembrane domain-binding thrombopoietin receptor agonist: proof of concept that selected receptor mutations in congenital amegakaryocytic thrombocytopenia can be stimulated with alternative thrombopoietic agents.

PubMed

Fox, Norma E; Lim, Jihyang; Chen, Rose; Geddis, Amy E

2010-05-01

To determine whether specific c-Mpl mutations might respond to thrombopoietin receptor agonists. We created cell line models of type II c-Mpl mutations identified in congenital amegakaryocytic thrombocytopenia. We selected F104S c-Mpl for further study because it exhibited surface expression of the receptor. We measured proliferation of cell lines expressing wild-type or F104S c-Mpl in response to thrombopoietin receptor agonists targeting the extracellular (m-AMP4) or transmembrane (LGD-4665) domains of the receptor by 1-methyltetrazole-5-thiol assay. We measured thrombopoietin binding to the mutant receptor using an in vitro thrombopoietin uptake assay and identified F104 as a potentially critical residue for the interaction between the receptor and its ligand by aligning thrombopoietin and erythropoietin receptors from multiple species. Cells expressing F104S c-Mpl proliferated in response to LGD-4665, but not thrombopoietin or m-AMP4. Compared to thrombopoietin, LGD-4665 stimulates signaling with delayed kinetics in both wild-type and F104S c-Mpl-expressing cells. Although F104S c-Mpl is expressed on the cell surface in our BaF3 cell line model, the mutant receptor does not bind thrombopoietin. Comparison to the erythropoietin receptor suggests that F104 engages in hydrogen-bonding interactions that are critical for binding to thrombopoietin. These findings suggest that a small subset of patients with congenital amegakaryocytic thrombocytopenia might respond to treatment with thrombopoietin receptor agonists, but that responsiveness will depend on the type of mutation and agonist used. We postulate that F104 is critical for thrombopoietin binding. The kinetics of signaling in response to a transmembrane domain-binding agonist are delayed in comparison to thrombopoietin. 2010 ISEH Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.

Constrained positive matrix factorization: Elemental ratios, spatial distinction, and chemical transport model source contributions

NASA Astrophysics Data System (ADS)

Sturtz, Timothy M.

Source apportionment models attempt to untangle the relationship between pollution sources and the impacts at downwind receptors. Two frameworks of source apportionment models exist: source-oriented and receptor-oriented. Source based apportionment models use presumed emissions and atmospheric processes to estimate the downwind source contributions. Conversely, receptor based models leverage speciated concentration data from downwind receptors and apply statistical methods to predict source contributions. Integration of both source-oriented and receptor-oriented models could lead to a better understanding of the implications sources have on the environment and society. The research presented here investigated three different types of constraints applied to the Positive Matrix Factorization (PMF) receptor model within the framework of the Multilinear Engine (ME-2): element ratio constraints, spatial separation constraints, and chemical transport model (CTM) source attribution constraints. PM10-2.5 mass and trace element concentrations were measured in Winston-Salem, Chicago, and St. Paul at up to 60 sites per city during two different seasons in 2010. PMF was used to explore the underlying sources of variability. Information on previously reported PM10-2.5 tire and brake wear profiles were used to constrain these features in PMF by prior specification of selected species ratios. We also modified PMF to allow for combining the measurements from all three cities into a single model while preserving city-specific soil features. Relatively minor differences were observed between model predictions with and without the prior ratio constraints, increasing confidence in our ability to identify separate brake wear and tire wear features. Using separate data, source contributions to total fine particle carbon predicted by a CTM were incorporated into the PMF receptor model to form a receptor-oriented hybrid model. The level of influence of the CTM versus traditional PMF was varied using a weighting parameter applied to an object function as implemented in ME-2. The resulting hybrid model was used to quantify the contributions of total carbon from both wildfires and biogenic sources at two Interagency Monitoring of Protected Visual Environment monitoring sites, Monture and Sula Peak, Montana, from 2006 through 2008.

Impact of input data uncertainty on environmental exposure assessment models: A case study for electromagnetic field modelling from mobile phone base stations.

PubMed

Beekhuizen, Johan; Heuvelink, Gerard B M; Huss, Anke; Bürgi, Alfred; Kromhout, Hans; Vermeulen, Roel

2014-11-01

With the increased availability of spatial data and computing power, spatial prediction approaches have become a standard tool for exposure assessment in environmental epidemiology. However, such models are largely dependent on accurate input data. Uncertainties in the input data can therefore have a large effect on model predictions, but are rarely quantified. With Monte Carlo simulation we assessed the effect of input uncertainty on the prediction of radio-frequency electromagnetic fields (RF-EMF) from mobile phone base stations at 252 receptor sites in Amsterdam, The Netherlands. The impact on ranking and classification was determined by computing the Spearman correlations and weighted Cohen's Kappas (based on tertiles of the RF-EMF exposure distribution) between modelled values and RF-EMF measurements performed at the receptor sites. The uncertainty in modelled RF-EMF levels was large with a median coefficient of variation of 1.5. Uncertainty in receptor site height, building damping and building height contributed most to model output uncertainty. For exposure ranking and classification, the heights of buildings and receptor sites were the most important sources of uncertainty, followed by building damping, antenna- and site location. Uncertainty in antenna power, tilt, height and direction had a smaller impact on model performance. We quantified the effect of input data uncertainty on the prediction accuracy of an RF-EMF environmental exposure model, thereby identifying the most important sources of uncertainty and estimating the total uncertainty stemming from potential errors in the input data. This approach can be used to optimize the model and better interpret model output. Copyright © 2014 Elsevier Inc. All rights reserved.

Dicholine succinate, the neuronal insulin sensitizer, normalizes behavior, REM sleep, hippocampal pGSK3 beta and mRNAs of NMDA receptor subunits in mouse models of depression.

PubMed

Cline, Brandon H; Costa-Nunes, Joao P; Cespuglio, Raymond; Markova, Natalyia; Santos, Ana I; Bukhman, Yury V; Kubatiev, Aslan; Steinbusch, Harry W M; Lesch, Klaus-Peter; Strekalova, Tatyana

2015-01-01

Central insulin receptor-mediated signaling is attracting the growing attention of researchers because of rapidly accumulating evidence implicating it in the mechanisms of plasticity, stress response, and neuropsychiatric disorders including depression. Dicholine succinate (DS), a mitochondrial complex II substrate, was shown to enhance insulin-receptor mediated signaling in neurons and is regarded as a sensitizer of the neuronal insulin receptor. Compounds enhancing neuronal insulin receptor-mediated transmission exert an antidepressant-like effect in several pre-clinical paradigms of depression; similarly, such properties for DS were found with a stress-induced anhedonia model. Here, we additionally studied the effects of DS on several variables which were ameliorated by other insulin receptor sensitizers in mice. Pre-treatment with DS of chronically stressed C57BL6 mice rescued normal contextual fear conditioning, hippocampal gene expression of NMDA receptor subunit NR2A, the NR2A/NR2B ratio and increased REM sleep rebound after acute predation. In 18-month-old C57BL6 mice, a model of elderly depression, DS restored normal sucrose preference and activated the expression of neural plasticity factors in the hippocampus as shown by Illumina microarray. Finally, young naïve DS-treated C57BL6 mice had reduced depressive- and anxiety-like behaviors and, similarly to imipramine-treated mice, preserved hippocampal levels of the phosphorylated (inactive) form of GSK3 beta that was lowered by forced swimming in pharmacologically naïve animals. Thus, DS can ameliorate behavioral and molecular outcomes under a variety of stress- and depression-related conditions. This further highlights neuronal insulin signaling as a new factor of pathogenesis and a potential pharmacotherapy of affective pathologies.

Allosteric modulation of the group III mGlu4 receptor provides functional neuroprotection in the 6-hydroxydopamine rat model of Parkinson's disease

PubMed Central

Betts, Matthew J; O'Neill, Michael J; Duty, Susan

2012-01-01

BACKGROUND AND PURPOSE We recently reported that broad spectrum agonist-induced activation of presynaptic group III metabotropic glutamate (mGlu) receptors within the substantia nigra pars compacta using L-2-amino-4-phosphonobutyrate provided functional neuroprotection in the 6-hydroxydopamine lesion rat model of Parkinson's disease. The aim of this study was to establish whether selective activation of the mGlu4 receptor alone could afford similar functional neuroprotection. EXPERIMENTAL APPROACH The neuroprotective effects of 8 days of supranigral treatment with a positive allosteric modulator of mGlu4 receptors, (+/−)-cis-2-(3,5-dichlorphenylcarbamoyl)cyclohexanecarboxylic acid (VU0155041), were investigated in rats with unilateral 6-hydroxydopamine lesions. The effects of VU0155041 treatment on motor function were assessed using both habitual (cylinder test) and forced (adjusted stepping, amphetamine-induced rotations) behavioural tests. Nigrostriatal tract integrity was examined by analysis of tyrosine hydroxylase, dopa decarboxylase or dopamine levels in the striatum and tyrosine hydroxylase-positive cell counts in the substantia nigra pars compacta. KEY RESULTS VU0155041 provided around 40% histological protection against a unilateral 6-hydroxydopamine lesion as well as significant preservation of motor function. These effects were inhibited by pre-treatment with (RS)-α-cyclopropyl-4-phosphonophenylglycine, confirming a receptor-mediated response. Reduced levels of inflammatory markers were also evident in the brains of VU0155041-treated animals. CONCLUSIONS AND IMPLICATIONS Allosteric potentiation of mGlu4 receptors in the substantia nigra pars compacta provided neuroprotective effects in the 6-hydroxydopamine rat model A reduced inflammatory response may contribute, in part, to this action. In addition to the reported symptomatic effects, activation of mGlu4 receptors may also offer a novel approach for slowing the progressive degeneration observed in Parkinson's disease. PMID:22404342

Dicholine succinate, the neuronal insulin sensitizer, normalizes behavior, REM sleep, hippocampal pGSK3 beta and mRNAs of NMDA receptor subunits in mouse models of depression

PubMed Central

Cline, Brandon H.; Costa-Nunes, Joao P.; Cespuglio, Raymond; Markova, Natalyia; Santos, Ana I.; Bukhman, Yury V.; Kubatiev, Aslan; Steinbusch, Harry W. M.; Lesch, Klaus-Peter; Strekalova, Tatyana

2015-01-01

Central insulin receptor-mediated signaling is attracting the growing attention of researchers because of rapidly accumulating evidence implicating it in the mechanisms of plasticity, stress response, and neuropsychiatric disorders including depression. Dicholine succinate (DS), a mitochondrial complex II substrate, was shown to enhance insulin-receptor mediated signaling in neurons and is regarded as a sensitizer of the neuronal insulin receptor. Compounds enhancing neuronal insulin receptor-mediated transmission exert an antidepressant-like effect in several pre-clinical paradigms of depression; similarly, such properties for DS were found with a stress-induced anhedonia model. Here, we additionally studied the effects of DS on several variables which were ameliorated by other insulin receptor sensitizers in mice. Pre-treatment with DS of chronically stressed C57BL6 mice rescued normal contextual fear conditioning, hippocampal gene expression of NMDA receptor subunit NR2A, the NR2A/NR2B ratio and increased REM sleep rebound after acute predation. In 18-month-old C57BL6 mice, a model of elderly depression, DS restored normal sucrose preference and activated the expression of neural plasticity factors in the hippocampus as shown by Illumina microarray. Finally, young naïve DS-treated C57BL6 mice had reduced depressive- and anxiety-like behaviors and, similarly to imipramine-treated mice, preserved hippocampal levels of the phosphorylated (inactive) form of GSK3 beta that was lowered by forced swimming in pharmacologically naïve animals. Thus, DS can ameliorate behavioral and molecular outcomes under a variety of stress- and depression-related conditions. This further highlights neuronal insulin signaling as a new factor of pathogenesis and a potential pharmacotherapy of affective pathologies. PMID:25767439

EFFECT OF SYSTEMIC BETA-BLOCKERS, ACE INHIBITORS, AND ANGIOTENSIN RECEPTOR BLOCKERS ON DEVELOPMENT OF CHOROIDAL NEOVASCULARIZATION IN PATIENTS WITH AGE-RELATED MACULAR DEGENERATION.

PubMed

Thomas, Akshay S; Redd, Travis; Hwang, Thomas

2015-10-01

Recent studies have suggested that the use of systemic beta-blockers, angiotensin-converting enzyme inhibitors, and angiotensin receptor blockers can induce regression of choroidal neovascularization in rodent models. The purpose of this study is to evaluate if these agents have a protective effect against the development of choroidal neovascularization in patients with age-related macular degeneration. In this single-center retrospective case-control study, the charts of 250 patients with neovascular age-related macular degeneration were compared with those of 250 controls with dry age-related macular degeneration. Charts were reviewed for current and past use of beta-blockers, angiotensin-converting enzyme inhibitors, and angiotensin receptor blockers. Frequency tables were generated, and associations were examined using chi-square tests, t-tests, and multivariate logistic regression. There was no statistically significant difference between rates of beta-blocker use (P = 0.57), angiotensin-converting enzyme inhibitors use (P = 0.20), or angiotensin receptor blockers use (P = 0.61) between the 2 groups. Additionally, there was no statistically significant difference between rates of use of combinations of the above drugs between the two groups. Although there is growing evidence that beta-blockers, angiotensin-converting enzyme inhibitors, and angiotensin receptor blockers can induce regression of choroidal neovascularization in rodent models, these medications do not seem to confer a protective effect against the development of choroidal neovascularization in patients with age-related macular degeneration.

Analogues of doxanthrine reveal differences between the dopamine D 1 receptor binding properties of chromanoisoquinolines and hexahydrobenzo[a]phenanthridines

USGS Publications Warehouse

Cueva, J.P.; Chemel, B.R.; Juncosa, J.I.; Lill, M.A.; Watts, V.J.; Nichols, D.E.

2012-01-01

Efforts to develop selective agonists for dopamine D 1-like receptors led to the discovery of dihydrexidine and doxanthrine, two bioisosteric ??-phenyldopamine-type full agonist ligands that display selectivity and potency at D 1-like receptors. We report herein an improved methodology for the synthesis of substituted chromanoisoquinolines (doxanthrine derivatives) and the evaluation of several new compounds for their ability to bind to D 1- and D 2-like receptors. Identical pendant phenyl ring substitutions on the dihydrexidine and doxanthrine templates surprisingly led to different effects on D 1-like receptor binding, suggesting important differences between the interactions of these ligands with the D 1 receptor. We propose, based on the biological results and molecular modeling studies, that slight conformational differences between the tetralin and chroman-based compounds lead to a shift in the location of the pendant ring substituents within the receptor. ?? 2011 Elsevier Ltd. All rights reserved.

The Ligand Binding Region of the Sigma-1 Receptor: Studies Utilizing Photoaffinity Probes, Sphingosine and N-Alkylamines

PubMed Central

Ruoho, Arnold E.; Chu, Uyen B.; Ramachandran, Subramaniam; Fontanilla, Dominique; Mavlyutov, Timur; Hajipour, Abdol R.

2015-01-01

The sigma-1 receptor is a 26 kDa endoplasmic reticulum resident membrane protein that has been shown to have chaperone activity in addition to its promiscuous binding to pharmacological agents. Ligand binding domain(s) of the sigma-1 receptor have been identified using the E. coli expressed and purified receptor protein and novel radioiodinated azido photoaffinity probes followed by pro-teolytic and chemical cleavage strategies. The outcome of these experiments indicates that the sigma-1 receptor ligand binding regions are formed primarily by juxtaposition of its second and third hydrophobic domains, regions where the protein shares considerable homology with the fungal enzyme, sterol isomerase that is essential for the biosynthesis of ergosterol. Data indicate that these hydrophobic steroid binding domain like (SBDL) regions on the sigma-1 receptor are likely to interact selectively with N-alkyl amines such as the endogenous sphingolipids and with synthetic N-alkylamines and N-aralkylamines derivatives. A proposed model for the sigma-1 receptor is presented. PMID:22288412

Nonerythropoietic Tissue Protective Compounds Are Highly Effective Facilitators of Wound Healing

PubMed Central

Erbayraktar, Zübeyde; Erbayraktar, Serhat; Yilmaz, Osman; Cerami, Anthony; Coleman, Thomas; Brines, Michael

2009-01-01

Erythropoietin (EPO) is a type I cytokine that utilizes different receptor isoforms either to maintain hematopoiesis or protect against injuries that arise from widely diverse etiologies. EPO also facilitates healing by reducing inflammation and mobilizing endothelial progenitor cells to participate in restorative neoangiogenesis, but it is unclear which EPO receptor isoform is responsible for healing and whether this receptor use varies according to the type of wound. In the present studies carried out in the rat, we have utilized receptor-selective derivatives of EPO to determine which receptor type operates in (i) a nonischemic wound (skin punch biopsy), (ii) a permanently ischemic wound (raised musculocutaneous flap), (iii) an intermittent ischemic reperfusion wound (pressure or decubitus ulcer), or (iv) wounds complicated by infection (cecal ligation and perforation). Using these models, we demonstrate that nonerythropoietic tissue protective compounds administered immediately following injury limit wound size and accelerate eschar closure independent of wound type. Moreover, in a model of peritonitis-induced adhesions, daily administration of the nonerythropoietic derivative carbamyl-EPO (10 μg/kg-bw) was associated with significantly lower serum TNFα concentration, illness scores, increased survival, as well as decreased adhesion formation. These results confirm that wound healing is mediated by the tissue protective receptor isoform and argue that nonerythropoietic tissue protective molecules constitute promising new PMID:19593407

Ionotropic and metabotropic glutamate receptor structure and pharmacology.

PubMed

Kew, James N C; Kemp, John A

2005-04-01

L: -Glutamate is the major excitatory neurotransmitter in the central nervous system (CNS) and mediates its actions via activation of both ionotropic and metabotropic receptor families. The development of selective ligands, including competitive agonists and antagonists and positive and negative allosteric modulators, has enabled investigation of the functional roles of glutamate receptor family members. In this review we describe the subunit structure and composition of the ionotropic and metabotropic glutamate receptors and discuss their pharmacology, particularly with respect to selective tools useful for investigation of their function in the CNS. A large number of ligands are now available that are selective either for glutamate receptor subfamilies or for particular receptor subtypes. Such ligands have enabled considerable advances in the elucidation of the physiological and pathophysiological roles of receptor family members. Furthermore, efficacy in animal models of neurological and psychiatric disorders has supported the progression of several glutamatergic ligands into clinical studies. These include ionotropic glutamate receptor antagonists, which have entered clinical trials for disorders including epilepsy and ischaemic stroke, alpha-amino-3-hydroxy-5-methyl-4-isoazolepropionic acid (AMPA) receptor positive allosteric modulators which are under evaluation as cognitive enhancers, and metabotropic glutamate receptor 2 (mGluR2) agonists which are undergoing clinical evaluation as anxiolytics. Furthermore, preclinical studies have illustrated therapeutic potential for ligands selective for other receptor subtypes in various disorders. These include mGluR1 antagonists in pain, mGluR5 antagonists in anxiety, pain and drug abuse and mGluR5 positive allosteric modulators in schizophrenia. Selective pharmacological tools have enabled the study of glutamate receptors. However, pharmacological coverage of the family is incomplete and considerable scope remains for the development of novel ligands, particularly those with in vivo utility, and for the their use together with existing tools for the further investigation of the roles of receptor family members in CNS function and as potentially novel therapeutics.

Neuroprotective mechanism of the novel melatonin derivative Neu-P11 in brain ischemia related models.

PubMed

Buendia, Izaskun; Gómez-Rangel, Vanessa; González-Lafuente, Laura; Parada, Esther; León, Rafael; Gameiro, Isabel; Michalska, Patrycja; Laudon, Moshe; Egea, Javier; López, Manuela G

2015-12-01

Stopping the ischemic cascade by targeting its components is a potential strategy for acute ischemic stroke treatment. During ischemia and especially over reperfusion, oxidative stress plays a major role in causing neuronal cell death. Melatonin has been previously reported to provide neuroprotective effects in in vivo models of stroke by a mechanism that implicates melatonin receptors. In this context, this study was planned to test the potential neuroprotective effects of the novel melatonin MT1/MT2 receptor agonist, Neu-P11, against brain ischemia in in vitro and in vivo models, and to elucidate its underlying mechanism of action. Neu-P11 proved to be a good antioxidant, to protect against glutamate-induced excitotoxicity and oxygen and glucose deprivation in hippocampal slices, and to reduce infarct volume in an in vivo stroke model. Regarding its mechanism of action, the protective effect of Neu-P11 was reverted by luzindole (melatonin receptor antagonist), AG490 (JAK2 inhibitor), LY294002 (PI3/AKT inhibitor) and PD98059 (MEK/ERK1/2 inhibitor). In conclusion, Neu-P11 affords neuroprotection against brain ischemia in in vitro and in vivo models by activating a pro-survival signaling pathway that involves melatonin receptors, JAK/STAT, PI3K/Akt and MEK/ERK1/2. Copyright © 2015 Elsevier Ltd. All rights reserved.

Integrated analysis reveals that STAT3 is central to the crosstalk between HER/ErbB receptor signaling pathways in human mammary epithelial cells

DOE PAGES

Gong, Chunhong; Zhang, Yi; Shankaran, Harish; ...

2014-10-02

Human epidermal growth factor receptors (HER, also known as ErbB) drive cellular proliferation, pro-survival and stress responses by activating several downstream kinases, in particular ERK, p38, JNK (SAPK), the PI3K/AKT, as well as various transcriptional regulators such as STAT3. When co-expressed, first three members of HER family (HER1-3) can form homo- and hetero-dimers. Based on the considerable evidence which suggest that every receptor dimer activates intracellular signaling pathways differentially, we hypothesized that the HER dimerization pattern is a better predictor of downstream signaling than the total receptor activation levels. We validated our hypothesis using a combination of model-based analysis tomore » quantify the HER dimerization patterns and multi-factorial experiments where HER dimerization patterns and signaling crosstalk were rationally perturbed. We have measured the activation of HER1-3 receptors and of the sentinel signaling proteins ERK, AKT, p38, JNK, STAT3 as a function of time in a panel of human mammary epithelial (HME) cells expressing different levels of HER1-3 stimulated with various ligand combinations. Our analysis using multiple ways of clustering the activation data has confirmed that the HER receptor dimer is a better predictor of the signaling through p38, ERK and AKT pathways than the total HER receptor expression and activation levels. Targeted inhibition studies to identify the causal effects allowed us to obtain a consensus regulatory interaction model, which revealed that STAT3 occupies a central role in the crosstalk between the studied pathways.« less

Rescue of deficient amygdala tonic γ-aminobutyric acidergic currents in the Fmr-/y mouse model of fragile X syndrome by a novel γ-aminobutyric acid type A receptor-positive allosteric modulator.

PubMed

Martin, Brandon S; Martinez-Botella, Gabriel; Loya, Carlos M; Salituro, Francesco G; Robichaud, Albert J; Huntsman, Molly M; Ackley, Mike A; Doherty, James J; Corbin, Joshua G

2016-06-01

Alterations in the ratio of excitatory to inhibitory transmission are emerging as a common component of many nervous system disorders, including autism spectrum disorders (ASDs). Tonic γ-aminobutyric acidergic (GABAergic) transmission provided by peri- and extrasynaptic GABA type A (GABAA ) receptors powerfully controls neuronal excitability and plasticity and, therefore, provides a rational therapeutic target for normalizing hyperexcitable networks across a variety of disorders, including ASDs. Our previous studies revealed tonic GABAergic deficits in principal excitatory neurons in the basolateral amygdala (BLA) in the Fmr1(-/y) knockout (KO) mouse model fragile X syndrome. To correct amygdala deficits in tonic GABAergic neurotransmission in Fmr1(-/y) KO mice, we developed a novel positive allosteric modulator of GABAA receptors, SGE-872, based on endogenously active neurosteroids. This study shows that SGE-872 is nearly as potent and twice as efficacious for positively modulating GABAA receptors as its parent molecule, allopregnanolone. Furthermore, at submicromolar concentrations (≤1 μM), SGE-872 is selective for tonic, extrasynaptic α4β3δ-containing GABAA receptors over typical synaptic α1β2γ2 receptors. We further find that SGE-872 strikingly rescues the tonic GABAergic transmission deficit in principal excitatory neurons in the Fmr1(-/y) KO BLA, a structure heavily implicated in the neuropathology of ASDs. Therefore, the potent and selective action of SGE-872 on tonic GABAA receptors containing α4 subunits may represent a novel and highly useful therapeutic avenue for ASDs and related disorders involving hyperexcitability of neuronal networks. © 2015 Wiley Periodicals, Inc.

Antiseizure Activity of Midazolam in Mice Lacking δ-Subunit Extrasynaptic GABA(A) Receptors.

PubMed

Reddy, Sandesh D; Younus, Iyan; Clossen, Bryan L; Reddy, Doodipala Samba

2015-06-01

Midazolam is a benzodiazepine anticonvulsant with rapid onset and short duration of action. Midazolam is the current drug of choice for acute seizures and status epilepticus, including those caused by organophosphate nerve agents. The antiseizure activity of midazolam is thought to result from its allosteric potentiation of synaptic GABA(A) receptors in the brain. However, there are indications that benzodiazepines promote neurosteroid synthesis via the 18-kDa cholesterol transporter protein (TSPO). Therefore, we investigated the role of neurosteroids and their extrasynaptic GABA(A) receptor targets in the antiseizure activity of midazolam. Here, we used δ-subunit knockout (DKO) mice bearing a targeted deletion of the extrasynaptic receptors to investigate the contribution of the extrasynaptic receptors to the antiseizure activity of midazolam using the 6-Hz and hippocampus kindling seizure models. In both models, midazolam produced rapid and dose-dependent protection against seizures (ED50, 0.4 mg/kg). Moreover, the antiseizure potency of midazolam was undiminished in DKO mice compared with control mice. Pretreatment with PK11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide], a TSPO blocker, or finasteride, a 5α-reductase neurosteroid inhibitor, did not affect the antiseizure effect of midazolam. The antiseizure activity of midazolam was significantly reversed by pretreatment with flumazenil, a benzodiazepine antagonist. Plasma and brain levels of the neurosteroid allopregnanolone were not significantly greater in midazolam-treated animals. These studies therefore provide strong evidence that neurosteroids and extrasynaptic GABA(A) receptors are not involved in the antiseizure activity of midazolam, which mainly occurs through synaptic GABA(A) receptors via direct binding to benzodiazepine sites. This study reaffirms midazolam's use for controlling acute seizures and status epilepticus. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Antiseizure Activity of Midazolam in Mice Lacking δ-Subunit Extrasynaptic GABAA Receptors

PubMed Central

Reddy, Sandesh D.; Younus, Iyan; Clossen, Bryan L.

2015-01-01

Midazolam is a benzodiazepine anticonvulsant with rapid onset and short duration of action. Midazolam is the current drug of choice for acute seizures and status epilepticus, including those caused by organophosphate nerve agents. The antiseizure activity of midazolam is thought to result from its allosteric potentiation of synaptic GABAA receptors in the brain. However, there are indications that benzodiazepines promote neurosteroid synthesis via the 18-kDa cholesterol transporter protein (TSPO). Therefore, we investigated the role of neurosteroids and their extrasynaptic GABAA receptor targets in the antiseizure activity of midazolam. Here, we used δ-subunit knockout (DKO) mice bearing a targeted deletion of the extrasynaptic receptors to investigate the contribution of the extrasynaptic receptors to the antiseizure activity of midazolam using the 6-Hz and hippocampus kindling seizure models. In both models, midazolam produced rapid and dose-dependent protection against seizures (ED50, 0.4 mg/kg). Moreover, the antiseizure potency of midazolam was undiminished in DKO mice compared with control mice. Pretreatment with PK11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide], a TSPO blocker, or finasteride, a 5α-reductase neurosteroid inhibitor, did not affect the antiseizure effect of midazolam. The antiseizure activity of midazolam was significantly reversed by pretreatment with flumazenil, a benzodiazepine antagonist. Plasma and brain levels of the neurosteroid allopregnanolone were not significantly greater in midazolam-treated animals. These studies therefore provide strong evidence that neurosteroids and extrasynaptic GABAA receptors are not involved in the antiseizure activity of midazolam, which mainly occurs through synaptic GABAA receptors via direct binding to benzodiazepine sites. This study reaffirms midazolam’s use for controlling acute seizures and status epilepticus. PMID:25784648

Mechanistic analysis of the function of agonists and allosteric modulators: reconciling two-state and operational models

PubMed Central

Roche, David; Gil, Debora; Giraldo, Jesús

2013-01-01

Two-state and operational models of both agonism and allosterism are compared to identify and characterize common pharmacological parameters. To account for the receptor-dependent basal response, constitutive receptor activity is considered in the operational models. By arranging two-state models as the fraction of active receptors and operational models as the fractional response relative to the maximum effect of the system, a one-by-one correspondence between parameters is found. The comparative analysis allows a better understanding of complex allosteric interactions. In particular, the inclusion of constitutive receptor activity in the operational model of allosterism allows the characterization of modulators able to lower the basal response of the system; that is, allosteric modulators with negative intrinsic efficacy. Theoretical simulations and overall goodness of fit of the models to simulated data suggest that it is feasible to apply the models to experimental data and constitute one step forward in receptor theory formalism. Linked Articles Another recent review on allosteric modulation can be found at: Kenakin, T (2013). New concepts in pharmacological efficacy at 7TM receptors: IUPHAR Review 2. British Journal of Pharmacology 168: 554–575. doi: 10.1111/j.1476-5381.2012.02223.x And in this issue of BJP there is an article on a new allosteric modulator: Newman AS, Batis N, Grafton G, Caputo F, Brady CA, Lambert J, Peters JA, Gordon J, Brain KL, Powell AD and Barnes NM (2013). 5-Chloroindole: a potent allosteric modulator of the 5-HT3 receptor. British Journal of Pharmacology 169: 1228–1238. doi: 10.1111/bph.12213 PMID:23647200

Computer-aided design of negative allosteric modulators of metabotropic glutamate receptor 5 (mGluR5): Comparative molecular field analysis of aryl ether derivatives.

PubMed

Selvam, Chelliah; Thilagavathi, Ramasamy; Narasimhan, Balasubramanian; Kumar, Pradeep; Jordan, Brian C; Ranganna, Kasturi

2016-02-15

The metabotropic glutamate receptors (mGlu receptors) have emerged as attractive targets for number of neurological and psychiatric disorders. Recently, mGluR5 negative allosteric modulators (NAMs) have gained considerable attention in pharmacological research. Comparative molecular field analysis (CoMFA) was performed on 73 analogs of aryl ether which were reported as mGluR5 NAMs. The study produced a statistically significant model with high correlation coefficient and good predictive abilities. Published by Elsevier Ltd.

Structure-Based Approach To Identify 5-[4-Hydroxyphenyl]pyrrole-2-carbonitrile Derivatives as Potent and Tissue Selective Androgen Receptor Modulators.

PubMed

Unwalla, Ray; Mousseau, James J; Fadeyi, Olugbeminiyi O; Choi, Chulho; Parris, Kevin; Hu, Baihua; Kenney, Thomas; Chippari, Susan; McNally, Christopher; Vishwanathan, Karthick; Kilbourne, Edward; Thompson, Catherine; Nagpal, Sunil; Wrobel, Jay; Yudt, Matthew; Morris, Carl A; Powell, Dennis; Gilbert, Adam M; Chekler, Eugene L Piatnitski

2017-07-27

In an effort to find new and safer treatments for osteoporosis and frailty, we describe a novel series of selective androgen receptor modulators (SARMs). Using a structure-based approach, we identified compound 7, a potent AR (ARE EC 50 = 0.34 nM) and selective (N/C interaction EC 50 = 1206 nM) modulator. In vivo data, an AR LBD X-ray structure of 7, and further insights from modeling studies of ligand receptor interactions are also presented.

Impacts of Lateral Boundary Conditions on US Ozone ...

EPA Pesticide Factsheets

Chemical boundary conditions are a key input to regional-scale photochemical models. In this study, we perform annual simulations over North America with chemical boundary conditions prepared from two global models (GEOS-CHEM and Hemispheric CMAQ). Results indicate that the impacts of different boundary conditions on ozone can be significant throughout the year. The National Exposure Research Laboratory (NERL) Computational Exposure Division (CED) develops and evaluates data, decision-support tools, and models to be applied to media-specific or receptor-specific problem areas. CED uses modeling-based approaches to characterize exposures, evaluate fate and transport, and support environmental diagnostics/forensics with input from multiple data sources. It also develops media- and receptor-specific models, process models, and decision support tools for use both within and outside of EPA.

Water quality assessment and apportionment of pollution sources using APCS-MLR and PMF receptor modeling techniques in three major rivers of South Florida.

PubMed

Haji Gholizadeh, Mohammad; Melesse, Assefa M; Reddi, Lakshmi

2016-10-01

In this study, principal component analysis (PCA), factor analysis (FA), and the absolute principal component score-multiple linear regression (APCS-MLR) receptor modeling technique were used to assess the water quality and identify and quantify the potential pollution sources affecting the water quality of three major rivers of South Florida. For this purpose, 15years (2000-2014) dataset of 12 water quality variables covering 16 monitoring stations, and approximately 35,000 observations was used. The PCA/FA method identified five and four potential pollution sources in wet and dry seasons, respectively, and the effective mechanisms, rules and causes were explained. The APCS-MLR apportioned their contributions to each water quality variable. Results showed that the point source pollution discharges from anthropogenic factors due to the discharge of agriculture waste and domestic and industrial wastewater were the major sources of river water contamination. Also, the studied variables were categorized into three groups of nutrients (total kjeldahl nitrogen, total phosphorus, total phosphate, and ammonia-N), water murkiness conducive parameters (total suspended solids, turbidity, and chlorophyll-a), and salt ions (magnesium, chloride, and sodium), and average contributions of different potential pollution sources to these categories were considered separately. The data matrix was also subjected to PMF receptor model using the EPA PMF-5.0 program and the two-way model described was performed for the PMF analyses. Comparison of the obtained results of PMF and APCS-MLR models showed that there were some significant differences in estimated contribution for each potential pollution source, especially in the wet season. Eventually, it was concluded that the APCS-MLR receptor modeling approach appears to be more physically plausible for the current study. It is believed that the results of apportionment could be very useful to the local authorities for the control and management of pollution and better protection of important riverine water quality. Copyright © 2016 Elsevier B.V. All rights reserved.

(−) Arctigenin and (+) Pinoresinol Are Antagonists of the Human Thyroid Hormone Receptor β

PubMed Central

2015-01-01

Lignans are important biologically active dietary polyphenolic compounds. Consumption of foods that are rich in lignans is associated with positive health effects. Using modeling tools to probe the ligand-binding pockets of molecular receptors, we found that lignans have high docking affinity for the human thyroid hormone receptor β. Follow-up experimental results show that lignans (−) arctigenin and (+) pinoresinol are antagonists of the human thyroid hormone receptor β. The modeled complexes show key plausible interactions between the two ligands and important amino acid residues of the receptor. PMID:25383984

Extraintestinal roles of bombesin-like peptides and their receptors: lung.

PubMed

Qin, Xiao-Qun; Qu, Xiangping

2013-02-01

Description of the recent findings of the biological roles of bombesin-like peptides and their receptors in lungs. Gastrin-releasing peptide (GRP) was involved in the airway inflammation in murine models of airway hyperreactivity. The circulating proGRP could serve as a valuable tumor marker for small-cell lung cancers, and the plasma level of proGRP is more stable compared with that of serum proGRP. Recent studies also shed light on the intracellular signaling pathways of bombesin receptor subtype-3 (BRS-3) activation in cultured human lung cancer cells. The relevant biology of BLPs and their receptors in lung cancers and other lung diseases still remains largely unknown. With the development of several highly specific BRS-3 agonists, recent studies provided some insights into the biological effects of BRS-3 in lungs.

Identification of the kainate receptor subunits underlying modulation of excitatory synaptic transmission in the CA3 region of the hippocampus.

PubMed

Contractor, A; Swanson, G T; Sailer, A; O'Gorman, S; Heinemann, S F

2000-11-15

To understand the physiological role of kainate receptors and their participation in seizure induction in animal models of epilepsy, it will be necessary to develop a comprehensive description of their action in the CA3 region of the hippocampus. Activation of presynaptic kainate receptors depresses excitatory synaptic transmission at mossy fiber and associational-commissural inputs to CA3 pyramidal neurons (Vignes et al., 1998; Bortolotto et al., 1999; Kamiya and Ozawa, 2000). In this study, we use gene-targeted mice lacking glutamate receptor 5 (GluR5) or GluR6 kainate receptor subunits to identify the receptor subunits that comprise the kainate receptors responsible for presynaptic modulation of CA3 transmission. We found that bath application of kainate (3 microm) profoundly reduced EPSCs at mossy fiber and collateral synapses in neurons from wild-type and GluR5(-/-) mice but had no effect on EPSCs in neurons from GluR6(-/-) mice. These results therefore contrast with previous studies that supported a role for GluR5-containing receptors at mossy fiber and associational-commissural synapses (Vignes et al., 1998; Bortolotto et al., 1999). Surprisingly, at perforant path synapses kainate receptor activation enhanced transmission; this potentiation was abolished in both GluR5 and GluR6 knock-out mice. Kainate receptors thus play multiple and complex roles to modulate excitatory synaptic transmission in the CA3 region of the hippocampus.

New techniques for positron emission tomography in the study of human neurological disorders: Progress report, 15 June 1992--31 October 1992

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

1992-01-01

During the past six months, we have continued work on the fronts of kinetic modeling of radioligands for studying neurotransmitter/receptor systems, iterative reconstruction techniques, and methodology for PET cerebral blood flow activation studies. Initial human PET studies have been performed and analyzed with many different kinetic model formulations to determine the quantitative potential of the neuronwsmitter/receptor ligand, ({sup 11}C)N-methyl piperidyl benzilate (NMPB), a muscarinic cholinergic antagonist. In addition, initial human studies using ({sup 11}C)tetrabenazine (TBZ), a marker for monoantine nerve terminal density. Results of the NWB studies have indicated that this new agent yields better estimates of receptor density thanmore » previous muscarinic ligands developed at our facility, ({sup 11}C)-TRB and ({sup 11}C)scopolamine. TRB and scopolamine have previously been shown to be only partially successful ligands due to sub-optimal values of the individual rate constants, causing varying degrees of flow limitation. This is found to be much less of a problem for NMPB due to the 2.0--2.5 fold increase in ligand transport observed in the human studies ({approximately}60% first pass extraction). A 2-parameter 2-compartment simplification had previously been implemented for the benzodiazepine ligand, (C-11)FMZ, and a similar model appears to be suitable for TBZ based on the preliminary human data.« less

New techniques for positron emission tomography in the study of human neurological disorders: Progress report, 15 June 1992--31 October 1992

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

1992-10-01

During the past six months, we have continued work on the fronts of kinetic modeling of radioligands for studying neurotransmitter/receptor systems, iterative reconstruction techniques, and methodology for PET cerebral blood flow activation studies. Initial human PET studies have been performed and analyzed with many different kinetic model formulations to determine the quantitative potential of the neuronwsmitter/receptor ligand, [{sup 11}C]N-methyl piperidyl benzilate (NMPB), a muscarinic cholinergic antagonist. In addition, initial human studies using [{sup 11}C]tetrabenazine (TBZ), a marker for monoantine nerve terminal density. Results of the NWB studies have indicated that this new agent yields better estimates of receptor density thanmore » previous muscarinic ligands developed at our facility, [{sup 11}C]-TRB and [{sup 11}C]scopolamine. TRB and scopolamine have previously been shown to be only partially successful ligands due to sub-optimal values of the individual rate constants, causing varying degrees of flow limitation. This is found to be much less of a problem for NMPB due to the 2.0--2.5 fold increase in ligand transport observed in the human studies ({approximately}60% first pass extraction). A 2-parameter 2-compartment simplification had previously been implemented for the benzodiazepine ligand, [C-11]FMZ, and a similar model appears to be suitable for TBZ based on the preliminary human data.« less

An evolutionary switch in ND2 enables Src kinase regulation of NMDA receptors

NASA Astrophysics Data System (ADS)

Scanlon, David P.; Bah, Alaji; Krzeminski, Mickaël; Zhang, Wenbo; Leduc-Pessah, Heather L.; Dong, Yi Na; Forman-Kay, Julie D.; Salter, Michael W.

2017-05-01

The non-receptor tyrosine kinase Src is a key signalling hub for upregulating the function of N-methyl D-aspartate receptors (NMDARs). Src is anchored within the NMDAR complex via NADH dehydrogenase subunit 2 (ND2), a mitochondrially encoded adaptor protein. The interacting regions between Src and ND2 have been broadly identified, but the interaction between ND2 and the NMDAR has remained elusive. Here we generate a homology model of ND2 and dock it onto the NMDAR via the transmembrane domain of GluN1. This interaction is enabled by the evolutionary loss of three helices in bilaterian ND2 proteins compared to their ancestral homologues. We experimentally validate our model and demonstrate that blocking this interaction with an ND2 fragment identified in our experimental studies prevents Src-mediated upregulation of NMDAR currents in neurons. Our findings establish the mode of interaction between an NMDAR accessory protein with one of the core subunits of the receptor.

Functional proteomics outlines the complexity of breast cancer molecular subtypes.

PubMed

Gámez-Pozo, Angelo; Trilla-Fuertes, Lucía; Berges-Soria, Julia; Selevsek, Nathalie; López-Vacas, Rocío; Díaz-Almirón, Mariana; Nanni, Paolo; Arevalillo, Jorge M; Navarro, Hilario; Grossmann, Jonas; Gayá Moreno, Francisco; Gómez Rioja, Rubén; Prado-Vázquez, Guillermo; Zapater-Moros, Andrea; Main, Paloma; Feliú, Jaime; Martínez Del Prado, Purificación; Zamora, Pilar; Ciruelos, Eva; Espinosa, Enrique; Fresno Vara, Juan Ángel

2017-08-30

Breast cancer is a heterogeneous disease comprising a variety of entities with various genetic backgrounds. Estrogen receptor-positive, human epidermal growth factor receptor 2-negative tumors typically have a favorable outcome; however, some patients eventually relapse, which suggests some heterogeneity within this category. In the present study, we used proteomics and miRNA profiling techniques to characterize a set of 102 either estrogen receptor-positive (ER+)/progesterone receptor-positive (PR+) or triple-negative formalin-fixed, paraffin-embedded breast tumors. Protein expression-based probabilistic graphical models and flux balance analyses revealed that some ER+/PR+ samples had a protein expression profile similar to that of triple-negative samples and had a clinical outcome similar to those with triple-negative disease. This probabilistic graphical model-based classification had prognostic value in patients with luminal A breast cancer. This prognostic information was independent of that provided by standard genomic tests for breast cancer, such as MammaPrint, OncoType Dx and the 8-gene Score.

Discovery of Indazoles as Potent, Orally Active Dual Neurokinin 1 Receptor Antagonists and Serotonin Transporter Inhibitors for the Treatment of Depression.

PubMed

Degnan, Andrew P; Tora, George O; Huang, Hong; Conlon, David A; Davis, Carl D; Hanumegowda, Umesh M; Hou, Xiaoping; Hsiao, Yi; Hu, Joanna; Krause, Rudolph; Li, Yu-Wen; Newton, Amy E; Pieschl, Rick L; Raybon, Joseph; Rosner, Thorsten; Sun, Jung-Hui; Taber, Matthew T; Taylor, Sarah J; Wong, Michael K; Zhang, Huiping; Lodge, Nicholas J; Bronson, Joanne J; Macor, John E; Gillman, Kevin W

2016-12-21

Combination studies of neurokinin 1 (NK1) receptor antagonists and serotonin-selective reuptake inhibitors (SSRIs) have shown promise in preclinical models of depression. Such a combination may offer important advantages over the current standard of care. Herein we describe the discovery and optimization of an indazole-based chemotype to provide a series of potent dual NK1 receptor antagonists/serotonin transporter (SERT) inhibitors to overcome issues of ion channel blockade. This effort culminated in the identification of compound 9, an analogue that demonstrated favorable oral bioavailability, excellent brain uptake, and robust in vivo efficacy in a validated depression model. Over the course of this work, a novel heterocycle-directed asymmetric hydrogenation was developed to facilitate installation of the key stereogenic center.

Estimating Air-Manganese Exposures in Two Ohio Towns ...

EPA Pesticide Factsheets

Manganese (Mn), a nutrient required for normal metabolic function, is also a persistent air pollutant and a known neurotoxin at high concentrations. Elevated exposures can result in a number of motor and cognitive deficits. Quantifying chronic personal exposures in residential populations studied by environmental epidemiologists can be time-consuming and expensive. We developed an approach for quantifying chronic exposures for two towns (Marietta and East Liverpool, Ohio) with elevated air Mn concentrations (air-Mn) related to ambient emissions from industrial processes. This was accomplished through the use of measured and modeled data in the communities studied. A novel approach was developed because one of the facilities lacked emissions data for the purposes of modeling. A unit emission rate was assumed over the surface area of both source facilities, and offsite concentrations at receptor residences and air monitoring sites were estimated with the American Meteorological Society/Environmental Protection Agency Regulatory Model (AERMOD). Ratios of all modeled receptor points were created, and a long-running air monitor was identified as a reference location. All ratios were normalized to the reference location. Long-term averages at all residential receptor points were calculated using modeled ratios and data from the reference monitoring location. Modeled five-year average air-Mn exposures ranged from 0.03-1.61 µg/m3 in Marietta and 0.01-6.32 µg/m3 in E

Quantification of source impact to PM using three-dimensional weighted factor model analysis on multi-site data

NASA Astrophysics Data System (ADS)

Shi, Guoliang; Peng, Xing; Huangfu, Yanqi; Wang, Wei; Xu, Jiao; Tian, Yingze; Feng, Yinchang; Ivey, Cesunica E.; Russell, Armistead G.

2017-07-01

Source apportionment technologies are used to understand the impacts of important sources of particulate matter (PM) air quality, and are widely used for both scientific studies and air quality management. Generally, receptor models apportion speciated PM data from a single sampling site. With the development of large scale monitoring networks, PM speciation are observed at multiple sites in an urban area. For these situations, the models should account for three factors, or dimensions, of the PM, including the chemical species concentrations, sampling periods and sampling site information, suggesting the potential power of a three-dimensional source apportionment approach. However, the principle of three-dimensional Parallel Factor Analysis (Ordinary PARAFAC) model does not always work well in real environmental situations for multi-site receptor datasets. In this work, a new three-way receptor model, called "multi-site three way factor analysis" model is proposed to deal with the multi-site receptor datasets. Synthetic datasets were developed and introduced into the new model to test its performance. Average absolute error (AAE, between estimated and true contributions) for extracted sources were all less than 50%. Additionally, three-dimensional ambient datasets from a Chinese mega-city, Chengdu, were analyzed using this new model to assess the application. Four factors are extracted by the multi-site WFA3 model: secondary source have the highest contributions (64.73 and 56.24 μg/m3), followed by vehicular exhaust (30.13 and 33.60 μg/m3), crustal dust (26.12 and 29.99 μg/m3) and coal combustion (10.73 and 14.83 μg/m3). The model was also compared to PMF, with general agreement, though PMF suggested a lower crustal contribution.

E-type prostanoid receptor 4 (EP4) in disease and therapy

PubMed Central

Konya, Viktoria; Marsche, Gunther; Schuligoi, Rufina; Heinemann, Akos

2013-01-01

The large variety of biological functions governed by prostaglandin (PG) E2 is mediated by signaling through four distinct E-type prostanoid (EP) receptors. The availability of mouse strains with genetic ablation of each EP receptor subtype and the development of selective EP agonists and antagonists have tremendously advanced our understanding of PGE2 as a physiologically and clinically relevant mediator. Moreover, studies using disease models revealed numerous conditions in which distinct EP receptors might be exploited therapeutically. In this context, the EP4 receptor is currently emerging as most versatile and promising among PGE2 receptors. Anti-inflammatory, anti-thrombotic and vasoprotective effects have been proposed for the EP4 receptor, along with its recently described unfavorable tumor-promoting and pro-angiogenic roles. A possible explanation for the diverse biological functions of EP4 might be the multiple signaling pathways switched on upon EP4 activation. The present review attempts to summarize the EP4 receptor-triggered signaling modules and the possible therapeutic applications of EP4-selective agonists and antagonists. PMID:23523686

Interferons and Their Receptors in Birds: A Comparison of Gene Structure, Phylogenetic Analysis, and Cross Modulation

PubMed Central

Zhou, Hao; Chen, Shun; Wang, Mingshu; Cheng, Anchun

2014-01-01

Interferon may be thought of as a key, with the interferon receptor as the signal lock: Crosstalk between them maintains their balance during viral infection. In this review, the protein structure of avian interferon and the interferon receptor are discussed, indicating remarkable similarity between different species. However, the structures of the interferon receptors are more sophisticated than those of the interferons, suggesting that the interferon receptor is a more complicated signal lock system and has considerable diversity in subtypes or structures. Preliminary evolutionary analysis showed that the subunits of the interferon receptor formed a distinct clade, and the orthologs may be derived from the same ancestor. Furthermore, the development of interferons and interferon receptors in birds may be related to an animal’s age and the maintenance of a balanced state. In addition, the equilibrium between interferon and its receptor during pathological and physiological states revealed that the virus and the host influence this equilibrium. Birds could represent an important model for studies on interferon’s antiviral activities and may provide the basis for new antiviral strategies. PMID:25405736

The β3-adrenergic receptor is dispensable for browning of adipose tissues.

PubMed

de Jong, Jasper M A; Wouters, René T F; Boulet, Nathalie; Cannon, Barbara; Nedergaard, Jan; Petrovic, Natasa

2017-06-01

Brown and brite/beige adipocytes are attractive therapeutic targets to treat metabolic diseases. To maximally utilize their functional potential, further understanding is required about their identities and their functional differences. Recent studies with β 3 -adrenergic receptor knockout mice reported that brite/beige adipocytes, but not classical brown adipocytes, require the β 3 -adrenergic receptor for cold-induced transcriptional activation of thermogenic genes. We aimed to further characterize this requirement of the β 3 -adrenergic receptor as a functional distinction between classical brown and brite/beige adipocytes. However, when comparing wild-type and β 3 -adrenergic receptor knockout mice, we observed no differences in cold-induced thermogenic gene expression ( Ucp1 , Pgc1a , Dio2 , and Cidea ) in brown or white (brite/beige) adipose tissues. Irrespective of the duration of the cold exposure or the sex of the mice, we observed no effect of the absence of the β 3 -adrenergic receptor. Experiments with the β 3 -adrenergic receptor agonist CL-316,243 verified the functional absence of β 3 -adrenergic signaling in these knockout mice. The β 3 -adrenergic receptor knockout model in the present study was maintained on a FVB/N background, whereas earlier reports used C57BL/6 and 129Sv mice. Thus our data imply background-dependent differences in adrenergic signaling mechanisms in response to cold exposure. Nonetheless, the present data indicate that the β 3 -adrenergic receptor is dispensable for cold-induced transcriptional activation in both classical brown and, as opposed to earlier studies, brite/beige cells. Copyright © 2017 the American Physiological Society.

The tertiary structures of porcine AhR and ARNT proteins and molecular interactions within the TCDD/AhR/ARNT complex.

PubMed

Orlowska, Karina; Molcan, Tomasz; Swigonska, Sylwia; Sadowska, Agnieszka; Jablonska, Monika; Nynca, Anna; Jastrzebski, Jan P; Ciereszko, Renata E

2016-06-01

The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that can be activated by structurally diverse synthetic and natural chemicals, including toxic environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In the present study, homology models of the porcine AhR-ligand binding domain (LBD) and the porcine aryl hydrocarbon receptor nuclear translocator-ligand binding domain (ARNT-LBD) were created on the basis of structures of closely related respective proteins i.e., human Hif-2α and ARNT. Molecular docking of TCDD to the porcine AhR-LBD model revealed high binding affinity (-8.8kcal/mol) between TCDD and the receptor. Moreover, formation of the TCDD/AhR-LBD complex was confirmed experimentally with the use of electrophoretic mobility shift assay (EMSA). It was found that TCDD (10nM, 2h of incubation) not only bound to the AhR in the porcine granulosa cells but also activated the receptor. The current study provides a framework for examining the key events involved in the ligand-dependent activation of the AhR. Copyright © 2016 Elsevier Inc. All rights reserved.

Unveiling the molecular mechanism of brassinosteroids: Insights from structure-based molecular modeling studies.

PubMed

Lei, Beilei; Liu, Jiyuan; Yao, Xiaojun

2015-12-01

Brassinosteroid (BR) phytohormones play indispensable roles in plant growth and development. Brassinolide (BL) and 24-epibrassinolide (24-epiBL) are the most active ones among the BRs reported thus far. Unfortunately, the extremely low natural content and intricate synthesis process limit their popularization in agricultural production. Earlier reports to discover alternative compounds have resulted in molecules with nearly same scaffold structure and without diversity in chemical space. In the present study, receptors structure based BRs regulation mechanism was analyzed. First, we examined the detailed binding interactions and their dynamic stability between BL and its receptor BRI1 and co-receptor BAK1. Then, the binding modes and binding free energies for 24-epiBL and a series of representative BRs binding with BRI1 and BRI1-BAK1 were carried out by molecular docking, energy minimization and MM-PBSA free energy calculation. The obtained binding structures and energetic results provided vital insights into the structural factors affecting the activity from both receptors and BRs aspects. Subsequently, the obtained knowledge will serve as valuable guidance to build pharmacophore models for rational screening of new scaffold alternative BRs. Copyright © 2015 Elsevier Inc. All rights reserved.

The effect of fenoterol stereochemistry on the β2 adrenergic receptor system: ligand directed chiral recognition

PubMed Central

Jozwiak, Krzysztof; Plazinska, Anita; Toll, Lawrence; Jimenez, Lucita; Woo, Anthony Yiu-Ho; Xiao, Rui-Ping; Wainer, Irving W.

2011-01-01

The β2 adrenergic receptor (β2-AR) is a model system for studying the ligand recognition process in G-protein coupled receptors. Fenoterol (FEN) is a β2-AR selective agonist that has two centers of chirality and exists as four stereoisomers. Radioligand binding studies determined that stereochemistry greatly influences the binding affinity. Subsequent Van’t Hoff analysis shows very different thermodynamics of binding depending on the stereoconfiguration of the molecule. The binding of (S,x’)-isomers is almost entirely enthalpy controlled whereas binding of (R,x’)-isomers is purely entropy driven. Stereochemistry of FEN molecule also affects the coupling of the receptor to different G proteins. In a rat cardiomyocyte contractility model, (R,R’)-FEN was shown to selectively activate Gs protein signaling while the (S,R’)- isomer activated both Gi and Gs protein. The overall data demonstrate that the chirality at the two chiral centers of the FEN molecule influences the magnitude of binding affinity, thermodynamics of local interactions within the binding site and the global mechanism of β2-AR activation. Differences in thermodynamic parameters and non-uniform G-protein coupling suggest a mechanism of chiral recognition in which observed enantioselectivities arise from the interaction of the (R,x’)-FEN stereoisomers with a different receptor conformation than the one with which the (S,x’)-isomer interacts. PMID:21618615

Effect of fenoterol stereochemistry on the β2 adrenergic receptor system: ligand-directed chiral recognition.

PubMed

Jozwiak, Krzysztof; Plazinska, Anita; Toll, Lawrence; Jimenez, Lucita; Woo, Anthony Yiu-Ho; Xiao, Rui-Ping; Wainer, Irving W

2011-01-01

The β(2) adrenergic receptor (β(2)-AR) is a model system for studying the ligand recognition process in G protein-coupled receptors. Fenoterol (FEN) is a β(2)-AR selective agonist that has two centers of chirality and exists as four stereoisomers. Radioligand binding studies determined that stereochemistry greatly influences the binding affinity. Subsequent Van't Hoff analysis shows very different thermodynamics of binding depending on the stereoconfiguration of the molecule. The binding of (S,x')-isomers is almost entirely enthalpy controlled whereas binding of (R,x')-isomers is purely entropy driven. Stereochemistry of FEN molecule also affects the coupling of the receptor to different G proteins. In a rat cardiomyocyte contractility model, (R,R')-FEN was shown to selectively activate G(s) protein signaling while the (S,R')-isomer activated both G(i) and G(s) protein. The overall data demonstrate that the chirality at the two chiral centers of the FEN molecule influences the magnitude of binding affinity, thermodynamics of local interactions within the binding site, and the global mechanism of β(2)-AR activation. Differences in thermodynamic parameters and nonuniform G-protein coupling suggest a mechanism of chiral recognition in which observed enantioselectivities arise from the interaction of the (R,x')-FEN stereoisomers with a different receptor conformation than the one with which the (S,x')-isomer interacts. Copyright © 2011 Wiley-Liss, Inc.

Effects of electroacupuncture on the levels of retinal gamma-aminobutyric acid and its receptors in a guinea pig model of lens-induced myopia.

PubMed

Sha, F; Ye, X; Zhao, W; Xu, C-L; Wang, L; Ding, M-H; Bi, A-L; Wu, J-F; Jiang, W-J; Guo, D-D; Guo, J-G; Bi, H-S

2015-02-26

Gamma-aminobutyric acid (GABA) is a major inhibitory neurotransmitter of the retina and affects myopic development. Electroacupuncture (EA) is widely utilized to treat myopia in clinical settings. However, there are few reports on whether EA affects the level of retinal GABA during myopic development. To study this issue, in the present study, we explored the changes of retinal GABA content and the expression of its receptor subtypes, and the effects of EA stimulation on them in a guinea pig model with lens-induced myopia (LIM). Our results showed that the content of GABA and the expression of GABAA and GABAC receptors of retina were up-regulated during the development of myopia, and this up-regulation was inhibited by applying EA to Hegu (LI4) and Taiyang (EX-HN5) acupoints. Moreover, these effects of EA show a positional specificity. While applying EA at a sham acupoint, no apparent change of myopic retinal GABA and its receptor subtypes was observed. Taken together, our findings suggest that LIM is effective to up-regulate the level of retinal GABA, GABAA and GABAC receptors in guinea pigs and the effect may be inhibited by EA stimulation at LI4 and EX-HN5 acupoints. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

Homology Modeling, Validation and Dynamics of the G Protein-coupled Estrogen Receptor 1 (GPER-1).

PubMed

Bruno, Agostino; Aiello, Francesca; Costantino, Gabriele; Radi, Marco

2016-09-01

Estrogens exert their action mainly by binding three receptors, namely estrogen receptors α and β (ERα and ERβ) and GPER-1 (G-protein coupled estrogen receptor 1). While the patho-physiological role of both ERα and ERβ has been deeply investigated, the role of GPER-1 in estrogens' signaling has not been clearly defined yet. Unfortunately, only few GPER-1 selective ligands were discovered so far, and the real efficiency of such compounds is still matter of debate. To better understand the physiological relevance of GPER-1, new selective chemical probes are higly needed. In this scenario, we report herein the generation and validation of a three-dimensional (3-D) GPER-1 homology model by means of docking studies and molecular dynamics simulations. The model thus generated was employed to (i) decipher the structural basis underlying the ability of estrogens and some Selective Estrogen Receptor Modulators (SERMs) to bind GPER-1 and classical ERα and ERβ, and (ii) generate a reliable G1/GPER-1 complex useful in rationalizing the pharmacological profile of G1 reported in the literature. The G1/GPER-1 complex herein reported could be further exploited in drug design approaches aimed at improving the pharmacological profile of G1 or at identifying new chemical entities (NCEs) as potential modulators of GPER-1. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Immunization with the Haemophilus ducreyi hemoglobin receptor HgbA protects against infection in the swine model of chancroid.

PubMed

Afonina, Galyna; Leduc, Isabelle; Nepluev, Igor; Jeter, Chrystina; Routh, Patty; Almond, Glen; Orndorff, Paul E; Hobbs, Marcia; Elkins, Christopher

2006-04-01

The etiologic agent of chancroid is Haemophilus ducreyi. To fulfill its obligate requirement for heme, H. ducreyi uses two TonB-dependent receptors: the hemoglobin receptor (HgbA) and a receptor for free heme (TdhA). Expression of HgbA is necessary for H. ducreyi to survive and initiate disease in a human model of chancroid. In this study, we used a swine model of H. ducreyi infection to demonstrate that an experimental HgbA vaccine efficiently prevents chancroid, as determined by several parameters. Histological sections of immunized animals lacked typical microscopic features of chancroid. All inoculated sites from mock-immunized pigs yielded viable H. ducreyi cells, whereas no viable H. ducreyi cells were recovered from inoculated sites of HgbA-immunized pigs. Antibodies from sera of HgbA-immunized animals bound to and initiated antibody-dependent bactericidal activity against homologous H. ducreyi strain 35000HP and heterologous strain CIP542 ATCC; however, an isogenic hgbA mutant of 35000HP was not killed, proving specificity. Anti-HgbA immunoglobulin G blocked hemoglobin binding to the HgbA receptor, suggesting a novel mechanism of protection through the limitation of heme/iron acquisition by H. ducreyi. Such a vaccine strategy might be applied to other bacterial pathogens with strict heme/iron requirements. Taken together, these data suggest continuing the development of an HgbA subunit vaccine to prevent chancroid.

Ligand and structure-based methodologies for the prediction of the activity of G protein-coupled receptor ligands

NASA Astrophysics Data System (ADS)

Costanzi, Stefano; Tikhonova, Irina G.; Harden, T. Kendall; Jacobson, Kenneth A.

2009-11-01

Accurate in silico models for the quantitative prediction of the activity of G protein-coupled receptor (GPCR) ligands would greatly facilitate the process of drug discovery and development. Several methodologies have been developed based on the properties of the ligands, the direct study of the receptor-ligand interactions, or a combination of both approaches. Ligand-based three-dimensional quantitative structure-activity relationships (3D-QSAR) techniques, not requiring knowledge of the receptor structure, have been historically the first to be applied to the prediction of the activity of GPCR ligands. They are generally endowed with robustness and good ranking ability; however they are highly dependent on training sets. Structure-based techniques generally do not provide the level of accuracy necessary to yield meaningful rankings when applied to GPCR homology models. However, they are essentially independent from training sets and have a sufficient level of accuracy to allow an effective discrimination between binders and nonbinders, thus qualifying as viable lead discovery tools. The combination of ligand and structure-based methodologies in the form of receptor-based 3D-QSAR and ligand and structure-based consensus models results in robust and accurate quantitative predictions. The contribution of the structure-based component to these combined approaches is expected to become more substantial and effective in the future, as more sophisticated scoring functions are developed and more detailed structural information on GPCRs is gathered.

PUFA diets alter the microRNA expression profiles in an inflammation rat model

PubMed Central

ZHENG, ZHENG; GE, YINLIN; ZHANG, JINYU; XUE, MEILAN; LI, QUAN; LIN, DONGLIANG; MA, WENHUI

2015-01-01

Omega-3 and -6 polyunsaturated fatty acids (PUFAs) can directly or indirectly regulate immune homeostasis via inflammatory pathways, and components of these pathways are crucial targets of microRNAs (miRNAs). However, no study has examined the changes in the miRNA transcriptome during PUFA-regulated inflammatory processes. Here, we established PUFA diet-induced autoimmune-prone (AP) and autoimmune-averse (AA) rat models, and studied their physical characteristics and immune status. Additionally, miRNA expression patterns in the rat models were compared using microarray assays and bioinformatic methods. A total of 54 miRNAs were differentially expressed in common between the AP and the AA rats, and the changes in rno-miR-19b-3p, -146b-5p and -183-5p expression were validated using stem-loop reverse transcription-quantitative polymerase chain reaction. To better understand the mechanisms underlying PUFA-regulated miRNA changes during inflammation, computational algorithms and biological databases were used to identify the target genes of the three validated miRNAs. Furthermore, Gene Ontology (GO) term annotation and KEGG pathway analyses of the miRNA targets further allowed to explore the potential implication of the miRNAs in inflammatory pathways. The predicted PUFA-regulated inflammatory pathways included the Toll-like receptor (TLR), T cell receptor (TCR), NOD-like receptor (NLR), RIG-I-like receptor (RLR), mitogen-activated protein kinase (MAPK) and the transforming growth factor-β (TGF-β) pathway. This study is the first report, to the best of our knowledge, on in vivo comparative profiling of miRNA transcriptomes in PUFA diet-induced inflammatory rat models using a microarray approach. The results provide a useful resource for future investigation of the role of PUFA-regulated miRNAs in immune homeostasis. PMID:25672643

VPAC receptors: structure, molecular pharmacology and interaction with accessory proteins.

PubMed

Couvineau, Alain; Laburthe, Marc

2012-05-01

The vasoactive intestinal peptide (VIP) is a neuropeptide with wide distribution in both central and peripheral nervous systems, where it plays important regulatory role in many physiological processes. VIP displays a large biological functions including regulation of exocrine secretions, hormone release, fetal development, immune responses, etc. VIP appears to exert beneficial effect in neuro-degenerative and inflammatory diseases. The mechanism of action of VIP implicates two subtypes of receptors (VPAC1 and VPAC2), which are members of class B receptors belonging to the super-family of GPCR. This article reviews the current knowledge regarding the structure and molecular pharmacology of VPAC receptors. The structure-function relationship of VPAC1 receptor has been extensively studied, allowing to understand the molecular basis for receptor affinity, specificity, desensitization and coupling to adenylyl cyclase. Those studies have clearly demonstrated the crucial role of the N-terminal ectodomain (N-ted) of VPAC1 receptor in VIP recognition. By using different approaches including directed mutagenesis, photoaffinity labelling, NMR, molecular modelling and molecular dynamic simulation, it has been shown that the VIP molecule interacts with the N-ted of VPAC1 receptor, which is itself structured as a 'Sushi' domain. VPAC1 receptor also interacts with a few accessory proteins that play a role in cell signalling of receptors. Recent advances in the structural characterization of VPAC receptor and more generally of class B GPCRs will lead to the design of new molecules, which could have considerable interest for the treatment of inflammatory and neuro-degenerative diseases. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Development of a Fluorescent Bodipy Probe for Visualization of the Serotonin 5-HT1A Receptor in Native Cells of the Immune System.

PubMed

Hernández-Torres, Gloria; Enríquez-Palacios, Ernesto; Mecha, Miriam; Feliú, Ana; Rueda-Zubiaurre, Ainoa; Angelina, Alba; Martín-Cruz, Leticia; Martín-Fontecha, Mar; Palomares, Oscar; Guaza, Carmen; Peña-Cabrera, Eduardo; López-Rodríguez, María L; Ortega-Gutiérrez, Silvia

2018-05-14

Serotonin (5-HT) modulates key aspects of the immune system. However, its precise function and the receptors involved in the observed effects have remained elusive. Among the different serotonin receptors, 5-HT 1A plays an important role in the immune system given its presence in cells involved in both the innate and adaptive immune responses, but its actual levels of expression under different conditions have not been comprehensively studied due to the lack of suitable tools. To further clarify the role of 5-HT 1A receptor in the immune system, we have developed a fluorescent small molecule probe that enables the direct study of the receptor levels in native cells. This probe allows direct profiling of the receptor expression in immune cells using flow cytometry. Our results show that important subsets of immune cells including human monocytes and dendritic cells express functional 5-HT 1A and that its activation is associated with anti-inflammatory signaling. Furthermore, application of the probe to the experimental autoimmune encephalomyelitis model of multiple sclerosis demonstrates its potential to detect the specific overexpression of the 5-HT 1A receptor in CD4+ T cells. Accordingly, the probe reported herein represents a useful tool whose use can be extended to study the levels of 5-HT 1A receptor in ex vivo samples of different immune system conditions.

The Spinal Antinociceptive Effects of Endomorphins in Rats: Behavioral and G Protein Functional Studies

PubMed Central

Xie, Hong; Woods, James H.; Traynor, John R.; Ko, Mei-Chuan

2008-01-01

BACKGROUND Endomorphin-1 and endomorphin-2 are endogenous peptides that are highly selective for μ-opioid receptors. However, studies of their functional efficacy and selectivity are controversial. In this study, we systematically compared the effects of intrathecal (i.t.) administration of endomorphin-1 and -2 on nociception assays and G protein activation with those of [d-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (DAMGO), a highly effective peptidic μ-opioid receptor agonist. METHODS Male Sprague-Dawley rats were used. Acute and inflammatory pain models were used to compare the duration and magnitude of antinociception. Agonist-stimulated [35S]GTPγS binding was used to observe the functional activity at the level of the receptor-G protein in both spinal cord and thalamic membranes. In addition, antagonists selective for each receptor type were used to verify the functional selectivity of endomorphins in the rat spinal cord. RESULTS After i.t. administration, endomorphin-1 and -2 produced less antinociceptive effects than DAMGO in the model of acute pain. Concentration–response curves for DAMGO-, endomorphin-1-, and endomorphin-2-stimulated [35S]GTPγS binding revealed that both endomorphin-1 and -2 produced less G protein activation (i.e., approximately 50%–60%) than DAMGO did in the membranes of spinal cord and thalamus. In addition, i.t. endomorphin-induced antinociception was blocked by μ-opioid receptor selective dose of naltrexone (P < 0.05), but not by δ- and κ-opioid receptor antagonists, naltrindole and nor-binaltorphimine (P > 0.05). CONCLUSIONS Endomorphins are partial agonists for G protein activation at spinal and thalamic μ-opioid receptors. Both in vivo and in vitro measurements together suggest that DAMGO is more effective than endomorphins. Spinal endomorphins’ antinociceptive efficacy may range between 53% and 84% depending on the intensity and modality of the nociceptive stimulus. PMID:18499626

Global and Regional Modeling of Long-Range Transport and Intercontinental Source-Receptor Linkages

EPA Science Inventory

In this study, we compare air quality over North America simulated by the C-IFS global model and the CMAQ regional model driven by boundary conditions from C-IFS against surface and upper air observations. Results indicate substantial differences in model performance for surface ...

LiCABEDS II. Modeling of ligand selectivity for G-protein-coupled cannabinoid receptors.

PubMed

Ma, Chao; Wang, Lirong; Yang, Peng; Myint, Kyaw Z; Xie, Xiang-Qun

2013-01-28

The cannabinoid receptor subtype 2 (CB2) is a promising therapeutic target for blood cancer, pain relief, osteoporosis, and immune system disease. The recent withdrawal of Rimonabant, which targets another closely related cannabinoid receptor (CB1), accentuates the importance of selectivity for the development of CB2 ligands in order to minimize their effects on the CB1 receptor. In our previous study, LiCABEDS (Ligand Classifier of Adaptively Boosting Ensemble Decision Stumps) was reported as a generic ligand classification algorithm for the prediction of categorical molecular properties. Here, we report extension of the application of LiCABEDS to the modeling of cannabinoid ligand selectivity with molecular fingerprints as descriptors. The performance of LiCABEDS was systematically compared with another popular classification algorithm, support vector machine (SVM), according to prediction precision and recall rate. In addition, the examination of LiCABEDS models revealed the difference in structure diversity of CB1 and CB2 selective ligands. The structure determination from data mining could be useful for the design of novel cannabinoid lead compounds. More importantly, the potential of LiCABEDS was demonstrated through successful identification of newly synthesized CB2 selective compounds.

Computer modeling of the neurotoxin binding site of acetylcholine receptor spanning residues 185 through 196

NASA Technical Reports Server (NTRS)

Garduno-Juarez, R.; Shibata, M.; Zielinski, T. J.; Rein, R.

1987-01-01

A model of the complex between the acetylcholine receptor and the snake neurotoxin, cobratoxin, was built by molecular model building and energy optimization techniques. The experimentally identified functionally important residues of cobratoxin and the dodecapeptide corresponding to the residues 185-196 of acetylcholine receptor alpha subunit were used to build the model. Both cis and trans conformers of cyclic L-cystine portion of the dodecapeptide were examined. Binding residues independently identified on cobratoxin are shown to interact with the dodecapeptide AChR model.

The dopamine hypothesis of bipolar affective disorder: the state of the art and implications for treatment

PubMed Central

Ashok, A H; Marques, T R; Jauhar, S; Nour, M M; Goodwin, G M; Young, A H; Howes, O D

2017-01-01

Bipolar affective disorder is a common neuropsychiatric disorder. Although its neurobiological underpinnings are incompletely understood, the dopamine hypothesis has been a key theory of the pathophysiology of both manic and depressive phases of the illness for over four decades. The increased use of antidopaminergics in the treatment of this disorder and new in vivo neuroimaging and post-mortem studies makes it timely to review this theory. To do this, we conducted a systematic search for post-mortem, pharmacological, functional magnetic resonance and molecular imaging studies of dopamine function in bipolar disorder. Converging findings from pharmacological and imaging studies support the hypothesis that a state of hyperdopaminergia, specifically elevations in D2/3 receptor availability and a hyperactive reward processing network, underlies mania. In bipolar depression imaging studies show increased dopamine transporter levels, but changes in other aspects of dopaminergic function are inconsistent. Puzzlingly, pharmacological evidence shows that both dopamine agonists and antidopaminergics can improve bipolar depressive symptoms and perhaps actions at other receptors may reconcile these findings. Tentatively, this evidence suggests a model where an elevation in striatal D2/3 receptor availability would lead to increased dopaminergic neurotransmission and mania, whilst increased striatal dopamine transporter (DAT) levels would lead to reduced dopaminergic function and depression. Thus, it can be speculated that a failure of dopamine receptor and transporter homoeostasis might underlie the pathophysiology of this disorder. The limitations of this model include its reliance on pharmacological evidence, as these studies could potentially affect other monoamines, and the scarcity of imaging evidence on dopaminergic function. This model, if confirmed, has implications for developing new treatment strategies such as reducing the dopamine synthesis and/or release in mania and DAT blockade in bipolar depression. PMID:28289283

Insulin growth factor-1 (IGF-1) enhances hippocampal excitatory and seizure activity through IGF-1 receptor-mediated mechanisms in the epileptic brain.

PubMed

Jiang, Guohui; Wang, Wei; Cao, Qingqing; Gu, Juan; Mi, Xiujuan; Wang, Kewei; Chen, Guojun; Wang, Xuefeng

2015-12-01

Insulin-like growth factor-1 (IGF-1) is known to promote neurogenesis and survival. However, recent studies have suggested that IGF-1 regulates neuronal firing and excitatory neurotransmission. In the present study, focusing on temporal lobe epilepsy, we found that IGF-1 levels and IGF-1 receptor activation are increased in human epileptogenic tissues, and pilocarpine- and pentylenetetrazole-treated rat models. Using an acute model of seizures, we showed that lateral cerebroventricular infusion of IGF-1 elevates IGF-1 receptor (IGF-1R) signalling before pilocarpine application had proconvulsant effects. In vivo electroencephalogram recordings and power spectrogram analysis of local field potential revealed that IGF-1 promotes epileptiform activities. This effect is diminished by co-application of an IGF-1R inhibitor. In an in vitro electrophysiological study, we demonstrated that IGF-1 enhancement of excitatory neurotransmission and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor- and N-methyl-D-aspartate receptor-mediated currents is inhibited by IGF-1R inhibitor. Finally, activation of extracellular signal-related kinase (ERK)-1/2 and protein kinase B (Akt) in seizures in rats is increased by exogenous IGF-1 and diminished by picropodophyllin. A behavioural study reveals that the ERK1/2 or Akt inhibitor attenuates seizure activity. These results indicate that increased IGF-1 levels after recurrent hippocampal neuronal firings might, in turn, promote seizure activity via IGF-1R-dependent mechanisms. The present study presents a previously unappreciated role of IGF-1R in the development of seizure activity. © 2015 Authors; published by Portland Press Limited.

Prediction of CNS occupancy of dopamine D2 receptor based on systemic exposure and in vitro experiments.

PubMed

Kanamitsu, Kayoko; Arakawa, Ryosuke; Sugiyama, Yuichi; Suhara, Tetsuya; Kusuhara, Hiroyuki

2016-12-01

The effect of drugs in the central nervous system (CNS) is closely related to occupancy of their target receptor. In this study, we integrated plasma concentrations, in vitro/in vivo data for receptor or protein binding, and in silico data, using a physiologically based pharmacokinetic model, to examine the predictability of receptor occupancy in humans. The occupancy of the dopamine D2 receptor and the plasma concentrations of the antipsychotic drugs quetiapine and perospirone in humans were collected from the literature or produced experimentally. Association and dissociation rate constants and unbound fractions in the serum and brain were determined in vitro/in vivo using human D2 receptor-expressing membrane fractions, human serum and mouse brain. The permeability of drugs across the blood-brain barrier was estimated based on their physicochemical properties. The effect of a metabolite of perospirone, ID-15036, was also considered. The time profiles of D2 receptor occupancy following oral dose of quetiapine and perospirone predicted were similar to the observed values. This approach could assist in the design of clinical studies for drug development and the prediction of the impact of drug-drug interactions on CNS function in clinical settings. Copyright © 2016 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

Application of agonist-receptor modeling to the sweetness synergy between high fructose corn syrup and sucralose, and between high-potency sweeteners.

PubMed

Wolf, P A; Bridges, J R; Wicklund, R

2010-03-01

The agonist-receptor-transducer model of D. Ennis is applied to beverage formulations sweetened with high fructose corn syrup, sucralose, and other high-potency sweeteners, confirming the utility of the model, and supports the growing volume of evidence for multiple binding sites on the sweetness receptor. The model is further simplified to require less parameters for other sweetener blend systems whenever potency information is available for the single sweeteners.

Biphalin preferentially recruits peripheral opioid receptors to facilitate analgesia in a mouse model of cancer pain - A comparison with morphine.

PubMed

Lesniak, Anna; Bochynska-Czyz, Marta; Sacharczuk, Mariusz; Benhye, Sandor; Misicka, Aleksandra; Bujalska-Zadrozny, Magdalena; Lipkowski, Andrzej W

2016-06-30

The search for new drugs for cancer pain management has been a long-standing goal in basic and clinical research. Classical opioid drugs exert their primary antinociceptive effect upon activating opioid receptors located in the central nervous system. A substantial body of evidence points to the relevance of peripheral opioid receptors as potential targets for cancer pain treatment. Peptides showing limited blood-brain-barrier permeability promote peripheral analgesia in many pain models. In the present study we examined the peripheral and central analgesic effect of intravenously administered biphalin - a dimeric opioid peptide in a mouse skin cancer pain model, developed by an intraplantar inoculation of B16F0 melanoma cells. The effect of biphalin was compared with morphine - a golden standard in cancer pain management. Biphalin produced profound, dose-dependent and naloxone sensitive spinal analgesia. Additionally, the effect in the tumor-bearing paw was largely mediated by peripheral opioid receptors, as it was readily attenuated by the blood-brain-barrier-restricted opioid receptor antagonist - naloxone methiodide. On the contrary, morphine facilitated its analgesic effect primarily by activating spinal opioid receptors. Both drugs induced tolerance in B16F0 - implanted paws after chronic treatment, however biphalin as opposed to morphine, showed little decrease in its activity at the spinal level. Our results indicate that biphalin may be considered a future alternative drug in cancer pain treatment due to an enhanced local analgesic activity as well as lower tolerance liability compared with morphine. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparison of nicotinic receptor binding and biotransformation of coniine in the rat and chick.

PubMed

Forsyth, C S; Speth, R C; Wecker, L; Galey, F D; Frank, A A

1996-12-31

Coniine, an alkaloid from Conium maculatum (poison hemlock), is a known teratogen in many domestic species with maternal ingestion resulting in arthrogryposis of the offspring. We have previously shown that rats are not susceptible and rabbits only weakly susceptible to coniine-induced arthrogryposis. However, the chick embryo does provide a reproducible laboratory animal model of coniine-induced teratogenesis. The reason for this cross-species variation is unknown. The purpose of this study was to evaluate coniine binding to nicotinic receptors and to measure coniine metabolism in vitro between susceptible and non-susceptible species. Using the chick model, neither the peripheral nicotinic receptor antagonist d-tubocurarine chloride nor the central nicotinic receptor antagonist trimethaphan camsylate blocked the teratogenesis or lethality of 1.5% coniine (50 microliters/egg). Trimethaphan camsylate enhanced coniine-induced lethality in a dose-dependent manner. Neither nicotinic receptor blocker prevented nicotine sulfate-induced malformations but d-tubocurarine chloride did block lethality in a dose-dependent manner. Competition by coniine for [125I]-alpha-bungarotoxin to nicotinic receptors isolated from adult rat diaphragm and chick thigh muscle and competition by coniine for [3H]-cytisine to receptors from rat and chick brain were used to assess coniine binding to nicotinic receptors. The IC50 for coniine in rat diaphragm was 314 microM while that for chick leg muscle was 70 microM. For neuronal nicotinic receptors, the IC50s of coniine for maternal rat brain, fetal rat brain, and chick brain were 1100 microM, 820 microM, and 270 microM, respectively. There were no differences in coniine biotransformation in vitro by microsomes from rat or chick livers. Differences in apparent affinity of coniine for nicotinic receptors or differences in the quantity of the nicotinic receptor between the rat and chick may explain, in part, the differences in susceptibility of coniine-induced teratogenesis between these two species.

A plurality of molecular targets: The receptor ecosystem for bisphenol-A (BPA).

PubMed

MacKay, Harry; Abizaid, Alfonso

2018-05-01

Bisphenol-A (BPA) is a well-known endocrine disrupting compound (EDC), capable of affecting the normal function and development of the reproductive system, brain, adipose tissue, and more. In spite of these diverse and well characterized effects, there is often comparatively little known about the molecular mechanisms which bring them about. BPA has traditionally been regarded as a primarily estrogenic EDC, and this perspective is often what guides research into the effects of BPA. However, emerging data from in-vitro and in-silico models show that BPA binds with a significant number of hormone receptors, including a number of nuclear and membrane-bound estrogen receptors, androgen receptors, as well as the thyroid hormone receptor, glucocorticoid receptor, and PPARγ. With this increased diversity of receptor targets, it may be possible to explain some of the more puzzling aspects of BPA pharmacology, including its non-monotonic dose-response curve, as well as experimental results which disagree with estrogenic positive controls. This paper reviews the receptors for which BPA has a known interaction, and discusses the implications of taking these receptors into account when studying the disruptive effects of BPA on growth and development. Copyright © 2017 Elsevier Inc. All rights reserved.

Behavioral characterization of mice lacking histamine H(3) receptors.

PubMed

Toyota, Hiroshi; Dugovic, Christine; Koehl, Muriel; Laposky, Aaron D; Weber, China; Ngo, Karen; Wu, Ying; Lee, Doo Hyun; Yanai, Kazuhiko; Sakurai, Eiko; Watanabe, Takehiko; Liu, Changlu; Chen, Jingcai; Barbier, Ann J; Turek, Fred W; Fung-Leung, Wai-Ping; Lovenberg, Timothy W

2002-08-01

Brain histamine H(3) receptors are predominantly presynaptic and serve an important autoregulatory function for the release of histamine and other neurotransmitters. They have been implicated in a variety of brain functions, including arousal, locomotor activity, thermoregulation, food intake, and memory. The recent cloning of the H(3) receptor in our laboratory has made it possible to create a transgenic line of mice devoid of H(3) receptors. This paper provides the first description of the H(3) receptor-deficient mouse (H(3)(-/-)), including molecular and pharmacologic verification of the receptor deletion as well as phenotypic screens. The H(3)(-/-) mice showed a decrease in overall locomotion, wheel-running behavior, and body temperature during the dark phase but maintained normal circadian rhythmicity. H(3)(-/-) mice were insensitive to the wake-promoting effects of the H(3) receptor antagonist thioperamide. We also observed a slightly decreased stereotypic response to the dopamine releaser, methamphetamine, and an insensitivity to the amnesic effects of the cholinergic receptor antagonist, scopolamine. These data indicate that the H(3) receptor-deficient mouse represents a valuable model for studying histaminergic regulation of a variety of behaviors and neurotransmitter systems, including dopamine and acetylcholine.

Role of tissue factor and protease-activated receptors in a mouse model of endotoxemia.

PubMed

Pawlinski, Rafal; Pedersen, Brian; Schabbauer, Gernot; Tencati, Michael; Holscher, Todd; Boisvert, William; Andrade-Gordon, Patricia; Frank, Rolf Dario; Mackman, Nigel

2004-02-15

Sepsis is associated with a systemic activation of coagulation and an excessive inflammatory response. Anticoagulants have been shown to inhibit both coagulation and inflammation in sepsis. In this study, we used both genetic and pharmacologic approaches to analyze the role of tissue factor and protease-activated receptors in coagulation and inflammation in a mouse endotoxemia model. We used mice expressing low levels of the procoagulant molecule, tissue factor (TF), to analyze the effects of TF deficiency either in all tissues or selectively in hematopoietic cells. Low TF mice had reduced coagulation, inflammation, and mortality compared with control mice. Similarly, a deficiency of TF expression by hematopoietic cells reduced lipopolysaccharide (LPS)-induced coagulation, inflammation, and mortality. Inhibition of the down-stream coagulation protease, thrombin, reduced fibrin deposition and prolonged survival without affecting inflammation. Deficiency of either protease activated receptor-1 (PAR-1) or protease activated receptor-2 (PAR-2) alone did not affect inflammation or survival. However, a combination of thrombin inhibition and PAR-2 deficiency reduced inflammation and mortality. These data demonstrate that hematopoietic cells are the major pathologic site of TF expression during endotoxemia and suggest that multiple protease-activated receptors mediate crosstalk between coagulation and inflammation.

Novel effects of diosgenin on skin aging.

PubMed

Tada, Yayoi; Kanda, Naoko; Haratake, Akinori; Tobiishi, Megumi; Uchiwa, Hideyo; Watanabe, Shinichi

2009-06-01

Extracts of Dioscorea coomposita or Dioscorea villosa are consumed as supplemental health foods at the time of climacteric. The extracts contain large amounts of the plant steroid, diosgenin. Here, we studied the safety and efficacy of diosgenin against skin aging at the time of climacteric. In vitro, diosgenin enhanced DNA synthesis in a human 3D skin equivalent model, and increased bromodeoxyuridine uptake and intracellular cAMP level in adult human keratinocytes. The increase of bromodeoxyuridine uptake by diosgenin was blocked by an adenylate cyclase inhibitor, but not by antisense oligonucleotides against estrogen receptor alpha, estrogen receptor beta or an orphan G-protein-coupled receptor, GPR30, indicating the involvement of cAMP but not estrogen receptor alpha, estrogen receptor beta or GPR30. In vivo, administration of diosgenin improved the epidermal thickness in the ovariectomized mice, a climacteric model, without altering the degree of fat accumulation. In order to examine the safety of diosgenin, diosgenin and 17beta-estradiol were administered to breast cancer-burdened mice. The results revealed that while 17beta-estradiol accelerated the tumor growth, diosgenin did not show this effect. Our finding, a restoration of keratinocyte proliferation in aged skin, suggests that diosgenin may have potential as a safe health food for climacteric.

The group II metabotropic glutamate receptor agonist LY354740 and the D2 receptor antagonist haloperidol reduce locomotor hyperactivity but fail to rescue spatial working memory in GluA1 knockout mice.

PubMed

Boerner, Thomas; Bygrave, Alexei M; Chen, Jingkai; Fernando, Anushka; Jackson, Stephanie; Barkus, Chris; Sprengel, Rolf; Seeburg, Peter H; Harrison, Paul J; Gilmour, Gary; Bannerman, David M; Sanderson, David J

2017-04-01

Group II metabotropic glutamate receptor agonists have been suggested as potential anti-psychotics, at least in part, based on the observation that the agonist LY354740 appeared to rescue the cognitive deficits caused by non-competitive N-methyl-d-aspartate receptor (NMDAR) antagonists, including spatial working memory deficits in rodents. Here, we tested the ability of LY354740 to rescue spatial working memory performance in mice that lack the GluA1 subunit of the AMPA glutamate receptor, encoded by Gria1, a gene recently implicated in schizophrenia by genome-wide association studies. We found that LY354740 failed to rescue the spatial working memory deficit in Gria1 -/- mice during rewarded alternation performance in the T-maze. In contrast, LY354740 did reduce the locomotor hyperactivity in these animals to a level that was similar to controls. A similar pattern was found with the dopamine receptor antagonist haloperidol, with no amelioration of the spatial working memory deficit in Gria1 -/- mice, even though the same dose of haloperidol reduced their locomotor hyperactivity. These results with LY354740 contrast with the rescue of spatial working memory in models of glutamatergic hypofunction using non-competitive NMDAR antagonists. Future studies should determine whether group II mGluR agonists can rescue spatial working memory deficits with other NMDAR manipulations, including genetic models and other pharmacological manipulations of NMDAR function. © 2017 The Authors. European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Synthesis, Modeling, and Pharmacological Evaluation of UMB 425, a Mixed μ Agonist/δ Antagonist Opioid Analgesic with Reduced Tolerance Liabilities

PubMed Central

2013-01-01

Opioid narcotics are used for the treatment of moderate-to-severe pain and primarily exert their analgesic effects through μ receptors. Although traditional μ agonists can cause undesired side effects, including tolerance, addition of δ antagonists can attenuate said side effects. Herein, we report 4a,9-dihydroxy-7a-(hydroxymethyl)-3-methyl-2,3,4,4a,5,6-hexahydro-1H-4,12-methanobenzofuro[3,2-e]isoquinolin-7(7aH)-one (UMB 425) a 5,14-bridged morphinan-based orvinol precursor synthesized from thebaine. Although UMB 425 lacks δ-specific motifs, conformationally sampled pharmacophore models for μ and δ receptors predict it to have efficacy similar to morphine at μ receptors and similar to naltrexone at δ receptors, due to the compound sampling conformations in which the hydroxyl moiety interacts with the receptors similar to orvinols. As predicted, UMB 425 exhibits a mixed μ agonist/δ antagonist profile as determined in receptor binding and [35S]GTPγS functional assays in CHO cells. In vivo studies in mice show that UMB 425 displays potent antinociception in the hot plate and tail-flick assays. The antinociceptive effects of UMB 425 are blocked by naloxone, but not by the κ-selective antagonist norbinaltorphimine. During a 6-day tolerance paradigm, UMB 425 maintains significantly greater antinociception compared to morphine. These studies thus indicate that, even in the absence of δ-specific motifs fused to the C-ring, UMB 425 has mixed μ agonist/δ antagonist properties in vitro that translate to reduced tolerance liabilities in vivo. PMID:23713721

Identifying binding modes of two synthetic derivatives of adrenalin to the α2C-adrenoceptor by using molecular modeling; insights into the α2C-adrenoceptor activation.

PubMed

Gholami, Samira; Bordbar, A Khalegh; Lohrasebi, Amir

2017-04-01

Although, α2C adrenergic receptor (AR) mediates a number of physiological functions in vivo and has great therapeutic potential, the absence of its crystal structure is a major difficulty in the activation mechanism studies and drug design endeavors. Here, a homology model of α2C AR has been presented by means of multiple sequence alignment. The used templates were the latest crystal structures of the other ARs (Protein Data Bank IDs: 2R4R, 2RH1, 4GPO, 3P0G, 4BVN and 4LDO) that have 38.4% identity with the query. We then conducted docking simulations to understand and analyze the binding of noradrenaline (NOR), and its derivatives, namely arachidonoyl adrenalin (AA-AD) and arachidonoyl noradrenalin (AA-NOR) to the receptor. The existence of H-bonds between the ligands and SER218 residue implies the same binding site of derivatives with respect to the NOR. AA-AD and AA-NOR bind to the receptor with the larger binding affinities. The presence of salt bridge between ARG149 and GLU377 in the free receptor, obtained from molecular dynamics studies proved that the receptor still is in its basal state before binding process take places. The activation process is characterized by increasing in the RMSD values of the backbone receptor in the bound state, increasing the RMSF of the transmembrane involved in the activation process and the disappearance of the ARG149-GLU377 salt bridge. Copyright © 2017 Elsevier B.V. All rights reserved.

Epileptogenesis causes an N-methyl-d-aspartate receptor/Ca2+-dependent decrease in Ca2+/calmodulin-dependent protein kinase II activity in a hippocampal neuronal culture model of spontaneous recurrent epileptiform discharges.

PubMed

Blair, Robert E; Sombati, Sompong; Churn, Severn B; Delorenzo, Robert J

2008-06-24

Alterations in the function of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) have been observed in both in vivo and in vitro models of epileptogenesis; however the molecular mechanism mediating the effects of epileptogenesis on CaM kinase II has not been elucidated. This study was initiated to evaluate the molecular pathways involved in causing the long-lasting decrease in CaM kinase II activity in the hippocampal neuronal culture model of low Mg2+-induced spontaneous recurrent epileptiform discharges (SREDs). We show here that the decrease in CaM kinase II activity associated with SREDs in hippocampal cultures involves a Ca2+/N-methyl-d-aspartate (NMDA) receptor-dependent mechanism. Low Mg2+-induced SREDs result in a significant decrease in Ca2+/calmodulin-dependent substrate phosphorylation of the synthetic peptide autocamtide-2. Reduction of extracellular Ca2+ levels (0.2 mM in treatment solution) or the addition of dl-2-amino-5-phosphonovaleric acid (APV) 25 microM blocked the low Mg2+-induced decrease in CaM kinase II-dependent substrate phosphorylation. Antagonists of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid receptor or L-type voltage sensitive Ca2+ channel had no effect on the low Mg2+-induced decrease in CaM kinase II-dependent substrate phosphorylation. The results of this study demonstrate that the decrease in CaM kinase II activity associated with this model of epileptogenesis involves a selective Ca2+/NMDA receptor-dependent mechanism and may contribute to the production and maintenance of SREDs in this model.

Epileptogenesis causes an N-methyl-d-aspartate receptor/Ca2+-dependent decrease in Ca2+/calmodulin-dependent protein kinase II activity in a hippocampal neuronal culture model of spontaneous recurrent epileptiform discharges

PubMed Central

Blair, Robert E.; Sombati, Sompong; Churn, Severn B.; DeLorenzo, Robert J.

2008-01-01

Alterations in the function of Ca2+/calmodulin-dependent protein kinase II (CaM Kinase II) have been observed in both in vivo and in vitro models of epileptogenesis; however the molecular mechanism mediating the effects of epileptogenesis on CaM Kinase II have not been elucidated. This study was initiated to evaluate the molecular pathways involved in causing the long lasting decrease in CaM Kinase II activity in the hippocampal neuronal culture model of low Mg2+ induced spontaneous recurrent epileptiform discharges (SREDs). We show here that the decrease in CaM kinase II activity associated with SREDs in hippocampal cultures involves a Ca2+/N-methyl-d-aspartate (NMDA) receptor-dependent mechanism. Low Mg2+ induced SREDs results in a significant decrease in Ca2+/calmodulin-dependent substrate phosphorylation of the synthetic peptide autocamtide-2. Reduction of extracellular Ca2+ levels (0.2 mM in treatment solution) or the addition of DL-2-amino-5-phosphonovaleric acid (APV) 25 µM blocked the low Mg2+ induced decrease in CaM kinase II-dependent substrate phosphorylation. Antagonists of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid receptor or L-type voltage sensitive Ca2+ channel had no effect on the low Mg2+ induced decrease in CaM kinase II-dependent substrate phosphorylation. The results of this study demonstrate that the decrease in CaM kinase II activity associated with this model of epileptogenesis involves a selective Ca2+/NMDA receptor-dependent mechanism and may contribute to the production and maintenance of SREDs in this model. PMID:18495112

The cognition-enhancing activity of E1R, a novel positive allosteric modulator of sigma-1 receptors.

PubMed

Zvejniece, L; Vavers, E; Svalbe, B; Vilskersts, R; Domracheva, I; Vorona, M; Veinberg, G; Misane, I; Stonans, I; Kalvinsh, I; Dambrova, M

2014-02-01

Here, we describe the in vitro and in vivo effects of (4R,5S)-2-(5-methyl-2-oxo-4-phenyl-pyrrolidin-1-yl)-acetamide (E1R), a novel positive allosteric modulator of sigma-1 receptors. E1R was tested for sigma receptor binding activity in a [³H](+)-pentazocine assay, in bradykinin (BK)-induced intracellular Ca²⁺ concentration ([Ca²⁺](i)) assays and in an electrically stimulated rat vas deferens model. E1R's effects on cognitive function were tested using passive avoidance (PA) and Y-maze tests in mice. A selective sigma-1 receptor antagonist (NE-100), was used to study the involvement of the sigma-1 receptor in the effects of E1R. The open-field test was used to detect the effects of E1R on locomotion. Pretreatment with E1R enhanced the selective sigma-1 receptor agonist PRE-084's stimulating effect during a model study employing electrically stimulated rat vasa deferentia and an assay measuring the BK-induced [Ca²⁺](i) increase. Pretreatment with E1R facilitated PA retention in a dose-related manner. Furthermore, E1R alleviated the scopolamine-induced cognitive impairment during the PA and Y-maze tests in mice. The in vivo and in vitro effects of E1R were blocked by treatment with the selective sigma-1 receptor antagonist NE-100. E1R did not affect locomotor activity. E1R is a novel 4,5-disubstituted derivative of piracetam that enhances cognition and demonstrates efficacy against scopolamine-induced cholinergic dysfunction in mice. These effects are attributed to its positive modulatory action on the sigma-1 receptor and this activity may be relevant when developing new drugs for treating cognitive symptoms related to neurodegenerative diseases. © 2013 The British Pharmacological Society.

The cognition-enhancing activity of E1R, a novel positive allosteric modulator of sigma-1 receptors

PubMed Central

Zvejniece, L; Vavers, E; Svalbe, B; Vilskersts, R; Domracheva, I; Vorona, M; Veinberg, G; Misane, I; Stonans, I; Kalvinsh, I; Dambrova, M

2014-01-01

Background and Purpose Here, we describe the in vitro and in vivo effects of (4R,5S)-2-(5-methyl-2-oxo-4-phenyl-pyrrolidin-1-yl)-acetamide (E1R), a novel positive allosteric modulator of sigma-1 receptors. Experimental Approach E1R was tested for sigma receptor binding activity in a [3H](+)-pentazocine assay, in bradykinin (BK)-induced intracellular Ca2+ concentration ([Ca2+]i) assays and in an electrically stimulated rat vas deferens model. E1R's effects on cognitive function were tested using passive avoidance (PA) and Y-maze tests in mice. A selective sigma-1 receptor antagonist (NE-100), was used to study the involvement of the sigma-1 receptor in the effects of E1R. The open-field test was used to detect the effects of E1R on locomotion. Key Results Pretreatment with E1R enhanced the selective sigma-1 receptor agonist PRE-084's stimulating effect during a model study employing electrically stimulated rat vasa deferentia and an assay measuring the BK-induced [Ca2+]i increase. Pretreatment with E1R facilitated PA retention in a dose-related manner. Furthermore, E1R alleviated the scopolamine-induced cognitive impairment during the PA and Y-maze tests in mice. The in vivo and in vitro effects of E1R were blocked by treatment with the selective sigma-1 receptor antagonist NE-100. E1R did not affect locomotor activity. Conclusion and Implications E1R is a novel 4,5-disubstituted derivative of piracetam that enhances cognition and demonstrates efficacy against scopolamine-induced cholinergic dysfunction in mice. These effects are attributed to its positive modulatory action on the sigma-1 receptor and this activity may be relevant when developing new drugs for treating cognitive symptoms related to neurodegenerative diseases. PMID:24490863

Ethylene Regulates Levels of Ethylene Receptor/CTR1 Signaling Complexes in Arabidopsis thaliana

DOE PAGES

Shakeel, Samina N.; Gao, Zhiyong; Amir, Madiha; ...

2015-03-26

The plant hormone ethylene is perceived by a five-member family of receptors in Arabidopsis thaliana. The receptors function in conjunction with the Raf-like kinase CTR1 to negatively regulate ethylene signal transduction. CTR1 interacts with multiple members of the receptor family based on co-purification analysis, interacting more strongly with receptors containing a receiver domain. Levels of membrane-associated CTR1 vary in response to ethylene, doing so in a post-transcriptional manner that correlates with ethylene-mediated changes in levels of the ethylene receptors ERS1, ERS2, EIN4, and ETR2. Interactions between CTR1 and the receptor ETR1 protect ETR1 from ethylene-induced turnover. Kinetic and dose-response analysesmore » support a model in which two opposing factors control levels of the ethylene receptor/CTR1 complexes. Ethylene stimulates the production of new complexes largely through transcriptional induction of the receptors. However, ethylene also induces turnover of receptors, such that levels of ethylene receptor/CTR1 complexes decrease at higher ethylene concentrations. Lastly, we discuss implications of this model for ethylene signaling.« less

Ethylene Regulates Levels of Ethylene Receptor/CTR1 Signaling Complexes in Arabidopsis thaliana*

PubMed Central

Shakeel, Samina N.; Gao, Zhiyong; Amir, Madiha; Chen, Yi-Feng; Rai, Muneeza Iqbal; Haq, Noor Ul; Schaller, G. Eric

2015-01-01

The plant hormone ethylene is perceived by a five-member family of receptors in Arabidopsis thaliana. The receptors function in conjunction with the Raf-like kinase CTR1 to negatively regulate ethylene signal transduction. CTR1 interacts with multiple members of the receptor family based on co-purification analysis, interacting more strongly with receptors containing a receiver domain. Levels of membrane-associated CTR1 vary in response to ethylene, doing so in a post-transcriptional manner that correlates with ethylene-mediated changes in levels of the ethylene receptors ERS1, ERS2, EIN4, and ETR2. Interactions between CTR1 and the receptor ETR1 protect ETR1 from ethylene-induced turnover. Kinetic and dose-response analyses support a model in which two opposing factors control levels of the ethylene receptor/CTR1 complexes. Ethylene stimulates the production of new complexes largely through transcriptional induction of the receptors. However, ethylene also induces turnover of receptors, such that levels of ethylene receptor/CTR1 complexes decrease at higher ethylene concentrations. Implications of this model for ethylene signaling are discussed. PMID:25814663

Identification of bicyclic hexafluoroisopropyl alcohol sulfonamides as retinoic acid receptor-related orphan receptor gamma (RORγ/RORc) inverse agonists. Employing structure-based drug design to improve pregnane X receptor (PXR) selectivity.

PubMed

Gong, Hua; Weinstein, David S; Lu, Zhonghui; Duan, James J-W; Stachura, Sylwia; Haque, Lauren; Karmakar, Ananta; Hemagiri, Hemalatha; Raut, Dhanya Kumar; Gupta, Arun Kumar; Khan, Javed; Camac, Dan; Sack, John S; Pudzianowski, Andrew; Wu, Dauh-Rurng; Yarde, Melissa; Shen, Ding-Ren; Borowski, Virna; Xie, Jenny H; Sun, Huadong; D'Arienzo, Celia; Dabros, Marta; Galella, Michael A; Wang, Faye; Weigelt, Carolyn A; Zhao, Qihong; Foster, William; Somerville, John E; Salter-Cid, Luisa M; Barrish, Joel C; Carter, Percy H; Dhar, T G Murali

2018-01-15

We disclose the optimization of a high throughput screening hit to yield benzothiazine and tetrahydroquinoline sulfonamides as potent RORγt inverse agonists. However, a majority of these compounds showed potent activity against pregnane X receptor (PXR) and modest activity against liver X receptor α (LXRα). Structure-based drug design (SBDD) led to the identification of benzothiazine and tetrahydroquinoline sulfonamide analogs which completely dialed out LXRα activity and were less potent at PXR. Pharmacodynamic (PD) data for compound 35 in an IL-23 induced IL-17 mouse model is discussed along with the implications of a high Y max in the PXR assay for long term preclinical pharmacokinetic (PK) studies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Structural modeling of G-protein coupled receptors: An overview on automatic web-servers.

PubMed

Busato, Mirko; Giorgetti, Alejandro

2016-08-01

Despite the significant efforts and discoveries during the last few years in G protein-coupled receptor (GPCR) expression and crystallization, the receptors with known structures to date are limited only to a small fraction of human GPCRs. The lack of experimental three-dimensional structures of the receptors represents a strong limitation that hampers a deep understanding of their function. Computational techniques are thus a valid alternative strategy to model three-dimensional structures. Indeed, recent advances in the field, together with extraordinary developments in crystallography, in particular due to its ability to capture GPCRs in different activation states, have led to encouraging results in the generation of accurate models. This, prompted the community of modelers to render their methods publicly available through dedicated databases and web-servers. Here, we present an extensive overview on these services, focusing on their advantages, drawbacks and their role in successful applications. Future challenges in the field of GPCR modeling, such as the predictions of long loop regions and the modeling of receptor activation states are presented as well. Copyright © 2016 Elsevier Ltd. All rights reserved.

Predictive model of thrombospondin-1 and vascular endothelial growth factor in breast tumor tissue.

PubMed

Rohrs, Jennifer A; Sulistio, Christopher D; Finley, Stacey D

2016-01-01

Angiogenesis, the formation of new blood capillaries from pre-existing vessels, is a hallmark of cancer. Thus far, strategies for reducing tumor angiogenesis have focused on inhibiting pro-angiogenic factors, while less is known about the therapeutic effects of mimicking the actions of angiogenesis inhibitors. Thrombospondin-1 (TSP1) is an important endogenous inhibitor of angiogenesis that has been investigated as an anti-angiogenic agent. TSP1 impedes the growth of new blood vessels in many ways, including crosstalk with pro-angiogenic factors. Due to the complexity of TSP1 signaling, a predictive systems biology model would provide quantitative understanding of the angiogenic balance in tumor tissue. Therefore, we have developed a molecular-detailed, mechanistic model of TSP1 and vascular endothelial growth factor (VEGF), a promoter of angiogenesis, in breast tumor tissue. The model predicts the distribution of the angiogenic factors in tumor tissue, revealing that TSP1 is primarily in an inactive, cleaved form due to the action of proteases, rather than bound to its cellular receptors or to VEGF. The model also predicts the effects of enhancing TSP1's interactions with its receptors and with VEGF. To provide additional predictions that can guide the development of new anti-angiogenic drugs, we simulate administration of exogenous TSP1 mimetics that bind specific targets. The model predicts that the CD47-binding TSP1 mimetic dramatically decreases the ratio of receptor-bound VEGF to receptor-bound TSP1, in favor of anti-angiogenesis. Thus, we have established a model that provides a quantitative framework to study the response to TSP1 mimetics.

A Compartment Model of VEGF Distribution in Humans in the Presence of Soluble VEGF Receptor-1 Acting as a Ligand Trap

PubMed Central

Wu, Florence T. H.; Stefanini, Marianne O.; Mac Gabhann, Feilim; Popel, Aleksander S.

2009-01-01

Vascular endothelial growth factor (VEGF), through its activation of cell surface receptor tyrosine kinases including VEGFR1 and VEGFR2, is a vital regulator of stimulatory and inhibitory processes that keep angiogenesis – new capillary growth from existing microvasculature – at a dynamic balance in normal physiology. Soluble VEGF receptor-1 (sVEGFR1) – a naturally-occurring truncated version of VEGFR1 lacking the transmembrane and intracellular signaling domains – has been postulated to exert inhibitory effects on angiogenic signaling via two mechanisms: direct sequestration of angiogenic ligands such as VEGF; or dominant-negative heterodimerization with surface VEGFRs. In pre-clinical studies, sVEGFR1 gene and protein therapy have demonstrated efficacy in inhibiting tumor angiogenesis; while in clinical studies, sVEGFR1 has shown utility as a diagnostic or prognostic marker in a widening array of angiogenesis–dependent diseases. Here we developed a novel computational multi-tissue model for recapitulating the dynamic systemic distributions of VEGF and sVEGFR1. Model features included: physiologically-based multi-scale compartmentalization of the human body; inter-compartmental macromolecular biotransport processes (vascular permeability, lymphatic drainage); and molecularly-detailed binding interactions between the ligand isoforms VEGF121 and VEGF165, signaling receptors VEGFR1 and VEGFR2, non-signaling co-receptor neuropilin-1 (NRP1), as well as sVEGFR1. The model was parameterized to represent a healthy human subject, whereupon we investigated the effects of sVEGFR1 on the distribution and activation of VEGF ligands and receptors. We assessed the healthy baseline stability of circulating VEGF and sVEGFR1 levels in plasma, as well as their reliability in indicating tissue-level angiogenic signaling potential. Unexpectedly, simulated results showed that sVEGFR1 – acting as a diffusible VEGF sink alone, i.e., without sVEGFR1-VEGFR heterodimerization – did not significantly lower interstitial VEGF, nor inhibit signaling potential in tissues. Additionally, the sensitivity of plasma VEGF and sVEGFR1 to physiological fluctuations in transport rates may partially account for the heterogeneity in clinical measurements of these circulating angiogenic markers, potentially hindering their diagnostic reliability for diseases. PMID:19352513

Cannabidiol: pharmacology and potential therapeutic role in epilepsy and other neuropsychiatric disorders.

PubMed

Devinsky, Orrin; Cilio, Maria Roberta; Cross, Helen; Fernandez-Ruiz, Javier; French, Jacqueline; Hill, Charlotte; Katz, Russell; Di Marzo, Vincenzo; Jutras-Aswad, Didier; Notcutt, William George; Martinez-Orgado, Jose; Robson, Philip J; Rohrback, Brian G; Thiele, Elizabeth; Whalley, Benjamin; Friedman, Daniel

2014-06-01

To present a summary of current scientific evidence about the cannabinoid, cannabidiol (CBD) with regard to its relevance to epilepsy and other selected neuropsychiatric disorders. We summarize the presentations from a conference in which invited participants reviewed relevant aspects of the physiology, mechanisms of action, pharmacology, and data from studies with animal models and human subjects. Cannabis has been used to treat disease since ancient times. Δ(9) -Tetrahydrocannabinol (Δ(9) -THC) is the major psychoactive ingredient and CBD is the major nonpsychoactive ingredient in cannabis. Cannabis and Δ(9) -THC are anticonvulsant in most animal models but can be proconvulsant in some healthy animals. The psychotropic effects of Δ(9) -THC limit tolerability. CBD is anticonvulsant in many acute animal models, but there are limited data in chronic models. The antiepileptic mechanisms of CBD are not known, but may include effects on the equilibrative nucleoside transporter; the orphan G-protein-coupled receptor GPR55; the transient receptor potential of vanilloid type-1 channel; the 5-HT1a receptor; and the α3 and α1 glycine receptors. CBD has neuroprotective and antiinflammatory effects, and it appears to be well tolerated in humans, but small and methodologically limited studies of CBD in human epilepsy have been inconclusive. More recent anecdotal reports of high-ratio CBD:Δ(9) -THC medical marijuana have claimed efficacy, but studies were not controlled. CBD bears investigation in epilepsy and other neuropsychiatric disorders, including anxiety, schizophrenia, addiction, and neonatal hypoxic-ischemic encephalopathy. However, we lack data from well-powered double-blind randomized, controlled studies on the efficacy of pure CBD for any disorder. Initial dose-tolerability and double-blind randomized, controlled studies focusing on target intractable epilepsy populations such as patients with Dravet and Lennox-Gastaut syndromes are being planned. Trials in other treatment-resistant epilepsies may also be warranted. A PowerPoint slide summarizing this article is available for download in the Supporting Information section here. Wiley Periodicals, Inc. © 2014 International League Against Epilepsy.

Preclinical Models in Chimeric Antigen Receptor-Engineered T-Cell Therapy.

PubMed

Siegler, Elizabeth Louise; Wang, Pin

2018-05-01

Cancer immunotherapy has enormous potential in inducing long-term remission in cancer patients, and chimeric antigen receptor (CAR)-engineered T cells have been largely successful in treating hematological malignancies in the clinic. CAR-T therapy has not been as effective in treating solid tumors, in part due to the immunosuppressive tumor microenvironment. Additionally, CAR-T therapy can cause dangerous side effects, including off-tumor toxicity, cytokine release syndrome, and neurotoxicity. Animal models of CAR-T therapy often fail to predict such adverse events and frequently overestimate the efficacy of the treatment. Nearly all preclinical CAR-T studies have been performed in mice, including syngeneic, xenograft, transgenic, and humanized mouse models. Recently, a few studies have used primate models to mimic clinical side effects better. To date, no single model perfectly recapitulates the human immune system and tumor microenvironment, and some models have revealed CAR-T limitations that were contradicted or missed entirely in other models. Careful model selection based on the primary goals of the study is a crucial step in evaluating CAR-T treatment. Advancements are being made in preclinical models, with the ultimate objective of providing safer, more effective CAR-T therapy to patients.

Probability model for atmospheric sulfur dioxide concentrations in the area of Venice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buttazzoni, C.; Lavagnini, I.; Marani, A.

1986-09-01

This paper deals with a comparative screening of existing air quality models based on their ability to simulate the distribution of sulfur dioxide data in the Venetian area. Investigations have been carried out on sulfur dioxide dispersion in the atmosphere of the Venetian area. The studies have been mainly focused on transport models (Gaussian, plume and K-models) aiming at meaningful correlations of sources and receptors. Among the results, a noteworthy disagreement of simulated and experimental data, due to the lack of thorough knowledge of source field conditions and of local meteorology of the sea-land transition area, has been shown. Investigationsmore » with receptor oriented models (based, e.g., on time series analysis, Fourier analysis, or statistical distributions) have also been performed.« less

Quantitative targeted absolute proteomic analysis of transporters, receptors and junction proteins for validation of human cerebral microvascular endothelial cell line hCMEC/D3 as a human blood-brain barrier model.

PubMed

Ohtsuki, Sumio; Ikeda, Chiemi; Uchida, Yasuo; Sakamoto, Yumi; Miller, Florence; Glacial, Fabienne; Decleves, Xavier; Scherrmann, Jean-Michel; Couraud, Pierre-Olivier; Kubo, Yoshiyuki; Tachikawa, Masanori; Terasaki, Tetsuya

2013-01-07

Human cerebral microvascular endothelial cell line hCMEC/D3 is an established model of the human blood-brain barrier (BBB). The purpose of the present study was to determine, by means of quantitative targeted absolute proteomics, the protein expression levels in hCMEC/D3 cells of multiple transporters, receptors and junction proteins for comparison with our previously reported findings in isolated human brain microvessels. Among 91 target molecules, 12 transporters, 2 receptors, 1 junction protein and 1 membrane marker were present at quantifiable levels in plasma membrane fraction of hCMEC/D3 cells. ABCA2, MDR1, MRP4, BCRP, GLUT1, 4F2hc, MCT1, ENT1, transferrin and insulin receptors and claudin-5 were detected in both hCMEC/D3 cells and human brain microvessels. After normalization based on Na(+)/K(+) ATPase expression, the differences in protein expression levels between hCMEC/D3 cells and human brain microvessels were within 4-fold for these proteins, with the exceptions of ENT1, transferrin receptor and claudin-5. ABCA8, LAT1, LRP1 and γ-GTP were below the limit of quantification in the cells, but were found in human brain microvessels. ABCA3, ABCA6, MRP1 and ATA1 were found only in hCMEC/D3 cells. Furthermore, compared with human umbilical vein endothelial cells (HUVECs) as reference nonbrain endothelial cells, MDR1 was found only in hCMEC/D3 cells, and GLUT1 expression was 15-fold higher in hCMEC/D3 cells than in HUVECs. In conclusion, this is the first study to examine the suitability and limitations of the hCMEC/D3 cell line as a BBB functional model in terms of quantitative expression levels of transporters, receptors and tight junction proteins.

Potential role for ET-2 acting through ETA receptors in experimental colitis in mice.

PubMed

Claudino, R F; Leite, D F; Bento, A F; Chichorro, J G; Calixto, J B; Rae, G A

2017-02-01

This study attempted to clarify the roles of endothelins and mechanisms associated with ET A /ET B receptors in mouse models of colitis. Colitis was induced by intracolonic administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS, 1.5 mg/animal) or dextran sulfate sodium (DSS, 3%). After colitis establishment, mice received Atrasentan (ET A receptor antagonist, 10 mg/kg), A-192621 (ET B receptor antagonist, 20 mg/kg) or Dexamethasone (1 mg/kg) and several inflammatory parameters were assessed, as well as mRNA levels for ET-1, ET-2 and ET receptors. Atrasentan treatment ameliorates TNBS- and DSS-induced colitis. In the TNBS model was observed reduction in macroscopic and microscopic score, colon weight, neutrophil influx, IL-1β, MIP-2 and keratinocyte chemoattractant (KC) levels, inhibition of adhesion molecules expression and restoration of IL-10 levels. However, A192621 treatment did not modify any parameter. ET-1 and ET-2 mRNA was decreased 24 h, but ET-2 mRNA was markedly increased at 48 h after TNBS. ET-2 was able to potentiate LPS-induced KC production in vitro. ET A and ET B receptors mRNA were increased at 24, 48 and 72 h after colitis induction. Atrasentan treatment was effective in reducing the severity of colitis in DSS- and TNBS-treated mice, suggesting that ET A receptors might be a potential target for inflammatory bowel diseases.