The effects of agonists of ionotropic GABA(A) and metabotropic GABA(B) receptors on learning.

PubMed

Zyablitseva, Evgeniya A; Kositsyn, Nikolay S; Shul'gina, Galina I

2009-05-01

The research described here investigates the role played by inhibitory processes in the discriminations made by the nervous system of humans and animals between familiar and unfamiliar and significant and nonsignificant events. This research compared the effects of two inhibitory mediators of gamma-aminobutyric acid (GABA): 1) phenibut, a nonselective agonist of ionotropic GABA(A) and metabotropic GABA(B) receptors and 2) gaboxadol a selective agonist of ionotropic GABA(A) receptors on the process of developing active defensive and inhibitory conditioned reflexes in alert non-immobilized rabbits. It was found that phenibut, but not gaboxadol, accelerates the development of defensive reflexes at an early stage of conditioning. Both phenibut and gaboxadol facilitate the development of conditioned inhibition, but the effect of gaboxadol occurs at later stages of conditioning and is less stable than that of phenibut. The earlier and more stable effects of phenibut, as compared to gaboxadol, on storage in memory of the inhibitory significance of a stimulus may occur because GABA(B) receptors play the dominant role in the development of internal inhibition during an early stage of conditioning. On the other hand this may occur because the participation of both GABA(A) and GABA(B) receptors are essential to the process. We discuss the polyfunctionality of GABA receptors as a function of their structure and the positions of the relevant neurons in the brain as this factor can affect regulation of various types of psychological processes.

GABAB Receptor Positive Modulation Decreases Selective Molecular and Behavioral Effects of Cocaine

PubMed Central

Lhuillier, Loic; Mombereau, Cedric; Cryan, John F.; Kaupmann, Klemens

2006-01-01

Exposure to cocaine induces selective behavioral and molecular adaptations. In rodents, acute cocaine induces increased locomotor activity whereas prolonged drug exposure results in behavioral locomotor sensitization, which is thought to be a consequence of drug–induced neuroadaptive changes. Recent attention has been given to compounds activating GABAB receptors as potential anti-addictive therapies. In particular the principle of allosteric positive GABAB receptor modulators is very promising in this respect, as positive modulators lack the sedative and muscle relaxant properties of full GABAB receptor agonists such as baclofen. Here we investigated the effects of systemic application of the GABAB receptor positive modulator GS39783 in animals treated with acute and chronic cocaine administration. Both GS39783 and baclofen dose-dependently attenuated acute cocaine-induced hyperlocomotion. Furthermore, both compounds also efficiently blocked cocaine-induced Fos induction in the striatal complex. In chronic studies GS39783 induced a modest attenuation of cocaine-induced locomotor sensitization. Chronic cocaine induces the accumulation of the transcription factor ΔFosB and up regulates cAMP-response-element-binding-protein (CREB) and dopamine-and-cAMP-regulated-phosphoprotein of 32 kd (DARPP-32). GS39783 blocked the induction/activation of DARPP-32 and CREB in the nucleus accumbens and dorsal striatum and partially inhibited ΔFosB accumulation in the dorsal striatum. In summary our data provide evidence that GS39783 attenuates the acute behavioral effects of cocaine exposure in rodents and in addition prevents the induction of selective long-term adaptive changes in dopaminergic signaling pathways. Further investigation of GABAB receptor positive modulation as a novel therapeutic strategy for the treatment of cocaine dependence and possibly other drugs of abuse is therefore warranted. PMID:16710312

General, kappa, delta and mu opioid receptor antagonists mediate feeding elicited by the GABA-B agonist baclofen in the ventral tegmental area and nucleus accumbens shell in rats: reciprocal and regional interactions.

PubMed

Miner, Patricia; Shimonova, Lyudmila; Khaimov, Arthur; Borukhova, Yaffa; Ilyayeva, Ester; Ranaldi, Robert; Bodnar, Richard J

2012-03-14

Food intake is significantly increased following administration of agonists of GABA and opioid receptors into the nucleus accumbens shell (NACs) and ventral tegmental area (VTA). GABA-A or GABA-B receptor antagonist pretreatment within the VTA or NACs differentially affects mu-opioid agonist-induced feeding elicited from the same site. Correspondingly, general or selective opioid receptor antagonist pretreatment within the VTA or NACs differentially affects GABA agonist-induced feeding elicited from the same site. Regional interactions have been evaluated in feeding studies by administering antagonists in one site prior to agonist administration in a second site. Thus, opioid antagonist-opioid agonist and GABA antagonist-GABA agonist feeding interactions have been identified between the VTA and NACs. However, pretreatment with GABA-A or GABA-B receptor antagonists in the VTA failed to affect mu opioid agonist-induced feeding elicited from the NACs, and correspondingly, these antagonists administered in the NACs failed to affect mu opioid-induced feeding elicited from the VTA. To evaluate whether regional and reciprocal VTA and NACs feeding interactions occur for opioid receptor modulation of GABA agonist-mediated feeding, the present study examined whether feeding elicited by the GABA-B agonist, baclofen microinjected into the NACs was dose-dependently blocked by pretreatment with general (naltrexone: NTX), mu (beta-funaltrexamine: BFNA), kappa (nor-binaltorphamine: NBNI) or delta (naltrindole: NTI) opioid antagonists in the VTA, and correspondingly, whether VTA baclofen-induced feeding was dose-dependently blocked by NACs pretreatment with NTX, BFNA, NBNI or NTI in rats. Bilateral pairs of cannulae aimed at the VTA and NACs were stereotaxically implanted in rats, and their food intakes were assessed following vehicle and baclofen (200 ng) in each site. Baclofen produced similar magnitudes of increased food intake following VTA and NACs treatment. Baclofen

The role of GABAB receptors in human reinforcement learning.

PubMed

Ort, Andres; Kometer, Michael; Rohde, Judith; Seifritz, Erich; Vollenweider, Franz X

2014-10-01

Behavioral evidence from human studies suggests that the γ-aminobutyric acid type B receptor (GABAB receptor) agonist baclofen modulates reinforcement learning and reduces craving in patients with addiction spectrum disorders. However, in contrast to the well established role of dopamine in reinforcement learning, the mechanisms by which the GABAB receptor influences reinforcement learning in humans remain completely unknown. To further elucidate this issue, a cross-over, double-blind, placebo-controlled study was performed in healthy human subjects (N=15) to test the effects of baclofen (20 and 50mg p.o.) on probabilistic reinforcement learning. Outcomes were the feedback-induced P2 component of the event-related potential, the feedback-related negativity, and the P300 component of the event-related potential. Baclofen produced a reduction of P2 amplitude over the course of the experiment, but did not modulate the feedback-related negativity. Furthermore, there was a trend towards increased learning after baclofen administration relative to placebo over the course of the experiment. The present results extend previous theories of reinforcement learning, which focus on the importance of mesolimbic dopamine signaling, and indicate that stimulation of cortical GABAB receptors in a fronto-parietal network leads to better attentional allocation in reinforcement learning. This observation is a first step in our understanding of how baclofen may improve reinforcement learning in healthy subjects. Further studies with bigger sample sizes are needed to corroborate this conclusion and furthermore, test this effect in patients with addiction spectrum disorder. Copyright © 2014 Elsevier B.V. and ECNP. All rights reserved.

Administration of growth hormone and nandrolone decanoate alters mRNA expression of the GABAB receptor subunits as well as of the GH receptor, IGF-1, and IGF-2 in rat brain.

PubMed

Grönbladh, Alfhild; Johansson, Jenny; Nyberg, Fred; Hallberg, Mathias

2014-01-01

The illicit use of anabolic androgenic steroids (AAS), especially among young adults, is of major concern. Among AAS users it is common to combine the AAS nandrolone decanoate (ND), with intake of growth hormone (GH) and a connection between gonadal steroids and the GH system has been suggested. Both AAS and GH affect functions in the brain, for example those associated with the hypothalamus and pituitary, and several GH actions are mediated by growth factors such as insulin-like growth factor 1 (IGF-1) and insulin-like growth factor 2 (IGF-2). The GABAergic system is implicated in actions induced by AAS and previous studies have provided evidence for a link between GH and GABAB receptors in the brain. Our aim was to examine the impact of AAS administration and a subsequent administration of GH, on the expression of GABAB receptors and important GH mediators in rat brain. The aim was to investigate the CNS effects of a high-dose ND, and to study if a low, but physiological relevant, dose of GH could reverse the ND-induced effects. In the present study, male rats were administered a high dose of ND every third day during three weeks, and subsequently the rats were given recombinant human GH (rhGH) during ten days. Quantitative PCR (qPCR) was used to analyze gene expression in hypothalamus, anterior pituitary, caudate putamen, nucleus accumbens, and amygdala. In the pituitary gland, the expression of GABAB receptor subunits was affected differently by the steroid treatment; the GABAB1 mRNA expression was decreased whereas a distinct elevation of the GABAB2 expression was found. Administration of ND also caused a decrease of GHR, IGF-1, and IGF-2 mRNA expression in the pituitary while the corresponding expression in the hypothalamus, caudate putamen, nucleus accumbens, and amygdala was unaffected. The rhGH administration did not alter the GABAB2 expression but increased the GABAB1 gene expression in the hypothalamus as compared to the AAS treated group. These results

GABAB receptor cell surface export is controlled by an endoplasmic reticulum gatekeeper

PubMed Central

Doly, Stéphane; Shirvani, Hamasseh; Gäta, Gabriel; Meye, Frank; Emerit, Michel-Boris; Enslen, Hervé; Achour, Lamia; Pardo-Lopez, Liliana; Kwon, Yang Seung; Armand, Vincent; Gardette, Robert; Giros, Bruno; Gassmann, Martin; Bettler, Bernhard; Mameli, Manuel; Darmon, Michèle; Marullo, Stefano

2016-01-01

Summary Endoplasmic reticulum (ER) release and cell surface export of many G protein-coupled receptors (GPCRs), are tightly regulated. For GABAB receptors of GABA, the major mammalian inhibitory neurotransmitter, the ligand-binding GB1 subunit is maintained in the ER by unknown mechanisms in the absence of hetero-dimerization with the GB2 subunit. We report that GB1 retention is regulated by a specific gatekeeper, PRAF2. This ER resident transmembrane protein binds to GB1, preventing its progression in the biosynthetic pathway. GB1 release occurs upon competitive displacement from PRAF2 by GB2. PRAF2 concentration, relative to that of GB1 and GB2, tightly controls cell surface receptor density and controls GABAB function in neurons. Experimental perturbation of PRAF2 levels in vivo caused marked hyperactivity disorders in mice. These data reveal an unanticipated major impact of specific ER gate-keepers on GPCR function and identify PRAF2 as a new molecular target with therapeutic potential for psychiatric and neurological diseases involving GABAB function. PMID:26033241

The anorexic agents, sibutramine and fenfluramine, depress GABAB-induced inhibitory postsynaptic potentials in rat mesencephalic dopaminergic cells

PubMed Central

Ledonne, Ada; Sebastianelli, Luca; Federici, Mauro; Bernardi, Giorgio; Mercuri, Nicola Biagio

2009-01-01

Background and purpose Nutrition is the result of a complex interaction among environmental, homeostatic and reward-related processes. Accumulating evidence supports key roles for the dopaminergic neurons of the ventral midbrain in regulating feeding behaviour. For this reason, in the present study, we have investigated the electrophysiological effects of two centrally acting anorexic agents, fenfluramine and sibutramine, on these cells. Experimental approach Rat midbrain slices were used to make intracellular recordings from dopaminergic neurons of the substantia nigra and the ventral tegmental area. Gamma-aminobutyric acid (GABA)-mediated synaptic transmission was assessed from the inhibitory postsynaptic potentials (IPSPs) mediated by GABAA and GABAB receptors. Key results Fenfluramine and sibutramine reduced, concentration-dependently, the GABAB IPSPs, without affecting the GABAA-mediated potentials. This effect is presynaptic, as postsynaptic membrane responses induced by application of a GABAB receptor agonist, baclofen, were not affected by the two drugs. Furthermore, the selective 5-hydroxytriptamine 1B (5-HT1B) receptor antagonist, SB216641, blocked the reduction of GABAB IPSPs caused by fenfluramine and sibutramine, indicating that the receptor mediating this effect is 5-HT1B. Conclusions and implications Two anorexic agents, fenfluramine and sibutramine, induced the activation of 5-HT1B receptors located on presynaptic GABAergic terminals, thus reducing the release of GABA. This action can alter the strength of synaptic afferents that modify the activity of dopaminergic neurons, inducing neuronal excitation. Our results reveal an additional mechanism of action for fenfluramine and sibutramine that might contribute to reducing food intake, by influencing the pleasurable and motor aspects of feeding behaviour. PMID:19298257

GABA(B) receptors, schizophrenia and sleep dysfunction: a review of the relationship and its potential clinical and therapeutic implications.

PubMed

Kantrowitz, Joshua; Citrome, Leslie; Javitt, Daniel

2009-08-01

Evidence for an intrinsic relationship between sleep, cognition and the symptomatic manifestations of schizophrenia is accumulating. This review presents evidence for the possible utility of GABA(B) receptor agonists for the treatment of subjective and objective sleep abnormalities related to schizophrenia. At the phenotypic level, sleep disturbance occurs in 16-30% of patients with schizophrenia and is related to reduced quality of life and poor coping skills. On the neurophysiological level, studies suggest that sleep deficits reflect a core component of schizophrenia. Specifically, slow-wave sleep deficits, which are inversely correlated with cognition scores, are seen. Moreover, sleep plays an increasingly well documented role in memory consolidation in schizophrenia. Correlations of slow-wave sleep deficits with impaired reaction time and declarative memory have also been reported. Thus, both behavioural insomnia and sleep architecture are critical therapeutic targets in patients with schizophrenia. However, long-term treatment with antipsychotics often results in residual sleep dysfunction and does not improve slow-wave sleep, and adjunctive GABA(A) receptor modulators, such as benzodiazepines and zolpidem, can impair sleep architecture and cognition in schizophrenia. GABA(B) receptor agonists have therapeutic potential in schizophrenia. These agents have minimal effect on rapid eye movement sleep while increasing slow-wave sleep. Preclinical associations with increased expression of genes related to slow-wave sleep production and circadian rhythm function have also been reported. GABA(B) receptor deficits result in a sustained hyperdopaminergic state and can be reversed by a GABA(B) receptor agonist. Genetic, postmortem and electrophysiological studies also associate GABA(B) receptors with schizophrenia. While studies thus far have not shown significant effects, prior focus on the use of GABA(B) receptor agonists has been on the positive symptoms of

The GABA(B) receptor positive modulator BHF177 attenuated anxiety, but not conditioned fear, in rats.

PubMed

Li, Xia; Kaczanowska, Katarzyna; Finn, M G; Markou, Athina; Risbrough, Victoria B

2015-10-01

GABAB (γ-aminobutyric acid B) receptors may be a therapeutic target for anxiety disorders. Here we characterized the effects of the GABAB receptor positive allosteric modulator (PAM) BHF177 on conditioned and unconditioned physiological responses to threat in the light-enhanced startle (LES), stress-induced hyperthermia, and fear-potentiated startle (FPS) procedures in rats. The effects of BHF177 on LES were compared with those of the GABAB receptor agonists baclofen and CGP44532, and the positive control buspirone, a 5-HT1A receptor partial agonist with anxiolytic activity in humans. Baclofen (0.4, 0.9 and 1.25 mg/kg) and CGP44532 (0.065, 0.125 and 0.25 mg/kg) administration had significant sedative, but not anxiolytic, activity reflected in overall decrease in the startle response in the LES tests. BHF177 (10, 20 and 40 mg/kg) had no effect on LES, nor did it produce an overall sedative effect. Interesting, however, when rats were grouped by high and low LES responses, BHF177 had anxiolytic-like effects only on LES in high, but not low, LES responding rats. BHF177 also blocked stress-induced hyperthermia, but had no effect on conditioned fear responses in the FPS test. Buspirone (1 and 3 mg/kg) had an anxiolytic-like profile in both LES and FPS tests. These results indicate that BHF177 may specifically attenuate unconditioned anxiety in individuals that exhibit a high anxiety state, and has fewer sedative effects than direct agonists. Thus, BHF177 or other GABAB receptor PAMs may be promising compounds for alleviating increased anxiety seen in various psychiatric disorders with a superior side-effect profile compared to GABAB receptor agonists. Published by Elsevier Ltd.

GABAB receptor ligands for the treatment of alcohol use disorder: preclinical and clinical evidence

PubMed Central

Agabio, Roberta; Colombo, Giancarlo

2014-01-01

The present paper summarizes the preclinical and clinical studies conducted to define the “anti-alcohol” pharmacological profile of the prototypic GABAB receptor agonist, baclofen, and its therapeutic potential for treatment of alcohol use disorder (AUD). Numerous studies have reported baclofen-induced suppression of alcohol drinking (including relapse- and binge-like drinking) and alcohol reinforcing, motivational, stimulating, and rewarding properties in rodents and monkeys. The majority of clinical surveys conducted to date—including case reports, retrospective chart reviews, and randomized placebo-controlled studies—suggest the ability of baclofen to suppress alcohol consumption, craving for alcohol, and alcohol withdrawal symptomatology in alcohol-dependent patients. The recent identification of a positive allosteric modulatory binding site, together with the synthesis of in vivo effective ligands, represents a novel, and likely more favorable, option for pharmacological manipulations of the GABAB receptor. Accordingly, data collected to date suggest that positive allosteric modulators of the GABAB receptor reproduce several “anti-alcohol” effects of baclofen and display a higher therapeutic index (with larger separation—in terms of doses—between “anti-alcohol” effects and sedation). PMID:24936171

Antagonism of GABA-B but not GABA-A receptors in the VTA prevents stress- and intra-VTA CRF-induced reinstatement of extinguished cocaine seeking in rats

PubMed Central

Blacktop, Jordan M.; Vranjkovic, Oliver; Mayer, Matthieu; Van Hoof, Matthew; Baker, David A.; Mantsch, John R.

2015-01-01

Stress-induced reinstatement of cocaine seeking requires corticotropin releasing factor (CRF) actions in the ventral tegmental area (VTA). However the mechanisms through which CRF regulates VTA function to promote cocaine use are not fully understood. Here we examined the role of GABAergic neurotransmission in the VTA mediated by GABA-A or GABA-B receptors in the reinstatement of extinguished cocaine seeking by a stressor, uncontrollable intermittent footshock, or bilateral intra-VTA administration of CRF. Rats underwent repeated daily cocaine self-administration (1.0 mg/kg/ing; 14 × 6 hrs/day) and extinction and were tested for reinstatement in response to footshock (0.5 mA, 0.5” duration, average every 40 sec; range 10–70 sec) or intra-VTA CRF delivery (500 ng/side) following intra-VTA pretreatment with the GABA-A antagonist, bicuculline, the GABA-B antagonist, 2-hydroxysaclofen or vehicle. Intra-VTA bicuculline (1, 10 or 20 ng/side) failed to block footshock- or CRF-induced cocaine seeking at either dose tested. By contrast, 2-hydroxysaclofen (0.2 or 2 µg/side) prevented reinstatement by both footshock and intra-VTA CRF at a concentration that failed to attenuate food-reinforced lever pressing (45 mg sucrose-sweetened pellets; FR4 schedule) in a separate group of rats. These data suggest that GABA-B receptor-dependent CRF actions in the VTA mediate stress-induced cocaine seeking and that GABA-B receptor antagonists may have utility for the management of stress-induced relapse in cocaine addicts. PMID:26596556

Effects of GABA(B) receptor agents on cocaine priming, discrete contextual cue and food induced relapses.

PubMed

Filip, Małgorzata; Frankowska, Małgorzata

2007-10-01

In the present study we investigated the effects of the GABA(B) receptor antagonist (2S)-(+)-5,5-dimethyl-2-morpholineacetic acid (SCH 50911), the agonists baclofen and 3-aminopropyl(methyl)phosphinic acid (SKF 97541), and the allosteric positive modulator 3,5-bis(1,1-dimethylethyl)-4-hydroxy-beta,beta-dimethylbenzenepropanol (CGP 7930) on cocaine seeking behavior. The effects of the above drugs on the reinstatement of responding induced by natural reinforcer (food) were also studied. Male Wistar rats were trained to self-administer either cocaine (0.5 mg/kg/infusion) or food (sweet milk) and responding on the reinforcer-paired lever was extinguished. Reinstatement of responding was induced by a noncontingent presentation of the self-administered reinforcer (10 mg/kg cocaine, i.p.), a discrete contextual cue, or a contingent presentation of food. SCH 50911 (3-10 mg/kg) dose-dependently attenuated responding on the previously cocaine-paired lever during both reinstatement conditions, with slightly greater efficacy at reducing conditioned cue reinstatement. At the same time, it failed to alter reinstatement of food-seeking behavior. Baclofen (1.25-5 mg/kg) and SKF 97541 (0.03-0.3 mg/kg) attenuated cocaine- or food-seeking behavior; the effect of the drug appeared more effective for cocaine-seeking than food-seeking. CGP 7930 (10-30 mg/kg) reduced cocaine seeking without affecting food-induced reinstatement on reward seeking. Our results indicate that tonic activation of GABA(B) receptors is required for cocaine seeking behavior in rats. Moreover, the GABA(B) receptor antagonist SCH 50911 was effective in reducing relapse to cocaine at doses that failed to alter reinstatement of food-seeking behavior (present study), basal locomotor activity, cocaine and food self-administration (Filip et al., submitted for publication), suggesting its selective effects on motivated drug-seeking behavior. The potent inhibitory responses on cocaine seeking behavior were also seen

The effects of intraperitoneal and intracerebroventricular administration of the GABAB receptor antagonist CGP 35348 on food intake in rats.

PubMed

Patel, Sunit M; Ebenezer, Ivor S

2004-10-25

In order to test the hypothesis that endogenous gamma-aminobutyric acid (GABA), acting at central GABAB receptors, plays a physiological role in the control of feeding behaviour, it was reasoned that blocking these receptors with a centrally active GABAB receptor antagonist should reduce food intake in hungry rats. In the present study, experiments were carried out to test this possibility using the GABAB receptor antagonist 3-aminopropyl-diethoxy-methyl-phosphinic acid (CGP 35348), which is water-soluble and can penetrate the blood-brain barrier from the systemic circulation. CGP 35348 (50 and 100 mg/kg, i.p.) had no effect on food intake in 22-h fasted rats, but a higher dose (i.e. 500 mg/kg., i.p.) significantly reduced cumulative food consumption. These findings are consistent with previous observations that high systemic doses of CGP 35348 are needed to block central GABAB receptors. However, to eliminate the possibility that the 500 mg/kg dose of CGP 35348 decreased food intake by a peripheral, rather than a central mode of action, further experiments were undertaken where the drug was given directly into the brain by the intracerebroventricular (i.c.v.) route. I.c.v. administration of CGP 35348 (5 and 10 microg) significantly decreased cumulative food intake food intake in rats that had been fasted for 22 h. By contrast, i.c.v. administration of CGP 35348 (10 microg) had no effect on water intake in 16-h water-deprived rats. The results indicate that CGP 35348 reduces food consumption in hungry rats by blocking central GABAB receptors in a behaviourally specific manner. These findings suggest that endogenous GABA acting at central GABAB receptors plays a physiological role in the regulation of feeding behaviour.

GABAB receptor modulation of the release of substance P from capsaicin-sensitive neurones in the rat trachea in vitro.

PubMed Central

Ray, N. J.; Jones, A. J.; Keen, P.

1991-01-01

1. The role of gamma-aminobutyric acid (GABA) as an inhibitory transmitter in the central nervous system is well documented. Recently, GABAA and GABAB receptors have been identified in the peripheral nervous system, notably on primary afferent neurones (PAN). We have utilised a multi-superfusion system to investigate the effect of selective GABA receptor agonists and antagonists on the release of substance P (SP) from the rat trachea in vitro. 2. GABA (1-100 microM) did not affect spontaneous release of SP-like immunoreactivity (LI) but caused dose-related inhibition of calcium-dependent potassium (60 mM)-stimulated SP-LI release. The greatest inhibition of 77.7 +/- 18.8% was observed at 100 microM. 3. The inhibitory effect of GABA was mimicked by the GABAB receptor agonist, (+/-)-baclofen (1-100 microM), but not the GABAA receptor agonist, 3-amino-1-propane-sulphonic acid (3-APS, 1-100 microM). Baclofen (100 microM) had no effect on SP-LI release stimulated by capsaicin (1 microM). 4. The inhibitory effect of baclofen (30 microM) was significantly reduced by prior and concomitant exposure to the GABAB receptor antagonist, phacolofen (100 microM) but not the GABAA receptor antagonist, bicuculline (10 microM). Neither antagonist, alone, affected spontaneous or potassium-stimulated SP-LI release. 5. We conclude that activation of pre-synaptic GABAB receptors on the peripheral termini of PANs in the rat trachea inhibits SP-LI release and suggest that GABAB receptor agonists may be of value in the therapeutic treatment of asthma. PMID:1713105

GABAB-receptor activation alters the firing pattern of dopamine neurons in the rat substantia nigra.

PubMed

Engberg, G; Kling-Petersen, T; Nissbrandt, H

1993-11-01

Previous electrophysiological experiments have emphasized the importance of the firing pattern for the functioning of midbrain dopamine (DA) neurons. In this regard, excitatory amino acid receptors appear to constitute an important modulatory control mechanism. In the present study, extracellular recording techniques were used to investigate the significance of GABAB-receptor activation for the firing properties of DA neurons in the substantia nigra (SN) in the rat. Intravenous administration of the GABAB-receptor agonist baclofen (1-16 mg/kg) was associated with a dose-dependent regularization of the firing pattern, concomitant with a reduction in burst firing. At higher doses (16-32 mg/kg), the firing rate of the DA neurons was dose-dependently decreased. Also, microiontophoretic application of baclofen regularized the firing pattern of nigral DA neurons, including a reduction of burst firing. Both the regularization of the firing pattern and inhibition of firing rate produced by systemic baclofen administration was antagonized by the GABAB-receptor antagonist CGP 35348 (200 mg/kg, i.v.). The GABAA-receptor agonist muscimol produced effects on the firing properties of DA neurons that were opposite to those observed following baclofen, i.e., an increase in firing rate accompanied by a decreased regularity. The NMDA receptor antagonist MK 801 (0.4-3.2 mg/kg, i.v.) produced a moderate, dose-dependent increase in the firing rate of the nigral DA neurons as well as a slightly regularized firing pattern. Pretreatment with MK 801 (3.2 mg/kg, i.v., 3-10 min) did neither promote nor prevent the regularization of the firing pattern or inhibition of firing rate on the nigral DA neurons produced by baclofen. The present results clearly show that GABAB-receptors can alter the firing pattern of nigral DA neurons, hereby counterbalancing the previously described ability of glutamate to induce burst firing activity on these neurons.

Human brain somatostatin release from isolated cortical nerve endings and its modulation through GABAB receptors.

PubMed Central

Bonanno, G.; Gemignani, A.; Schmid, G.; Severi, P.; Cavazzani, P.; Raiteri, M.

1996-01-01

1. The release of somatostatin-like immunoreactivity (SRIF-LI) in the human brain was studied in synaptosomal preparations from fresh neocortical specimens obtained from patients undergoing neurosurgery to remove deeply sited tumours. 2. The basal outflow of SRIF-LI from superfused synaptosomes was increased about 3 fold during exposure to a depolarizing medium containing 15 mM KCl. The K(+)-evoked overflow of SRIF-LI was almost totally dependent on the presence of Ca2+ in the superfusion medium. 3. The GABAB receptor agonist, (-)-baclofen (0.3 - 100 microM), inhibited the overflow of SRIF-LI in a concentration-dependent manner (EC50 = 1.84 +/- 0.20 microM; maximal effect: about 50%). The novel GABAB receptor ligand, 3-aminopropyl(difluoromethyl)phosphinic acid (CGP 47656) mimicked (-)-baclofen in inhibiting the SRIF-LI overflow (EC50 = 3.06 +/- 0.52 microM; maximal effect: about 50%), whereas the GABAA receptor agonist, muscimol, was ineffective up to 100 microM. 4. The inhibition by 10 microM (-)-baclofen of the K(+)-evoked SRIF-LI overflow was concentration-dependently prevented by two selective GABAB receptor antagonists, 3-amino-propyl (diethoxymethyl)-phosphinic acid (CGP 35348) (IC50 = 24.40 +/- 2.52 microM) and [3-[[(3,4-dichlorophenyl) methyl]amino]propyl] (diethoxymethyl) phosphinic acid (CGP 52432) (IC50 = 0.06 +/- 0.005 microM). 5. The inhibition of SRIF-LI overflow caused by 10 microM CGP 47656 was abolished by 1 microM CGP 52432. 6. When human synaptosomes were labelled with [3H]-GABA and depolarized in superfusion with 15 mM KCl, the inhibition by 10 microM (-)-baclofen of the depolarization-evoked [3H]-GABA overflow was largely prevented by 10 microM CGP 47656 which therefore behaved as an autoreceptor antagonist. 7. In conclusion: (a) the characteristics of SRIF-LI release from synaptosomal preparations of human neocortex are compatible with a neuronal origin; (b) the nerve terminals releasing the neuropeptide possess inhibitory receptors of the

Spinal GABA-B receptor modulates neutrophil recruitment to the knee joint in zymosan-induced arthritis.

PubMed

Bassi, Gabriel S; do C Malvar, David; Cunha, Thiago M; Cunha, Fernando Q; Kanashiro, Alexandre

2016-08-01

Recent studies have demonstrated that the central nervous system controls inflammatory responses by activating complex efferent neuroimmune pathways. The present study was designed to evaluate the role that central gamma-aminobutyric acid type B (GABA-B) receptor plays in neutrophil migration in a murine model of zymosan-induced arthritis by using different pharmacological tools. We observed that intrathecal administration of baclofen, a selective GABA-B agonist, exacerbated the inflammatory response in the knee after zymosan administration characterized by an increase in the neutrophil recruitment and knee joint edema, whereas saclofen, a GABA-B antagonist, exerted the opposite effect. Intrathecal pretreatment of the animals with SB203580 (an inhibitor of p38 mitogen-activated protein kinase) blocked the pro-inflammatory effect of baclofen. On the other hand, systemic administration of guanethidine, a sympatholytic drug that inhibits catecholamine release, and nadolol, a beta-adrenergic receptor antagonist, reversed the effect of saclofen. Moreover, saclofen suppressed the release of the pro-inflammatory cytokines into the knee joint (ELISA) and pain-related behaviors (open field test). Since the anti-inflammatory effect of saclofen depends on the sympathetic nervous system integrity, we observed that isoproterenol, a beta-adrenergic receptor agonist, mimics the central GABA-B blockade decreasing knee joint neutrophil recruitment. Together, these results demonstrate that the pharmacological manipulation of spinal GABAergic transmission aids control of neutrophil migration to the inflamed joint by modulating the activation of the knee joint-innervating sympathetic terminal fibers through a mechanism dependent on peripheral beta-adrenergic receptors and central components, such as p38 MAPK.

Antagonism of GABA-B but not GABA-A receptors in the VTA prevents stress- and intra-VTA CRF-induced reinstatement of extinguished cocaine seeking in rats.

PubMed

Blacktop, Jordan M; Vranjkovic, Oliver; Mayer, Matthieu; Van Hoof, Matthew; Baker, David A; Mantsch, John R

2016-03-01

Stress-induced reinstatement of cocaine seeking requires corticotropin releasing factor (CRF) actions in the ventral tegmental area (VTA). However the mechanisms through which CRF regulates VTA function to promote cocaine use are not fully understood. Here we examined the role of GABAergic neurotransmission in the VTA mediated by GABA-A or GABA-B receptors in the reinstatement of extinguished cocaine seeking by a stressor, uncontrollable intermittent footshock, or bilateral intra-VTA administration of CRF. Rats underwent repeated daily cocaine self-administration (1.0 mg/kg/ing; 14 × 6 h/day) and extinction and were tested for reinstatement in response to footshock (0.5 mA, 0.5" duration, average every 40 s; range 10-70 s) or intra-VTA CRF delivery (500 ng/side) following intra-VTA pretreatment with the GABA-A antagonist, bicuculline, the GABA-B antagonist, 2-hydroxysaclofen or vehicle. Intra-VTA bicuculline (1, 10 or 20 ng/side) failed to block footshock- or CRF-induced cocaine seeking at either dose tested. By contrast, 2-hydroxysaclofen (0.2 or 2 μg/side) prevented reinstatement by both footshock and intra-VTA CRF at a concentration that failed to attenuate food-reinforced lever pressing (45 mg sucrose-sweetened pellets; FR4 schedule) in a separate group of rats. These data suggest that GABA-B receptor-dependent CRF actions in the VTA mediate stress-induced cocaine seeking and that GABA-B receptor antagonists may have utility for the management of stress-induced relapse in cocaine addicts. Copyright © 2015 Elsevier Ltd. All rights reserved.

Comparison of the effects of the GABAB receptor positive modulator BHF177 and the GABAB receptor agonist baclofen on anxiety-like behavior, learning, and memory in mice

PubMed Central

Li, Xia; Risbrough, Victoria B.; Cates-Gatto, Chelsea; Kaczanowska, Katarzyna; Finn, M. G.; Roberts, Amanda J; Markou, Athina

2013-01-01

γ-Aminobutyric acid B (GABAB) receptor activation is a potential therapeutic approach for the treatment of drug addiction, pain, anxiety, and depression. However, full agonists of this receptor induce side-effects, such as sedation, muscle relaxation, tolerance, and cognitive disruption. Positive allosteric modulators (PAMs) of the GABAB receptor may have similar therapeutic effects as agonists with superior side-effect profiles. The present study behaviorally characterized N-([1R,2R,4S]-bicyclo[2.2.1]hept-2-yl)-2-methyl-5-(4-[trifluoromethyl]phenyl)-4-pyrimidinamine (BHF177), a GABAB receptor PAM, in mouse models of anxiety-like behavior, learning and memory. In addition, the effects of BHF177 were compared with the agonist baclofen. Unlike the anxiolytic chlordiazepoxide, baclofen (0.5, 1.5, and 2.5 mg/kg, intraperitoneally) and BHF177 (10, 20, and 40 mg/kg, orally) had no effect on anxiety-like behavior in the elevated plus maze, light/dark box, or the Vogel conflict test. Baclofen increased punished drinking in the Vogel conflict test, however this effect may be attributable to analgesic actions of baclofen. At the highest dose tested (2.5 mg/kg), baclofen-treated mice exhibited sedation-like effects (i.e., reduced locomotor activity) across many of the tests, whereas BHF177-treated mice exhibited no sedation-like effects. BHF177 exhibited pro-convulsion properties only in mice, but not in rats, indicating that this effect may be species-specific. At doses that were not sedative or pro-convulsant, baclofen and BHF177 had no selective effects on fear memory retrieval in contextual and cued fear conditioning or spatial learning and memory in the Barnes maze. These data suggest that BHF177 has little sedative activity, no anxiolytic-like profile, and minimal impairment of learning and memory in mice. PMID:23376712

Reversal of inhibition of putative dopaminergic neurons of the ventral tegmental area: Interaction of GABAB and D2 receptors

PubMed Central

Nimitvilai, Sudarat; Arora, Devinder S.; McElvain, Maureen A.; Brodie, Mark S.

2012-01-01

Neurons of the ventral tegmental area (VTA) are critical in the rewarding and reinforcing properties of drugs of abuse. Desensitization of VTA neurons to moderate extracellular concentrations of dopamine (DA) is dependent on protein kinase C (PKC) and intracellular calcium levels. This desensitization is called DA inhibition reversal (DIR), as it requires concurrent activation of D2 and D1-like receptors; activation of D2 receptors alone does not result in desensitization. Activation of other G-protein linked receptors can substitute for D1 activation. Like D2 receptors, GABAB receptors in the VTA are coupled to G-protein-linked potassium channels. In the present study, we examined interactions between a GABAB agonist, baclofen, and dopamine agonists, dopamine and quinpirole, to determine whether there was some interaction in the processes of desensitization of GABAB and D2 responses. Long-duration administration of baclofen alone produced reversal of the baclofen-induced inhibition indicative of desensitization, and this desensitization persisted for at least 60 min after baclofen washout. Desensitization to baclofen was dependent on protein kinase C. Dopamine inhibition was reduced for 30 min after baclofen-induced desensitization and conversely, the magnitude of baclofen inhibition was reduced for 30 min by long-duration application of dopamine, but not quinpirole. These results indicate that D2 and GABAB receptors share some protein kinase C-dependent mechanisms of receptor desensitization. PMID:22986166

Different in vitro and in vivo profiles of substituted 3-aminopropylphosphinate and 3-aminopropyl(methyl)phosphinate GABAB receptor agonists as inhibitors of transient lower oesophageal sphincter relaxation

PubMed Central

Lehmann, A; Antonsson, M; Aurell-Holmberg, A; Blackshaw, LA; Brändén, L; Elebring, T; Jensen, J; Kärrberg, L; Mattsson, JP; Nilsson, K; Oja, SS; Saransaari, P; von Unge, S

2012-01-01

BACKGROUND AND PURPOSE Gastro-oesophageal reflux is predominantly caused by transient lower oesophageal sphincter relaxation (TLOSR) and GABAB receptor stimulation inhibits TLOSR. Lesogaberan produces fewer CNS side effects than baclofen, which has been attributed to its affinity for the GABA transporter (GAT), the action of which limits stimulation of central GABAB receptors. To understand the structure–activity relationship for analogues of lesogaberan (3-aminopropylphosphinic acids), and corresponding 3-aminopropyl(methyl)phosphinic acids, we have compared representatives of these classes in different in vitro and in vivo models. EXPERIMENTAL APPROACH The compounds were characterized in terms of GABAB agonism in vitro. Binding to GATs and cellular uptake was done using rat brain membranes and slices respectively. TLOSR was measured in dogs, and CNS side effects were evaluated as hypothermia in mice and rats. KEY RESULTS 3-Aminopropylphosphinic acids inhibited TLOSR with a superior therapeutic index compared to 3-aminopropyl(methyl)phosphinic acids. This difference was most likely due to differential GAT-mediated uptake into brain cells of the former but not latter. In agreement, 3-aminopropyl(methyl)phosphinic acids were much more potent in producing hypothermia in rats even when administered i.c.v. CONCLUSIONS AND IMPLICATIONS An enhanced therapeutic window for 3-aminopropylphosphinic acids compared with 3-aminopropyl(methyl)phosphinic acids with respect to inhibition of TLOSR was observed and is probably mechanistically linked to neural cell uptake of the former but not latter group of compounds. These findings offer a platform for discovery of new GABAB receptor agonists for the treatment of reflux disease and other conditions where selective peripheral GABAB receptor agonism may afford therapeutic effects. PMID:21950457

GABAB-ergic motor cortex dysfunction in SSADH deficiency

PubMed Central

Cohen, Leonardo G.; Pearl, Phillip L.; Fritsch, Brita; Jung, Nikolai H.; Dustin, Irene; Theodore, William H.

2012-01-01

Objective: Succinic semialdehyde dehydrogenase (SSADH) deficiency is a rare autosomal recessive disorder of GABA degradation leading to elevations in brain GABA and γ-hydroxybutyric acid (GHB). The effect of chronically elevated GABA and GHB on cortical excitability is unknown. We hypothesized that use-dependent downregulation of GABA receptor expression would promote cortical disinhibition rather than inhibition, predominantly via presynaptic GABAergic mechanisms. Methods: We quantified the magnitude of excitation and inhibition in primary motor cortex (M1) in patients with SSADH deficiency, their parents (obligate heterozygotes), age-matched healthy young controls, and healthy adults using single and paired pulse transcranial magnetic stimulation (TMS). Results: Long interval intracortical inhibition was significantly reduced and the cortical silent period was significantly shortened in patients with SSADH deficiency compared to heterozygous parents and control groups. Conclusions: Since long interval intracortical inhibition and cortical silent period are thought to reflect GABAB receptor–mediated inhibitory circuits, our results point to a particularly GABAB-ergic motor cortex dysfunction in patients with SSADH deficiency. This human phenotype is consistent with the proposed mechanism of use-dependent downregulation of postsynaptic GABAB receptors in SSADH deficiency animal models. Additionally, the results suggest autoinhibition of GABAergic neurons. This first demonstration of altered GABAB-ergic function in patients with SSADH deficiency may help to explain clinical features of the disease, and suggest pathophysiologic mechanisms in other neurotransmitter-related disorders. Neurology® 2012;79:47–54 PMID:22722631

Electrophysiological actions of GABAB agonists and antagonists in rat dorso-lateral septal neurones in vitro.

PubMed

Bon, C; Galvan, M

1996-06-01

1. The actions of GABAB-receptor agonists and antagonists on rat dorso-lateral septal neurones in vitro were recorded with intracellular microelectrodes. 2. In the presence of 1 microM tetrodotoxin to prevent indirect neuronal effects caused by action potential-dependent neurotransmitter release, bath application of baclofen (0.1-30 microM) or SK&F 97541 (0.01-3 microM) evoked concentration-dependent hyperpolarizations which reversed close to the potassium equilibrium potential; the EC50S were 0.55 and 0.05 microM, respectively. No significant desensitization was observed during prolonged agonist exposure (< or = 10 min). 3. Hyperpolarizations induced by baclofen were antagonized in a competitive manner by the following GABAB-receptors antagonists (calculated pA2 values in parentheses): CGP 36742 (4.0), 2-OH saclofen (4.2), CGP 35348 (4.5), CGP 52432 (6.7) and CGP 55845A (8.3). Responses to SK&F 97541 were also antagonized by CGP 55845A (pA2 = 8.4). 4. The amplitude of the late, GABAB receptor-mediated inhibitory postsynaptic potential (i.p.s.p.) was reduced by the GABAB antagonists as follows (means +/- s.e.mean): CGP 55845A (1 microM) 91 +/- 5%, CGP 52432 (1 microM) 64 +/- 5%, CGP 35348 (100 microM) 82 +/- 5%, CGP 36742 (100 microM) 76 +/- 8%, and 2-OH saclofen (100 microM) 68 +/- 3%. 5. It is concluded that neurones in the rat dorso-lateral septal nucleus express conventional GABAB receptors, which are involved in the generation of slow inhibitory postsynaptic potentials. CGP 55845A is the most potent GABAB receptor antagonist described in this brain area.

Inhibition of presynaptic calcium transients in cortical inputs to the dorsolateral striatum by metabotropic GABAB and mGlu2/3 receptors

PubMed Central

Kupferschmidt, David A; Lovinger, David M

2015-01-01

Cortical inputs to the dorsolateral striatum (DLS) are dynamically regulated during skill learning and habit formation, and are dysregulated in disorders characterized by impaired action control. Therefore, a mechanistic investigation of the processes regulating corticostriatal transmission is key to understanding DLS-associated circuit function, behaviour and pathology. Presynaptic GABAB and group II metabotropic glutamate (mGlu2/3) receptors exert marked inhibitory control over corticostriatal glutamate release in the DLS, yet the signalling pathways through which they do so are unclear. We developed a novel approach using the genetically encoded calcium (Ca2+) indicator GCaMP6 to assess presynaptic Ca2+ in corticostriatal projections to the DLS. Using simultaneous photometric presynaptic Ca2+ and striatal field potential recordings, we report that relative to P/Q-type Ca2+ channels, N-type channels preferentially contributed to evoked presynaptic Ca2+ influx in motor cortex projections to, and excitatory transmission in, the DLS. Activation of GABAB or mGlu2/3 receptors inhibited both evoked presynaptic Ca2+ transients and striatal field potentials. mGlu2/3 receptor-mediated depression did not require functional N-type Ca2+ channels, but was attenuated by blockade of P/Q-type channels. These findings reveal presynaptic mechanisms of inhibitory modulation of corticostriatal function that probably contribute to the selection and shaping of behavioural repertoires. Key points Plastic changes at cortical inputs to the dorsolateral striatum (DLS) underlie skill learning and habit formation, so characterizing the mechanisms by which these inputs are regulated is important for understanding the neural basis of action control. We developed a novel approach using the genetically encoded calcium (Ca2+) indicator GCaMP6 and brain slice photometry to assess evoked presynaptic Ca2+ transients in cortical inputs to the DLS and study their regulation by GABAB and mGlu2

Different in vitro and in vivo profiles of substituted 3-aminopropylphosphinate and 3-aminopropyl(methyl)phosphinate GABA(B) receptor agonists as inhibitors of transient lower oesophageal sphincter relaxation.

PubMed

Lehmann, A; Antonsson, M; Aurell-Holmberg, A; Blackshaw, L A; Brändén, L; Elebring, T; Jensen, J; Kärrberg, L; Mattsson, J P; Nilsson, K; Oja, S S; Saransaari, P; von Unge, S

2012-03-01

Gastro-oesophageal reflux is predominantly caused by transient lower oesophageal sphincter relaxation (TLOSR) and GABA(B) receptor stimulation inhibits TLOSR. Lesogaberan produces fewer CNS side effects than baclofen, which has been attributed to its affinity for the GABA transporter (GAT), the action of which limits stimulation of central GABA(B) receptors. To understand the structure-activity relationship for analogues of lesogaberan (3-aminopropylphosphinic acids), and corresponding 3-aminopropyl(methyl)phosphinic acids, we have compared representatives of these classes in different in vitro and in vivo models. The compounds were characterized in terms of GABA(B) agonism in vitro. Binding to GATs and cellular uptake was done using rat brain membranes and slices respectively. TLOSR was measured in dogs, and CNS side effects were evaluated as hypothermia in mice and rats. 3-Aminopropylphosphinic acids inhibited TLOSR with a superior therapeutic index compared to 3-aminopropyl(methyl)phosphinic acids. This difference was most likely due to differential GAT-mediated uptake into brain cells of the former but not latter. In agreement, 3-aminopropyl(methyl)phosphinic acids were much more potent in producing hypothermia in rats even when administered i.c.v. An enhanced therapeutic window for 3-aminopropylphosphinic acids compared with 3-aminopropyl(methyl)phosphinic acids with respect to inhibition of TLOSR was observed and is probably mechanistically linked to neural cell uptake of the former but not latter group of compounds. These findings offer a platform for discovery of new GABA(B) receptor agonists for the treatment of reflux disease and other conditions where selective peripheral GABA(B) receptor agonism may afford therapeutic effects. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Kindling alters entorhinal cortex-hippocampal interaction by increased efficacy of presynaptic GABA(B) autoreceptors in layer III of the entorhinal cortex.

PubMed

Gloveli, Tengis; Behr, Joachim; Dugladze, Tamar; Kokaia, Zaal; Kokaia, Merab; Heinemann, Uwe

2003-08-01

We studied the effect of kindling, a model of temporal lobe epilepsy, on the frequency-dependent information transfer from the entorhinal cortex to the hippocampus in vitro. In control rats repetitive synaptic activation of layer III projection cells resulted in a frequency dependent depression of the synaptic transfer of action potentials to the hippocampus. One-to-two-days after kindling this effect was strongly reduced. Although no substantial change in synaptic inhibition upon single electrical stimulation was detected in kindled rats, there was a significant depression in the prolonged inhibition following high frequency stimulation. In kindled animals, paired-pulse depression (PPD) of stimulus-evoked IPSCs in layer III neurons was significantly stronger than in control rats. The increase of PPD is most likely caused by an increased presynaptic GABA(B) receptor-mediated autoinhibition. In kindled animals activation of presynaptic GABA(B) receptors by baclofen (10 microM) suppressed monosynaptic IPSCs significantly more than in control rats. In contrast, activation of postsynaptic GABA(B) receptors by baclofen was accompanied by comparable changes of the membrane conductance in both animal groups. Thus, in kindled animals activation of the layer III-CA1 pathway is facilitated by an increased GABA(B) receptor-mediated autoinhibition leading to an enhanced activation of the monosynaptic EC-CA1 pathway.

Pharmacologic Treatment with GABAB Receptor Agonist of Methamphetamine-Induced Cognitive Impairment in Mice

PubMed Central

Mizoguchi, Hiroyuki; Yamada, Kiyofumi

2011-01-01

Methamphetamine (METH) is a highly addictive drug, and addiction to METH has increased to epidemic proportions worldwide. Chronic use of METH causes psychiatric symptoms, such as hallucinations and delusions, and long-term cognitive deficits, which are indistinguishable from paranoid schizophrenia. The GABA receptor system is known to play a significant role in modulating the dopaminergic neuronal system, which is related to behavioral changes induced by drug abuse. However, few studies have investigated the effects of GABA receptor agonists on cognitive deficits induced by METH. In the present review, we show that baclofen, a GABA receptor agonist, is effective in treating METH-induced impairment of object recognition memory and prepulse inhibition (PPI) of the startle reflex, a measure of sensorimotor gating in mice. Acute and repeated treatment with METH induced a significant impairment of PPI. Furthermore, repeated but not acute treatment of METH resulted in a long-lasting deficit of object recognition memory. Baclofen, a GABAB receptor agonist, dose-dependently ameliorated the METH-induced PPI deficits and object recognition memory impairment in mice. On the other hand, THIP, a GABAA receptor agonist, had no effect on METH-induced cognitive deficits. These results suggest that GABAB receptors may constitute a putative new target in treating cognitive deficits in chronic METH users. PMID:21886573

The effects of acute multiple intraperitoneal injections of the GABAB receptor agonist baclofen on food intake in rats.

PubMed

Patel, Sunit M; Ebenezer, Ivor S

2008-12-28

This study was undertaken to examine the effects of acute repeated administration of the GABA(B) receptor agonist baclofen on food intake in rats. In Experiment 1, the effects of repeated intraperitoneal (i.p.) injections of the GABA(B) receptor agonist baclofen (1 and 2 mg/kg) at 2 h intervals were investigated on food intake in non-deprived male Wistar rats. Both doses of baclofen significantly increased food intake after the 1st injection (P<0.05), but had no effects on intake following the 2nd and 3rd injections. By contrast, in Experiment 2, diazepam (1 and 2 mg/kg, i.p.) significantly increased food intake (at least, P<0.05) after each of 3 injection separated by 2 h in non-deprived rats. These data show that tolerance occurs to the hyperphagic effects of baclofen with acute multiple injections, and may have important implications for future studies investigating the effects of GABA(B) receptor agonists on food intake and energy homeostasis.

L-baclofen-sensitive GABAB binding sites in the medial vestibular nucleus localized by immunocytochemistry

NASA Technical Reports Server (NTRS)

Holstein, G. R.; Martinelli, G. P.; Cohen, B.

1992-01-01

L-Baclofen-sensitive GABAB binding sites in the medial vestibular nucleus (MVN) were identified immunocytochemically and visualized ultrastructurally in L-baclofen-preinjected rats and monkeys, using a mouse monoclonal antibody with specificity for the p-chlorophenyl moiety of baclofen. Saline-preinjected animals showed no immunostain. In drug-injected animals, there was evidence for both pre- and postsynaptic GABAergic inhibition in MVN mediated by GABAB receptors. These neural elements could be utilized in control of velocity storage in the vestibulo-ocular reflex.

Behavioral impact of neurotransmitter-activated GPCRs: Muscarinic and GABAB receptors regulate C. elegans locomotion

PubMed Central

Dittman, Jeremy S; Kaplan, Joshua M

2008-01-01

Neurotransmitter released from presynaptic terminals activates both ligand-gated ion channels (ionotropic receptors) and a variety of G protein-coupled receptors (GPCRs). These neurotransmitter receptors are expressed on both pre- and postsynaptic cells. Thus, each neurotransmitter acts on multiple receptor classes, generating a large repertoire of physiological responses. The impact of many ionotropic receptors on neuronal activity and behavior has been clearly elucidated; however, much less is known about how neurotransmitter-gated GPCRs regulate neurons and circuits. In C. elegans, both Acetylcholine (ACh) and GABA are released in the nerve cord and mediate fast neuromuscular excitation and inhibition during locomotion. Here we identify a muscarinic receptor (GAR-2) and the GABAB receptor dimer (GBB-1/2) that detect synaptically released ACh and GABA, respectively. Both GAR-2 and GBB-1/2 inhibited cholinergic motor neurons when ACh and GABA levels were enhanced. Loss of either GPCR resulted in movement defects, suggesting that these receptors are activated during locomotion. When the negative feedback provided by GAR-2 was replaced with positive feedback, animals became highly sensitive to ACh levels and locomotion was severely impaired. Thus, conserved GPCRs act in the nematode motor circuit to provide negative feedback and to regulate locomotory behaviors that underlie navigation. PMID:18614679

Lateral Orbitofrontal Cortical Modulation on the Medial Prefrontal Cortex-Amygdala Pathway: Differential Regulation of Intra-Amygdala GABAA and GABAB Receptors.

PubMed

Chang, Chun-Hui

2017-07-01

The basolateral complex of the amygdala receives inputs from neocortical areas, including the medial prefrontal cortex and lateral orbitofrontal cortex. Earlier studies have shown that lateral orbitofrontal cortex activation exerts an inhibitory gating on medial prefrontal cortex-amygdala information flow. Here we examined the individual role of GABAA and GABAB receptors in this process. In vivo extracellular single-unit recordings were done in anesthetized rats. We searched amygdala neurons that fire in response to medial prefrontal cortex activation, tested lateral orbitofrontal cortex gating at different delays (lateral orbitofrontal cortex-medial prefrontal cortex delays: 25, 50, 100, 250, 500, and 1000 milliseconds), and examined differential contribution of GABAA and GABAB receptors with iontophoresis. Relative to baseline, lateral orbitofrontal cortex stimulation exerted an inhibitory modulatory gating on the medial prefrontal cortex-amygdala pathway and was effective up to a long delay of 500 ms (long-delay latencies at 100, 250, and 500 milliseconds). Moreover, blockade of intra-amygdala GABAA receptors with bicuculline abolished the lateral orbitofrontal cortex inhibitory gating at both short- (25 milliseconds) and long-delay (100 milliseconds) intervals, while blockade of GABAB receptors with saclofen reversed the inhibitory gating at long delay (100 milliseconds) only. Among the majority of the neurons examined (8 of 9), inactivation of either GABAA or GABAB receptors during baseline did not change evoked probability per se, suggesting that local feed-forward inhibitory mechanism is pathway specific. Our results suggest that the effect of lateral orbitofrontal cortex inhibitory modulatory gating was effective up to 500 milliseconds and that intra-amygdala GABAA and GABAB receptors differentially modulate the short- and long-delay lateral orbitofrontal cortex inhibitory gating on the medial prefrontal cortex-amygdala pathway. © The Author 2017

Brain region-selective cellular redistribution of mGlu5 but not GABA(B) receptors following methamphetamine-induced associative learning.

PubMed

Herrold, Amy A; Voigt, Robin M; Napier, T Celeste

2011-12-01

Alterations in receptor expression and distribution between cell surface and cytoplasm are means by which psychostimulants regulate neurotransmission. Metabotropic glutamate receptor group I, subtype 5 (mGluR5) and GABA(B) receptors (GABA(B) R) are critically involved in the development and expression of stimulant-induced behaviors, including conditioned place preference (CPP), an index of drug-seeking. However, it is not known if psychostimulant-induced CPP alters the trafficking of these receptors. To fill this gap, this study used methamphetamine (Meth)-induced CPP in rats to ascertain if receptor changes occur in limbic brain regions that regulate drug-seeking, the medial prefrontal cortex (mPFC), nucleus accumbens (NAc), and ventral pallidum (VP). To do so, ex vivo tissue was assessed for changes in expression and surface vs. intracellular distribution of mGluR5 and GABA(B) Rs. There was a decrease in the surface to intracellular ratio of mGluR5 in the mPFC in Meth-conditioned rats, commensurate with an increase in intracellular levels. mGluR5 levels in the NAc or the VP were unaltered. There were no changes for GABA(B) R in any brain region assayed. This ex vivo snapshot of metabotropic glutamate and GABA receptor cellular distribution following induction of Meth-induced CPP is the first report to determine if these receptors are differentially altered after Meth-induced CPP. The results suggest that this Meth treatment paradigm likely induced a compensatory change in mGluR5 surface to intracellular ratio such that the surface remains unaltered while an increase in intracellular protein occurred. Copyright © 2011 Wiley-Liss, Inc.

Selective Effects of Baclofen on Use-Dependent Modulation of GABAB Inhibition after Tetraplegia

PubMed Central

Barry, Melissa D.; Bunday, Karen L.; Chen, Robert

2013-01-01

Baclofen is a GABAB receptor agonist commonly used to relief spasticity related to motor disorders. The effects of baclofen on voluntary motor output are limited and not yet understood. Using noninvasive transcranial magnetic and electrical stimulation techniques, we examined electrophysiological measures probably involving GABAB (long-interval intracortical inhibition and the cortical silent period) and GABAA (short-interval intracortical inhibition) receptors, which are inhibitory effects mediated by subcortical and cortical mechanisms. We demonstrate increased active long-interval intracortical inhibition and prolonged cortical silent period during voluntary activity of an intrinsic finger muscle in humans with chronic incomplete cervical spinal cord injury (SCI) compared with age-matched controls, whereas resting long-interval intracortical inhibition was unchanged. However, long-term (∼6 years) use of baclofen decreased active long-interval intracortical inhibition to similar levels as controls but did not affect the duration of the cortical silent period. We found a correlation between signs of spasticity and long-interval intracortical inhibition in patients with SCI. Short-interval intracortical inhibition was decreased during voluntary contraction compared with rest but there was no effect of SCI or baclofen use. Together, these results demonstrate that baclofen selectively maintains use-dependent modulation of largely subcortical but not cortical GABAB neuronal pathways after human SCI. Thus, cortical GABAB circuits may be less sensitive to baclofen than spinal GABAB circuits. This may contribute to the limited effects of baclofen on voluntary motor output in subjects with motor disorders affected by spasticity. PMID:23904624

GABAB receptor-mediated, layer-specific synaptic plasticity reorganizes gamma-frequency neocortical response to stimulation

PubMed Central

Ainsworth, Matthew; Lee, Shane; Kaiser, Marcus; Simonotto, Jennifer; Kopell, Nancy J.

2016-01-01

Repeated presentations of sensory stimuli generate transient gamma-frequency (30–80 Hz) responses in neocortex that show plasticity in a task-dependent manner. Complex relationships between individual neuronal outputs and the mean, local field potential (population activity) accompany these changes, but little is known about the underlying mechanisms responsible. Here we show that transient stimulation of input layer 4 sufficient to generate gamma oscillations induced two different, lamina-specific plastic processes that correlated with lamina-specific changes in responses to further, repeated stimulation: Unit rates and recruitment showed overall enhancement in supragranular layers and suppression in infragranular layers associated with excitatory or inhibitory synaptic potentiation onto principal cells, respectively. Both synaptic processes were critically dependent on activation of GABAB receptors and, together, appeared to temporally segregate the cortical representation. These data suggest that adaptation to repetitive sensory input dramatically alters the spatiotemporal properties of the neocortical response in a manner that may both refine and minimize cortical output simultaneously. PMID:27118845

GABAB receptor-mediated, layer-specific synaptic plasticity reorganizes gamma-frequency neocortical response to stimulation.

PubMed

Ainsworth, Matthew; Lee, Shane; Kaiser, Marcus; Simonotto, Jennifer; Kopell, Nancy J; Whittington, Miles A

2016-05-10

Repeated presentations of sensory stimuli generate transient gamma-frequency (30-80 Hz) responses in neocortex that show plasticity in a task-dependent manner. Complex relationships between individual neuronal outputs and the mean, local field potential (population activity) accompany these changes, but little is known about the underlying mechanisms responsible. Here we show that transient stimulation of input layer 4 sufficient to generate gamma oscillations induced two different, lamina-specific plastic processes that correlated with lamina-specific changes in responses to further, repeated stimulation: Unit rates and recruitment showed overall enhancement in supragranular layers and suppression in infragranular layers associated with excitatory or inhibitory synaptic potentiation onto principal cells, respectively. Both synaptic processes were critically dependent on activation of GABAB receptors and, together, appeared to temporally segregate the cortical representation. These data suggest that adaptation to repetitive sensory input dramatically alters the spatiotemporal properties of the neocortical response in a manner that may both refine and minimize cortical output simultaneously.

The tuberal lateral hypothalamus is a major target for GABAA--but not GABAB-mediated control of food intake.

PubMed

Turenius, Christine I; Charles, Jonathan R; Tsai, Donna H; Ebersole, Priscilla L; Htut, Myat H; Ngo, Phuong T; Lara, Raul N; Stanley, B Glenn

2009-08-04

The lateral hypothalamus (LH) is a site of integration for control mechanisms of feeding behavior as it has extensive reciprocal connections with multiple intrahypothalamic and extrahypothalamic brain areas. Evidence suggests that blockade of ionotropric gamma-aminobutyric acid (GABA) receptors in the LH elicits eating in satiated rats. To determine whether this GABA(A) receptor antagonist effect is specific to the LH, the antagonist picrotoxin was injected into one of six nearby sites and food intake was measured. Picrotoxin at 133 pmol elicited eating in the LH, but not in surrounding sites (thalamus, lateral preoptic area, ventral tegmental area, dorsomedial hypothalamus, and entopeduncular nucleus). More specifically, picrotoxin injected into the tuberal LH (tLH) elicited eating, but was ineffective when injected into the anterior or posterior LH. We also investigated whether GABA(B) receptors in the LH participated in the control of food intake and found that neither blockade nor activation of these receptors under multiple conditions changed food intake. Collectively, our findings suggest that GABA(A) but not GABA(B) receptors in the tLH act to suppress feeding behavior.

Effects of acute administration of the GABA(B) receptor agonist baclofen on behavioral flexibility in rats

PubMed Central

Beas, B. Sofia; Setlow, Barry; Bizon, Jennifer L.

2016-01-01

RATIONALE The ability to adjust response strategies when faced with changes in the environment is critical for normal adaptive behavior. Such behavioral flexibility is compromised by experimental disruption of cortical GABAergic signaling, as well as in conditions such as schizophrenia and normal aging that are characterized by cortical hyperexcitability. The current studies were designed to determine whether stimulation of GABAergic signaling using the GABA(B) receptor agonist baclofen can facilitate behavioral flexibility. METHODS Male Fischer 344 rats were trained in a set-shifting task in which they learned to discriminate between two response levers to obtain a food reward. Correct levers were signaled in accordance with two distinct response rules (Rule 1: correct lever signaled by a cue light; Rule 2: correct lever signaled by its left/right position). The order of rule presentation varied, but they were always presented sequentially, with the trials and errors to reach criterion performance on the second (set shift) rule providing the measure of behavioral flexibility. Experiments determined the effects of the GABA(B) receptor agonist baclofen (i.p., 0, 1.0, 2.5 and 4.0 mg/kg) administered acutely before the shift to the second rule. RESULTS Baclofen enhanced set-shifting performance. Control experiments demonstrated that this enhancement was not simply due to improved discrimination learning, nor was it due to impaired recall of the initial discrimination rule. CONCLUSIONS The results demonstrate that baclofen can facilitate behavioral flexibility, suggesting that GABA(B) receptor agonists may have utility for treating behavioral dysfunction in neuropsychiatric disorders. PMID:27256354

Effects of acute administration of the GABA(B) receptor agonist baclofen on behavioral flexibility in rats.

PubMed

Beas, B Sofia; Setlow, Barry; Bizon, Jennifer L

2016-07-01

The ability to adjust response strategies when faced with changes in the environment is critical for normal adaptive behavior. Such behavioral flexibility is compromised by experimental disruption of cortical GABAergic signaling, as well as in conditions such as schizophrenia and normal aging that are characterized by cortical hyperexcitability. The current studies were designed to determine whether stimulation of GABAergic signaling using the GABA(B) receptor agonist baclofen can facilitate behavioral flexibility. Male Fischer 344 rats were trained in a set-shifting task in which they learned to discriminate between two response levers to obtain a food reward. Correct levers were signaled in accordance with two distinct response rules (rule 1: correct lever signaled by a cue light; rule 2: correct lever signaled by its left/right position). The order of rule presentation varied, but they were always presented sequentially, with the trials and errors to reach criterion performance on the second (set shift) rule providing the measure of behavioral flexibility. Experiments determined the effects of the GABA(B) receptor agonist baclofen (intraperitoneal, 0, 1.0, 2.5, and 4.0 mg/kg) administered acutely before the shift to the second rule. Baclofen enhanced set-shifting performance. Control experiments demonstrated that this enhancement was not simply due to improved discrimination learning, nor was it due to impaired recall of the initial discrimination rule. The results demonstrate that baclofen can facilitate behavioral flexibility, suggesting that GABA(B) receptor agonists may have utility for treating behavioral dysfunction in neuropsychiatric disorders.

Effects of intraperitoneal administration of the GABAB receptor positive allosteric modulator 2,6-di tert-butyl-4-(2-hydroxy-2,2-dimethyl-propyl)-phenol (CGP7930) on food intake in non-deprived rats.

PubMed

Ebenezer, Ivor S

2012-09-05

γ-Aminobutyric acid-(B) (GABA(B)) receptor positive allosteric modulators (PAMs) act on an allosteric site on the GABA(B) receptor to potentiate the effects of GABA and GABA(B) receptor agonists. It has previously been demonstrated that the GABA(B) receptor agonist baclofen increases food intake in non-deprived rats. The aim of this study was to investigate whether the GABA(B) receptor PAM 2,6-di tert-butyl-4-(2-hydroxy-2,2-dimethyl-propyl)-phenol (CGP7930) would (i) increase food intake, and (ii) potentiate the hyperphagic effects of baclofen in rats. In Experiment 1, the effects of intraperitoneal (i.p.) administration of CGP7930 (1, 6 and 12 mg/kg) was investigated on food intake in non-deprived male Wistar rats. The 12 mg/kg dose of CGP7930 significantly increased cumulative food intake 30, 60 and 120 min (P<0.05, in each case) after administration. The 1 and 6 mg/kg doses were without effect. In Experiment 2, the effects of pretreatment with CGP7930 (6 mg/kg; i.p.) 5 min prior to administration of baclofen (2mg/kg, i.p.) was investigated on 30min cumulative food intake in non-deprived male Wistar rats. Baclofen (2mg/kg) significantly increased food intake compared with vehicle treatment (P<0.01). CGP7930 (6 mg/kg) had no effect on feeding. However, pretreatment with CGP7930 (6 mg/kg) significantly potentiated the hyperphagic effects of baclofen (2mg/kg) (P<0.01). These findings show that CGP7930 increases food intake and enhances the hyperphagic effects of baclofen, and are consistent with in vitro studies that suggest that it potentiates the effects of endogenous GABA and GABA(B) receptor agonists by allosteric modulation of the GABA(B) receptor. Copyright © 2012 Elsevier B.V. All rights reserved.

Reversal of disease-related pathologies in the fragile X mouse model by selective activation of GABAB receptors with arbaclofen.

PubMed

Henderson, Christina; Wijetunge, Lasani; Kinoshita, Mika Nakamoto; Shumway, Matthew; Hammond, Rebecca S; Postma, Friso R; Brynczka, Christopher; Rush, Roger; Thomas, Alexia; Paylor, Richard; Warren, Stephen T; Vanderklish, Peter W; Kind, Peter C; Carpenter, Randall L; Bear, Mark F; Healy, Aileen M

2012-09-19

Fragile X syndrome (FXS), the most common inherited cause of intellectual disability and autism, results from the transcriptional silencing of FMR1 and loss of the mRNA translational repressor protein fragile X mental retardation protein (FMRP). Patients with FXS exhibit changes in neuronal dendritic spine morphology, a pathology associated with altered synaptic function. Studies in the mouse model of fragile X have shown that loss of FMRP causes excessive synaptic protein synthesis, which results in synaptic dysfunction and altered spine morphology. We tested whether the pharmacologic activation of the γ-aminobutyric acid type B (GABA(B)) receptor could correct or reverse these phenotypes in Fmr1-knockout mice. Basal protein synthesis, which is elevated in the hippocampus of Fmr1-knockout mice, was corrected by the in vitro application of the selective GABA(B) receptor agonist STX209 (arbaclofen, R-baclofen). STX209 also reduced to wild-type values the elevated AMPA receptor internalization in Fmr1-knockout cultured neurons, a known functional consequence of increased protein synthesis. Acute administration of STX209 in vivo, at doses that modify behavior, decreased mRNA translation in the cortex of Fmr1-knockout mice. Finally, the chronic administration of STX209 in juvenile mice corrected the increased spine density in Fmr1-knockout mice without affecting spine density in wild-type mice. Thus, activation of the GABA(B) receptor with STX209 corrected synaptic abnormalities considered central to fragile X pathophysiology, a finding that suggests that STX209 may be a potentially effective therapy to treat the core symptoms of FXS.

The interaction between hippocampal GABA-B and cannabinoid receptors upon spatial change and object novelty discrimination memory function.

PubMed

Nasehi, Mohammad; Alaghmandan-Motlagh, Niyousha; Ebrahimi-Ghiri, Mohaddeseh; Nami, Mohammad; Zarrindast, Mohammad-Reza

2017-10-01

Previous studies have postulated functional links between GABA and cannabinoid systems in the hippocampus. The aim of the present study was to investigate any possible interaction between these systems in spatial change and object novelty discrimination memory consolidation in the dorsal hippocampus (CA1 region) of NMRI mice. Assessment of the spatial change and object novelty discrimination memory function was carried out in a non-associative task. The experiment comprised mice exposure to an open field containing five objects followed by the examination of their reactivity to object displacement (spatial change) and object substitution (object novelty) after three sessions of habituation. Our results showed that the post-training intraperitoneal administration of the higher dose of ACPA (0.02 mg/kg) impaired both spatial change and novelty discrimination memory functions. Meanwhile, the higher dose of GABA-B receptor agonist, baclofen, impaired the spatial change memory by itself. Moreover, the post-training intra-CA1 microinjection of a subthreshold dose of baclofen increased the ACPA effect on spatial change and novelty discrimination memory at a lower and higher dose, respectively. On the other hand, the lower and higher but not mid-level doses of GABA-B receptor antagonist, phaclofen, could reverse memory deficits induced by ACPA. However, phaclofen at its mid-level dose impaired the novelty discrimination memory and whereas the higher dose impaired the spatial change memory. Based on our findings, GABA-B receptors in the CA1 region appear to modulate the ACPA-induced cannabinoid CB1 signaling upon spatial change and novelty discrimination memory functions.

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.

Comparison of the effect of the GABAB receptor agonist, baclofen, and the positive allosteric modulator of the GABAB receptor, GS39783, on alcohol self-administration in three different lines of alcohol-preferring rats

PubMed Central

Maccioni, Paola; Zaru, Alessandro; Loi, Barbara; Lobina, Carla; Carai, Mauro A.M.; Gessa, Gian Luigi; Capra, Alessandro; Mugnaini, Claudia; Pasquini, Serena; Corelli, Federico; Hyytiä, Petri; Lumeng, Lawrence; Colombo, Giancarlo

2012-01-01

may resemble the differential effectiveness of pharmacotherapies among the different typologies of human alcoholics; (d) the GABAB receptor is part of the neural substrate mediating the reinforcing and motivational properties of alcohol. PMID:22486245

Epigenetic regulation of dorsal raphe GABA(B1a) associated with isolation-induced abnormal responses to social stimulation in mice.

PubMed

Araki, Ryota; Hiraki, Yosuke; Nishida, Shoji; Kuramoto, Nobuyuki; Matsumoto, Kinzo; Yabe, Takeshi

2016-02-01

In isolation-reared mice, social encounter stimulation induces locomotor hyperactivity and activation of the dorsal raphe nucleus (DRN), suggesting that dysregulation of dorsal raphe function may be involved in abnormal behaviors. In this study, we examined the involvement of dorsal raphe GABAergic dysregulation in the abnormal behaviors of isolation-reared mice. We also studied an epigenetic mechanism underlying abnormalities of the dorsal raphe GABAergic system. Both mRNA and protein levels of GABA(B1a), a GABA(B) receptor subunit, were increased in the DRN of isolation-reared mice, compared with these levels in group-reared mice. In contrast, mRNA levels for other GABAergic system-related genes (GABA(A) receptor α1, β2 and γ2 subunits, GABA(B) receptor 1b and 2 subunits, and glutamate decarboxylase 67 and 65) were unchanged. Intra-DRN microinjection of 0.06 nmol baclofen (a GABA(B) receptor agonist) exacerbated encounter-induced hyperactivity and aggressive behavior, while microinjection of 0.3 nmol phaclofen (a GABA(B) receptor antagonist) attenuated encounter-induced hyperactivity and aggressive behavior in isolation-reared mice. Furthermore, microinjection of 0.06 nmol baclofen elicited encounter-induced hyperactivity in group-reared mice. Neither baclofen nor phaclofen affected immobility time in the forced swim test and hyperactivity in a novel environment of isolation reared mice. Bisulfite sequence analyses revealed that the DNA methylation level of the CpG island around the transcription start site (TSS) of GABA(B1a) was decreased in the DRN of isolation-reared mice. Chromatin immunoprecipitation analysis showed that histone H3 was hyperacetylated around the TSS of GABA(B1a) in the DRN of isolation-reared mice. These findings indicate that an increase in dorsal raphe GABA(B1a) expression via epigenetic regulation is associated with abnormal responses to social stimulation such as encounter-induced hyperactivity and aggressive behavior in isolation

Neuroendocrine response to GABA-B receptor agonism in alcohol-dependent individuals: Results from a combined outpatient and human laboratory experiment.

PubMed

Farokhnia, Mehdi; Sheskier, Mikela B; Lee, Mary R; Le, April N; Singley, Erick; Bouhlal, Sofia; Ton, Timmy; Zhao, Zhen; Leggio, Lorenzo

2018-04-14

Gamma-aminobutyric acid (GABA), the main inhibitory neurotransmitter in the nervous system, plays an important role in biobehavioral processes that regulate alcohol seeking, food intake, and stress response. The metabotropic GABA-B receptor has been investigated as a potential therapeutic target for alcohol use disorder, by using orthosteric agonists (e.g., baclofen) and positive allosteric modulators. Whether and how pharmacological manipulation of the GABA-B receptor, in combination with alcohol intake, may affect feeding- and stress-related neuroendocrine pathways remains unknown. In the present randomized, double-blind, placebo-controlled study, thirty-four alcohol-dependent individuals received baclofen (30 mg/day) or placebo in a naturalistic outpatient setting for one week, and then performed a controlled laboratory experiment which included alcohol cue-reactivity, fixed-dose priming, and self-administration procedures. Blood samples were collected, and the following neuroendocrine markers were measured: ghrelin, leptin, amylin, glucagon-like peptide-1 (GLP-1), insulin, prolactin, thyroid-stimulating hormone, growth hormone, cortisol, and adrenocorticotropic hormone (ACTH). During the outpatient phase, baclofen significantly increased blood concentrations of acyl-ghrelin (p = 0.01), leptin (p = 0.01), amylin (p = 0.004), and GLP-1 (p = 0.02). Significant drug × time-point interaction effects for amylin (p = 0.001) and insulin (p = 0.03), and trend-level interaction effects for GLP-1 (p = 0.06) and ACTH (p = 0.10) were found during the laboratory experiment. Baclofen, compared to placebo, had no effect on alcohol drinking in this study (p's ≥ 0.05). Together with previous studies, these findings shed light on the role of the GABAergic system and GABA-B receptors in the shared neurobiology of alcohol-, feeding-, and stress-related behaviors. Copyright © 2018. Published by Elsevier Ltd.

GABAB Receptor Agonist R-Baclofen Reverses Social Deficits and Reduces Repetitive Behavior in Two Mouse Models of Autism

PubMed Central

Silverman, J L; Pride, M C; Hayes, J E; Puhger, K R; Butler-Struben, H M; Baker, S; Crawley, J N

2015-01-01

Autism spectrum disorder (ASD) is diagnosed by two core behavioral criteria, unusual reciprocal social interactions and communication, and stereotyped, repetitive behaviors with restricted interests. Excitatory/inhibitory imbalance is a prominent hypothesis for the etiology of autism. The selective GABAB receptor agonist R-baclofen previously reversed social deficits and reduced repetitive behaviors in a mouse model of Fragile X syndrome, and Arbaclofen improved some clinical symptoms in some Fragile X and ASD patients. To evaluate R-baclofen in a broader range of mouse models of ASD, we tested both the R-baclofen enantiomer and the less potent S-baclofen enantiomer in two inbred strains of mice that display low sociability and/or high repetitive or stereotyped behaviors. R-baclofen treatment reversed social approach deficits in BTBR T+ Itpr3tf/J (BTBR), reduced repetitive self-grooming and high marble burying scores in BTBR, and reduced stereotyped jumping in C58/J (C58), at nonsedating doses. S-baclofen produced minimal effects at the same doses. These findings encourage investigations of R-baclofen in other preclinical model systems. Additional clinical studies may be warranted to further evaluate the hypothesis that the GABAB receptor represents a promising pharmacological target for treating appropriately stratified subsets of individuals with ASD. PMID:25754761

GABAB Receptor Agonist R-Baclofen Reverses Social Deficits and Reduces Repetitive Behavior in Two Mouse Models of Autism.

PubMed

Silverman, J L; Pride, M C; Hayes, J E; Puhger, K R; Butler-Struben, H M; Baker, S; Crawley, J N

2015-08-01

Autism spectrum disorder (ASD) is diagnosed by two core behavioral criteria, unusual reciprocal social interactions and communication, and stereotyped, repetitive behaviors with restricted interests. Excitatory/inhibitory imbalance is a prominent hypothesis for the etiology of autism. The selective GABAB receptor agonist R-baclofen previously reversed social deficits and reduced repetitive behaviors in a mouse model of Fragile X syndrome, and Arbaclofen improved some clinical symptoms in some Fragile X and ASD patients. To evaluate R-baclofen in a broader range of mouse models of ASD, we tested both the R-baclofen enantiomer and the less potent S-baclofen enantiomer in two inbred strains of mice that display low sociability and/or high repetitive or stereotyped behaviors. R-baclofen treatment reversed social approach deficits in BTBR T+ Itpr3tf/J (BTBR), reduced repetitive self-grooming and high marble burying scores in BTBR, and reduced stereotyped jumping in C58/J (C58), at nonsedating doses. S-baclofen produced minimal effects at the same doses. These findings encourage investigations of R-baclofen in other preclinical model systems. Additional clinical studies may be warranted to further evaluate the hypothesis that the GABAB receptor represents a promising pharmacological target for treating appropriately stratified subsets of individuals with ASD.

The GABAB receptor agonist, baclofen, contributes to three distinct varieties of amnesia in the human brain - A detailed case report.

PubMed

Zeman, Adam; Hoefeijzers, Serge; Milton, Fraser; Dewar, Michaela; Carr, Melanie; Streatfield, Claire

2016-01-01

We describe a patient in whom long-term, therapeutic infusion of the selective gamma-amino-butyric acid type B (GABAB) receptor agonist, baclofen, into the cerebrospinal fluid (CSF) gave rise to three distinct varieties of memory impairment: i) repeated, short periods of severe global amnesia, ii) accelerated long-term forgetting (ALF), evident over intervals of days and iii) a loss of established autobiographical memories. This pattern of impairment has been reported in patients with temporal lobe epilepsy (TLE), in particular the subtype of Transient Epileptic Amnesia (TEA). The amnesic episodes and accelerated forgetting remitted on withdrawal of baclofen, while the autobiographical amnesia (AbA) persisted. This exceptional case highlights the occurrence of 'non-standard' forms of human amnesia, reflecting the biological complexity of memory processes. It suggests a role for GABAB signalling in the modulation of human memory over multiple time-scales and hints at its involvement in 'epileptic amnesia'. Copyright © 2015 Elsevier Ltd. All rights reserved.

Separate and combined effects of the GABAB agonist baclofen and Δ9-THC in humans discriminating Δ9-THC

PubMed Central

Lile, Joshua A.; Kelly, Thomas H.; Hays, Lon R.

2012-01-01

Background Our previous research with the GABA reuptake inhibitor tiagabine suggested the involvement GABA in the interoceptive effects of Δ9-THC. The aim of the present study was to determine the potential involvement of the GABAB receptor subtype by assessing the separate and combined effects of the GABAB-selective agonist baclofen and Δ9-THC using pharmacologically specific drug-discrimination procedures. Methods Eight cannabis users learned to discriminate 30 mg oral Δ9-THC from placebo and then received baclofen (25 and 50 mg), Δ9-THC (5, 15 and 30 mg) and placebo, alone and in combination. Self-report, task performance and physiological measures were also collected. Results Δ9-THC functioned as a discriminative stimulus, produced subjective effects typically associated with cannabinoids (e.g., High, Stoned, Like Drug), elevated heart rate and impaired rate and accuracy on a psychomotor performance task. Baclofen alone (50 mg) substituted for the Δ9-THC discriminative stimulus, and both baclofen doses shifted the discriminative-stimulus effects of Δ9-THC leftward/upward. Similar results were observed on other cannabinoid-sensitive outcomes, although baclofen generally did not engender Δ9-THC-like subjective responses when administered alone. Conclusions These results suggest that the GABAB receptor subtype is involved in the abuse-related effects of Δ9-THC, and that GABAB receptors were responsible, at least in part, for the effects of tiagabine-induced elevated GABA on cannabinoid-related behaviors in our previous study. Future research should test GABAergic compounds selective for other GABA receptor subtypes (i.e., GABAA) to determine the contribution of the different GABA receptors in the effects of Δ9-THC, and by extension cannabis, in humans. PMID:22699093

Chronic baclofen desensitizes GABA(B)-mediated G-protein activation and stimulates phosphorylation of kinases in mesocorticolimbic rat brain.

PubMed

Keegan, Bradley M T; Beveridge, Thomas J R; Pezor, Jeffrey J; Xiao, Ruoyu; Sexton, Tammy; Childers, Steven R; Howlett, Allyn C

2015-08-01

The GABAB receptor is a therapeutic target for CNS and neuropathic disorders; however, few preclinical studies have explored effects of chronic stimulation. This study evaluated acute and chronic baclofen treatments on GABAB-activated G-proteins and signaling protein phosphorylation as indicators of GABAB signaling capacity. Brain sections from rats acutely administered baclofen (5 mg/kg, i.p.) showed no significant differences from controls in GABAB-stimulated GTPγS binding in any brain region, but displayed significantly greater phosphorylation/activation of focal adhesion kinase (pFAK(Tyr397)) in mesocorticolimbic regions (caudate putamen, cortex, hippocampus, thalamus) and elevated phosphorylated/activated glycogen synthase kinase 3-β (pGSK3β(Tyr216)) in the prefrontal cortex, cerebral cortex, caudate putamen, nucleus accumbens, thalamus, septum, and globus pallidus. In rats administered chronic baclofen (5 mg/kg, t.i.d. for five days), GABAB-stimulated GTPγS binding was significantly diminished in the prefrontal cortex, septum, amygdala, and parabrachial nucleus compared to controls. This effect was specific to GABAB receptors: there was no effect of chronic baclofen treatment on adenosine A1-stimulated GTPγS binding in any region. Chronically-treated rats also exhibited increases in pFAK(Tyr397) and pGSK3β(Tyr216) compared to controls, and displayed wide-spread elevations in phosphorylated dopamine- and cAMP-regulated phosphoprotein-32 (pDARPP-32(Thr34)) compared to acutely-treated or control rats. We postulate that those neuroadaptive effects of GABAB stimulation mediated by G-proteins and their sequelae correlate with tolerance to several of baclofen's effects, whereas sustained signaling via kinase cascades points to cross-talk between GABAB receptors and alternative mechanisms that are resistant to desensitization. Both desensitized and sustained signaling pathways should be considered in the development of pharmacotherapies targeting the GABA

Gi-Coupled γ-Aminobutyric Acid–B Receptors Cross-Regulate Phospholipase C and Calcium in Airway Smooth Muscle

PubMed Central

Mizuta, Kentaro; Mizuta, Fumiko; Xu, Dingbang; Masaki, Eiji; Panettieri, Reynold A.

2011-01-01

γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system, and exerts its actions via both ionotropic (GABAA) and metabotropic (GABAB) receptors. Although the functional expression of GABAB receptors coupled to the Gi protein was reported for airway smooth muscle, the role of GABAB receptors in airway responsiveness remains unclear. We investigated whether Gi-coupled GABAB receptors cross-regulate phospholipase C (PLC), an enzyme classically regulated by Gq-coupled receptors in human airway smooth muscle cells. Both the GABAB-selective agonist baclofen and the endogenous ligand GABA significantly increased the synthesis of inositol phosphate, whereas GABAA receptor agonists, muscimol, and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol exerted no effect. The baclofen-induced synthesis of inositol phosphate and transient increases in [Ca2+]i were blocked by CGP35348 and CGP55845 (selective GABAB antagonists), pertussis toxin (PTX, which inactivates the Gi protein), gallein (a Gβγ signaling inhibitor), U73122 (an inhibitor of PLC-β), and xestospongin C, an inositol 1,4,5-triphosphate receptor blocker. Baclofen also potentiated the bradykinin-induced synthesis of inositol phosphate and transient increases in [Ca2+]i, which were blocked by CGP35348 or PTX. Moreover, baclofen potentiated the substance P–induced contraction of airway smooth muscle in isolated guinea pig tracheal rings. In conclusion, the stimulation of GABAB receptors in human airway smooth muscle cells rapidly mobilizes intracellular Ca2+ stores by the synthesis of inositol phosphate via the activation of PLC-β, which is stimulated by Gβγ protein liberated from Gi proteins coupled to GABAB receptors. Furthermore, crosstalk between GABAB receptors and Gq-coupled receptors potentiates the synthesis of inositol phosphate, transient increases in [Ca2+]i, and smooth muscle contraction through Gi proteins. PMID:21719794

The effects of intraperitoneal administration of the GABA(B) receptor agonist baclofen on food intake in CFLP and C57BL/6 mice.

PubMed

Ebenezer, Ivor S; Prabhaker, Monika

2007-08-13

The effects of the GABA(B) receptor agonist baclofen were investigated on food intake in non-deprived CFLP and C57BL/6 mice. In Experiment 1, baclofen (1-8 mg /kg) administered i.p. to CFLP mice, produced a dose-related increase in food intake. The 4 and 8 mg/kg doses produced significant increases in cumulative feeding when measure 120 min after administration (at least P < 0.05, in each case). In Experiment 2, baclofen (1-10 mg/kg), administered intraperitoneally (i.p.) to C57BL/6 mice, also produced a dose-related increase in food intake. The 4 mg/kg dose of baclofen significantly increased cumulative food intake at 60 min (P < 0.05), while the 2 and 4 mg/kg doses significantly increased cumulative food intake at 120 min (P < 0.01, in each case). The 10mg/kg dose was without effect. These data show that systemic administration of the GABA(B) agonist baclofen produces an increase in food consumption in two different strains of mice and extend previous observations made in rat to another rodent species.

Distribution and properties of GABA(B) antagonist [3H]CGP 62349 binding in the rhesus monkey thalamus and basal ganglia and the influence of lesions in the reticular thalamic nucleus.

PubMed

Ambardekar, A V; Ilinsky, I A; Forestl, W; Bowery, N G; Kultas-Ilinsky, K

1999-01-01

GABA(B) receptors are believed to be associated with the efferents of the nucleus reticularis thalami, which is implicated in the regulation of activity in the thalamocortical-corticothalamic circuit and plays a role in absence seizures. Yet, the distribution of GABA(B) receptors in the thalamus has only been studied in the rat, and there is no comparable information in primates. The potent GABA(B) receptor antagonist [3H]CGP 62349 was used to study the distribution and binding properties of the receptor in control monkeys and those with small ibotenic acid lesions in the anterodorsal segment of the nucleus reticularis thalami. Eight-micrometer-thick cryostat sections of the fresh frozen brains were incubated in the presence of varying concentrations of the ligand. Autoradiographs were analysed using a quantitative image analysis technique, and binding parameters were calculated for select thalamic nuclei as well as basal ganglia structures present in the same sections. The overall number of GABA(B) binding sites in the monkey thalamus and basal ganglia was several-fold higher than previously reported values for the rat. In the thalamus, the receptors were distributed rather uniformly and the binding densities and affinities were high (Bmax range of 245.5-437.9 fmol/ mg of tissue, Kd range of 0.136-0.604 nM). In the basal ganglia, the number of binding sites and the affinities were lower (Bmax range of 51.1-244.2 fmol/mg of tissue; K(d) range of 0.416-1.394 nM), and the differences between nuclei were more pronounced, with striatum and substantia nigra pars compacta displaying the highest binding densities. Seven days post-lesion, a 20-30% decrease in Bmax values (P < 0.05) was found in the nuclei receiving input from the lesioned nucleus reticularis thalami sector (the mediodorsal nucleus and densicellular and magnocellular parts of the ventral anterior nucleus) without changes in affinity. No significant changes were detected in any other structures. The results

Gi-coupled γ-aminobutyric acid-B receptors cross-regulate phospholipase C and calcium in airway smooth muscle.

PubMed

Mizuta, Kentaro; Mizuta, Fumiko; Xu, Dingbang; Masaki, Eiji; Panettieri, Reynold A; Emala, Charles W

2011-12-01

γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system, and exerts its actions via both ionotropic (GABA(A)) and metabotropic (GABA(B)) receptors. Although the functional expression of GABA(B) receptors coupled to the G(i) protein was reported for airway smooth muscle, the role of GABA(B) receptors in airway responsiveness remains unclear. We investigated whether G(i)-coupled GABA(B) receptors cross-regulate phospholipase C (PLC), an enzyme classically regulated by G(q)-coupled receptors in human airway smooth muscle cells. Both the GABA(B)-selective agonist baclofen and the endogenous ligand GABA significantly increased the synthesis of inositol phosphate, whereas GABA(A) receptor agonists, muscimol, and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol exerted no effect. The baclofen-induced synthesis of inositol phosphate and transient increases in [Ca(2+)](i) were blocked by CGP35348 and CGP55845 (selective GABA(B) antagonists), pertussis toxin (PTX, which inactivates the G(i) protein), gallein (a G(βγ) signaling inhibitor), U73122 (an inhibitor of PLC-β), and xestospongin C, an inositol 1,4,5-triphosphate receptor blocker. Baclofen also potentiated the bradykinin-induced synthesis of inositol phosphate and transient increases in [Ca(2+)](i), which were blocked by CGP35348 or PTX. Moreover, baclofen potentiated the substance P-induced contraction of airway smooth muscle in isolated guinea pig tracheal rings. In conclusion, the stimulation of GABA(B) receptors in human airway smooth muscle cells rapidly mobilizes intracellular Ca(2+) stores by the synthesis of inositol phosphate via the activation of PLC-β, which is stimulated by G(βγ) protein liberated from G(i) proteins coupled to GABA(B) receptors. Furthermore, crosstalk between GABA(B) receptors and G(q)-coupled receptors potentiates the synthesis of inositol phosphate, transient increases in [Ca(2+)](i), and smooth muscle contraction through G

Effect of GABAB receptor antagonists (CGP 35348 and CGP 55845) on serum interleukin 6 and 18 concentrations in albino mice following neonatal hypoxia ischemia insult.

PubMed

Gillani, Quratulane; Ali, Muhammad; Iqbal, Furhan

2016-09-01

Interleukin (IL) 6 and 18 plays an important role in inflammatory response following hypoxia ischemia encephalopathy (HIE). Present study was designed to demonstrate the effect of two GABAB receptor antagonists (CGP 35348 and 55845), respectively, on the serum IL6 and IL 18 concentrations in albino mice. Albino mice pups (of both genders) were subjected to Murine model of hypoxia-ischemia encephalopathy on postnatal day 10 (right common carotid artery was ligated followed by 8% hypoxia for 25 minutes). After neonatal brain damage and following weaning, mice were divided in three groups, in gender specific manner, and fed on normal rodent diet till they were 13 week old. At this time point, group 1 received intraperitonial saline solution (control group), group 2 was supplemented with CGP 35348 (1mg/ml solvent/Kg body weight) and group 3 with CGP 55845 (1mg/ml solvent/Kg body weight), intraperitonially, for 12 days and IL 6 and 18 concentrations were determined in serum by ELISA. It was observed that CGP 35348 supplementation resulted in reduced interlukin-6 and interlukin-18 concentrations in male albino mice. While CGP 55845 supplementation increased IL-6 and IL-18 concentrations in female albino mice following HIE. Our results are indicating that GABAB receptor antagonist's supplementation affects IL concentrations in albino mice in a gender specific manner following neonatal brain damage and can be further explored for the treatments of hypoxia ischemia associated neurological ailments.

Anticonvulsant effects of structurally diverse GABA(B) positive allosteric modulators in the DBA/2J audiogenic seizure test: Comparison to baclofen and utility as a pharmacodynamic screening model.

PubMed

Brown, Jordan W; Moeller, Achim; Schmidt, Martin; Turner, Sean C; Nimmrich, Volker; Ma, Junli; Rueter, Lynne E; van der Kam, Elizabeth; Zhang, Min

2016-02-01

The GABA(B) receptor has been indicated as a promising target for multiple CNS-related disorders. Baclofen, a prototypical orthosteric agonist, is used clinically for the treatment of spastic movement disorders, but is associated with unwanted side-effects, such as sedation and motor impairment. Positive allosteric modulators (PAM), which bind to a topographically-distinct site apart from the orthosteric binding pocket, may provide an improved side-effect profile while maintaining baclofen-like efficacy. GABA, the major inhibitory neurotransmitter in the CNS, plays an important role in the etiology and treatment of seizure disorders. Baclofen is known to produce anticonvulsant effects in the DBA/2J mouse audiogenic seizure test (AGS), suggesting it may be a suitable assay for assessing pharmacodynamic effects. Little is known about the effects of GABA(B) PAMs, however. The studies presented here sought to investigate the AGS test as a pharmacodynamic (PD) screening model for GABA(B) PAMs by comparing the profile of structurally diverse PAMs to baclofen. GS39783, rac-BHFF, CMPPE, A-1295120 (N-(3-(4-(4-chloro-3-fluorobenzyl)-6-methoxy-3,5-dioxo-4,5-dihydro-1,2,4-triazin-2(3H)-yl)phenyl)acetamide), and A-1474713 (N-(3-(4-(4-chlorobenzyl)-3,5-dioxo-4,5-dihydro-1,2,4-triazin-2(3H)-yl)phenyl)acetamide) all produced robust, dose-dependent anticonvulsant effects; a similar profile was observed with baclofen. Pre-treatment with the GABA(B) antagonist SCH50911 completely blocked the anticonvulsant effects of baclofen and CMPPE in the AGS test, indicating such effects are likely mediated by the GABA(B) receptor. In addition to the standard anticonvulsant endpoint of the AGS test, video tracking software was employed to assess potential drug-induced motor side-effects during the acclimation period of the test. This analysis was sensitive to detecting drug-induced changes in total distance traveled, which was used to establish a therapeutic index (TI = hypoactivity

The novel, peripherally restricted GABAB receptor agonist lesogaberan (AZD3355) inhibits acid reflux and reduces esophageal acid exposure as measured with 24-h pHmetry in dogs.

PubMed

Brändén, Lena; Fredriksson, Anita; Harring, Emelie; Jensen, Jörgen; Lehmann, Anders

2010-05-25

While patients with symptoms of gastroesophageal reflux disease generally respond well to proton pump inhibitors, 20-30% continue to experience troublesome symptoms. In such cases, agents that target transient lower esophageal sphincter (LES) relaxation may be useful as add-on therapy to proton pump inhibitors. The GABAB receptor agonist baclofen inhibits transient LES relaxation but it is not an ideal agent due to central nervous system activity. Lesogaberan (AZD3355) is a peripherally restricted GABAB receptor agonist with limited central nervous system activity that inhibits transient LES relaxation in dogs. In the present study, the comparative effects of lesogaberan (7 micromol/kg) and baclofen (2.8 micromol/kg) on reflux were studied in dogs using 24-h pHmetry. Drugs (or vehicle control) were administered orally prior to the first meal of the day, and the number of reflux episodes (pH<4 for > or = 5 s) and acid exposure time were computed for the 24-h monitoring period. The mean (S.E.M.) number of reflux episodes/24 h was 4.6 (0.4) and 6.4 (0.6) for lesogaberan and baclofen, respectively, versus 10.7 (0.5) for control (P<0.0001 for both). Acid exposure time was 51.2 (4.5) min for control versus 23.6 (3.8) min for lesogaberan (P<0.0001) and 35.4 (6.5) min with baclofen (P=0.05). It is concluded that lesogaberan significantly reduces acid reflux in dogs, with comparable efficacy to baclofen. Copyright 2010 Elsevier B.V. All rights reserved.

Differential Cav2.1 and Cav2.3 channel inhibition by baclofen and α-conotoxin Vc1.1 via GABAB receptor activation

PubMed Central

McArthur, Jeffrey R.; Cuny, Hartmut; Clark, Richard J.; Adams, David J.

2014-01-01

Neuronal Cav2.1 (P/Q-type), Cav2.2 (N-type), and Cav2.3 (R-type) calcium channels contribute to synaptic transmission and are modulated through G protein–coupled receptor pathways. The analgesic α-conotoxin Vc1.1 acts through γ-aminobutyric acid type B (GABAB) receptors (GABABRs) to inhibit Cav2.2 channels. We investigated GABABR-mediated modulation by Vc1.1, a cyclized form of Vc1.1 (c-Vc1.1), and the GABABR agonist baclofen of human Cav2.1 or Cav2.3 channels heterologously expressed in human embryonic kidney cells. 50 µM baclofen inhibited Cav2.1 and Cav2.3 channel Ba2+ currents by ∼40%, whereas c-Vc1.1 did not affect Cav2.1 but potently inhibited Cav2.3, with a half-maximal inhibitory concentration of ∼300 pM. Depolarizing paired pulses revealed that ∼75% of the baclofen inhibition of Cav2.1 was voltage dependent and could be relieved by strong depolarization. In contrast, baclofen or Vc1.1 inhibition of Cav2.3 channels was solely mediated through voltage-independent pathways that could be disrupted by pertussis toxin, guanosine 5′-[β-thio]diphosphate trilithium salt, or the GABABR antagonist CGP55845. Overexpression of the kinase c-Src significantly increased inhibition of Cav2.3 by c-Vc1.1. Conversely, coexpression of a catalytically inactive double mutant form of c-Src or pretreatment with a phosphorylated pp60c-Src peptide abolished the effect of c-Vc1.1. Site-directed mutational analyses of Cav2.3 demonstrated that tyrosines 1761 and 1765 within exon 37 are critical for inhibition of Cav2.3 by c-Vc1.1 and are involved in baclofen inhibition of these channels. Remarkably, point mutations introducing specific c-Src phosphorylation sites into human Cav2.1 channels conferred c-Vc1.1 sensitivity. Our findings show that Vc1.1 inhibition of Cav2.3, which defines Cav2.3 channels as potential targets for analgesic α-conotoxins, is caused by specific c-Src phosphorylation sites in the C terminus. PMID:24688019

Virus Infection and Death Receptor-Mediated Apoptosis.

PubMed

Zhou, Xingchen; Jiang, Wenbo; Liu, Zhongshun; Liu, Shuai; Liang, Xiaozhen

2017-10-27

Virus infection can trigger extrinsic apoptosis. Cell-surface death receptors of the tumor necrosis factor family mediate this process. They either assist persistent viral infection or elicit the elimination of infected cells by the host. Death receptor-mediated apoptosis plays an important role in viral pathogenesis and the host antiviral response. Many viruses have acquired the capability to subvert death receptor-mediated apoptosis and evade the host immune response, mainly by virally encoded gene products that suppress death receptor-mediated apoptosis. In this review, we summarize the current information on virus infection and death receptor-mediated apoptosis, particularly focusing on the viral proteins that modulate death receptor-mediated apoptosis.

Virus Infection and Death Receptor-Mediated Apoptosis

PubMed Central

Zhou, Xingchen; Jiang, Wenbo; Liu, Zhongshun; Liu, Shuai; Liang, Xiaozhen

2017-01-01

Virus infection can trigger extrinsic apoptosis. Cell-surface death receptors of the tumor necrosis factor family mediate this process. They either assist persistent viral infection or elicit the elimination of infected cells by the host. Death receptor-mediated apoptosis plays an important role in viral pathogenesis and the host antiviral response. Many viruses have acquired the capability to subvert death receptor-mediated apoptosis and evade the host immune response, mainly by virally encoded gene products that suppress death receptor-mediated apoptosis. In this review, we summarize the current information on virus infection and death receptor-mediated apoptosis, particularly focusing on the viral proteins that modulate death receptor-mediated apoptosis. PMID:29077026

Mediator-dependent Nuclear Receptor Functions

PubMed Central

Chen, Wei; Roeder, Robert

2011-01-01

As gene-specific transcription factors, nuclear hormone receptors are broadly involved in many important biological processes. Their function on target genes requires the stepwise assembly of different coactivator complexes that facilitate chromatin remodeling and subsequent preinitiation complex (PIC) formation and function. Mediator has proved to be a crucial, and general, nuclear receptor-interacting coactivator, with demonstrated functions in transcription steps ranging from chromatin remodeling to subsequent PIC formation and function. Here we discuss (i) our current understanding of pathways that nuclear receptors and other interacting cofactors employ to recruit Mediator to target gene enhancers and promoters, including conditional requirements for the strong NR-Mediator interactions mediated by the NR AF2 domain and the MED1 LXXLLL motifs and (ii) mechanisms by which Mediator acts to transmit signals from enhancer-bound nuclear receptors to the general transcription machinery at core promoters to effect PIC formation and function. PMID:21854863

GABA, its receptors, and GABAergic inhibition in mouse taste buds

PubMed Central

Dvoryanchikov, Gennady; Huang, Yijen A; Barro-Soria, Rene; Chaudhari, Nirupa; Roper, Stephen D.

2012-01-01

Taste buds consist of at least three principal cell types that have different functions in processing gustatory signals — glial-like Type I cells, Receptor (Type II) cells, and Presynaptic (Type III) cells. Using a combination of Ca2+ imaging, single cell RT-PCR, and immunostaining, we show that γ-amino butyric acid (GABA) is an inhibitory transmitter in mouse taste buds, acting on GABA-A and GABA-B receptors to suppress transmitter (ATP) secretion from Receptor cells during taste stimulation. Specifically, Receptor cells express GABA-A receptor subunits β2, δ, π, as well as GABA-B receptors. In contrast, Presynaptic cells express the GABA-Aβ3 subunit and only occasionally GABA-B receptors. In keeping with the distinct expression pattern of GABA receptors in Presynaptic cells, we detected no GABAergic suppression of transmitter release from Presynaptic cells. We suggest that GABA may serve function(s) in taste buds in addition to synaptic inhibition. Finally, we also defined the source of GABA in taste buds: GABA is synthesized by GAD65 in Type I taste cells as well as by GAD67 in Presynaptic (Type III) taste cells and is stored in both those two cell types. We conclude that GABA is released during taste stimulation and possibly also during growth and differentiation of taste buds. PMID:21490220

GABA, its receptors, and GABAergic inhibition in mouse taste buds.

PubMed

Dvoryanchikov, Gennady; Huang, Yijen A; Barro-Soria, Rene; Chaudhari, Nirupa; Roper, Stephen D

2011-04-13

Taste buds consist of at least three principal cell types that have different functions in processing gustatory signals: glial-like (type I) cells, receptor (type II) cells, and presynaptic (type III) cells. Using a combination of Ca2+ imaging, single-cell reverse transcriptase-PCR and immunostaining, we show that GABA is an inhibitory transmitter in mouse taste buds, acting on GABA(A) and GABA(B) receptors to suppress transmitter (ATP) secretion from receptor cells during taste stimulation. Specifically, receptor cells express GABA(A) receptor subunits β2, δ, and π, as well as GABA(B) receptors. In contrast, presynaptic cells express the GABA(A) β3 subunit and only occasionally GABA(B) receptors. In keeping with the distinct expression pattern of GABA receptors in presynaptic cells, we detected no GABAergic suppression of transmitter release from presynaptic cells. We suggest that GABA may serve function(s) in taste buds in addition to synaptic inhibition. Finally, we also defined the source of GABA in taste buds: GABA is synthesized by GAD65 in type I taste cells as well as by GAD67 in presynaptic (type III) taste cells and is stored in both those two cell types. We conclude that GABA is an inhibitory transmitter released during taste stimulation and possibly also during growth and differentiation of taste buds.

Opiate-induced motor stimulation is regulated by gamma-aminobutyric acid type B receptors found in the ventral tegmental area in mice.

PubMed

Leite-Morris, Kimberly A; Fukudome, Eugene Y; Kaplan, Gary B

2002-01-14

Recent studies suggest that gamma-aminobutyric acid type B (GABA(B)) receptors located on dopaminergic cells in the ventral tegmental area (VTA) regulate mesolimbic dopaminergic (A10) activity. In the current study, we identified GABA(B) receptor subtypes in the area of the VTA and examined their role in modulating acute opiate actions. We studied the effects of intra-VTA infusions of the selective GABA(B) agonist baclofen on morphine-induced locomotor stimulation and A10 neuronal activation. Drug treatments were followed by ambulatory activity monitoring for 180 min. Intra-VTA baclofen treatment produced a 70% inhibition of morphine-stimulated locomotor activity. Furthermore, functional activation of A10 neurons was assessed by immunohistochemical staining of c-Fos in the nucleus accumbens (NAc), where A10 neurons terminate. We found that morphine treatment increased the levels of Fos-positive nuclei in the NAc, while intra-VTA baclofen treatment reversed morphine's effects. Finally, GABA(B) receptor subtypes and isoforms were identified in the ventromedial mesencephalon using immunoblotting. We demonstrated the presence of GABA(B)R1a (130 kDa), GABA(B)R1b (100 kDa), and GABA(B)R2 (120 kDa) receptor subtypes in this region. These results suggest that GABA(B) receptor isoforms are found in the VTA and their activation results in the blockade of behavioral effects of opiates via inhibition of dopaminergic neurotransmission.

Multiple functions of GABA A and GABA B receptors during pattern processing in the zebrafish olfactory bulb.

PubMed

Tabor, Rico; Yaksi, Emre; Friedrich, Rainer W

2008-07-01

gamma-Aminobutyric acid (GABA)ergic synapses are thought to play pivotal roles in the processing of activity patterns in the olfactory bulb (OB), but their functions have been difficult to study during odor responses in the intact system. We pharmacologically manipulated GABA(A) and GABA(B) receptors in the OB of zebrafish and analysed the effects on odor responses of the output neurons, the mitral cells (MCs), by electrophysiological recordings and temporally deconvolved two-photon Ca2+ imaging. The blockade of GABA(B) receptors enhanced presynaptic Ca2+ influx into afferent axon terminals, and changed the amplitude and time course of a subset of MC responses, indicating that GABA(B) receptors have a modulatory influence on OB output activity. The blockade of GABA(A) receptors induced epileptiform firing, enhanced excitatory responses and abolished fast oscillations in the local field potential. Moreover, the topological reorganization and decorrelation of MC activity patterns during the initial phase of the response was perturbed. These results indicate that GABA(A) receptor-containing circuits participate in the balance of excitation and inhibition, the regulation of total OB output activity, the synchronization of odor-dependent neuronal ensembles, and the reorganization of odor-encoding activity patterns. GABA(A) and GABA(B) receptors are therefore differentially involved in multiple functions of neuronal circuits in the OB.

Particle aggregation during receptor-mediated endocytosis

NASA Astrophysics Data System (ADS)

Mao, Sheng; Kosmrlj, Andrej

Receptor-mediated endocytosis of particles is driven by large binding energy between ligands on particles and receptors on a membrane, which compensates for the membrane bending energy and for the cost due to the mixing entropy of receptors. While the receptor-mediated endocytosis of individual particle is well understood, much less is known about the joint entry of multiple particles. Here, we demonstrate that the endocytosis of multiple particles leads to a kinetically driven entropic attraction, which may cause the aggregation of particles observed in experiments. During the endocytosis particles absorb nearby receptors and thus produce regions, which are depleted of receptors. When such depleted regions start overlapping, the corresponding particles experience osmotic-like attractive entropic force. If the attractive force between particles is large enough to overcome the repulsive interaction due to membrane bending, then particles tend to aggregate provided that they are sufficiently close, such that they are not completely engulfed before they come in contact. We discuss the necessary conditions for the aggregation of cylindrical particles during receptor-mediated endocytosis and comment on the generalization to spherical particles.

Receptor-mediated cell mechanosensing

PubMed Central

Chen, Yunfeng; Ju, Lining; Rushdi, Muaz; Ge, Chenghao; Zhu, Cheng

2017-01-01

Mechanosensing describes the ability of a cell to sense mechanical cues of its microenvironment, including not only all components of force, stress, and strain but also substrate rigidity, topology, and adhesiveness. This ability is crucial for the cell to respond to the surrounding mechanical cues and adapt to the changing environment. Examples of responses and adaptation include (de)activation, proliferation/apoptosis, and (de)differentiation. Receptor-mediated cell mechanosensing is a multistep process that is initiated by binding of cell surface receptors to their ligands on the extracellular matrix or the surface of adjacent cells. Mechanical cues are presented by the ligand and received by the receptor at the binding interface; but their transmission over space and time and their conversion into biochemical signals may involve other domains and additional molecules. In this review, a four-step model is described for the receptor-mediated cell mechanosensing process. Platelet glycoprotein Ib, T-cell receptor, and integrins are used as examples to illustrate the key concepts and players in this process. PMID:28954860

Behavioural endophenotypes in mice lacking the auxiliary GABAB receptor subunit KCTD16.

PubMed

Cathomas, Flurin; Sigrist, Hannes; Schmid, Luca; Seifritz, Erich; Gassmann, Martin; Bettler, Bernhard; Pryce, Christopher R

2017-01-15

Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the brain and is implicated in the pathophysiology of a number of neuropsychiatric disorders. The GABA B receptors are G-protein coupled receptors consisting of principle subunits and auxiliary potassium channel tetramerization domain (KCTD) subunits. The KCTD subunits 8, 12, 12b and 16 are cytosolic proteins that determine the kinetics of the GABA B receptor response. Previously, we demonstrated that Kctd12 null mutant mice (Kctd12 -/- ) exhibit increased auditory fear learning and that Kctd12 +/- mice show altered circadian activity, as well as increased intrinsic excitability in hippocampal pyramidal neurons. KCTD16 has been demonstrated to influence neuronal excitability by regulating GABA B receptor-mediated gating of postsynaptic ion channels. In the present study we investigated for behavioural endophenotypes in Kctd16 -/- and Kctd16 +/- mice. Compared with wild-type (WT) littermates, auditory and contextual fear conditioning were normal in both Kctd16 -/- and Kctd16 +/- mice. When fear memory was tested on the following day, Kctd16 -/- mice exhibited less extinction of auditory fear memory relative to WT and Kctd16 +/- mice, as well as more contextual fear memory relative to WT and, in particular, Kctd16 +/- mice. Relative to WT, both Kctd16 +/- and Kctd16 -/- mice exhibited normal circadian activity. This study adds to the evidence that auxillary KCTD subunits of GABA B receptors contribute to the regulation of behaviours that could constitute endophenotypes for hyper-reactivity to aversive stimuli in neuropsychiatric disorders. Copyright © 2016 Elsevier B.V. All rights reserved.

Behavioral characterization of escalated aggression induced by GABAB receptor activation in the dorsal raphé nucleus

PubMed Central

Takahashi, Aki; Schilit, Arielle N.; Kim, Jisoo; DeBold, Joseph F.; Koide, Tsuyoshi; Miczek, Klaus A.

2013-01-01

Rationale Pharmacological activation of GABAB receptors in the dorsal raphé nucleus (DRN) can escalate territorial aggression in male mice. Objectives We characterized this escalated aggression in terms of its behavioral and environmental determinants. Methods Aggressive behavior of resident male (CFW or ICR mouse) was assessed in confrontations with a group-housed intruder. Either baclofen (0.06 nmol/0.2 μl) or vehicle (saline) was microinjected into the DRN ten minutes before the confrontation. We examined baclofen-heightened aggression in five situations: aggression in a neutral arena and after social instigation (experiment 1), aggression during the light phase of the cycle (experiment 2), aggression without prior fighting experience (experiment 3), aggression toward a female (experiment 4), and aggression after defeat experiences (experiment 5). In addition, we examined the body targets towards which bites are directed and the duration of aggressive bursts after baclofen treatment. Results Regardless of the past social experience, baclofen escalated aggressive behaviors. Even in the neutral arena and after defeat experiences, where aggressive behaviors were inhibited, baclofen significantly increased aggression. Baclofen increased attack bites directed at vulnerable body areas of male intruders but not toward a female and only in the dark. Also, baclofen prolonged the duration of aggressive bursts. Conclusions For baclofen to escalate aggression, specific stimulation (male intruder) and tonic level of serotonin (dark cycle) are required. Once aggressive behavior is triggered, intra-DRN baclofen escalates the level of aggression to abnormal levels and renders it difficult to terminate. Also, baclofen counteracts the effects of novelty or past experiences of defeat. PMID:22395428

Effect of GABA receptor agonists or antagonists injected spinally on the blood glucose level in mice.

PubMed

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

2013-05-01

The possible roles of gamma-amino butyric acid (GABA) receptors located in the spinal cord for the regulation of the blood glucose level were studied in ICR mice. We found in the present study that intrathecal (i.t.) injection with baclofen (a GABAB receptor agonist; 1-10 μg/5 μl) or bicuculline (a GABAA receptor antagonist; 1-10 μg/5 μl) caused an elevation of the blood glucose level in a dose-dependent manner. The hyperglycemic effect induced by baclofen was more pronounced than that induced by bicuculline. However, muscimol (a GABAA receptor agonist; 1-5 μg/5 μl) or phaclofen (a GABAB receptor antagonist; 5-10 μg/5 μl) administered i.t. did not affect the blood glucose level. Baclofen-induced elevation of the blood glucose was dose-dependently attenuated by phaclofen. Furthermore, i.t. pretreatment with pertussis toxin (PTX; 0.05 or 0.1 μg/5 μl) for 6 days dose-dependently reduced the hyperglycemic effect induced by baclofen. Our results suggest that GABAB receptors located in the spinal cord play important roles for the elevation of the blood glucose level. Spinally located PTX-sensitive G-proteins appear to be involved in hyperglycemic effect induced by baclofen. Furthermore, inactivation of GABAA receptors located in the spinal cord appears to be responsible for tonic up-regulation of the blood glucose level.

Inactivation of the maternal fragile X gene results in sensitization of GABAB receptor function in the offspring.

PubMed

Zupan, Bojana; Toth, Miklos

2008-12-01

Fragile X syndrome is an X-linked disorder caused by the inactivation of the FMR1 gene, with symptoms ranging from impaired cognitive functions to seizures, anxiety, sensory abnormalities, and hyperactivity. Although fragile X syndrome is considered a typical Mendelian disorder, we have recently reported that the environment, specifically the fmr1(+/-) or fmr1(-/-) [H or knockout (KO)] maternal environment, elicits on its own a partial fragile X-like phenotype and can contribute to the overall phenotype of fmr1(-/0) (KO) male offspring. Genetically fmr1(+/0) (WT) males born to H females (H(maternal) > WT(offspring)), similar to KO male offspring born to H and KO mothers (H > KO and KO > KO), exhibit locomotor hyperactivity. These mice also showed reduced D(2) autoreceptor function, indicating a possible diminished feedback inhibition of dopamine (DA) release in the nigrostriatal and mesolimbic systems. The GABAergic system also regulates DA release, in part via presynaptic GABA(B) receptors (Rs) located on midbrain dopaminergic neurons. Here, we show that the locomotor inhibitory effect of the GABA(B)R agonist baclofen [4-amino-3-(4-chlorophenyl)-butanoic acid] is enhanced in all progeny of mutant mothers (H > WT, H > KO, and KO > KO) compared with WT > WT mice, irrespective of their own genotype. However, increased sensitivity to baclofen was selective and limited to the locomotor response because the muscle-relaxant and sedative effects of the drug were not altered by the maternal environment. These data show that GABA(B)R sensitization, traditionally induced pharmacologically, can also be elicited by the fmr1-deficient maternal environment.

Activation of postsynaptic GABAB receptors modulates the bursting pattern and synaptic activity of olfactory bulb juxtaglomerular neurons.

PubMed

Karpuk, Nikolay; Hayar, Abdallah

2008-01-01

Olfactory bulb glomeruli are formed by a network of three major types of neurons collectively called juxtaglomerular (JG) cells, which include external tufted (ET), periglomerular (PG), and short axon (SA) cells. There is solid evidence that gamma-aminobutyric acid (GABA) released from PG neurons presynaptically inhibits glutamate release from olfactory nerve terminals via activation of GABA(B) receptors (GABA(B)-Rs). However, it is still unclear whether ET cells have GABA(B)-Rs. We have investigated whether ET cells have functional postsynaptic GABA(B)-Rs using extracellular and whole cell recordings in olfactory bulb slices. In the presence of fast synaptic blockers (CNQX, APV, and gabazine), the GABA(B)-R agonist baclofen either completely inhibited the bursting or reduced the bursting frequency and increased the burst duration and the number of spikes/burst in ET cells. In the presence of fast synaptic blockers and tetrodotoxin, baclofen induced an outward current in ET cells, suggesting a direct postsynaptic effect. Baclofen reduced the frequency and amplitude of spontaneous EPSCs in PG and SA cells. In the presence of sodium and potassium channel blockers, baclofen reduced the frequency of miniature EPSCs, which were inhibited by the calcium channel blocker cadmium. All baclofen effects were reversed by application of the GABA(B)-R antagonist CGP55845. We suggest that activation of GABA(B)-Rs directly inhibits ET cell bursting and decreases excitatory dendrodendritic transmission from ET to PG and SA cells. Thus the postsynaptic GABA(B)-Rs on ET cells may play an important role in shaping the activation pattern of the glomeruli during olfactory coding.

Differential association of GABAB receptors with their effector ion channels in Purkinje cells.

PubMed

Luján, Rafael; Aguado, Carolina; Ciruela, Francisco; Cózar, Javier; Kleindienst, David; de la Ossa, Luis; Bettler, Bernhard; Wickman, Kevin; Watanabe, Masahiko; Shigemoto, Ryuichi; Fukazawa, Yugo

2018-04-01

Metabotropic GABA B receptors mediate slow inhibitory effects presynaptically and postsynaptically through the modulation of different effector signalling pathways. Here, we analysed the distribution of GABA B receptors using highly sensitive SDS-digested freeze-fracture replica labelling in mouse cerebellar Purkinje cells. Immunoreactivity for GABA B1 was observed on presynaptic and, more abundantly, on postsynaptic compartments, showing both scattered and clustered distribution patterns. Quantitative analysis of immunoparticles revealed a somato-dendritic gradient, with the density of immunoparticles increasing 26-fold from somata to dendritic spines. To understand the spatial relationship of GABA B receptors with two key effector ion channels, the G protein-gated inwardly rectifying K + (GIRK/Kir3) channel and the voltage-dependent Ca 2+ channel, biochemical and immunohistochemical approaches were performed. Co-immunoprecipitation analysis demonstrated that GABA B receptors co-assembled with GIRK and Ca V 2.1 channels in the cerebellum. Using double-labelling immunoelectron microscopic techniques, co-clustering between GABA B1 and GIRK2 was detected in dendritic spines, whereas they were mainly segregated in the dendritic shafts. In contrast, co-clustering of GABA B1 and Ca V 2.1 was detected in dendritic shafts but not spines. Presynaptically, although no significant co-clustering of GABA B1 and GIRK2 or Ca V 2.1 channels was detected, inter-cluster distance for GABA B1 and GIRK2 was significantly smaller in the active zone than in the dendritic shafts, and that for GABA B1 and Ca V 2.1 was significantly smaller in the active zone than in the dendritic shafts and spines. Thus, GABA B receptors are associated with GIRK and Ca V 2.1 channels in different subcellular compartments. These data provide a better framework for understanding the different roles played by GABA B receptors and their effector ion channels in the cerebellar network.

Bombesin receptor-mediated imaging and cytotoxicity: review and current status

PubMed Central

Sancho, Veronica; Di Florio, Alessia; Moody, Terry W.; Jensen, Robert T.

2010-01-01

The three mammalian bombesin (Bn) receptors (gastrin-releasing peptide [GRP] receptor, neuromedin B [NMB] receptor, BRS-3) are one of the classes of G protein-coupled receptors that are most frequently over-express/ectopically expressed by common, important malignancies. Because of the clinical success of somatostatin receptor-mediated imaging and cytotoxicity with neuroendocrine tumors, there is now increasing interest in pursuing a similar approach with Bn receptors. In the last few years then have been more than 200 studies in this area. In the present paper, the in vitro and in vivo results, as well as results of human studies from many of these studies are reviewed and the current state of Bn receptor-mediated imaging or cytotoxicity is discussed. Both Bn receptor-mediated imaging studies as well as Bn receptor-mediated tumoral cytotoxic studies using radioactive and non-radioactive Bn-based ligands are covered. PMID:21034419

Autoantibodies to Synaptic Receptors and Neuronal Cell Surface Proteins in Autoimmune Diseases of the Central Nervous System

PubMed Central

Geis, Christian; Graus, Francesc

2017-01-01

Investigations in the last 10 years have revealed a new category of neurological diseases mediated by antibodies against cell surface and synaptic proteins. There are currently 16 such diseases all characterized by autoantibodies against neuronal proteins involved in synaptic signaling and plasticity. In clinical practice these findings have changed the diagnostic and treatment approach to potentially lethal, but now treatable, neurological and psychiatric syndromes previously considered idiopathic or not even suspected to be immune-mediated. Studies show that patients' antibodies can impair the surface dynamics of the target receptors eliminating them from synapses (e.g., NMDA receptor), block the function of the antigens without changing their synaptic density (e.g., GABAb receptor), interfere with synaptic protein-protein interactions (LGI1, Caspr2), alter synapse formation (e.g., neurexin-3α), or by unclear mechanisms associate to a new form of tauopathy (IgLON5). Here we first trace the process of discovery of these diseases, describing the triggers and symptoms related to each autoantigen, and then review in detail the structural and functional alterations caused by the autoantibodies with special emphasis in those (NMDA receptor, amphiphysin) that have been modeled in animals. PMID:28298428

Selective antagonism of AMPA receptors unmasks kainate receptor-mediated responses in hippocampal neurons.

PubMed

Paternain, A V; Morales, M; Lerma, J

1995-01-01

Although both protein and mRNAs for kainate receptor subunits are abundant in several brain regions, the responsiveness of AMPA receptors to kainate has made it difficult to demonstrate the presence of functional kainate-type receptors in native cells. Recently, however, we have shown that many hippocampal neurons in culture express glutamate receptors of the kainate type. The large nondesensitizing response that kainate induces at AMPA receptors precludes detection and analysis of smaller, rapidly desensitizing currents induced by kainate at kainate receptors. Consequently, the functional significance of these strongly desensitizing glutamate receptors remains enigmatic. We report here that the family of new noncompetitive antagonists of AMPA receptors (GYKI 52466 and 53655) minimally affects kainate-induced responses at kainate receptors while completely blocking AMPA receptor-mediated currents, making it possible to separate the responses mediated by each receptor. These compounds will allow determination of the role played by kainate receptors in synaptic transmission and plasticity in the mammalian brain, as well as evaluation of their involvement in neurotoxicity.

Alpha-1A Adrenergic receptor activation increases inhibitory tone in CA1 hippocampus

PubMed Central

Hillman, Kristin L.; Lei, Saobo; Doze, Van A.

2009-01-01

The endogenous catecholamine norepinephrine (NE) exhibits anti-epileptic properties, however it is not well understood which adrenergic receptor (AR) mediates this effect. The aim of this study was to investigate α1-adrenergic receptor (AR) activation in region CA1 of the hippocampus, a subcortical structure often implicated in temporal lobe epilepsies. Using cell-attached and whole-cell recordings in rat hippocampal slices, we confirmed that selective α1-AR activation increases action potential firing in a subpopulation of CA1 interneurons. We found that this response is mediated via the α1A-AR subtype, initiated by sodium influx, and appears independent of second messenger signaling. In CA1 pyramidal cells, α1A-AR activation decreases activity due to increased pre-synaptic GABA and somatostatin release. Examination of post-synaptic receptor involvement revealed that while GABAA receptors mediate the majority of α1A-adrenergic effects on CA1 pyramidal cells, significant contributions are also made by GABAB and somatostatin receptors. Finally, to test whether α1A-AR activation could have potential therapeutic implications, we performed AR agonist challenges using two in vitro epileptiform models. When GABAA receptors were available, α1A-AR activation significantly decreased epileptiform bursting in CA1. Together, our findings directly link stimulation of the α1A-AR subtype to release of GABA and somatostatin at the single cell level and suggest that α1A-AR activation may represent one mechanism by which NE exerts anti-epileptic effects within the hippocampus. PMID:19201164

Bidirectional enantioselective effects of the GABAB receptor agonist baclofen in two mouse models of excessive ethanol consumption

PubMed Central

Kasten, Chelsea R.; Blasingame, Shelby N.; Boehm, Stephen L.

2014-01-01

The GABAB receptor agonist baclofen has been studied extensively in preclinical models of alcohol use disorders, yet results on its efficacy have been uncertain. Racemic baclofen, which is used clinically, can be broken down into separate enantiomers of the drug. Baclofen has been shown to produce enantioselective effects in behavioral assays including those modeling reflexive and sexual behavior. The current studies sought to characterize the enantioselective effects of baclofen in two separate models of ethanol consumption. The first was a Drinking-in-the-Dark procedure that provides “binge-like” ethanol access to mice by restricting access to a two hour period, three hours into the dark cycle. The second was a two-bottle choice procedure that utilized selectively bred High Alcohol Preferring 1 (HAP1) mice to model chronic ethanol access. HAP1 mice are selectively bred to consume pharmacologically relevant amounts of ethanol in a 24-hour two-bottle choice paradigm. The results showed that baclofen yields enantioselective effects on ethanol intake in both models, and that these effects are bidirectional. Total ethanol intake was decreased by R(+)- baclofen, while total intake was increased by S(-)-baclofen in the binge-like and chronic drinking models. Whereas overall binge-like saccharin intake was significantly reduced by R(+)- baclofen, chronic intake was not significantly altered. S(-)- baclofen did not significantly alter saccharin intake. Neither enantiomer significantly affected locomotion during binge-like reinforcer consumption. Collectively, these results demonstrate that baclofen produces enantioselective effects on ethanol consumption. More importantly, the modulation of consumption is bidirectional. The opposing enantioselective effects may explain some of the variance seen in published baclofen literature. PMID:25557834

Bidirectional enantioselective effects of the GABAB receptor agonist baclofen in two mouse models of excessive ethanol consumption.

PubMed

Kasten, Chelsea R; Blasingame, Shelby N; Boehm, Stephen L

2015-02-01

The GABAB receptor agonist baclofen has been studied extensively in preclinical models of alcohol-use disorders, yet results on its efficacy have been uncertain. Racemic baclofen, which is used clinically, can be broken down into separate enantiomers of the drug. Baclofen has been shown to produce enantioselective effects in behavioral assays, including those modeling reflexive and sexual behavior. The current studies sought to characterize the enantioselective effects of baclofen in two separate models of ethanol consumption. The first was a Drinking-in-the-Dark procedure that provides "binge-like" ethanol access to mice by restricting access to a 2-h period, 3 h into the dark cycle. The second was a two-bottle choice procedure that utilized selectively bred High Alcohol Preferring 1 (HAP1) mice to model chronic ethanol access. HAP1 mice are selectively bred to consume pharmacologically relevant amounts of ethanol in a 24-h two-bottle choice paradigm. The results showed that baclofen yields enantioselective effects on ethanol intake in both models, and that these effects are bidirectional. Total ethanol intake was decreased by R(+)-baclofen, while total intake was increased by S(-)-baclofen in the binge-like and chronic drinking models. Whereas overall binge-like saccharin intake was significantly reduced by R(+)-baclofen, chronic intake was not significantly altered. S(-)-baclofen did not significantly alter saccharin intake. Neither enantiomer significantly affected locomotion during binge-like reinforcer consumption. Collectively, these results demonstrate that baclofen produces enantioselective effects on ethanol consumption. More importantly, the modulation of consumption is bidirectional. The opposing enantioselective effects may explain some of the variance seen in published baclofen literature. Copyright © 2015 Elsevier Inc. All rights reserved.

Muscarinic Receptors Modulate Dendrodendritic Inhibitory Synapses to Sculpt Glomerular Output

PubMed Central

Shao, Zuoyi; Puche, Adam; Wachowiak, Matt; Rothermel, Markus

2015-01-01

Cholinergic [acetylcholine (ACh)] axons from the basal forebrain innervate olfactory bulb glomeruli, the initial site of synaptic integration in the olfactory system. Both nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors (mAChRs) are expressed in glomeruli. The activation of nAChRs directly excites both mitral/tufted cells (MTCs) and external tufted cells (ETCs), the two major excitatory neurons that transmit glomerular output. The functional roles of mAChRs in glomerular circuits are unknown. We show that the restricted glomerular application of ACh causes rapid, brief nAChR-mediated excitation of both MTCs and ETCs in the mouse olfactory bulb. This excitation is followed by mAChR-mediated inhibition, which is blocked by GABAA receptor antagonists, indicating the engagement of periglomerular cells (PGCs) and/or short axon cells (SACs), the two major glomerular inhibitory neurons. Indeed, selective activation of glomerular mAChRs, with ionotropic GluRs and nAChRs blocked, increased IPSCs in MTCs and ETCs, indicating that mAChRs recruit glomerular inhibitory circuits. Selective activation of glomerular mAChRs in the presence of tetrodotoxin increased IPSCs in all glomerular neurons, indicating action potential-independent enhancement of GABA release from PGC and/or SAC dendrodendritic synapses. mAChR-mediated enhancement of GABA release also presynaptically suppressed the first synapse of the olfactory system via GABAB receptors on sensory terminals. Together, these results indicate that cholinergic modulation of glomerular circuits is biphasic, involving an initial excitation of MTC/ETCs mediated by nAChRs followed by inhibition mediated directly by mAChRs on PGCs/SACs. This may phasically enhance the sensitivity of glomerular outputs to odorants, an action that is consistent with recent in vivo findings. PMID:25855181

Muscarinic receptors modulate dendrodendritic inhibitory synapses to sculpt glomerular output.

PubMed

Liu, Shaolin; Shao, Zuoyi; Puche, Adam; Wachowiak, Matt; Rothermel, Markus; Shipley, Michael T

2015-04-08

Cholinergic [acetylcholine (ACh)] axons from the basal forebrain innervate olfactory bulb glomeruli, the initial site of synaptic integration in the olfactory system. Both nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors (mAChRs) are expressed in glomeruli. The activation of nAChRs directly excites both mitral/tufted cells (MTCs) and external tufted cells (ETCs), the two major excitatory neurons that transmit glomerular output. The functional roles of mAChRs in glomerular circuits are unknown. We show that the restricted glomerular application of ACh causes rapid, brief nAChR-mediated excitation of both MTCs and ETCs in the mouse olfactory bulb. This excitation is followed by mAChR-mediated inhibition, which is blocked by GABAA receptor antagonists, indicating the engagement of periglomerular cells (PGCs) and/or short axon cells (SACs), the two major glomerular inhibitory neurons. Indeed, selective activation of glomerular mAChRs, with ionotropic GluRs and nAChRs blocked, increased IPSCs in MTCs and ETCs, indicating that mAChRs recruit glomerular inhibitory circuits. Selective activation of glomerular mAChRs in the presence of tetrodotoxin increased IPSCs in all glomerular neurons, indicating action potential-independent enhancement of GABA release from PGC and/or SAC dendrodendritic synapses. mAChR-mediated enhancement of GABA release also presynaptically suppressed the first synapse of the olfactory system via GABAB receptors on sensory terminals. Together, these results indicate that cholinergic modulation of glomerular circuits is biphasic, involving an initial excitation of MTC/ETCs mediated by nAChRs followed by inhibition mediated directly by mAChRs on PGCs/SACs. This may phasically enhance the sensitivity of glomerular outputs to odorants, an action that is consistent with recent in vivo findings. Copyright © 2015 the authors 0270-6474/15/355680-13$15.00/0.

The gamma-aminobutyric acid type B (GABAB) receptor agonist baclofen inhibits morphine sensitization by decreasing the dopamine level in rat nucleus accumbens

PubMed Central

2012-01-01

Background Repeated morphine exposure can induce behavioral sensitization. There are evidences have shown that central gamma-aminobutyric acid (GABA) system is involved in morphine dependence. However, the effect of a GABAB receptor agonist baclofen on morphine-induced behavioral sensitization in rats is unclear. Methods We used morphine-induced behavioral sensitization model in rat to investigate the effects of baclofen on behavioral sensitization. Moreover, dopamine release in the shell of the nucleus accumbens was evaluated using microdialysis assay in vivo. Results The present study demonstrated that morphine challenge (3 mg/kg, s.c.) obviously enhanced the locomotor activity following 4-day consecutive morphine administration and 3-day withdrawal period, which indicated the expression of morphine sensitization. In addition, chronic treatment with baclofen (2.5, 5 mg/kg) significantly inhibited the development of morphine sensitization. It was also found that morphine challenge 3 days after repeated morphine administration produced a significant increase of extracellular dopamine release in nucleus accumbens. Furthermore, chronic treatment with baclofen decreased the dopamine release induced by morphine challenge. Conclusions Our results indicated that gamma-aminobutyric acid system plays an important role in the morphine sensitization in rat and suggested that behavioral sensitization is a promising model to study the mechanism underlying drug abuse. PMID:22559224

The γ-aminobutyric acid type B (GABAB) receptor agonist baclofen inhibits morphine sensitization by decreasing the dopamine level in rat nucleus accumbens.

PubMed

Fu, Zhenyu; Yang, Hongfa; Xiao, Yuqiang; Zhao, Gang; Huang, Haiyan

2012-07-10

Repeated morphine exposure can induce behavioral sensitization. There are evidences have shown that central gamma-aminobutyric acid (GABA) system is involved in morphine dependence. However, the effect of a GABAB receptor agonist baclofen on morphine-induced behavioral sensitization in rats is unclear. We used morphine-induced behavioral sensitization model in rat to investigate the effects of baclofen on behavioral sensitization. Moreover, dopamine release in the shell of the nucleus accumbens was evaluated using microdialysis assay in vivo. The present study demonstrated that morphine challenge (3 mg/kg, s.c.) obviously enhanced the locomotor activity following 4-day consecutive morphine administration and 3-day withdrawal period, which indicated the expression of morphine sensitization. In addition, chronic treatment with baclofen (2.5, 5 mg/kg) significantly inhibited the development of morphine sensitization. It was also found that morphine challenge 3 days after repeated morphine administration produced a significant increase of extracellular dopamine release in nucleus accumbens. Furthermore, chronic treatment with baclofen decreased the dopamine release induced by morphine challenge. Our results indicated that gamma-aminobutyric acid system plays an important role in the morphine sensitization in rat and suggested that behavioral sensitization is a promising model to study the mechanism underlying drug abuse.

Coexistence of Lambert-Eaton myasthenic syndrome and autoimmune encephalitis with anti-CRMP5/CV2 and anti-GABAB receptor antibodies in small cell lung cancer: A case report.

PubMed

Li, Hongfang; Zhang, Aimei; Hao, Yanlei; Guan, Hongzhi; Lv, Zhanyun

2018-05-01

Autoimmune encephalitis and Lambert-Eaton myasthenic syndrome are classic paraneoplastic neurological conditions common in patients with small cell lung cancer. The patient complained of tiredness, fluctuating recent memory loss, and inability to find his home. His family members reported a change in character, irritability, and paranoia. One month later, the patient had 1 grand mal seizure lasting 5 minutes. The patient was diagnosed with limbic encephalitis combined with Lambert-Eaton myasthenic syndrome. The gamma-aminobutyric acid B (GABAB) receptor and collapsin response mediator protein 5 (CRMP5, also called CV2) antibody test results were positive. Nine months after the onset of symptoms, the patient was diagnosed with small cell lung cancer. The patient was administered intravenous immunoglobulin for 5 days. He was then treated with 60 mg prednisone once per day. The prednisone dose was gradually reduced by 1 tablet every 2 weeks. After the diagnosis, the patient underwent 6 courses of chemotherapy with cisplatin combined with sequential chemoradiation therapy. The patient was able to take care of himself. Neurological examination revealed a lower limb proximal muscle strength level of 4 and a reduced limb tendon reflex. The patient had deficits in short-term memory, a Mini-Mental State Examination score of 26, Montreal Cognitive Assessment score of 24, Self-rating Depression Scale score of 54 (mild depression), and Self-Rating Anxiety Scale score of 42 (normal). Autoimmune diseases of the peripheral and central nervous systems can be observed at the same time in patients with small cell lung cancer, even when magnetic resonance imaging findings are negative and immune therapy is effective.

Receptor-Mediated Drug Delivery Systems Targeting to Glioma

PubMed Central

Wang, Shanshan; Meng, Ying; Li, Chengyi; Qian, Min; Huang, Rongqin

2015-01-01

Glioma has been considered to be the most frequent primary tumor within the central nervous system (CNS). The complexity of glioma, especially the existence of the blood-brain barrier (BBB), makes the survival and prognosis of glioma remain poor even after a standard treatment based on surgery, radiotherapy, and chemotherapy. This provides a rationale for the development of some novel therapeutic strategies. Among them, receptor-mediated drug delivery is a specific pattern taking advantage of differential expression of receptors between tumors and normal tissues. The strategy can actively transport drugs, such as small molecular drugs, gene medicines, and therapeutic proteins to glioma while minimizing adverse reactions. This review will summarize recent progress on receptor-mediated drug delivery systems targeting to glioma, and conclude the challenges and prospects of receptor-mediated glioma-targeted therapy for future applications. PMID:28344260

Structure, Dynamics, and Allosteric Potential of Ionotropic Glutamate Receptor N-Terminal Domains

PubMed Central

Krieger, James; Bahar, Ivet; Greger, Ingo H.

2015-01-01

Ionotropic glutamate receptors (iGluRs) are tetrameric cation channels that mediate synaptic transmission and plasticity. They have a unique modular architecture with four domains: the intracellular C-terminal domain (CTD) that is involved in synaptic targeting, the transmembrane domain (TMD) that forms the ion channel, the membrane-proximal ligand-binding domain (LBD) that binds agonists such as L-glutamate, and the distal N-terminal domain (NTD), whose function is the least clear. The extracellular portion, comprised of the LBD and NTD, is loosely arranged, mediating complex allosteric regulation and providing a rich target for drug development. Here, we briefly review recent work on iGluR NTD structure and dynamics, and further explore the allosteric potential for the NTD in AMPA-type iGluRs using coarse-grained simulations. We also investigate mechanisms underlying the established NTD allostery in NMDA-type iGluRs, as well as the fold-related metabotropic glutamate and GABAB receptors. We show that the clamshell motions intrinsically favored by the NTD bilobate fold are coupled to dimeric and higher-order rearrangements that impact the iGluR LBD and ultimately the TMD. Finally, we explore the dynamics of intact iGluRs and describe how it might affect receptor operation in a synaptic environment. PMID:26255587

Regulation of N-formyl peptide-mediated degranulation by receptor phosphorylation.

PubMed

Vines, Charlotte M; Xue, Mei; Maestas, Diane C; Cimino, Daniel F; Prossnitz, Eric R

2002-12-15

One of the major functions of the N-formyl peptide receptor (FPR) is to mediate leukocyte degranulation. Phosphorylation of the C-terminal domain of the FPR is required for receptor internalization and desensitization. Although arrestins mediate phosphorylation-dependent desensitization, internalization, and initiation of novel signaling cascades for a number of G protein-coupled receptors, their roles in FPR regulation and signaling remain unclear. CXCR1-mediated degranulation of RBL-2H3 cells is promoted by arrestin binding. To determine whether receptor phosphorylation or arrestin binding is required to promote FPR-mediated degranulation, we used RBL-2H3 cells stably transfected with either the wild-type FPR or a mutant form, DeltaST, which is incapable of undergoing ligand-stimulated phosphorylation. We observed that stimulation of wild-type FPR resulted in very low levels of degranulation compared with that mediated by cross-linking of the Fc(epsilon)RI receptor. Stimulation of the DeltaST mutant, however, resulted in levels of degranulation comparable to those of the Fc(epsilon)RI receptor, demonstrating that neither receptor phosphorylation nor arrestin binding was necessary to initiate FPR-mediated degranulation. Degranulation initiated by the DeltaST mutant was proportional to the level of active cell surface receptor, suggesting that either receptor internalization or desensitization may be responsible for terminating degranulation of the wild-type FPR. To distinguish between these possibilities, we used a partially phosphorylation-deficient mutant of the FPR that can undergo internalization, but not desensitization. Degranulation by this mutant FPR was indistinguishable from that of the DeltaST mutant, indicating that FPR phosphorylation or binding of arrestin but not internalization terminates the degranulation response.

Dose-response approaches for nuclear receptor-mediated ...

EPA Pesticide Factsheets

A public workshop, organized by a Steering Committee of scientists from government, industry, universities, and research organizations, was held at the National Institute of Environmental Health Sciences (NIEHS) in September, 2010. The workshop explored the dose-response implications of toxicant modes of action (MOA) mediated by nuclear receptors. The dominant paradigm in human health risk assessment has been linear extrapolation without a threshold for cancer, and estimation of sub-threshold doses for non-cancer and (in appropriate cases) cancer endpoints. However, recent publications question the application of dose-response modeling approaches with a threshold. The growing body of molecular toxicology information and computational toxicology tools has allowed for exploration of the presence or absence of subthreshold doses for a number of receptor-mediated MOPs. The workshop explored the development of dose-response approaches for nuclear receptor-mediated liver cancer, within a MOA Human Relevance framework (HRF). Case studies addressed activation of the AHR; the CAR/PXR, and the PPARa. This paper describes the workshop process, key issues discussed, and conclusions. The value of an interactive workshop approach to apply current MOA/HRF frameworks was demonstrated. The results may help direct research on the MOA and dose-response of receptor-based toxicity, since there are commonalities for many receptors in the basic pathways involved for late steps in the

Activity-dependent switch of GABAergic inhibition into glutamatergic excitation in astrocyte-neuron networks

PubMed Central

Perea, Gertrudis; Gómez, Ricardo; Mederos, Sara; Covelo, Ana; Ballesteros, Jesús J; Schlosser, Laura; Hernández-Vivanco, Alicia; Martín-Fernández, Mario; Quintana, Ruth; Rayan, Abdelrahman; Díez, Adolfo; Fuenzalida, Marco; Agarwal, Amit; Bergles, Dwight E; Bettler, Bernhard; Manahan-Vaughan, Denise; Martín, Eduardo D; Kirchhoff, Frank; Araque, Alfonso

2016-01-01

Interneurons are critical for proper neural network function and can activate Ca2+ signaling in astrocytes. However, the impact of the interneuron-astrocyte signaling into neuronal network operation remains unknown. Using the simplest hippocampal Astrocyte-Neuron network, i.e., GABAergic interneuron, pyramidal neuron, single CA3-CA1 glutamatergic synapse, and astrocytes, we found that interneuron-astrocyte signaling dynamically affected excitatory neurotransmission in an activity- and time-dependent manner, and determined the sign (inhibition vs potentiation) of the GABA-mediated effects. While synaptic inhibition was mediated by GABAA receptors, potentiation involved astrocyte GABAB receptors, astrocytic glutamate release, and presynaptic metabotropic glutamate receptors. Using conditional astrocyte-specific GABAB receptor (Gabbr1) knockout mice, we confirmed the glial source of the interneuron-induced potentiation, and demonstrated the involvement of astrocytes in hippocampal theta and gamma oscillations in vivo. Therefore, astrocytes decode interneuron activity and transform inhibitory into excitatory signals, contributing to the emergence of novel network properties resulting from the interneuron-astrocyte interplay. DOI: http://dx.doi.org/10.7554/eLife.20362.001 PMID:28012274

Tachykinin receptors mediating airway marcomolecular secretion

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

Gentry, S.E.

1991-01-01

Three tachykinin receptor types, termed NK1, NK2, and NK3, can be distinguished by the relative potency of various peptides in eliciting tissue responses. Airway macromolecular secretion is stimulated by the tachykinin substance P (SP). The purposes of this study were to determine the tachykinin receptor subtype responsible for this stimulation, and to examine the possible involvement of other neurotransmitters in mediating this effect. Ferret tracheal explants maintained in organ culture were labeled with {sup 3}H-glucosamine, a precursor of high molecular weight glycoconjugates (HMWG) which are released by airway secretory cells. Secretion of labeled HMWG then was determined in the absencemore » and presence of the tachykinins SP, neurokinin A (NKA), neurokinin B (NKB), physalaemin (PHY), and eledoisin (ELE). To evaluate the possible contribution of other mediators, tachykinin stimulation was examined in the presence of several receptor blockers.« less

Central pipecolic acid increases food intake under ad libitum feeding conditions in the neonatal chick.

PubMed

Takagi, Tomo; Tachibana, Tetsuya; Saito, Ei-Suke; Tomonaga, Shouzou; Saito, Shin; Bungo, Takashi; Denbow, D Michael; Furuse, Mitsuhiro

2003-08-21

It has been demonstrated that L-pipecolic acid (L-PA) is a major metabolic intermediate of L-lysine in the mammalian and chicken brain. A previous study showed that intracerebroventricular (i.c.v.) injection of L-PA suppressed feeding in neonatal chicks, and the actions were associated with gamma-aminobutyric acid (GABA)-B receptor activation. It has been reported that endogenous L-PA in the brain fluctuated under different feeding conditions. In the present study, we investigated the effect of i.c.v. injection of L-PA on food intake in the neonatal chick under ad libitum feeding conditions. The food intake was increased by 0.5 or 1.0 mg L-PA under ad libitum feeding conditions contrary to previous studies using fasted birds. A hyperphagic effect of L-PA (0.5 mg) was attenuated by both GABA-A receptor antagonist (picrotoxin, 0.5 microg) and GABA-B receptor antagonist (CGP54626, 21.0 ng). These results indicate that a hyperphagic effect of L-PA is mediated by both GABA-A and GABA-B receptors and L-PA differentially affects food intake under different feeding conditions in the neonatal chick.

Postsynaptic Synaptotagmins Mediate AMPA Receptor Exocytosis During LTP

PubMed Central

Wu, Dick; Bacaj, Taulant; Morishita, Wade; Goswami, Debanjan; Arendt, Kristin L.; Xu, Wei; Chen, Lu; Malenka, Robert C.; Südhof, Thomas C.

2017-01-01

Strengthening of synaptic connections by NMDA-receptor-dependent long-term potentiation (LTP) shapes neural circuits and mediates learning and memory. During NMDA-receptor-dependent LTP induction, Ca2+-influx stimulates recruitment of synaptic AMPA-receptors, thereby strengthening synapses. How Ca2+ induces AMPA-receptor recruitment, however, remains unclear. Here we show that, in pyramidal neurons of the hippocampal CA1-region, blocking postsynaptic expression of both synaptotagmin-1 and synaptotagmin-7, but not of synaptotagmin-1 or synaptotagmin-7 alone, abolished LTP. LTP was rescued by wild-type but not by Ca2+-binding-deficient mutant synaptotagmin-7. Blocking postsynaptic synaptotagmin-1/7 expression did not impair basal synaptic transmission, synaptic or extrasynaptic AMPA-receptor levels, or other AMPA-receptor trafficking events. Moreover, expression of dominant-negative mutant synaptotagmin-1 that inhibited Ca2+-dependent presynaptic vesicle exocytosis also blocked Ca2+-dependent postsynaptic AMPA-receptor exocytosis, thereby abolishing LTP. Our results suggest that postsynaptic synaptotagmin-1 and synaptotagmin-7 act as redundant Ca2+-sensors for Ca2+-dependent exocytosis of AMPA-receptors during LTP, thus delineating a simple mechanism for the recruitment of AMPA-receptors that mediates LTP. PMID:28355182

Mechanics of receptor-mediated endocytosis

NASA Astrophysics Data System (ADS)

Gao, Huajian; Shi, Wendong; Freund, Lambert B.

2005-07-01

Most viruses and bioparticles endocytosed by cells have characteristic sizes in the range of tens to hundreds of nanometers. The process of viruses entering and leaving animal cells is mediated by the binding interaction between ligand molecules on the viral capid and their receptor molecules on the cell membrane. How does the size of a bioparticle affect receptor-mediated endocytosis? Here, we study how a cell membrane containing diffusive mobile receptors wraps around a ligand-coated cylindrical or spherical particle. It is shown that particles in the size range of tens to hundreds of nanometers can enter or exit cells via wrapping even in the absence of clathrin or caveolin coats, and an optimal particles size exists for the smallest wrapping time. This model can also be extended to include the effect of clathrin coat. The results seem to show broad agreement with experimental observations. Author contributions: H.G. and L.B.F. designed research; H.G., W.S., and L.B.F. performed research; and H.G., W.S., and L.B.F. wrote the paper.Abbreviations: CNT, carbon nanotube; SWNT, single-walled nanotube.

Ionotropic and metabotropic receptor mediated airway sensory nerve activation.

PubMed

Lee, Min-Goo; Kollarik, Marian; Chuaychoo, Benjamas; Undem, Bradley J

2004-01-01

There are several receptors capable of inducing activating generator potentials in cough-associated afferent terminals in the airways. The chemical receptors leading to generator potentials can be subclassified into ionotropic and metabotropic types. An ionotropic receptor has an agonist-binding domain, and also serves directly as an ion channel that is opened upon binding of the agonist. Examples of ionotropic receptors found in airway sensory nerve terminals include receptors for serotonin (5-HT3 receptors), ATP (P2X receptors), acetylcholine (nicotinic receptors), receptors for capsaicin and related vanilloids (TRPV1 receptors), and acid receptors (acid sensing ion channels). Afferent nerve terminals can also be depolarized via activation of metabotropic or G-protein coupled receptors (GPCRs). Among the GPCRs that can lead to activation of airway afferent fibers include bradykinin B2 and adenosine A1 receptors. The signaling events leading to GPCR-mediated membrane depolarization are more complex than that seen with ionotropic receptors. The GPCR-mediated effects are thought to occur through classical second messenger systems such as activation of phospholipase C. This may lead to membrane depolarization through interaction with specific ionotropic receptors (such as TRPV1) and/or various types of calcium activated channels.

Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

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

Xu, Yuan, E-mail: yuan.xu@ki.se; Cardell, Lars-Olaf

Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B{sub 2} receptor agonist) and des-Arg{sup 9}-bradykinin-more » (selective B{sub 1} receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE{sub 2}. The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg{sup 9}-bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B{sub 2} receptors, but not those on B{sub 1}. Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance

Structure, Dynamics, and Allosteric Potential of Ionotropic Glutamate Receptor N-Terminal Domains.

PubMed

Krieger, James; Bahar, Ivet; Greger, Ingo H

2015-09-15

Ionotropic glutamate receptors (iGluRs) are tetrameric cation channels that mediate synaptic transmission and plasticity. They have a unique modular architecture with four domains: the intracellular C-terminal domain (CTD) that is involved in synaptic targeting, the transmembrane domain (TMD) that forms the ion channel, the membrane-proximal ligand-binding domain (LBD) that binds agonists such as L-glutamate, and the distal N-terminal domain (NTD), whose function is the least clear. The extracellular portion, comprised of the LBD and NTD, is loosely arranged, mediating complex allosteric regulation and providing a rich target for drug development. Here, we briefly review recent work on iGluR NTD structure and dynamics, and further explore the allosteric potential for the NTD in AMPA-type iGluRs using coarse-grained simulations. We also investigate mechanisms underlying the established NTD allostery in NMDA-type iGluRs, as well as the fold-related metabotropic glutamate and GABAB receptors. We show that the clamshell motions intrinsically favored by the NTD bilobate fold are coupled to dimeric and higher-order rearrangements that impact the iGluR LBD and ultimately the TMD. Finally, we explore the dynamics of intact iGluRs and describe how it might affect receptor operation in a synaptic environment. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Targeting receptor-mediated endocytotic pathways with nanoparticles: rationale and advances

PubMed Central

Xu, Shi; Olenyuk, Bogdan Z.; Okamoto, Curtis T.; Hamm-Alvarez, Sarah F.

2012-01-01

Targeting of drugs and their carrier systems by using receptor-mediated endocytotic pathways was in its nascent stages 25 years ago. In the intervening years, an explosion of knowledge focused on design and synthesis of nanoparticulate delivery systems as well as elucidation of the cellular complexity of what was previously-termed receptor-mediated endocytosis has now created a situation when it has become possible to design and test the feasibility of delivery of highly specific nanoparticle drug carriers to specific cells and tissue. This review outlines the mechanisms governing the major modes of receptor-mediated endocytosis used in drug delivery and highlights recent approaches using these as targets for in vivo drug delivery of nanoparticles. The review also discusses some of the inherent complexity associated with the simple shift from a ligand-drug conjugate versus a ligand-nanoparticle conjugate, in terms of ligand valency and its relationship to the mode of receptor-mediated internalization. PMID:23026636

Glucocorticoid Receptor-Mediated Repression of Pro-Inflammatory Genes in Rheumatoid Arthritis

DTIC Science & Technology

2015-10-01

1 AWARD NUMBER: W81XWH-14-1-0314 TITLE: Glucocorticoid Receptor-Mediated Repression of Pro-Inflammatory Genes in Rheumatoid Arthritis ...19 Sep 2015 4. TITLE AND SUBTITLE Glucocorticoid Receptor-Mediated Repression of Pro- Inflammatory Genes in Rheumatoid Arthritis 5a. CONTRACT NUMBER...SUBJECT TERMS Rheumatoid arthritis , inflammation and autoimmunity, macrophages, glucocorticoid receptor, transcriptional regulation, coactivators and

Stronger Dopamine D1 Receptor-Mediated Neurotransmission in Dyskinesia.

PubMed

Farré, Daniel; Muñoz, Ana; Moreno, Estefanía; Reyes-Resina, Irene; Canet-Pons, Júlia; Dopeso-Reyes, Iria G; Rico, Alberto J; Lluís, Carme; Mallol, Josefa; Navarro, Gemma; Canela, Enric I; Cortés, Antonio; Labandeira-García, José L; Casadó, Vicent; Lanciego, José L; Franco, Rafael

2015-12-01

Radioligand binding assays to rat striatal dopamine D1 receptors showed that brain lateralization of the dopaminergic system were not due to changes in expression but in agonist affinity. D1 receptor-mediated striatal imbalance resulted from a significantly higher agonist affinity in the left striatum. D1 receptors heteromerize with dopamine D3 receptors, which are considered therapeutic targets for dyskinesia in parkinsonian patients. Expression of both D3 and D1-D3 receptor heteromers were increased in samples from 6-hydroxy-dopamine-hemilesioned rats rendered dyskinetic by treatment with 3, 4-dihydroxyphenyl-L-alanine (L-DOPA). Similar findings were obtained using striatal samples from primates. Radioligand binding studies in the presence of a D3 agonist led in dyskinetic, but not in lesioned or L-DOPA-treated rats, to a higher dopamine sensitivity. Upon D3-receptor activation, the affinity of agonists for binding to the right striatal D1 receptor increased. Excess dopamine coming from L-DOPA medication likely activates D3 receptors thus making right and left striatal D1 receptors equally responsive to dopamine. These results show that dyskinesia occurs concurrently with a right/left striatal balance in D1 receptor-mediated neurotransmission.

Gaba mediated long-term depression (LTD) in the rat medial vestibular nuclei.

PubMed

Grassi, S; Della Torre, G; Zampolini, M; Pettorossi, V E

1995-01-01

As previously demonstrated, high frequency stimulation (HFS) of the primary vestibular afferents always induces a clear, long lasting depression of the polysynaptic (N2) component of the field potentials recorded in the dorsal portion of the medial vestibular nuclei (MVN). The induction of the HFS effect was mediated by the activation of glutamate NMDA receptors, since it was blocked by AP5. The mechanisms at the basis of such a depression were studied. Our results demonstrate that Gaba, acting on both GabaA and GabaB receptors, is involved in mediating this phenomenon. In fact, HFS applied during Bicuculline and Saclofen perfusion, was no longer able to induce an N2 depression, but provoked a slight potentiation. However, the N2 depression clearly emerged after drug wash-out. Furthermore, Bicuculline and Saclofen fully abolished the N2 depression and highlighted the potentiation, when administered after HFS. The possibility that the N2 depression is the result of a homosynaptic LTD can be excluded on the basis of our results. On the contrary, our findings suggest that the depression is due to an enhancement of the Gaba inhibitory effect due to an HFS dependent increase in gabaergic interneuron activity, which resets vestibular neuron excitability at a lower level.

Milrinone attenuates thromboxane receptor-mediated hyperresponsiveness in hypoxic pulmonary arterial myocytes.

PubMed

Santhosh, K T; Elkhateeb, O; Nolette, N; Outbih, O; Halayko, A J; Dakshinamurti, S

2011-07-01

Neonatal pulmonary hypertension (PPHN) is characterized by pulmonary vasoconstriction, due in part to dysregulation of the thromboxane prostanoid (TP) receptor. Hypoxia induces TP receptor-mediated hyperresponsiveness, whereas serine phosphorylation mediates desensitization of TP receptors. We hypothesized that prostacyclin (IP) receptor activity induces TP receptor phosphorylation and decreases ligand affinity; that TP receptor sensitization in hypoxic myocytes is due to IP receptor inactivation; and that this would be reversible by the cAMP-specific phosphodiesterase inhibitor milrinone. We examined functional regulation of TP receptors by serine phosphorylation and effects of IP receptor stimulation and protein kinase A (PKA) activity on TP receptor sensitivity in myocytes from neonatal porcine resistance pulmonary arteries after 72 h hypoxia in vitro. Ca(2+) response curves to U46619 (TP receptor agonist) were determined in hypoxic and normoxic myocytes incubated with or without iloprost (IP receptor agonist), forskolin (adenylyl cyclase activator), H8 (PKA inhibitor) or milrinone. TP and IP receptor saturation binding kinetics were measured in presence of iloprost or 8-bromo-cAMP. Ligand affinity for TP receptors was normalized in vitro by IP receptor signalling intermediates. However, IP receptor affinity was compromised in hypoxic myocytes, decreasing cAMP production. Milrinone normalized TP receptor sensitivity in hypoxic myocytes by restoring PKA-mediated regulatory TP receptor phosphorylation. TP receptor sensitivity and EC(50) for TP receptor agonists was regulated by PKA, as TP receptor serine phosphorylation by PKA down-regulated Ca(2+) mobilization. Hypoxia decreased IP receptor activity and cAMP generation, inducing TP receptor hyperresponsiveness, which was reversed by milrinone. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Milrinone attenuates thromboxane receptor-mediated hyperresponsiveness in hypoxic pulmonary arterial myocytes

PubMed Central

Santhosh, KT; Elkhateeb, O; Nolette, N; Outbih, O; Halayko, AJ; Dakshinamurti, S

2011-01-01

BACKGROUND AND PURPOSE Neonatal pulmonary hypertension (PPHN) is characterized by pulmonary vasoconstriction, due in part to dysregulation of the thromboxane prostanoid (TP) receptor. Hypoxia induces TP receptor–mediated hyperresponsiveness, whereas serine phosphorylation mediates desensitization of TP receptors. We hypothesized that prostacyclin (IP) receptor activity induces TP receptor phosphorylation and decreases ligand affinity; that TP receptor sensitization in hypoxic myocytes is due to IP receptor inactivation; and that this would be reversible by the cAMP-specific phosphodiesterase inhibitor milrinone. EXPERIMENTAL APPROACH We examined functional regulation of TP receptors by serine phosphorylation and effects of IP receptor stimulation and protein kinase A (PKA) activity on TP receptor sensitivity in myocytes from neonatal porcine resistance pulmonary arteries after 72 h hypoxia in vitro. Ca2+ response curves to U46619 (TP receptor agonist) were determined in hypoxic and normoxic myocytes incubated with or without iloprost (IP receptor agonist), forskolin (adenylyl cyclase activator), H8 (PKA inhibitor) or milrinone. TP and IP receptor saturation binding kinetics were measured in presence of iloprost or 8-bromo-cAMP. KEY RESULTS Ligand affinity for TP receptors was normalized in vitro by IP receptor signalling intermediates. However, IP receptor affinity was compromised in hypoxic myocytes, decreasing cAMP production. Milrinone normalized TP receptor sensitivity in hypoxic myocytes by restoring PKA-mediated regulatory TP receptor phosphorylation. CONCLUSIONS AND IMPLICATIONS TP receptor sensitivity and EC50 for TP receptor agonists was regulated by PKA, as TP receptor serine phosphorylation by PKA down-regulated Ca2+ mobilization. Hypoxia decreased IP receptor activity and cAMP generation, inducing TP receptor hyperresponsiveness, which was reversed by milrinone. PMID:21385177

MicroRNA and receptor mediated signaling pathways as potential therapeutic targets in heart failure.

PubMed

Tuttolomondo, Antonino; Simonetta, Irene; Pinto, Antonio

2016-11-01

Cardiac remodelling is a complex pathogenetic pathway involving genome expression, molecular, cellular, and interstitial changes that cause changes in size, shape and function of the heart after cardiac injury. Areas covered: We will review recent advances in understanding the role of several receptor-mediated signaling pathways and micro-RNAs, in addition to their potential as candidate target pathways in the pathogenesis of heart failure. The myocyte is the main target cell involved in the remodelling process via ischemia, cell necrosis and apoptosis (by means of various receptor pathways), and other mechanisms mediated by micro-RNAs. We will analyze the role of some receptor mediated signaling pathways such as natriuretic peptides, mediators of glycogen synthase kinase 3 and ERK1/2 pathways, beta-adrenergic receptor subtypes and relaxin receptor signaling mechanisms, TNF/TNF receptor family and TWEAK/Fn14 axis, and some micro-RNAs as candidate target pathways in pathogenesis of heart failure. These mediators of receptor-mediated pathways and micro-RNA are the most addressed targets of emerging therapies in modern heart failure treatment strategies. Expert opinion: Future treatment strategies should address mediators involved in multiple steps within heart failure pathogenetic pathways.

Insulin Regulates GABAA Receptor-Mediated Tonic Currents in the Prefrontal Cortex.

PubMed

Trujeque-Ramos, Saraí; Castillo-Rolón, Diego; Galarraga, Elvira; Tapia, Dagoberto; Arenas-López, Gabina; Mihailescu, Stefan; Hernández-López, Salvador

2018-01-01

Recent studies, have shown that insulin increases extrasynaptic GABA A receptor-mediated currents in the hippocampus, causing alterations of neuronal excitability. The prefrontal cortex (PFC) is another brain area which is involved in cognition functions and expresses insulin receptors. Here, we used electrophysiological, molecular, and immunocytochemical techniques to examine the effect of insulin on the extrasynaptic GABA A receptor-mediated tonic currents in brain slices. We found that insulin (20-500 nM) increases GABA A -mediated tonic currents. Our results suggest that insulin promotes the trafficking of extrasynaptic GABA A receptors from the cytoplasm to the cell membrane. Western blot analysis and immunocytochemistry showed that PFC extrasynaptic GABA A receptors contain α-5 and δ subunits. Insulin effect on tonic currents decreased the firing rate and neuronal excitability in layer 5-6 PFC cells. These effects of insulin were dependent on the activation of the PI3K enzyme, a key mediator of the insulin response within the brain. Taken together, these results suggest that insulin modulation of the GABA A -mediated tonic currents can modify the activity of neural circuits within the PFC. These actions could help to explain the alterations of cognitive processes associated with changes in insulin signaling.

Baclofen mediates neuroprotection on hippocampal CA1 pyramidal cells through the regulation of autophagy under chronic cerebral hypoperfusion

PubMed Central

Liu, Li; Li, Chang-jun; Lu, Yun; Zong, Xian-gang; Luo, Chao; Sun, Jun; Guo, Lian-jun

2015-01-01

GABA receptors play an important role in ischemic brain injury. Studies have indicated that autophagy is closely related to neurodegenerative diseases. However, during chronic cerebral hypoperfusion, the changes of autophagy in the hippocampal CA1 area, the correlation between GABA receptors and autophagy, and their influences on hippocampal neuronal apoptosis have not been well established. Here, we found that chronic cerebral hypoperfusion resulted in rat hippocampal atrophy, neuronal apoptosis, enhancement and redistribution of autophagy, down-regulation of Bcl-2/Bax ratio, elevation of cleaved caspase-3 levels, reduction of surface expression of GABAA receptor α1 subunit and an increase in surface and mitochondrial expression of connexin 43 (CX43) and CX36. Chronic administration of GABAB receptors agonist baclofen significantly alleviated neuronal damage. Meanwhile, baclofen could up-regulate the ratio of Bcl-2/Bax and increase the activation of Akt, GSK-3β and ERK which suppressed cytodestructive autophagy. The study also provided evidence that baclofen could attenuate the decrease in surface expression of GABAA receptor α1 subunit, and down-regulate surface and mitochondrial expression of CX43 and CX36, which might enhance protective autophagy. The current findings suggested that, under chronic cerebral hypoperfusion, the effects of GABAB receptors activation on autophagy regulation could reverse neuronal damage. PMID:26412641

Decreased GABA receptor in the cerebral cortex of epileptic rats: effect of Bacopa monnieri and Bacoside-A.

PubMed

Mathew, Jobin; Balakrishnan, Savitha; Antony, Sherin; Abraham, Pretty Mary; Paulose, C S

2012-02-24

Gamma amino butyric acid (GABA), the principal inhibitory neurotransmitter in the cerebral cortex, maintains the inhibitory tones that counter balances neuronal excitation. When this balance is perturbed, seizures may ensue. In the present study, alterations of the general GABA, GABAA and GABAB receptors in the cerebral cortex of the epileptic rat and the therapeutic application of Bacopa monnieri were investigated. Scatchard analysis of [3H]GABA, [3H]bicuculline and [3H]baclofen in the cerebral cortex of the epileptic rat showed significant decrease in Bmax (P < 0.001) compared to control. Real Time PCR amplification of GABA receptor subunits such as GABAAά1, GABAAγ, GABAAδ, GABAB and GAD where down regulated (P < 0.001) in epileptic rats. GABAAά5 subunit and Cyclic AMP responsible element binding protein were up regulated. Confocal imaging study confirmed the decreased GABA receptors in epileptic rats. Epileptic rats have deficit in radial arm and Y maze performance. Bacopa monnieri and Bacoside-A treatment reverses epilepsy associated changes to near control suggesting that decreased GABA receptors in the cerebral cortex have an important role in epileptic occurrence; Bacopa monnieri and Bacoside-A have therapeutic application in epilepsy management.

SOFI of GABAB neurotransmitter receptors in hippocampal neurons elucidates intracellular receptor trafficking and assembly

NASA Astrophysics Data System (ADS)

Huss, Anja; Ramírez, Omar; Santibáñez, Felipe; Couve, Andrés.; Härtel, Steffen; Enderlein, Jörg

2013-02-01

The synaptic efficacy of neurons depends on the number of neurotransmitter receptors in the plasma membrane. The availability of these receptors is controlled by their specific intracellular trafficking routes. γ-Aminobutyric acid type B receptors (GABABRs) are heteromeric proteins consisting of GABABR1 and GABABR2 subunits. These receptors are found at the plasma membrane of somatodendritic postsynaptic sites and in axons. It is unknown whether the assembly of the subunits occurs directly in the somatic endoplasmic reticulum (ER) followed by vesicular transport, or whether the assembly occurs after the separate transport of the subunits to the dendritic ER compartment. To address this question we have studied the assembly of the GABABRs in hippocampal neurons with dual-color, 3D super-resolution optical fluctuation imaging (SOFI). SOFI is a fluorescence imaging modality which yields superresolved spatial resolution, 3D-sectioning and high image contrast. We will use the SOFI images to quantify the distribution of the GABABR subunits in the plasma membrane and in the dendritic intracellular compartments. Finally, we want to apply quantitative co-localization analysis to determine the compartments in which the assembly of the GABABR subunits occurs.

Prostanoids and their receptors that modulate dendritic cell-mediated immunity.

PubMed

Gualde, Norbert; Harizi, Hedi

2004-08-01

Dendritic cells (DC) are essential for the initiation of immune responses by capturing, processing and presenting antigens to T cells. In addition to their important role as professional APC, they are able to produce immunosuppressive and pro-inflammatory prostanoids from arachidonic acid (AA) by the action of cyclooxygenase (COX) enzymes. In an autocrine and paracrine fashion, the secreted lipid mediators subsequently modulate the maturation, cytokine production, Th-cell polarizing ability, chemokine receptor expression, migration, and apoptosis of these extremely versatile APC. The biological actions of prostanoids, including their effects on APC-mediated immunity and acute inflammatory responses, are exerted by G protein-coupled receptors on plasma membrane. Some COX metabolites act as anti-inflammatory lipid mediators by binding to nuclear receptors and modulating DC functions. Although the role of cytokines in DC function has been studied extensively, the effects of prostanoids on DC biology have only recently become the focus of investigation. This review summarizes the current knowledge about the role of prostanoids and their receptors in modulating DC function and the subsequent immune responses.

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

Roles of OA1 octopamine receptor and Dop1 dopamine receptor in mediating appetitive and aversive reinforcement revealed by RNAi studies

PubMed Central

Awata, Hiroko; Wakuda, Ryo; Ishimaru, Yoshiyasu; Matsuoka, Yuji; Terao, Kanta; Katata, Satomi; Matsumoto, Yukihisa; Hamanaka, Yoshitaka; Noji, Sumihare; Mito, Taro; Mizunami, Makoto

2016-01-01

Revealing reinforcing mechanisms in associative learning is important for elucidation of brain mechanisms of behavior. In mammals, dopamine neurons are thought to mediate both appetitive and aversive reinforcement signals. Studies using transgenic fruit-flies suggested that dopamine neurons mediate both appetitive and aversive reinforcements, through the Dop1 dopamine receptor, but our studies using octopamine and dopamine receptor antagonists and using Dop1 knockout crickets suggested that octopamine neurons mediate appetitive reinforcement and dopamine neurons mediate aversive reinforcement in associative learning in crickets. To fully resolve this issue, we examined the effects of silencing of expression of genes that code the OA1 octopamine receptor and Dop1 and Dop2 dopamine receptors by RNAi in crickets. OA1-silenced crickets exhibited impairment in appetitive learning with water but not in aversive learning with sodium chloride solution, while Dop1-silenced crickets exhibited impairment in aversive learning but not in appetitive learning. Dop2-silenced crickets showed normal scores in both appetitive learning and aversive learning. The results indicate that octopamine neurons mediate appetitive reinforcement via OA1 and that dopamine neurons mediate aversive reinforcement via Dop1 in crickets, providing decisive evidence that neurotransmitters and receptors that mediate appetitive reinforcement indeed differ among different species of insects. PMID:27412401

Roles of OA1 octopamine receptor and Dop1 dopamine receptor in mediating appetitive and aversive reinforcement revealed by RNAi studies.

PubMed

Awata, Hiroko; Wakuda, Ryo; Ishimaru, Yoshiyasu; Matsuoka, Yuji; Terao, Kanta; Katata, Satomi; Matsumoto, Yukihisa; Hamanaka, Yoshitaka; Noji, Sumihare; Mito, Taro; Mizunami, Makoto

2016-07-14

Revealing reinforcing mechanisms in associative learning is important for elucidation of brain mechanisms of behavior. In mammals, dopamine neurons are thought to mediate both appetitive and aversive reinforcement signals. Studies using transgenic fruit-flies suggested that dopamine neurons mediate both appetitive and aversive reinforcements, through the Dop1 dopamine receptor, but our studies using octopamine and dopamine receptor antagonists and using Dop1 knockout crickets suggested that octopamine neurons mediate appetitive reinforcement and dopamine neurons mediate aversive reinforcement in associative learning in crickets. To fully resolve this issue, we examined the effects of silencing of expression of genes that code the OA1 octopamine receptor and Dop1 and Dop2 dopamine receptors by RNAi in crickets. OA1-silenced crickets exhibited impairment in appetitive learning with water but not in aversive learning with sodium chloride solution, while Dop1-silenced crickets exhibited impairment in aversive learning but not in appetitive learning. Dop2-silenced crickets showed normal scores in both appetitive learning and aversive learning. The results indicate that octopamine neurons mediate appetitive reinforcement via OA1 and that dopamine neurons mediate aversive reinforcement via Dop1 in crickets, providing decisive evidence that neurotransmitters and receptors that mediate appetitive reinforcement indeed differ among different species of insects.

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.

Plant cell surface receptor-mediated signaling - a common theme amid diversity.

PubMed

He, Yunxia; Zhou, Jinggeng; Shan, Libo; Meng, Xiangzong

2018-01-29

Sessile plants employ a diverse array of plasma membrane-bound receptors to perceive endogenous and exogenous signals for regulation of plant growth, development and immunity. These cell surface receptors include receptor-like kinases (RLKs) and receptor-like proteins (RLPs) that harbor different extracellular domains for perception of distinct ligands. Several RLK and RLP signaling pathways converge at the somatic embryogenesis receptor kinases (SERKs), which function as shared co-receptors. A repertoire of receptor-like cytoplasmic kinases (RLCKs) associate with the receptor complexes to relay intracellular signaling. Downstream of the receptor complexes, mitogen-activated protein kinase (MAPK) cascades are among the key signaling modules at which the signals converge, and these cascades regulate diverse cellular and physiological responses through phosphorylation of different downstream substrates. In this Review, we summarize the emerging common theme that underlies cell surface receptor-mediated signaling pathways in Arabidopsis thaliana : the dynamic association of RLKs and RLPs with specific co-receptors and RLCKs for signal transduction. We further discuss how signaling specificities are maintained through modules at which signals converge, with a focus on SERK-mediated receptor signaling. © 2018. Published by The Company of Biologists Ltd.

Ionotropic AMPA-type glutamate and metabotropic GABAB receptors: determining cellular physiology by proteomes.

PubMed

Bettler, Bernhard; Fakler, Bernd

2017-08-01

Ionotropic AMPA-type glutamate receptors and G-protein-coupled metabotropic GABA B receptors are key elements of neurotransmission whose cellular functions are determined by their protein constituents. Over the past couple of years unbiased proteomic approaches identified comprehensive sets of protein building blocks of these two types of neurotransmitter receptors in the brain (termed receptor proteomes). This provided the opportunity to match receptor proteomes with receptor physiology and to study the structural organization, regulation and function of native receptor complexes in an unprecedented manner. In this review we discuss the principles of receptor architecture and regulation emerging from the functional characterization of the proteomes of AMPA and GABA B receptors. We also highlight progress in unraveling the role of unexpected protein components for receptor physiology. Copyright © 2017 Elsevier Ltd. All rights reserved.

Long-term depression of inhibitory synaptic transmission induced by spike-timing dependent plasticity requires coactivation of endocannabinoid and muscarinic receptors.

PubMed

Ahumada, Juan; Fernández de Sevilla, David; Couve, Alejandro; Buño, Washington; Fuenzalida, Marco

2013-12-01

The precise timing of pre-postsynaptic activity is vital for the induction of long-term potentiation (LTP) or depression (LTD) at many central synapses. We show in synapses of rat CA1 pyramidal neurons in vitro that spike timing dependent plasticity (STDP) protocols that induce LTP at glutamatergic synapses can evoke LTD of inhibitory postsynaptic currents or STDP-iLTD. The STDP-iLTD requires a postsynaptic Ca(2+) increase, a release of endocannabinoids (eCBs), the activation of type-1 endocananabinoid receptors and presynaptic muscarinic receptors that mediate a decreased probability of GABA release. In contrast, the STDP-iLTD is independent of the activation of nicotinic receptors, GABAB Rs and G protein-coupled postsynaptic receptors at pyramidal neurons. We determine that the downregulation of presynaptic Cyclic adenosine monophosphate/protein Kinase A pathways is essential for the induction of STDP-iLTD. These results suggest a novel mechanism by which the activation of cholinergic neurons and retrograde signaling by eCBs can modulate the efficacy of GABAergic synaptic transmission in ways that may contribute to information processing and storage in the hippocampus. Copyright © 2013 Wiley Periodicals, Inc.

A functional assay to measure postsynaptic gamma-aminobutyric acidB responses in cultured spinal cord neurons: Heterologous regulation of the same K+ channel

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

Kamatchi, G.L.; Ticku, M.K.

1991-02-01

The stimulation of postsynaptic gamma-aminobutyric acid (GABA)B receptors leads to slow inhibitory postsynaptic potentials due to the influx of K(+)-ions. This was studied biochemically, in vitro in mammalian cultured spinal cord neurons by using 86Rb as a substitute for K+. (-)-Baclofen, a GABAB receptor agonist, produced a concentration-dependent increase in the 86Rb-influx. This effect was stereospecific and blocked by GABAB receptor antagonists like CGP 35 348 (3-aminopropyl-diethoxymethyl-phosphonic acid) and phaclofen. Apart from the GABAB receptors, both adenosine via adenosine1 receptors and 5-hydroxytryptamine (5-HT) via 5-HT1 alpha agonists also increased the 86Rb-influx. These agonists failed to show any additivity between themmore » when they were combined in their maximal concentration. In addition, their effect was antagonized specifically by their respective antagonists without influencing the others. These findings suggest the presence of GABAB, adenosine1 and 5-HT1 alpha receptors in the cultured spinal cord neurons, which exhibit a heterologous regulation of the same K(+)-channel. The effect of these agonists were antagonized by phorbol 12,13-didecanoate, an activator of protein kinase C, and pretreatment with pertussis toxin. This suggests that these agonists by acting on their own receptors converge on the same K(+)-channel through the Gi/Go proteins. In summary, we have developed a biochemical functional assay for studying and characterizing GABAB synaptic pharmacology in vitro, using spinal cord neurons.« less

Control of transient lower oesophageal sphincter relaxations and reflux by the GABAB agonist baclofen in patients with gastro-oesophageal reflux disease

PubMed Central

Zhang, Q; Lehmann, A; Rigda, R; Dent, J; Holloway, R H

2002-01-01

Background and aims: Transient lower oesophageal sphincter relaxations (TLOSRs) are the major cause of gastro-oesophageal reflux in normal subjects and in most patients with reflux disease. The gamma aminobutyric acid (GABA) receptor type B agonist, baclofen, is a potent inhibitor of TLOSRs in normal subjects. The aim of this study was to investigate the effect of baclofen on TLOSRs and postprandial gastro-oesophageal reflux in patients with reflux disease. Methods: In 20 patients with reflux disease, oesophageal motility and pH were measured, with patients in the sitting position, for three hours after a 3000 kJ mixed nutrient meal. On separate days at least one week apart, 40 mg oral baclofen or placebo was given 90 minutes before the meal. Results: Baclofen reduced the rate of TLOSRs by 40% from 15 (13.8–18.3) to 9 (5.8–13.3) per three hours (p<0.0002) and increased basal lower oesophageal sphincter pressure. Baclofen also significantly reduced the rate of reflux episodes by 43% from 7.0 (4.0–12.0) to 4.0 (1.5–9) per three hours (median (interquartile range); p<0.02). However, baclofen had no effect on oesophageal acid exposure (baclofen 4.9% (1.7–12.4) v placebo 5.0% (2.7–15.5)). Conclusions: In patients with reflux disease, the GABAB agonist baclofen significantly inhibits gastro-oesophageal reflux episodes by inhibition of TLOSRs. These findings suggest that GABAB agonists may be useful as therapeutic agents for the management of reflux disease. PMID:11772961

Extrasynaptic GABAA receptors in the crosshairs of hormones and ethanol

PubMed Central

Mody, Istvan

2008-01-01

Gamma-aminobutyric acid (GABA) is the main chemical inhibitory neurotransmitter in the brain. In the central nervous system (CNS) it acts on two distinct types of receptor: an ion channel, i.e., an “ionotropic” receptor permeable to Cl− and HCO3− (GABAA receptors) and a G-protein coupled “metabotropic” receptor that is linked to various effector mechanisms (GABAB receptors). This review will summarize novel developments in the physiology and pharmacology of GABAA receptors (GABAARs), specifically those found outside synapses. The focus will be on a particular combination of GABAAR subunits sensitive to ovarian and adrenal cortical steroid hormone metabolites that are synthesized in the brain (neurosteroids) and to sobriety impairing concentrations of ethanol. These receptors may be the final common pathway for interactions between ethanol and ovarian and stress-related neurosteroids. PMID:17714830

Decreased GABA receptor in the cerebral cortex of epileptic rats: effect of Bacopa monnieri and Bacoside-A

PubMed Central

2012-01-01

Abstact Background Gamma amino butyric acid (GABA), the principal inhibitory neurotransmitter in the cerebral cortex, maintains the inhibitory tones that counter balances neuronal excitation. When this balance is perturbed, seizures may ensue. Methods In the present study, alterations of the general GABA, GABAA and GABAB receptors in the cerebral cortex of the epileptic rat and the therapeutic application of Bacopa monnieri were investigated. Results Scatchard analysis of [3H]GABA, [3H]bicuculline and [3H]baclofen in the cerebral cortex of the epileptic rat showed significant decrease in Bmax (P < 0.001) compared to control. Real Time PCR amplification of GABA receptor subunits such as GABAAά1, GABAAγ, GABAAδ, GABAB and GAD where down regulated (P < 0.001) in epileptic rats. GABAAά5 subunit and Cyclic AMP responsible element binding protein were up regulated. Confocal imaging study confirmed the decreased GABA receptors in epileptic rats. Epileptic rats have deficit in radial arm and Y maze performance. Conclusions Bacopa monnieri and Bacoside-A treatment reverses epilepsy associated changes to near control suggesting that decreased GABA receptors in the cerebral cortex have an important role in epileptic occurrence; Bacopa monnieri and Bacoside-A have therapeutic application in epilepsy management. PMID:22364254

Selectivity and specificity of sphingosine-1-phosphate receptor ligands: caveats and critical thinking in characterizing receptor-mediated effects.

PubMed

Salomone, Salvatore; Waeber, Christian

2011-01-01

Receptors for sphingosine-1-phosphate (S1P) have been identified only recently. Their medicinal chemistry is therefore still in its infancy, and few selective agonists or antagonists are available. Furthermore, the selectivity of S1P receptor agonists or antagonists is not well established. JTE-013 and BML-241 (also known as CAY10444), used extensively as specific S1P(2) and S1P(3) receptors antagonists respectively, are cases in point. When analyzing S1P-induced vasoconstriction in mouse basilar artery, we observed that JTE-013 inhibited not only the effect of S1P, but also the effect of U46619, endothelin-1 or high KCl; JTE-013 strongly inhibited responses to S1P in S1P(2) receptor knockout mice. Similarly, BML-241 has been shown to inhibit increases in intracellular Ca(2+) concentration via P(2) receptor or α(1A)-adrenoceptor stimulation and α(1A)-adrenoceptor-mediated contraction of rat mesenteric artery, while it did not affect S1P(3)-mediated decrease of forskolin-induced cyclic AMP accumulation. Another putative S1P(1/3) receptor antagonist, VPC23019, does not inhibit S1P(3)-mediated vasoconstriction. With these examples in mind, we discuss caveats about relying on available pharmacological tools to characterize receptor subtypes.

The proteinase-activated receptor-2 mediates phagocytosis in a Rho-dependent manner in human keratinocytes.

PubMed

Scott, Glynis; Leopardi, Sonya; Parker, Lorelle; Babiarz, Laura; Seiberg, Miri; Han, Rujiing

2003-09-01

Recent work shows that the G-protein-coupled receptor proteinase activated receptor-2 activates signals that stimulate melanosome uptake in keratinocytes in vivo and in vitro. The Rho family of GTP-binding proteins is involved in cytoskeletal remodeling during phagocytosis. We show that proteinase-activated receptor-2 mediated phagocytosis in human keratinocytes is Rho dependent and that proteinase-activated receptor-2 signals to activate Rho. In contrast, Rho activity did not affect either proteinase-activated receptor-2 activity or mRNA and protein levels. We explored the signaling mechanisms of proteinase-activated receptor-2 mediated Rho activation in human keratinocytes and show that activation of proteinase-activated receptor-2, either through specific proteinase-activated receptor-2 activating peptides or through trypsinization, elevates cAMP in keratinocytes. Proteinase-activated receptor-2 mediated Rho activation was pertussis toxin insensitive and independent of the protein kinase A signaling pathway. These data are the first to show that proteinase-activated receptor-2 mediated phagocytosis is Rho dependent and that proteinase-activated receptor-2 signals to Rho and cAMP in keratinocytes. Because phagocytosis of melanosomes is recognized as an important mechanism for melanosome transfer to keratinocytes, these results suggest that Rho is a critical signaling intermediate in melanosome uptake in keratinocytes.

Menthol enhances phasic and tonic GABAA receptor-mediated currents in midbrain periaqueductal grey neurons

PubMed Central

Lau, Benjamin K; Karim, Shafinaz; Goodchild, Ann K; Vaughan, Christopher W; Drew, Geoffrey M

2014-01-01

Background and Purpose Menthol, a naturally occurring compound in the essential oil of mint leaves, is used for its medicinal, sensory and fragrant properties. Menthol acts via transient receptor potential (TRPM8 and TRPA1) channels and as a positive allosteric modulator of recombinant GABAA receptors. Here, we examined the actions of menthol on GABAA receptor-mediated currents in intact midbrain slices. Experimental Approach Whole-cell voltage-clamp recordings were made from periaqueductal grey (PAG) neurons in midbrain slices from rats to determine the effects of menthol on GABAA receptor-mediated phasic IPSCs and tonic currents. Key Results Menthol (150–750 μM) produced a concentration-dependent prolongation of spontaneous GABAA receptor-mediated IPSCs, but not non-NMDA receptor-mediated EPSCs throughout the PAG. Menthol actions were unaffected by TRPM8 and TRPA1 antagonists, tetrodotoxin and the benzodiazepine antagonist, flumazenil. Menthol also enhanced a tonic current, which was sensitive to the GABAA receptor antagonists, picrotoxin (100 μM), bicuculline (30 μM) and Zn2+ (100 μM), but unaffected by gabazine (10 μM) and a GABAC receptor antagonist, 1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid hydrate (TPMPA; 50 μM). In addition, menthol potentiated currents induced by the extrasynaptic GABAA receptor agonist THIP/gaboxadol (10 μM). Conclusions and Implications These results suggest that menthol positively modulates both synaptic and extrasynaptic populations of GABAA receptors in native PAG neurons. The development of agents that potentiate GABAA-mediated tonic currents and phasic IPSCs in a manner similar to menthol could provide a basis for novel GABAA-related pharmacotherapies. PMID:24460753

Heterogeneity of prejunctional NPY receptor-mediated inhibition of cardiac neurotransmission

PubMed Central

Serone, Adrian P; Wright, Christine E; Angus, James A

1999-01-01

Neuropeptide Y (NPY) has been proposed as the candidate inhibitory peptide mediating interactions between sympathetic and vagal neurotransmission in several species, including man. Here, we have defined the NPY receptors involved in modulation of cardiac autonomic neurotransmission using receptor-selective agonists and antagonists in the rabbit and guinea-pig isolated right atria.In isolated atrial preparations, sympathetically-mediated tachycardia (ST; with atropine 1 μM) or vagally-mediated bradycardia (VB; with propranolol 0.1–1 μM) in response to electrical field stimulation (EFS, 1–4 pulses) were tested 0–30 min after incubation with single concentrations of vehicle, NPY (0.01–10 μM), the Y2 receptor agonist N-Acetyl-[Leu28,31]NPY(24–36) (termed N-A[L]NPY(24–36)) or the Y1 receptor agonist [Leu31,Pro34]NPY (LP). The effect of NPY on the concentration-chronotropic response curves to isoprenaline and bethanechol were also assessed.Guinea-pig atria: NPY and N-A[L]NPY(24–36) caused concentration-dependent inhibition of VB and ST to EFS. Both peptides caused maximal inhibition of VB and ST within 10 min incubation and this remained constant. LP caused a concentration-dependent, transient inhibition of ST which was antagonized by the Y1-receptor antagonist GR231118 (0.3 μM), with apparent competitive kinetics. Rabbit atria: NPY (1 or 10 μM) had no effect on VB at any time point, but both NPY and LP caused a transient (∼10 min) inhibition of sympathetic tachycardia. This inhibition could be prevented by 0.3 μM GR231118. N-A[L]NPY(24–36) had no effect on ST. NPY had no effect on the response to β-adrenoceptor stimulation by isoprenaline nor muscarinic-receptor stimulation by bethanechol in either species.Thus, in the guinea-pig, NPY causes a stable inhibition of both VB and ST to EFS via Y2 receptors and transient inhibition of ST via Y1 receptors. In contrast in the rabbit, NPY has no effect on the cardiac vagus and

Cell type specificity of GABA(A) receptor mediated signaling in the hippocampus.

PubMed

Semyanov, A

2003-08-01

Inhibitory signaling mediated by ionotropic GABA(1) receptors generally acts as a major brake against excessive excitability in the brain. This is especially relevant in epilepsy-prone structures such as the hippocampus, in which GABA(A) receptor mediated inhibition is critical in suppressing epileptiform activity. Indeed, potentiating GABA(A) receptor mediated signaling is an important target for antiepileptic drug therapy. GABA(A) receptor mediated inhibition has different roles in the network dependent on the target neuron. Inhibiting principal cells will thus reduce network excitability, whilst inhibiting interneurons will increase network excitability; GABAergic therapeutic agents do not distinguish between these two alternatives, which may explain why, on occasion, GABAergic antiepileptic drugs can be proconvulsant. The importance of the target-cell for the effect of neuroactive drugs has emerged from a number of recent studies. Immunocytochemical data have suggested non-uniform distribution of GABA(A) receptor subunits among hippocampal interneurons and pyramidal cells. This has been confirmed by subsequent electropharmacological data. These have demonstrated that compounds which act on GABA(A) receptors or the extracellular GABA concentration can have distinct effects in different neuronal populations. Recently, it has also been discovered that presynaptic glutamate heteroreceptors can modulate GABA release in the hippocampus in a postsynaptic cell-specific manner. Since systemically administrated drugs may act on different neuronal subtypes, they can exhibit paradoxical effects. Distinguishing compounds that have target specific effects on GABAergic signaling may lead to novel and more effective treatments against epilepsy.

GABAB receptor attenuation of GABAA currents in neurons of the mammalian central nervous system.

PubMed

Shen, Wen; Nan, Changlong; Nelson, Peter T; Ripps, Harris; Slaughter, Malcolm M

2017-03-01

Ionotropic receptors are tightly regulated by second messenger systems and are often present along with their metabotropic counterparts on a neuron's plasma membrane. This leads to the hypothesis that the two receptor subtypes can interact, and indeed this has been observed in excitatory glutamate and inhibitory GABA receptors. In both systems the metabotropic pathway augments the ionotropic receptor response. However, we have found that the metabotropic GABA B receptor can suppress the ionotropic GABA A receptor current, in both the in vitro mouse retina and in human amygdala membrane fractions. Expression of amygdala membrane microdomains in Xenopus oocytes by microtransplantation produced functional ionotropic and metabotropic GABA receptors. Most GABA A receptors had properties of α -subunit containing receptors, with ~5% having ρ -subunit properties. Only GABA A receptors with α -subunit-like properties were regulated by GABA B receptors. In mouse retinal ganglion cells, where only α -subunit-containing GABA A receptors are expressed, GABA B receptors suppressed GABA A receptor currents. This suppression was blocked by GABA B receptor antagonists, G-protein inhibitors, and GABA B receptor antibodies. Based on the kinetic differences between metabotropic and ionotropic receptors, their interaction would suppress repeated, rapid GABAergic inhibition. © 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

The TIM and TAM families of phosphatidylserine receptors mediate dengue virus entry.

PubMed

Meertens, Laurent; Carnec, Xavier; Lecoin, Manuel Perera; Ramdasi, Rasika; Guivel-Benhassine, Florence; Lew, Erin; Lemke, Greg; Schwartz, Olivier; Amara, Ali

2012-10-18

Dengue viruses (DVs) are responsible for the most medically relevant arboviral diseases. However, the molecular interactions mediating DV entry are poorly understood. We determined that TIM and TAM proteins, two receptor families that mediate the phosphatidylserine (PtdSer)-dependent phagocytic removal of apoptotic cells, serve as DV entry factors. Cells poorly susceptible to DV are robustly infected after ectopic expression of TIM or TAM receptors. Conversely, DV infection of susceptible cells is inhibited by anti-TIM or anti-TAM antibodies or knockdown of TIM and TAM expression. TIM receptors facilitate DV entry by directly interacting with virion-associated PtdSer. TAM-mediated infection relies on indirect DV recognition, in which the TAM ligand Gas6 acts as a bridging molecule by binding to PtdSer within the virion. This dual mode of virus recognition by TIM and TAM receptors reveals how DVs usurp the apoptotic cell clearance pathway for infectious entry. Copyright © 2012 Elsevier Inc. All rights reserved.

Effects of the GABA(B) receptor agonist baclofen administered orally on normal food intake and intraperitoneally on fat intake in non-deprived rats.

PubMed

Bains, Rasneer S; Ebenezer, Ivor S

2013-01-05

It has been previously reported that the GABA(B) receptor agonist baclofen decreases food intake after oral administration and fat intake after intraperitoneal administration. The aim of the study was to investigate the effects of baclofen (1-4 mg/ kg) administered orally (Experiment 1) on food intake in non-deprived rats (n=6) and intraperitoneally (Experiment 2) on fat intake in non-deprived rats (n=8) that were naïve to baclofen (1st set of trials) and in the same group of rats after they were sub-chronically exposed to baclofen (2nd set of trials). The results from Experiment 1 show that baclofen had no effects on food intake during the 1st set of trials, but the 2 and 4 mg/kg doses significantly increased food consumption during the 2nd set of trials. Baclofen produced sedation during the 1st set of trials, but tolerance occurred to this effect and was not apparent during the 2nd set of trials. These observations suggest that the motor effects may have competed with the hyperphagic effects of baclofen during the 1st set of trials. The data from Experiment 2 show that baclofen had no effects on fat intake during either the 1st or 2nd set of trials. The results of the study thus indicate that orally administrated baclofen increases food intake and intraperitoneal administration has no effect on fat intake in non-deprived rats under the conditions used in this study. These findings may have important implications for research on the use of baclofen in studies concerned with ingestive behaviours. Copyright © 2012 Elsevier B.V. All rights reserved.

GABA(C) receptors: a molecular view.

PubMed

Enz, R

2001-08-01

In the central nervous system inhibitory neurotransmission is primarily achieved through activation of receptors for gamma-aminobutyric acid (GABA). Three types of GABA receptors have been identified on the basis of their pharmacological and electrophysiological properties. The predominant type, termed GABA(A), and a recently identified GABA(C) type, form ligand-gated chloride channels, whereas GABA(B) receptors activate separate cation channels via G proteins. Based on their homology to nicotinic acetylcholine receptors, GABA(C) receptors are believed to be oligomeric protein complexes composed of five subunits in a pentameric arrangement. To date up to five different GABA(C) receptors subunits have been identified in various species. Recent studies have shed new light on the biological characteristics of GABA(C) receptors, including the chromosomal localization of its subunit genes and resulting links to deseases, the cloning of new splice variants, the identification of GABA(C) receptor-associated proteins, the identification of domains involved in subunit assembly, and finally structure/function studies examining functional consequences of introduced mutations. This review summarizes recent data in view of the molecular structure of GABA(C) receptors and presents new insights into the biological function of this protein in the retina.

Ethanol and Mesolimbic Serotonin/Dopamine Interactions Via 5-HT1B Receptors

DTIC Science & Technology

2006-03-01

baclofen , a GABAB receptor agonist, into the VTA probe and the response of extracellular DA in the ipsilateral NACC was determined. A significant...decrease (50% deduction) in extracellular DA in the ipsilateral NACC after perfusion with baclofen was considered an appropriate implantation of the...the VTA with baclofen were included in data analyses. Approximately 70% of the animals that had undergone surgery had both probes correctly implanted

Receptors, mediators, and mechanisms involved in bacterial sepsis and septic shock.

PubMed

Van Amersfoort, Edwin S; Van Berkel, Theo J C; Kuiper, Johan

2003-07-01

Bacterial sepsis and septic shock result from the overproduction of inflammatory mediators as a consequence of the interaction of the immune system with bacteria and bacterial wall constituents in the body. Bacterial cell wall constituents such as lipopolysaccharide, peptidoglycans, and lipoteichoic acid are particularly responsible for the deleterious effects of bacteria. These constituents interact in the body with a large number of proteins and receptors, and this interaction determines the eventual inflammatory effect of the compounds. Within the circulation bacterial constituents interact with proteins such as plasma lipoproteins and lipopolysaccharide binding protein. The interaction of the bacterial constituents with receptors on the surface of mononuclear cells is mainly responsible for the induction of proinflammatory mediators by the bacterial constituents. The role of individual receptors such as the toll-like receptors and CD14 in the induction of proinflammatory cytokines and adhesion molecules is discussed in detail. In addition, the roles of a number of other receptors that bind bacterial compounds such as scavenger receptors and their modulating role in inflammation are described. Finally, the therapies for the treatment of bacterial sepsis and septic shock are discussed in relation to the action of the aforementioned receptors and proteins.

Pharmacological significance of the interplay between angiotensin receptors: MAS receptors as putative final mediators of the effects elicited by angiotensin AT1 receptors antagonists.

PubMed

Pernomian, Larissa; Pernomian, Laena; Gomes, Mayara S; da Silva, Carlos H T P

2015-12-15

The interplay between angiotensin AT1 receptors and MAS receptors relies on several inward regulatory mechanisms from renin-angiotensin system (RAS) including the functional crosstalk between angiotensin II and angiotensin-(1-7), the competitive AT1 antagonism exhibited by angiotensin-(1-7), the antagonist feature assigned to AT1/MAS heterodimerization on AT1 signaling and the AT1-mediated downregulation of angiotensin-converting enzyme 2 (ACE2). Recently, such interplay has acquired an important significance to RAS Pharmacology since a few studies have supporting strong evidences that MAS receptors mediate the effects elicited by AT1 antagonists. The present Perspective provides an overview of the regulatory mechanisms involving AT1 and MAS receptors, their significance to RAS Pharmacology and the future directions on the interplay between angiotensin receptors. Copyright © 2015 Elsevier B.V. All rights reserved.

Azadirachtin Interacts with Retinoic Acid Receptors and Inhibits Retinoic Acid-mediated Biological Responses*

PubMed Central

Thoh, Maikho; Babajan, Banaganapalli; Raghavendra, Pongali B.; Sureshkumar, Chitta; Manna, Sunil K.

2011-01-01

Considering the role of retinoids in regulation of more than 500 genes involved in cell cycle and growth arrest, a detailed understanding of the mechanism and its regulation is useful for therapy. The extract of the medicinal plant Neem (Azadirachta indica) is used against several ailments especially for anti-inflammatory, anti-itching, spermicidal, anticancer, and insecticidal activities. In this report we prove the detailed mechanism on the regulation of retinoic acid-mediated cell signaling by azadirachtin, active components of neem extract. Azadirachtin repressed all trans-retinoic acid (ATRA)-mediated nuclear transcription factor κB (NF-κB) activation, not the DNA binding but the NF-κB-dependent gene expression. It did not inhibit IκBα degradation, IκBα kinase activity, or p65 phosphorylation and its nuclear translocation but inhibited NF-κB-dependent reporter gene expression. Azadirachtin inhibited TRAF6-mediated, but not TRAF2-mediated NF-κB activation. It inhibited ATRA-induced Sp1 and CREB (cAMP-response element-binding protein) DNA binding. Azadirachtin inhibited ATRA binding with retinoid receptors, which is supported by biochemical and in silico evidences. Azadirachtin showed strong interaction with retinoid receptors. It suppressed ATRA-mediated removal of retinoid receptors, bound with DNA by inhibiting ATRA binding to its receptors. Overall, our data suggest that azadirachtin interacts with retinoic acid receptors and suppresses ATRA binding, inhibits falling off the receptors, and activates transcription factors like CREB, Sp1, NF-κB, etc. Thus, azadirachtin exerts anti-inflammatory and anti-metastatic responses by a novel pathway that would be beneficial for further anti-inflammatory and anti-cancer therapies. PMID:21127062

Azadirachtin interacts with retinoic acid receptors and inhibits retinoic acid-mediated biological responses.

PubMed

Thoh, Maikho; Babajan, Banaganapalli; Raghavendra, Pongali B; Sureshkumar, Chitta; Manna, Sunil K

2011-02-11

Considering the role of retinoids in regulation of more than 500 genes involved in cell cycle and growth arrest, a detailed understanding of the mechanism and its regulation is useful for therapy. The extract of the medicinal plant Neem (Azadirachta indica) is used against several ailments especially for anti-inflammatory, anti-itching, spermicidal, anticancer, and insecticidal activities. In this report we prove the detailed mechanism on the regulation of retinoic acid-mediated cell signaling by azadirachtin, active components of neem extract. Azadirachtin repressed all trans-retinoic acid (ATRA)-mediated nuclear transcription factor κB (NF-κB) activation, not the DNA binding but the NF-κB-dependent gene expression. It did not inhibit IκBα degradation, IκBα kinase activity, or p65 phosphorylation and its nuclear translocation but inhibited NF-κB-dependent reporter gene expression. Azadirachtin inhibited TRAF6-mediated, but not TRAF2-mediated NF-κB activation. It inhibited ATRA-induced Sp1 and CREB (cAMP-response element-binding protein) DNA binding. Azadirachtin inhibited ATRA binding with retinoid receptors, which is supported by biochemical and in silico evidences. Azadirachtin showed strong interaction with retinoid receptors. It suppressed ATRA-mediated removal of retinoid receptors, bound with DNA by inhibiting ATRA binding to its receptors. Overall, our data suggest that azadirachtin interacts with retinoic acid receptors and suppresses ATRA binding, inhibits falling off the receptors, and activates transcription factors like CREB, Sp1, NF-κB, etc. Thus, azadirachtin exerts anti-inflammatory and anti-metastatic responses by a novel pathway that would be beneficial for further anti-inflammatory and anti-cancer therapies.

Opioid receptors mediate direct predictive fear learning: evidence from one-trial blocking.

PubMed

Cole, Sindy; McNally, Gavan P

2007-04-01

Pavlovian fear learning depends on predictive error, so that fear learning occurs when the actual outcome of a conditioning trial exceeds the expected outcome. Previous research has shown that opioid receptors, including mu-opioid receptors in the ventrolateral quadrant of the midbrain periaqueductal gray (vlPAG), mediate such predictive fear learning. Four experiments reported here used a within-subject one-trial blocking design to study whether opioid receptors mediate a direct or indirect action of predictive error on Pavlovian association formation. In Stage I, rats were trained to fear conditioned stimulus (CS) A by pairing it with shock. In Stage II, CSA and CSB were co-presented once and co-terminated with shock. Two novel stimuli, CSC and CSD, were also co-presented once and co-terminated with shock in Stage II. The results showed one-trial blocking of fear learning (Experiment 1) as well as one-trial unblocking of fear learning when Stage II training employed a higher intensity footshock than was used in Stage I (Experiment 2). Systemic administrations of the opioid receptor antagonist naloxone (Experiment 3) or intra-vlPAG administrations of the selective mu-opioid receptor antagonist CTAP (Experiment 4) prior to Stage II training prevented one-trial blocking. These results show that opioid receptors mediate the direct actions of predictive error on Pavlovian association formation.

Nuclear receptor-mediated regulation of carboxylesterase expression and activity.

PubMed

Staudinger, Jeff L; Xu, Chenshu; Cui, Yue J; Klaassen, Curtis D

2010-03-01

Emerging evidence demonstrates that several nuclear receptor (NR) family members regulate drug-inducible expression and activity of several important carboxylesterase (CES) enzymes in mammalian liver and intestine. Numerous clinically prescribed anticancer prodrugs, carbamate and pyrethroid insecticides, environmental toxicants and procarcinogens are substrates for CES enzymes. Moreover, a key strategy used in rational drug design frequently utilizes an ester linkage methodology to selectively target a prodrug, or to improve the water solubility of a novel compound. This review summarizes the current state of knowledge regarding NR-mediated regulation of CES enzymes in mammals and highlights their importance in drug metabolism, drug-drug interactions and toxicology. New knowledge regarding the transcriptional regulation of CES enzymes by NR proteins pregnane x receptor (NR1I2) and constitutive androstane receptor (NR1I3) has recently come to light through the use of knockout and transgenic mouse models. Novel insights regarding the species-specific cross-regulation of glucocorticoid receptor (NR3C1) and PPAR-alpha (NR1C1) signaling and CES gene expression are discussed. Elucidation of the role of NR-mediated regulation of CES enzymes in liver and intestine will have a significant impact on rational drug design and the development of novel prodrugs, especially for patients on combination therapy.

Biological Functionalization of Drug Delivery Carriers to Bypass Size Restrictions of Receptor-Mediated Endocytosis Independently from Receptor Targeting

PubMed Central

Ansar, Maria; Serrano, Daniel; Papademetriou, Iason; Bhowmick, Tridib Kumar; Muro, Silvia

2014-01-01

Targeting of drug carriers to cell-surface receptors involved in endocytosis is commonly used for intracellular drug delivery. However, most endocytic receptors mediate uptake via clathrin or caveolar pathways associated with ≤200-nm vesicles, restricting carrier design. We recently showed that endocytosis mediated by intercellular adhesion molecule 1 (ICAM-1), which differs from clathrin- and caveolar-mediated pathways, allows uptake of nano- and micro-carriers in cell culture and in vivo due to recruitment of cellular sphingomyelinases to the plasmalemma. This leads to ceramide generation at carrier binding sites and formation of actin stress-fibers, enabling engulfment and uptake of a wide size-range of carriers. Here we adapted this paradigm to enhance uptake of drug carriers targeted to receptors associated with size-restricted pathways. We coated sphingomyelinase onto model (polystyrene) submicro- and micro-carriers targeted to clathrin-associated mannose-6-phosphate receptor. In endothelial cells, this provided ceramide enrichment at the cell surface and actin stress-fiber formation, modifying the uptake pathway and enhancing carrier endocytosis without affecting targeting, endosomal transport, cell-associated degradation, or cell viability. This improvement depended on the carrier size and enzyme dose, and similar results were observed for other receptors (transferrin receptor) and cell types (epithelial cells). This phenomenon also enhanced tissue accumulation of carriers after intravenous injection in mice. Hence, it is possible to maintain targeting toward a selected receptor while bypassing natural size-restrictions of its associated endocytic route by functionalization of drug carriers with biological elements mimicking the ICAM-1 pathway. This strategy holds considerable promise to enhance flexibility of design of targeted drug delivery systems. PMID:24237309

Biological functionalization of drug delivery carriers to bypass size restrictions of receptor-mediated endocytosis independently from receptor targeting.

PubMed

Ansar, Maria; Serrano, Daniel; Papademetriou, Iason; Bhowmick, Tridib Kumar; Muro, Silvia

2013-12-23

Targeting of drug carriers to cell-surface receptors involved in endocytosis is commonly used for intracellular drug delivery. However, most endocytic receptors mediate uptake via clathrin or caveolar pathways associated with ≤200-nm vesicles, restricting carrier design. We recently showed that endocytosis mediated by intercellular adhesion molecule 1 (ICAM-1), which differs from clathrin- and caveolae-mediated pathways, allows uptake of nano- and microcarriers in cell culture and in vivo due to recruitment of cellular sphingomyelinases to the plasmalemma. This leads to ceramide generation at carrier binding sites and formation of actin stress-fibers, enabling engulfment and uptake of a wide size-range of carriers. Here we adapted this paradigm to enhance uptake of drug carriers targeted to receptors associated with size-restricted pathways. We coated sphingomyelinase onto model (polystyrene) submicro- and microcarriers targeted to clathrin-associated mannose-6-phosphate receptor. In endothelial cells, this provided ceramide enrichment at the cell surface and actin stress-fiber formation, modifying the uptake pathway and enhancing carrier endocytosis without affecting targeting, endosomal transport, cell-associated degradation, or cell viability. This improvement depended on the carrier size and enzyme dose, and similar results were observed for other receptors (transferrin receptor) and cell types (epithelial cells). This phenomenon also enhanced tissue accumulation of carriers after intravenous injection in mice. Hence, it is possible to maintain targeting toward a selected receptor while bypassing natural size restrictions of its associated endocytic route by functionalization of drug carriers with biological elements mimicking the ICAM-1 pathway. This strategy holds considerable promise to enhance flexibility of design of targeted drug delivery systems.

Systematic study of association of four GABAergic genes: glutamic acid decarboxylase 1 gene, glutamic acid decarboxylase 2 gene, GABA(B) receptor 1 gene and GABA(A) receptor subunit beta2 gene, with schizophrenia using a universal DNA microarray.

PubMed

Zhao, Xu; Qin, Shengying; Shi, Yongyong; Zhang, Aiping; Zhang, Jing; Bian, Li; Wan, Chunling; Feng, Guoyin; Gu, Niufan; Zhang, Guangqi; He, Guang; He, Lin

2007-07-01

Several studies have suggested the dysfunction of the GABAergic system as a risk factor in the pathogenesis of schizophrenia. In the present study, case-control association analysis was conducted in four GABAergic genes: two glutamic acid decarboxylase genes (GAD1 and GAD2), a GABA(A) receptor subunit beta2 gene (GABRB2) and a GABA(B) receptor 1 gene (GABBR1). Using a universal DNA microarray procedure we genotyped a total of 20 SNPs on the above four genes in a study involving 292 patients and 286 controls of Chinese descent. Statistically significant differences were observed in the allelic frequencies of the rs187269C/T polymorphism in the GABRB2 gene (P=0.0450, chi(2)=12.40, OR=1.65) and the -292A/C polymorphism in the GAD1 gene (P=0.0450, chi(2)=14.64 OR=1.77). In addition, using an electrophoretic mobility shift assay (EMSA), we discovered differences in the U251 nuclear protein binding to oligonucleotides representing the -292 SNP on the GAD1 gene, which suggests that the -292C allele has reduced transcription factor binding efficiency compared with the 292A allele. Using the multifactor-dimensionality reduction method (MDR), we found that the interactions among the rs187269C/T polymorphism in the GABRB2 gene, the -243A/G polymorphism in the GAD2 gene and the 27379C/T and 661C/T polymorphisms in the GAD1 gene revealed a significant association with schizophrenia (P<0.001). These findings suggest that the GABRB2 and GAD1 genes alone and the combined effects of the polymorphisms in the four GABAergic system genes may confer susceptibility to the development of schizophrenia in the Chinese population.

Protein phosphatase 2A mediates resensitization of the neurokinin 1 receptor

PubMed Central

Murphy, Jane E.; Roosterman, Dirk; Cottrell, Graeme S.; Padilla, Benjamin E.; Feld, Micha; Brand, Eva; Cedron, Wendy J.; Steinhoff, Martin

2011-01-01

Activated G protein-coupled receptors (GPCRs) are phosphorylated and interact with β-arrestins, which mediate desensitization and endocytosis. Endothelin-converting enzyme-1 (ECE-1) degrades neuropeptides in endosomes and can promote recycling. Although endocytosis, dephosphorylation, and recycling are accepted mechanisms of receptor resensitization, a large proportion of desensitized receptors can remain at the cell surface. We investigated whether reactivation of noninternalized, desensitized (phosphorylated) receptors mediates resensitization of the substance P (SP) neurokinin 1 receptor (NK1R). Herein, we report a novel mechanism of resensitization by which protein phosphatase 2A (PP2A) is recruited to dephosphorylate noninternalized NK1R. A desensitizing concentration of SP reduced cell-surface SP binding sites by only 25%, and SP-induced Ca2+ signals were fully resensitized before cell-surface binding sites started to recover, suggesting resensitization of cell-surface-retained NK1R. SP induced association of β-arrestin1 and PP2A with noninternalized NK1R. β-Arrestin1 small interfering RNA knockdown prevented SP-induced association of cell-surface NK1R with PP2A, indicating that β-arrestin1 mediates this interaction. ECE-1 inhibition, by trapping β-arrestin1 in endosomes, also impeded SP-induced association of cell-surface NK1R with PP2A. Resensitization of NK1R signaling required both PP2A and ECE-1 activity. Thus, after stimulation with SP, PP2A interacts with noninternalized NK1R and mediates resensitization. PP2A interaction with NK1R requires β-arrestin1. ECE-1 promotes this process by releasing β-arrestin1 from NK1R in endosomes. These findings represent a novel mechanism of PP2A- and ECE-1-dependent resensitization of GPCRs. PMID:21795521

Cross-talk between an activator of nuclear receptors-mediated transcription and the D1 dopamine receptor signaling pathway.

PubMed

Schmidt, Azriel; Vogel, Robert; Rutledge, Su Jane; Opas, Evan E; Rodan, Gideon A; Friedman, Eitan

2005-03-01

Nuclear receptors are transcription factors that usually interact, in a ligand-dependent manner, with specific DNA sequences located within promoters of target genes. The nuclear receptors can also be controlled in a ligand-independent manner via the action of membrane receptors and cellular signaling pathways. 5-Tetradecyloxy-2-furancarboxylic acid (TOFA) was shown to stimulate transcription from the MMTV promoter via chimeric receptors that consist of the DNA binding domain of GR and the ligand binding regions of the PPARbeta or LXRbeta nuclear receptors (GR/PPARbeta and GR/LXRbeta). TOFA and hydroxycholesterols also modulate transcription from NF-kappaB- and AP-1-controlled reporter genes and induce neurite differentiation in PC12 cells. In CV-1 cells that express D(1) dopamine receptors, D(1) dopamine receptor stimulation was found to inhibit TOFA-stimulated transcription from the MMTV promoter that is under the control of chimeric GR/PPARbeta and GR/LXRbeta receptors. Treatment with the D(1) dopamine receptor antagonist, SCH23390, prevented dopamine-mediated suppression of transcription, and by itself increased transcription controlled by GR/LXRbeta. Furthermore, combined treatment of CV-1 cells with TOFA and SCH23390 increased transcription controlled by the GR/LXRbeta chimeric receptor synergistically. The significance of this in vitro synergy was demonstrated in vivo, by the observation that SCH23390 (but not haloperidol)-mediated catalepsy in rats was potentiated by TOFA, thus showing that an agent that mimics the in vitro activities of compounds that activate members of the LXR and PPAR receptor families can influence D1 dopamine receptor elicited responses.

G protein-coupled estrogen receptor inhibits the P2Y receptor-mediated Ca(2+) signaling pathway in human airway epithelia.

PubMed

Hao, Yuan; Chow, Alison W; Yip, Wallace C; Li, Chi H; Wan, Tai F; Tong, Benjamin C; Cheung, King H; Chan, Wood Y; Chen, Yangchao; Cheng, Christopher H; Ko, Wing H

2016-08-01

P2Y receptor activation causes the release of inflammatory cytokines in the bronchial epithelium, whereas G protein-coupled estrogen receptor (GPER), a novel estrogen (E2) receptor, may play an anti-inflammatory role in this process. We investigated the cellular mechanisms underlying the inhibitory effect of GPER activation on the P2Y receptor-mediated Ca(2+) signaling pathway and cytokine production in airway epithelia. Expression of GPER in primary human bronchial epithelial (HBE) or 16HBE14o- cells was confirmed on both the mRNA and protein levels. Stimulation of HBE or 16HBE14o- cells with E2 or G1, a specific agonist of GPER, attenuated the nucleotide-evoked increases in [Ca(2+)]i, whereas this effect was reversed by G15, a GPER-specific antagonist. G1 inhibited the secretion of two proinflammatory cytokines, interleukin (IL)-6 and IL-8, in cells stimulated by adenosine 5'-(γ-thio)triphosphate (ATPγS). G1 stimulated a real-time increase in cAMP levels in 16HBE14o- cells, which could be inhibited by adenylyl cyclase inhibitors. The inhibitory effects of E2 or G1 on P2Y receptor-induced increases in Ca(2+) were reversed by treating the cells with a protein kinase A (PKA) inhibitor. These results demonstrated that the inhibitory effects of G1 or E2 on P2Y receptor-mediated Ca(2+) mobilization and cytokine secretion were due to GPER-mediated activation of a cAMP-dependent PKA pathway. This study has reported, for the first time, the expression and function of GPER as an anti-inflammatory component in human bronchial epithelia, which may mediate through its opposing effects on the pro-inflammatory pathway activated by the P2Y receptors in inflamed airway epithelia.

Arsenic as an endocrine disruptor: arsenic disrupts retinoic acid receptor-and thyroid hormone receptor-mediated gene regulation and thyroid hormone-mediated amphibian tail metamorphosis.

PubMed

Davey, Jennifer C; Nomikos, Athena P; Wungjiranirun, Manida; Sherman, Jenna R; Ingram, Liam; Batki, Cavus; Lariviere, Jean P; Hamilton, Joshua W

2008-02-01

Chronic exposure to excess arsenic in drinking water has been strongly associated with increased risks of multiple cancers, diabetes, heart disease, and reproductive and developmental problems in humans. We previously demonstrated that As, a potent endocrine disruptor at low, environmentally relevant levels, alters steroid signaling at the level of receptor-mediated gene regulation for all five steroid receptors. The goal of this study was to determine whether As can also disrupt gene regulation via the retinoic acid (RA) receptor (RAR) and/or the thyroid hormone (TH) receptor (TR) and whether these effects are similar to previously observed effects on steroid regulation. Human embryonic NT2 or rat pituitary GH3 cells were treated with 0.01-5 microM sodium arsenite for 24 hr, with or without RA or TH, respectively, to examine effects of As on receptor-mediated gene transcription. At low, noncytotoxic doses, As significantly altered RAR-dependent gene transcription of a transfected RAR response element-luciferase construct and the native RA-inducible cytochrome P450 CYP26A gene in NT2 cells. Likewise, low-dose As significantly altered expression of a transfected TR response element-luciferase construct and the endogenous TR-regulated type I deiodinase (DIO1) gene in a similar manner in GH3 cells. An amphibian ex vivo tail metamorphosis assay was used to examine whether endocrine disruption by low-dose As could have specific pathophysiologic consequences, because tail metamorphosis is tightly controlled by TH through TR. TH-dependent tail shrinkage was inhibited in a dose-dependent manner by 0.1- 4.0 microM As. As had similar effects on RAR- and TR-mediated gene regulation as those previously observed for the steroid receptors, suggesting a common mechanism or action. Arsenic also profoundly affected a TR-dependent developmental process in a model animal system at very low concentrations. Because RAR and TH are critical for both normal human development and adult

Stimulation of accumbal GABAA receptors inhibits delta2-, but not delta1-, opioid receptor-mediated dopamine efflux in the nucleus accumbens of freely moving rats.

PubMed

Aono, Yuri; Kiguchi, Yuri; Watanabe, Yuriko; Waddington, John L; Saigusa, Tadashi

2017-11-15

The nucleus accumbens contains delta-opioid receptors that may reduce inhibitory neurotransmission. Reduction in GABA A receptor-mediated inhibition of accumbal dopamine release due to delta-opioid receptor activation should be suppressed by stimulating accumbal GABA A receptors. As delta-opioid receptors are divided into delta2- and delta1-opioid receptors, we analysed the effects of the GABA A receptor agonist muscimol on delta2- and delta1-opioid receptor-mediated accumbal dopamine efflux in freely moving rats using in vivo microdialysis. Drugs were administered intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 25-50min infusions. The delta2-opioid receptor agonist deltorphin II (25.0nmol)- and delta1-opioid receptor agonist DPDPE (5.0nmol)-induced increases in dopamine efflux were inhibited by the delta2-opioid receptor antagonist naltriben (1.5nmol) and the delta1-opioid receptor antagonist BNTX (150.0pmol), respectively. Muscimol (250.0pmol) inhibited deltorphin II (25.0nmol)-induced dopamine efflux. The GABA A receptor antagonist bicuculline (50.0pmol), which failed to affect deltorphin II (25.0nmol)-induced dopamine efflux, counteracted the inhibitory effect of muscimol on deltorphin II-induced dopamine efflux. Neither muscimol (250.0pmol) nor bicuculline (50.0 and 500.0pmol) altered DPDPE (5.0nmol)-induced dopamine efflux. The present results show that reduction in accumbal GABA A receptor-mediated inhibition of dopaminergic activity is necessary to produce delta2-opioid receptor-induced increase in accumbal dopamine efflux. This study indicates that activation of delta2- but not delta1-opioid receptors on the cell bodies and/or terminals of accumbal GABAergic interneurons inhibits GABA release and, accordingly, decreases GABA A receptor-mediated inhibition of dopaminergic terminals, resulting in enhanced accumbal dopamine efflux. Copyright © 2017 Elsevier B.V. All rights reserved.

ZINC-INDUCED EGF RECEPTOR SIGNALING REQUIRES SRC-MEDIATED PHOSPHORYLATION OF THE EGF RECEPTOR ON TYROSINE 845 (Y845)

EPA Science Inventory

ZINC-INDUCED EGF RECEPTOR SIGNALING REQUIRES Src-MEDIATED PHOSPHORYLATION OF THE EGF RECEPTOR ON TYROSINE 845 (Y845)
Weidong Wu1, Lee M. Graves2, Gordon N. Gill3 and James M. Samet4 1Center for Environmental Medicine and Lung Biology; 2Department of Pharmacology, University o...

In vitro binding and receptor-mediated activity of terlipressin at vasopressin receptors V1 and V2

PubMed Central

Jamil, Khurram; Pappas, Stephen Chris; Devarakonda, Krishna R

2018-01-01

Terlipressin, a synthetic, systemic vasoconstrictor with selective activity at vasopressin-1 (V1) receptors, is a pro-drug for the endogenous/natural porcine hormone [Lys8]-vasopressin (LVP). We investigated binding and receptor-mediated cellular activities of terlipressin, LVP, and endogenous human hormone [Arg8]-vasopressin (AVP) at V1 and vasopressin-2 (V2) receptors. Cell membrane homogenates of Chinese hamster ovary cells expressing human V1 and V2 receptors were used in competitive binding assays to measure receptor-binding activity. These cells were used in functional assays to measure receptor-mediated cellular activity of terlipressin, LVP, and AVP. Binding was measured by [3H]AVP counts, and the activity was measured by fluorometric detection of intracellular calcium mobilization (V1) and cyclic adenosine monophosphate (V2). Binding potency at V1 and V2 was AVP>LVP>>terlipressin. LVP and terlipressin had approximately sixfold higher affinity for V1 than for V2. Cellular activity potency was also AVP>LVP>>terlipressin. Terlipressin was a partial agonist at V1 and a full agonist at V2; LVP was a full agonist at both V1 and V2. The in vivo response to terlipressin is likely due to the partial V1 agonist activity of terlipressin and full V1 agonist activity of its metabolite, LVP. These results provide supportive evidence for previous findings and further establish terlipressin pharmacology for vasopressin receptors. PMID:29302194

In vitro binding and receptor-mediated activity of terlipressin at vasopressin receptors V1 and V2.

PubMed

Jamil, Khurram; Pappas, Stephen Chris; Devarakonda, Krishna R

2018-01-01

Terlipressin, a synthetic, systemic vasoconstrictor with selective activity at vasopressin-1 (V 1 ) receptors, is a pro-drug for the endogenous/natural porcine hormone [Lys 8 ]-vasopressin (LVP). We investigated binding and receptor-mediated cellular activities of terlipressin, LVP, and endogenous human hormone [Arg 8 ]-vasopressin (AVP) at V 1 and vasopressin-2 (V 2 ) receptors. Cell membrane homogenates of Chinese hamster ovary cells expressing human V 1 and V 2 receptors were used in competitive binding assays to measure receptor-binding activity. These cells were used in functional assays to measure receptor-mediated cellular activity of terlipressin, LVP, and AVP. Binding was measured by [ 3 H]AVP counts, and the activity was measured by fluorometric detection of intracellular calcium mobilization (V 1 ) and cyclic adenosine monophosphate (V 2 ). Binding potency at V 1 and V 2 was AVP>LVP>terlipressin. LVP and terlipressin had approximately sixfold higher affinity for V 1 than for V 2 . Cellular activity potency was also AVP>LVP>terlipressin. Terlipressin was a partial agonist at V 1 and a full agonist at V 2 ; LVP was a full agonist at both V 1 and V 2 . The in vivo response to terlipressin is likely due to the partial V 1 agonist activity of terlipressin and full V 1 agonist activity of its metabolite, LVP. These results provide supportive evidence for previous findings and further establish terlipressin pharmacology for vasopressin receptors.

SLP-76 is required for high-affinity IgE receptor- and IL-3 receptor-mediated activation of basophils.

PubMed

Hidano, Shinya; Kitamura, Daisuke; Kumar, Lalit; Geha, Raif S; Goitsuka, Ryo

2012-11-01

Basophils have been reported to play a critical role in allergic inflammation by secreting IL-4 in response to IL-3 or high-affinity IgE receptor (FcεRI)-cross-linking. However, the signaling pathways downstream of FcεRI and the IL-3 receptor in basophils have yet to be determined. In the present study, we used mice deficient in SLP-76 (Src homology 2 domain-containing leukocyte phosphoprotein of 76kDa) to demonstrate critical functions of this adaptor molecule in transducing FcεRI- and IL-3 receptor-mediated signals that induce basophil activation. Although SLP-76 was dispensable for in vivo differentiation, as well as IL-3-induced in vitro proliferation of basophils, IL-4 production induced by both stimuli was completely ablated by SLP-76 deficiency. Biochemical analyses revealed that IL-3-induced phosphorylation of phospholipase C (PLC) γ2 and Akt, but not STAT5, was severely reduced in SLP-76-deficient basophils, whereas FcεRI cross-linking phosphorylation of PLCγ2, but not Akt, was abrogated by SLP-76 deficiency, suggesting important differences in the requirement of SLP-76 for Akt activation between FcεRI- and IL-3 receptor-mediated signaling pathways in basophils. Because IL-3-induced IL-4 production was sensitive to calcineurin inhibitors and an intracellular calcium chelator, in addition to PI3K inhibitors, SLP-76 appears to regulate FcεRI- and IL-3 receptor-induced IL-4 production via mediating PLCγ2 activation in basophils. Taken together, these findings indicate that SLP-76 is an essential signaling component for basophil activation downstream of both FcεRI and the IL-3 receptor.

Effects of GABA-B receptor positive modulator on ketamine-induced psychosis-relevant behaviors and hippocampal electrical activity in freely moving rats.

PubMed

Ma, Jingyi; Stan Leung, L

2017-10-01

Decreased GABA B receptor function is proposed to mediate some symptoms of schizophrenia. In this study, we tested the effect of CGP7930, a GABA B receptor positive allosteric modulator, on ketamine-induced psychosis-relevant behaviors and hippocampal electrical activity in behaving rats. Electrodes were bilaterally implanted into the hippocampus, and cannulae were placed into the lateral ventricles of Long-Evans rats. CGP7930 or vehicle was injected intraperitoneally (i.p.) or intracerebroventricularly (i.c.v.), alone or 15 min prior to ketamine (3 mg/kg, subcutaneous) injection. Paired click auditory evoked potentials in the hippocampus (AEP), prepulse inhibition (PPI), and locomotor activity were recorded before and after drug injection. CGP7930 at doses of 1 mg/kg (i.p.) prevented ketamine-induced deficit of PPI. CGP7930 (1 mg/kg i.p.) also prevented the decrease in gating of hippocampal AEP and the increase in hippocampal gamma (65-100 Hz) waves induced by ketamine. Unilateral i.c.v. infusion of CGP7930 (0.3 mM/1 μL) also prevented the decrease in gating of hippocampal AEP induced by ketamine. Ketamine-induced behavioral hyperlocomotion was suppressed by 5 mg/kg i.p. CGP7930. CGP7930 alone, without ketamine, did not significantly affect integrated PPI, locomotion, gating of hippocampal AEP, or gamma waves. CGP7930 (1 mg/kg i.p.) increased heterosynaptically mediated paired pulse depression in the hippocampus, a measure of GABA B receptor function in vivo. CGP7930 reduces the behavioral and electrophysiological disruptions induced by ketamine in animals, and the hippocampus may be one of the neural targets where CGP7930 exerts its actions.

Glucose Deprivation Induces ATF4-Mediated Apoptosis through TRAIL Death Receptors

PubMed Central

Iurlaro, Raffaella; Püschel, Franziska; León-Annicchiarico, Clara Lucía; O'Connor, Hazel; Martin, Seamus J.; Palou-Gramón, Daniel; Lucendo, Estefanía

2017-01-01

ABSTRACT Metabolic stress occurs frequently in tumors and in normal tissues undergoing transient ischemia. Nutrient deprivation triggers, among many potential cell death-inducing pathways, an endoplasmic reticulum (ER) stress response with the induction of the integrated stress response transcription factor ATF4. However, how this results in cell death remains unknown. Here we show that glucose deprivation triggered ER stress and induced the unfolded protein response transcription factors ATF4 and CHOP. This was associated with the nontranscriptional accumulation of TRAIL receptor 1 (TRAIL-R1) (DR4) and with the ATF4-mediated, CHOP-independent induction of TRAIL-R2 (DR5), suggesting that cell death in this context may involve death receptor signaling. Consistent with this, the ablation of TRAIL-R1, TRAIL-R2, FADD, Bid, and caspase-8 attenuated cell death, although the downregulation of TRAIL did not, suggesting ligand-independent activation of TRAIL receptors. These data indicate that stress triggered by glucose deprivation promotes the ATF4-dependent upregulation of TRAIL-R2/DR5 and TRAIL receptor-mediated cell death. PMID:28242652

Potentiation of NMDA receptor-mediated transmission in striatal cholinergic interneurons

PubMed Central

Oswald, Manfred J.; Schulz, Jan M.; Kelsch, Wolfgang; Oorschot, Dorothy E.; Reynolds, John N. J.

2015-01-01

Pauses in the tonic firing of striatal cholinergic interneurons (CINs) emerge during reward-related learning in response to conditioning of a neutral cue. We have previously reported that augmenting the postsynaptic response to cortical afferents in CINs is coupled to the emergence of a cell-intrinsic afterhyperpolarization (AHP) underlying pauses in tonic activity. Here we investigated in a bihemispheric rat-brain slice preparation the mechanisms of synaptic plasticity of excitatory afferents to CINs and the association with changes in the AHP. We found that high frequency stimulation (HFS) of commissural corticostriatal afferents from the contralateral hemisphere induced a robust long-term depression (LTD) of postsynaptic potentials (PSP) in CINs. Depression of the PSP of smaller magnitude and duration was observed in response to HFS of the ipsilateral white matter or cerebral cortex. In Mg2+-free solution HFS induced NMDA receptor-dependent potentiation of the PSP, evident in both the maximal slope and amplitude of the PSP. The increase in maximal slope corroborates previous findings, and was blocked by antagonism of either D1-like dopamine receptors with SCH23390 or D2-like dopamine receptors with sulpiride during HFS in Mg2+-free solution. Potentiation of the slower PSP amplitude component was due to augmentation of the NMDA receptor-mediated potential as this was completely reversed on subsequent application of the NMDA receptor antagonist AP5. HFS similarly potentiated NMDA receptor currents isolated by blockade of AMPA/kainate receptors with CNQX. The plasticity-induced increase in the slow PSP component was directly associated with an increase in the subsequent AHP. Thus plasticity of cortical afferent synapses is ideally suited to influence the cue-induced firing dynamics of CINs, particularly through potentiation of NMDA receptor-mediated synaptic transmission. PMID:25914618

Potentiation of NMDA receptor-mediated transmission in striatal cholinergic interneurons.

PubMed

Oswald, Manfred J; Schulz, Jan M; Kelsch, Wolfgang; Oorschot, Dorothy E; Reynolds, John N J

2015-01-01

Pauses in the tonic firing of striatal cholinergic interneurons (CINs) emerge during reward-related learning in response to conditioning of a neutral cue. We have previously reported that augmenting the postsynaptic response to cortical afferents in CINs is coupled to the emergence of a cell-intrinsic afterhyperpolarization (AHP) underlying pauses in tonic activity. Here we investigated in a bihemispheric rat-brain slice preparation the mechanisms of synaptic plasticity of excitatory afferents to CINs and the association with changes in the AHP. We found that high frequency stimulation (HFS) of commissural corticostriatal afferents from the contralateral hemisphere induced a robust long-term depression (LTD) of postsynaptic potentials (PSP) in CINs. Depression of the PSP of smaller magnitude and duration was observed in response to HFS of the ipsilateral white matter or cerebral cortex. In Mg(2+)-free solution HFS induced NMDA receptor-dependent potentiation of the PSP, evident in both the maximal slope and amplitude of the PSP. The increase in maximal slope corroborates previous findings, and was blocked by antagonism of either D1-like dopamine receptors with SCH23390 or D2-like dopamine receptors with sulpiride during HFS in Mg(2+)-free solution. Potentiation of the slower PSP amplitude component was due to augmentation of the NMDA receptor-mediated potential as this was completely reversed on subsequent application of the NMDA receptor antagonist AP5. HFS similarly potentiated NMDA receptor currents isolated by blockade of AMPA/kainate receptors with CNQX. The plasticity-induced increase in the slow PSP component was directly associated with an increase in the subsequent AHP. Thus plasticity of cortical afferent synapses is ideally suited to influence the cue-induced firing dynamics of CINs, particularly through potentiation of NMDA receptor-mediated synaptic transmission.

The influences of metabotropic receptor activation on cellular signaling and synaptic function in amacrine cells.

PubMed

Gleason, Evanna

2012-01-01

Amacrine cells receive glutamatergic input from bipolar cells and GABAergic, glycinergic, cholinergic, and dopaminergic input from other amacrine cells. Glutamate, GABA, glycine, and acetylcholine (ACh) interact with ionotropic receptors and it is these interactions that form much of the functional circuitry in the inner retina. However, glutamate, GABA, ACh, and dopamine also activate metabotropic receptors linked to second messenger pathways that have the potential to modify the function of individual cells as well as retinal circuitry. Here, the physiological effects of activating dopamine receptors, metabotropic glutamate receptors, GABAB receptors, and muscarinic ACh receptors on amacrine cells will be discussed. The retina also expresses metabotropic receptors and the biochemical machinery associated with the synthesis and degradation of endocannabinoids and sphingosine-1-phosphate (S1P). The effects of activating cannabinoid receptors and S1P receptors on amacrine cell function will also be addressed. Copyright © Cambridge University Press, 2012

A slow excitatory postsynaptic current mediated by a novel metabotropic glutamate receptor in CA1 pyramidal neurons.

PubMed

Sheng, Nengyin; Yang, Jing; Silm, Katlin; Edwards, Robert H; Nicoll, Roger A

2017-03-15

Slow excitatory postsynaptic currents (EPSCs) mediated by metabotropic glutamate receptors (mGlu receptors) have been reported in several neuronal subtypes, but their presence in hippocampal pyramidal neurons remains elusive. Here we find that in CA1 pyramidal neurons a slow EPSC is induced by repetitive stimulation while ionotropic glutamate receptors and glutamate-uptake are blocked whereas it is absent in the VGLUT1 knockout mouse in which presynaptic glutamate is lost, suggesting the slow EPSC is mediated by glutamate activating mGlu receptors. However, it is not inhibited by known mGlu receptor antagonists. These findings suggest that this slow EPSC is mediated by a novel mGlu receptor, and that it may be involved in neurological diseases associated with abnormal high-concentration of extracellular glutamate. This article is part of the Special Issue entitled 'Metabotropic Glutamate Receptors, 5 years on'. Copyright © 2016 Elsevier Ltd. All rights reserved.

Ethanol and Mesolimbic Serotonin/Dopamine Interactions via 5-HT-1B Receptors

DTIC Science & Technology

2005-03-01

finished with After completion of the dialysis, the animals were given infusion of 50 [tM of baclofen , a GABAB receptor agonist, an intracardiac...in the quickly, and 40-[tm-thick coronal sections were cut on a ipsilateral NACC after perfusion with baclofen was consid- freezing microtome, stained...triethylamine, 11.5% and appropriate accumbal DA responses to perfusion of the acetonitrile and 11.5% methanol (pH 5.6 with H3 PO4 ), VTA with baclofen were

Modulation of NMDA and AMPA-Mediated Synaptic Transmission by CB1 Receptors in Frontal Cortical Pyramidal Cells

PubMed Central

Li, Qiang; Yan, Haidun; Wilson, Wilkie A.; Swartzwelder, H. Scott

2010-01-01

Although the endogenous cannabinoid system modulates a variety of physiological and pharmacological processes, the specific role of cannabinoid CB1 receptors in the modulation of glutamatergic neurotransmission and neural plasticity is not well understood. Using whole-cell patch clamp recording techniques, evoked or spontaneous excitatory postsynaptic currents (eEPSCs or sEPSCs) were recorded from visualized, layer II/III pyramidal cells in frontal cortical slices from rat brain. Bath application of the CB1 receptor agonist, WIN 55212-2 (WIN), reduced the amplitude of NMDA receptor-mediated EPSCs in a concentration-dependent manner. When co-applied with the specific CB1 antagonists, AM251 or AM281, WIN did not suppress NMDA receptor mediated EPSCs. WIN also reduced the amplitude of evoked AMPA receptor-mediated EPSCs, an effect that was also reversed by AM251. Both the frequency and amplitude of spontaneous AMPA receptor-mediated EPSCs were significantly reduced by WIN. In contrast, WIN reduced the frequency, but not the amplitude of miniature EPSCs, suggesting that the suppression of glutmatergic activity by CB1 receptors in the frontal neocortex is mediated by a pre-synaptic mechanism. Taken together, these data indicate a critical role for endocannabinoid signaling in the regulation of excitatory synaptic transmission in frontal neocortex, and suggest a possible neuronal mechanism whereby THC regulates cortical function. PMID:20420813

GABA-A Receptors Mediate Tonic Inhibition and Neurosteroid Sensitivity in the Brain.

PubMed

Reddy, Doodipala Samba

2018-01-01

Neurosteroids like allopregnanolone (AP) are positive allosteric modulators of synaptic and extrasynaptic GABA-A receptors. AP and related neurosteroids exhibit a greater potency for δ-containing extrasynaptic receptors. The δGABA-A receptors, which are expressed extrasynaptically in the dentate gyrus and other regions, contribute to tonic inhibition, promoting network shunting as well as reducing seizure susceptibility. Levels of endogenous neurosteroids fluctuate with ovarian cycle. Natural and synthetic neurosteroids maximally potentiate tonic inhibition in the hippocampus and provide robust protection against a variety of limbic seizures and status epilepticus. Recently, a consensus neurosteroid pharmacophore model has been proposed at extrasynaptic δGABA-A receptors based on structure-activity relationship for functional activation of tonic currents and seizure protection. Aside from anticonvulsant actions, neurosteroids have been found to be powerful anxiolytic and anesthetic agents. Neurosteroids and Zn 2+ have preferential affinity for δ-containing receptors. Thus, Zn 2+ can prevent neurosteroid activation of extrasynaptic δGABA-A receptor-mediated tonic inhibition. Recently, we demonstrated that Zn 2+ selectively inhibits extrasynaptic δGABA-A receptors and thereby fully prevents AP activation of tonic inhibition and seizure protection. We confirmed that neurosteroids exhibit greater sensitivity at extrasynaptic δGABA-A receptors. Overall, extrasynaptic GABA-A receptors are primary mediators of tonic inhibition in the brain and play a key role in the pathophysiology of epilepsy and other neurological disorders. © 2018 Elsevier Inc. All rights reserved.

Constitutive arrestin-mediated desensitization of a human vasopressin receptor mutant associated with nephrogenic diabetes insipidus.

PubMed

Barak, L S; Oakley, R H; Laporte, S A; Caron, M G

2001-01-02

Agonist-dependent desensitization and internalization of G protein-coupled receptors (GPCR) are mediated by the binding of arrestins to phosphorylated receptors. The affinity of arrestins for the phosphorylated GPCR regulates the ability of the internalized receptor to be dephosphorylated and recycled back to the plasma membrane. In this study, we show that the naturally occurring loss of function vasopressin receptor mutation R137H, which is associated with familial nephrogenic diabetes insipidus, induces constitutive arrestin-mediated desensitization. In contrast to the wild-type vasopressin receptor, the nonsignaling R137H receptor is phosphorylated and sequestered in arrestin-associated intracellular vesicles even in the absence of agonist. Eliminating molecular determinants on the receptor that promote high affinity arrestin-receptor interaction reestablishes plasma membrane localization and the ability of the mutated receptors to signal. These findings suggest that unregulated desensitization can contribute to the etiology of a GPCR-based disease, implying that pharmacological targeting of GPCR desensitization may be therapeutically beneficial.

Constitutive arrestin-mediated desensitization of a human vasopressin receptor mutant associated with nephrogenic diabetes insipidus

PubMed Central

Barak, Larry S.; Oakley, Robert H.; Laporte, Stéphane A.; Caron, Marc G.

2001-01-01

Agonist-dependent desensitization and internalization of G protein-coupled receptors (GPCR) are mediated by the binding of arrestins to phosphorylated receptors. The affinity of arrestins for the phosphorylated GPCR regulates the ability of the internalized receptor to be dephosphorylated and recycled back to the plasma membrane. In this study, we show that the naturally occurring loss of function vasopressin receptor mutation R137H, which is associated with familial nephrogenic diabetes insipidus, induces constitutive arrestin-mediated desensitization. In contrast to the wild-type vasopressin receptor, the nonsignaling R137H receptor is phosphorylated and sequestered in arrestin-associated intracellular vesicles even in the absence of agonist. Eliminating molecular determinants on the receptor that promote high affinity arrestin–receptor interaction reestablishes plasma membrane localization and the ability of the mutated receptors to signal. These findings suggest that unregulated desensitization can contribute to the etiology of a GPCR-based disease, implying that pharmacological targeting of GPCR desensitization may be therapeutically beneficial. PMID:11134505

Contribution of ventral tegmental GABA receptors to cocaine self-administration in rats.

PubMed

Backes, E N; Hemby, S E

2008-03-01

Recent evidence has suggested that compounds affecting GABAergic transmission may provide useful pharmacological tools for the treatment of cocaine addiction. Using a rat model of self-administration, the present study examined the effects of GABA agonists and antagonists injected directly into the ventral tegmental area (VTA) on cocaine intake in rats trained to self-administer cocaine (0, 125, 250 and 500 microg/infusion) under an FR5 schedule of reinforcement. Separate groups of rats received bilateral intra-VTA injections of the GABA-A antagonist picrotoxin (34 ng/side, n = 7; 68 ng/side, n = 8), GABA-A agonist muscimol (14 ng/side, n = 8), GABA-B agonist baclofen (56 ng/side, n = 7; 100 ng/side, n = 6), picrotoxin (68 ng/side) co-injected with the GABA-B antagonist 2-hydroxysaclofen (100 ng/side, n = 7; 2 microg/side, n = 8) or artificial cerebrospinal fluid (aCSF, n = 6) to assess the effects of the various compounds on the cocaine self-administration dose-response curve. Both picrotoxin and baclofen reduced responding maintained by cocaine, whereas muscimol had no effect on responding. In contrast, neither picrotoxin (n = 6) nor baclofen (n = 8) affected responding maintained by food. Interestingly, 2-hydroxysaclofen effectively blocked the suppression of responding produced by picrotoxin, suggesting that both picrotoxin and baclofen exert their effects via activation of GABA-B receptors. Additionally, these effects appear to be specific to cocaine reinforcement, supporting current investigation of baclofen as a treatment for cocaine addiction.

Contribution of ventral tegmental GABA receptors to cocaine self-administration in rats

PubMed Central

Backes, E.N.; Hemby, S.E.

2008-01-01

Recent evidence has suggested that compounds affecting GABAergic transmission may provide useful pharmacological tools for the treatment of cocaine addiction. Using a rat model of self-administration, the present study examined the effects of GABA agonists and antagonists injected directly into the ventral tegmental area (VTA) on cocaine intake in rats trained to self-administer cocaine (0, 125, 250 and 500 µg/infusion) under an FR5 schedule of reinforcement. Separate groups of rats received bilateral intra-VTA injections of the GABA-A antagonist picrotoxin (34 ng/side, n=7; 68 ng/side, n=8), GABA-A agonist muscimol (14 ng/side, n=8), GABA-B agonist baclofen (56 ng/side, n=7; 100 ng/side, n=6), picrotoxin (68 ng/side) co-injected with the GABA-B antagonist 2-hydroxysaclofen (100 ng/side, n=7; 2 µg/side, n=8) or artificial cerebrospinal fluid (aCSF, n=6) to assess the effects of the various compounds on the cocaine self-administration dose-response curve. Both picrotoxin and baclofen reduced responding maintained by cocaine, whereas muscimol had no effect on responding. In contrast, neither picrotoxin (n=6) nor baclofen (n=8) affected responding maintained by food. Interestingly, 2-hydroxysaclofen effectively blocked the suppression of responding produced by picrotoxin, suggesting that both picrotoxin and baclofen exert their effects via activation of GABA-B receptors. Additionally, these effects appear to be specific to cocaine reinforcement, supporting current investigation of baclofen as a treatment for cocaine addiction. PMID:17943439

Presynaptic Kainate Receptor Mediation of Frequency Facilitation at Hippocampal Mossy Fiber Synapses

NASA Astrophysics Data System (ADS)

Schmitz, Dietmar; Mellor, Jack; Nicoll, Roger A.

2001-03-01

Inhibition of transmitter release by presynaptic receptors is widespread in the central nervous system and is typically mediated via metabotropic receptors. In contrast, very little is known about facilitatory receptors, and synaptic activation of a facilitatory autoreceptor has not been established. Here we show that activation of presynaptic kainate receptors can facilitate transmitter release from hippocampal mossy fiber synapses. Synaptic activation of these presumed ionotropic kainate receptors is very fast (<10 ms) and lasts for seconds. Thus, these presynaptic kainate receptors contribute to the short-term plasticity characteristics of mossy fiber synapses, which were previously thought to be an intrinsic property of the synapse.

The Mas receptor mediates modulation of insulin signaling by angiotensin-(1-7).

PubMed

Muñoz, Marina C; Giani, Jorge F; Burghi, Valeria; Mayer, Marcos A; Carranza, Andrea; Taira, Carlos A; Dominici, Fernando P

2012-08-20

Angiotensin (Ang)-(1-7) stimulates proteins belonging to the insulin signaling pathway and ameliorates the Ang II negative effects at this level. However, up to date, receptors involved and mechanisms behind these observations remain unknown. Accordingly, in the present study, we explored the in vivo effects of antagonism of the Ang-(1-7) specific Mas receptor on insulin signal transduction in rat insulin-target tissues. We evaluated the acute modulation of insulin-stimulated phosphorylation of Akt, GSK-3β (Glycogen synthase kinase-3β) and AS160 (Akt substrate of 160kDa) by Ang-(1-7) and/or Ang II in the presence and absence of the selective Mas receptor antagonist A-779 in insulin-target tissues of normal rats. Also using A-779, we determined whether the Mas receptor mediates the improvement of insulin sensitivity exerted by chronic Ang-(1-7) treatment in fructose-fed rats (FFR), a model of insulin resistance, dyslipidemia and mild hypertension. The two major findings of the present work are as follows; 1) Ang-(1-7) attenuates acute Ang II-mediated inhibition of insulin signaling components in normal rats via a Mas receptor-dependent mechanism; and 2). The Mas receptor appears to be involved in beneficial effects of Ang-(1-7) on the phosphorylation of crucial insulin signaling mediators (Akt, GSK-3β and AS160), in liver, skeletal muscle and adipose tissue of FFR. These results shed light into the mechanism by which Ang-(1-7) exerts its positive physiological modulation of insulin actions in classical metabolic tissues and reinforces the central role of Akt in these effects. Copyright © 2012 Elsevier B.V. All rights reserved.

Testin, a novel binding partner of the calcium-sensing receptor, enhances receptor-mediated Rho-kinase signalling

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

Magno, Aaron L.; Department of Endocrinology and Diabetes, Sir Charles Gairdner Hospital, Hospital Avenue, Nedlands, Western Australia 6009; Ingley, Evan

Highlights: {yields} A yeast two-hybrid screen revealed testin bound to the calcium-sensing receptor. {yields} The second zinc finger of LIM domain 1 of testin is critical for interaction. {yields} Testin bound to a region of the receptor tail important for cell signalling. {yields} Testin and receptor interaction was confirmed in mammalian (HEK293) cells. {yields} Overexpression of testin enhanced receptor-mediated Rho signalling in HEK293 cells. -- Abstract: The calcium-sensing receptor (CaR) plays an integral role in calcium homeostasis and the regulation of other cellular functions including cell proliferation and cytoskeletal organisation. The multifunctional nature of the CaR is manifested through ligand-dependentmore » stimulation of different signalling pathways that are also regulated by partner binding proteins. Following a yeast two-hybrid library screen using the intracellular tail of the CaR as bait, we identified several novel binding partners including the focal adhesion protein, testin. Testin has not previously been shown to interact with cell surface receptors. The sites of interaction between the CaR and testin were mapped to the membrane proximal region of the receptor tail and the second zinc-finger of LIM domain 1 of testin, the integrity of which was found to be critical for the CaR-testin interaction. The CaR-testin association was confirmed in HEK293 cells by coimmunoprecipitation and confocal microscopy studies. Ectopic expression of testin in HEK293 cells stably expressing the CaR enhanced CaR-stimulated Rho activity but had no effect on CaR-stimulated ERK signalling. These results suggest an interplay between the CaR and testin in the regulation of CaR-mediated Rho signalling with possible effects on the cytoskeleton.« less

Opioid Receptors Mediate Direct Predictive Fear Learning: Evidence from One-Trial Blocking

ERIC Educational Resources Information Center

Cole, Sindy; McNally, Gavan P.

2007-01-01

Pavlovian fear learning depends on predictive error, so that fear learning occurs when the actual outcome of a conditioning trial exceeds the expected outcome. Previous research has shown that opioid receptors, including [mu]-opioid receptors in the ventrolateral quadrant of the midbrain periaqueductal gray (vlPAG), mediate such predictive fear…

G-protein mediates voltage regulation of agonist binding to muscarinic receptors: effects on receptor-Na/sup +/ channel interaction

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

Cohen-Armon, M.; Garty, H.; Sokolovsky, M.

1988-01-12

The authors previous experiments in membranes prepared from rat heart and brain led them to suggest that the binding of agonist to the muscarinic receptors and to the Na/sup +/ channels is a coupled event mediated by guanine nucleotide binding protein(s) (G-protein(s)). These in vitro findings prompted us to employ synaptoneurosomes from brain stem tissue to examine (i) the binding properties of (/sup 3/H) acetylcholine at resting potential and under depolarization conditions in the absence and presence of pertussis toxin; (ii) the binding of (/sup 3/H)batrachotoxin to Na/sup +/ channel(s) in the presence of the muscarinic agonists; and (iii) muscarinicallymore » induced /sup 22/Na/sup +/ uptake in the presence and absence of tetrodotoxin, which blocks Na/sup +/ channels. The findings indicate that agonist binding to muscarinic receptors is voltage dependent, that this process is mediated by G-protein(s), and that muscarinic agonists induce opening of Na/sup +/channels. The latter process persists even after pertussis toxin treatment, indicating that it is not likely to be mediated by pertussis toxin sensitive G-protein(s). The system with its three interacting components-receptor, G-protein, and Na/sup +/ channel-is such that at resting potential the muscarinic receptor induces opening of Na/sup +/ channels; this property may provide a possible physiological mechanism for the depolarization stimulus necessary for autoexcitation or repetitive firing in heart or brain tissues.« less

Virally mediated increased neurotensin 1 receptor in the nucleus accumbens decreases behavioral effects of mesolimbic system activation.

PubMed

Cáceda, Ricardo; Kinkead, Becky; Owens, Michael J; Nemeroff, Charles B

2005-12-14

Dopamine receptor agonist and NMDA receptor antagonist activation of the mesolimbic dopamine system increases locomotion and disrupts prepulse inhibition of the acoustic startle response (PPI), paradigms frequently used to study both the pharmacology of antipsychotic drugs and drugs of abuse. In rats, virally mediated overexpression of the neurotensin 1 (NT1) receptor in the nucleus accumbens antagonized d-amphetamine- and dizocilpine-induced PPI disruption, hyperlocomotion, and D-amphetamine-induced rearing. The NT receptor antagonist SR 142948A [2-[[5-(2,6-dimethoxyphenyl)-1-(4-N-(3-dimethylaminopropyl)-N-methylcarbamoyl)-2-isopropylphenyl)-1H-pyrazole-3-carbonyl]amino] adamantane-2-carboxylic acid, hydrochloride] blocked inhibition of dizocilpine-induced hyperlocomotion mediated by overexpression of the NT1 receptor. Together, these results suggest that increased nucleus accumbens NT neurotransmission, via the NT1 receptor, can decrease the effects of activation of the mesolimbic dopamine system and disruption of the glutamatergic input from limbic cortices, resembling the action of the atypical antipsychotic drug clozapine. In contrast to clozapine, virally mediated overexpression of the NT1 receptor in the nucleus accumbens had prolonged protective effects (up to 4 weeks after viral injection) without perturbing baseline PPI and locomotor behaviors. These data further confirm the NT1 receptor as the receptor mediating the antistimulant- and antipsychotic-like properties of NT and provide rationale for the development of NT1 receptor agonists as novel antipsychotic drugs. In addition, the NT1 receptor vector might be a valuable tool for understanding the mechanism of action of antipsychotic drugs and drugs of abuse and may have potential therapeutic applications.

Of pheromones and kairomones: what receptors mediate innate emotional responses?

PubMed

Fortes-Marco, Lluis; Lanuza, Enrique; Martinez-Garcia, Fernando

2013-09-01

Some chemicals elicit innate emotionally laden behavioral responses. Pheromones mediate sexual attraction, parental care or agonistic confrontation, whereas predators' kairomones elicit defensive behaviors in their preys. This essay explores the hypothesis that the detection of these semiochemicals relies on highly specific olfactory and/or vomeronasal receptors. The V1R, V2R, and formyl-peptide vomeronasal receptors bind their ligands in highly specific and sensitive way, thus being good candidates for pheromone- or kairomone-detectors (e.g., secreted and excreted proteins, peptides and lipophilic volatiles). The olfactory epithelium also expresses specific receptors, for example trace amine-associated receptors (TAAR) and guanylyl cyclase receptors (GC-D and other types), some of which bind kairomones and putative pheromones. However, most of the olfactory neurons express canonical olfactory receptors (ORs) that bind many ligands with different affinity, being not suitable for mediating responses to pheromones and kairomones. In this respect, trimethylthiazoline (TMT) is considered a fox-derived kairomone for mice and rats, but it seems to be detected by canonical ORs. Therefore, we have reassessed the kairomonal nature of TMT by analyzing the behavioral responses of outbred (CD1) and inbred mice (C57BL/J6) to TMT. Our results confirm that both mouse strains avoid TMT, which increases immobility in C57BL/J6, but not CD1 mice. However, mice of both strains sniff at TMT throughout the test and show no trace of TMT-induced contextual conditioning (immobility or avoidance). This suggests that TMT is not a kairomone but, similar to a loud noise, in high concentrations it induces aversion and stress as unspecific responses to a strong olfactory stimulation. Copyright © 2013 Wiley Periodicals, Inc.

Unique allosteric regulation of 5-hydroxytryptamine receptor-mediated signal transduction by oleamide

PubMed Central

Thomas, Elizabeth A.; Carson, Monica J.; Neal, Michael J.; Sutcliffe, J. Gregor

1997-01-01

The effects of oleamide, an amidated lipid isolated from the cerebrospinal fluid of sleep-deprived cats, on serotonin receptor-mediated responses were investigated in cultured mammalian cells. In rat P11 cells, which endogenously express the 5-hydroxytryptamine2A (5HT2A) receptor, oleamide significantly potentiated 5HT-induced phosphoinositide hydrolysis. In HeLa cells expressing the 5HT7 receptor subtype, oleamide caused a concentration-dependent increase in cAMP accumulation but with lower efficacy than that observed by 5HT. This effect was not observed in untransfected HeLa cells. Clozapine did not prevent the increase in cAMP elicited by oleamide, and ketanserin caused an ≈65% decrease. In the presence of 5HT, oleamide had the opposite effect on cAMP, causing insurmountable antagonism of the concentration-effect curve to 5HT, but had no effect on cAMP levels elicited by isoproterenol or forskolin. These results indicate that oleamide can modulate 5HT-mediated signal transduction at different subtypes of mammalian 5HT receptors. Additionally, our data indicate that oleamide acts at an apparent allosteric site on the 5HT7 receptor and elicits functional responses via activation of this site. This represents a unique mechanism of activation for 5HT G protein-coupled receptors and suggests that G protein-coupled neurotransmitter receptors may act like their iontropic counterparts (i.e., γ-aminobutyric acid type A receptors) in that there may be several binding sites on the receptor that regulate functional activity with varying efficacies. PMID:9391162

Thrombin-mediated proteoglycan synthesis utilizes both protein-tyrosine kinase and serine/threonine kinase receptor transactivation in vascular smooth muscle cells.

PubMed

Burch, Micah L; Getachew, Robel; Osman, Narin; Febbraio, Mark A; Little, Peter J

2013-03-08

G protein-coupled receptor signaling is mediated by three main mechanisms of action; these are the classical pathway, β-arrestin scaffold signaling, and the transactivation of protein-tyrosine kinase receptors such as those for EGF and PDGF. Recently, it has been demonstrated that G protein-coupled receptors can also mediate signals via transactivation of serine/threonine kinase receptors, most notably the transforming growth factor-β receptor family. Atherosclerosis is characterized by the development of lipid-laden plaques in blood vessel walls. Initiation of plaque development occurs via low density lipoprotein retention in the neointima of vessels due to binding with modified proteoglycans secreted by vascular smooth muscle cells. Here we show that transactivation of protein-tyrosine kinase receptors is mediated by matrix metalloproteinase triple membrane bypass signaling. In contrast, serine/threonine kinase receptor transactivation is mediated by a cytoskeletal rearrangement-Rho kinase-integrin system, and both protein-tyrosine kinase and serine/threonine kinase receptor transactivation concomitantly account for the total proteoglycan synthesis stimulated by thrombin in vascular smooth muscle. This work provides evidence of thrombin-mediated proteoglycan synthesis and paves the way for a potential therapeutic target for plaque development and atherosclerosis.

Estrogen-mediated inactivation of FOXO3a by the G protein-coupled estrogen receptor GPER.

PubMed

Zekas, Erin; Prossnitz, Eric R

2015-10-15

Estrogen (17β-estradiol) promotes the survival and proliferation of breast cancer cells and its receptors represent important therapeutic targets. The cellular actions of estrogen are mediated by the nuclear estrogen receptors ERα and ERβ as well as the 7-transmembrane spanning G protein-coupled estrogen receptor (GPER). We previously reported that estrogen activates the phosphoinositide 3-kinase (PI3Kinase) pathway via GPER, resulting in phosphatidylinositol (3,4,5)-trisphosphate (PIP3) production within the nucleus of breast cancer cells; however, the mechanisms and consequences of this activity remained unclear. MCF7 breast cancer cells were transfected with GFP-fused Forkhead box O3 (FOXO3) as a reporter to assess localization in response to estrogen stimulation. Inhibitors of PI3Kinases and EGFR were employed to determine the mechanisms of estrogen-mediated FOXO3a inactivation. Receptor knockdown with siRNA and the selective GPER agonist G-1 elucidated the estrogen receptor(s) responsible for estrogen-mediated FOXO3a inactivation. The effects of selective estrogen receptor modulators and downregulators (SERMs and SERDs) on FOXO3a in MCF7 cells were also determined. Cell survival (inhibition of apoptosis) was assessed by caspase activation. In the estrogen-responsive breast cancer cell line MCF7, FOXO3a inactivation occurs on a rapid time scale as a result of GPER, but not ERα, stimulation by estrogen, established by the GPER-selective agonist G-1 and knockdown of GPER and ERα. GPER-mediated inactivation of FOXO3a is effected by the p110α catalytic subunit of PI3Kinase as a result of transactivation of the EGFR. The SERMs tamoxifen and raloxifene, as well as the SERD ICI182,780, were active in mediating FOXO3a inactivation in a GPER-dependent manner. Additionally, estrogen-and G-1-mediated stimulation of MCF7 cells results in a decrease in caspase activation under proapoptotic conditions. Our results suggest that non-genomic signaling by GPER contributes

Old dance with a new partner: EGF receptor as the phenobarbital receptor mediating Cyp2B expression.

PubMed

Meyer, Sharon A; Jirtle, Randy L

2013-05-07

The decades-long quest for the phenobarbital (PhB) receptor that mediates activation of Cyp2B would appear fulfilled with the discovery by Mutoh et al., who found that PhB binds with pharmacological affinity to the epidermal growth factor receptor (EGFR). This finding provides a molecular basis for the suppression of hepatocyte EGFR signaling observed with PhB treatment, as previously noted in the context of tumor promotion. Although the PhB-mediated induction of Cyp2B expression through the association of a canonical nuclear receptor with the 5'-enhancer PBREM of Cyp2B is well known, direct binding of PhB to constitutive active androstane receptor (CAR, also known as NR1I3) typical of other xenobiotic-activated nuclear receptors has eluded detection. One EGF-activated pathway affected by the PhB-EGFR interaction is the loss of tyrosine phosphorylation of the scaffold protein RACK1. Dephosphorylated RACK1 provides the mechanistic link between the binding of PhB to EGFR and its effects on CAR by facilitating the interaction of serine/threonine phosphatase PP2A with inactive phosphorylated CAR. The dephosphorylation of CAR enables its translocation to the nucleus and activation of Cyp2B expression. Because EGFR and transducers RACK1, PP2A, and other partners are highly networked in numerous cellular pathways, this newly discovered partnership will surely reveal new fundamental roles for PhB beyond the regulation of drug metabolism.

Fast Modulation of μ-Opioid Receptor (MOR) Recycling Is Mediated by Receptor Agonists*

PubMed Central

Roman-Vendrell, Cristina; Yu, Y. Joy; Yudowski, Guillermo Ariel

2012-01-01

The μ-opioid receptor (MOR) is a member of the G protein-coupled receptor family and the main target of endogenous opioid neuropeptides and morphine. Upon activation by ligands, MORs are rapidly internalized via clathrin-coated pits in heterologous cells and dissociated striatal neurons. After initial endocytosis, resensitized receptors recycle back to the cell surface by vesicular delivery for subsequent cycles of activation. MOR trafficking has been linked to opioid tolerance after acute exposure to agonist, but it is also involved in the resensitization process. Several studies describe the regulation and mechanism of MOR endocytosis, but little is known about the recycling of resensitized receptors to the cell surface. To study this process, we induced internalization of MOR with [d-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO) and morphine and imaged in real time single vesicles recycling receptors to the cell surface. We determined single vesicle recycling kinetics and the number of receptors contained in them. Then we demonstrated that rapid vesicular delivery of recycling MORs to the cell surface was mediated by the actin-microtubule cytoskeleton. Recycling was also dependent on Rab4, Rab11, and the Ca2+-sensitive motor protein myosin Vb. Finally, we showed that recycling is acutely modulated by the presence of agonists and the levels of cAMP. Our work identifies a novel trafficking mechanism that increases the number of cell surface MORs during acute agonist exposure, effectively reducing the development of opioid tolerance. PMID:22378794

M2 and M3 muscarinic receptors are involved in enteric nerve-mediated contraction of the mouse ileum: Findings obtained with muscarinic-receptor knockout mouse.

PubMed

Takeuchi, Tadayoshi; Tanaka, Keisuke; Nakajima, Hidemitsu; Matsui, Minoru; Azuma, Yasu-Taka

2007-01-01

The involvement of muscarinic receptors in neurogenic responses of the ileum was studied in wild-type and muscarinic-receptor (M-receptor) knockout (KO) mice. Electrical field stimulation to the wild-type mouse ileum induced a biphasic response, a phasic and sustained contraction that was abolished by tetrodotoxin. The sustained contraction was prolonged for an extended period after the termination of electrical field stimulation. The phasic contraction was completely inhibited by atropine. In contrast, the sustained contraction was enhanced by atropine. Ileal strips prepared from M2-receptor KO mice exhibited a phasic contraction similar to that seen in wild-type mice and a sustained contraction that was larger than that in wild-type mice. In M3-receptor KO mice, the phasic contraction was smaller than that observed in wild-type mice. Acetylcholine exogenously administrated induced concentration-dependent contractions in strips isolated from wild-type, M2- and M3-receptor KO mice. However, contractions in M3-receptor KO mice shifted to the right. The sustained contraction was inhibited by capsaicin and neurokinin NK2 receptor antagonist, suggesting that it is mediated by substance P (SP). SP-induced contraction of M2-receptor KO mice did not differ from that of wild-type mice. SP immunoreactivity was located in enteric neurons, colocalized with M2 receptor immunoreactivity. These results suggest that atropine-sensitive phasic contraction is mainly mediated via the M3 receptor, and SP-mediated sustained contraction is negatively regulated by the M2 receptor at a presynaptic level.

Muscarinic Cholinergic Modulation of Long-Lasting Synaptic Plasticity in the Rat Dentate Gyrus

DTIC Science & Technology

1990-12-14

ability to block GABAB-mediated responses, which are PTx-sensitive. The effects of the GABAg receptor agonist baclofen on evoked responses were analyzed...both in slices previously exposed to 10/iM muscarine (n=4), and nonexposed slices (n=2). The disinhibitory effects of baclofen usually seen in...20 min washout of muscarine always preceeded the baclofen exposure, to allow for washout of muscarine. There were no differences in the responses to

Identification of the Muscarinic Acetylcholine Receptor Subtype Mediating Cholinergic Vasodilation in Murine Retinal Arterioles

PubMed Central

Sniatecki, Jan J.; Goloborodko, Evgeny; Steege, Andreas; Zavaritskaya, Olga; Vetter, Jan M.; Grus, Franz H.; Patzak, Andreas; Wess, Jürgen; Pfeiffer, Norbert

2011-01-01

Purpose. To identify the muscarinic acetylcholine receptor subtype that mediates cholinergic vasodilation in murine retinal arterioles. Methods. Muscarinic receptor gene expression was determined in murine retinal arterioles using real-time PCR. To assess the functional relevance of muscarinic receptors for mediating vascular responses, retinal vascular preparations from muscarinic receptor–deficient mice were studied in vitro. Changes in luminal arteriole diameter in response to muscarinic and nonmuscarinic vasoactive substances were measured by video microscopy. Results. Only mRNA for the M3 receptor was detected in retinal arterioles. Thus, M3 receptor–deficient mice (M3R−/−) and respective wild-type controls were used for functional studies. Acetylcholine concentration-dependently dilated retinal arterioles from wild-type mice. In contrast, vasodilation to acetylcholine was almost completely abolished in retinal arterioles from M3R−/− mice, whereas responses to the nitric oxide (NO) donor nitroprusside were retained. Carbachol, an acetylcholinesterase-resistant analog of acetylcholine, also evoked dilation in retinal arterioles from wild-type, but not from M3R−/−, mice. Vasodilation responses from wild-type mice to acetylcholine were negligible after incubation with the non–subtype-selective muscarinic receptor blocker atropine or the NO synthase inhibitor Nω-nitro-l-arginine methyl ester, and were even reversed to contraction after endothelial damage with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. Conclusions. These findings provide evidence that endothelial M3 receptors mediate cholinergic vasodilation in murine retinal arterioles via activation of NO synthase. PMID:21873683

Activation of EGF Receptor Kinase by L1-mediated Homophilic Cell Interactions

PubMed Central

Islam, Rafique; Kristiansen, Lars V.; Romani, Susana; Garcia-Alonso, Luis; Hortsch, Michael

2004-01-01

Neural cell adhesion molecules (CAMs) are important players during neurogenesis and neurite outgrowth as well as axonal fasciculation and pathfinding. Some of these developmental processes entail the activation of cellular signaling cascades. Pharmacological and genetic evidence indicates that the neurite outgrowth-promoting activity of L1-type CAMs is at least in part mediated by the stimulation of neuronal receptor tyrosine kinases (RTKs), especially FGF and EGF receptors. It has long been suspected that neural CAMs might physically interact with RTKs, but their activation by specific cell adhesion events has not been directly demonstrated. Here we report that gain-of-function conditions of the Drosophila L1-type CAM Neuroglian result in profound sensory axon pathfinding defects in the developing Drosophila wing. This phenotype can be suppressed by decreasing the normal gene dosage of the Drosophila EGF receptor gene. Furthermore, in Drosophila S2 cells, cell adhesion mediated by human L1-CAM results in the specific activation of human EGF tyrosine kinase at cell contact sites and EGF receptors engage in a physical interaction with L1-CAM molecules. Thus L1-type CAMs are able to promote the adhesion-dependent activation of EGF receptor signaling in vitro and in vivo. PMID:14718570

Activation of EGF receptor kinase by L1-mediated homophilic cell interactions.

PubMed

Islam, Rafique; Kristiansen, Lars V; Romani, Susana; Garcia-Alonso, Luis; Hortsch, Michael

2004-04-01

Neural cell adhesion molecules (CAMs) are important players during neurogenesis and neurite outgrowth as well as axonal fasciculation and pathfinding. Some of these developmental processes entail the activation of cellular signaling cascades. Pharmacological and genetic evidence indicates that the neurite outgrowth-promoting activity of L1-type CAMs is at least in part mediated by the stimulation of neuronal receptor tyrosine kinases (RTKs), especially FGF and EGF receptors. It has long been suspected that neural CAMs might physically interact with RTKs, but their activation by specific cell adhesion events has not been directly demonstrated. Here we report that gain-of-function conditions of the Drosophila L1-type CAM Neuroglian result in profound sensory axon pathfinding defects in the developing Drosophila wing. This phenotype can be suppressed by decreasing the normal gene dosage of the Drosophila EGF receptor gene. Furthermore, in Drosophila S2 cells, cell adhesion mediated by human L1-CAM results in the specific activation of human EGF tyrosine kinase at cell contact sites and EGF receptors engage in a physical interaction with L1-CAM molecules. Thus L1-type CAMs are able to promote the adhesion-dependent activation of EGF receptor signaling in vitro and in vivo.

Reduction by the Positive Allosteric Modulator of the GABAB Receptor, GS39783, of Alcohol Self-Administration in Sardinian Alcohol-Preferring Rats Exposed to the “Sipper” Procedure

PubMed Central

Maccioni, Paola; Flore, Paolo; Carai, Mauro A. M.; Mugnaini, Claudia; Pasquini, Serena; Corelli, Federico; Gessa, Gian Luigi; Colombo, Giancarlo

2010-01-01

The present study was designed to evaluate (a) alcohol self-administration behavior of selectively bred, Sardinian alcohol-preferring (sP) rats exposed to the so-called “sipper” procedure (characterized by the temporal separation between alcohol-seeking and -taking phases), and (b) the effect of the positive allosteric modulator of the GABAB receptor, GS39783, on alcohol self-administration in sP rats exposed to this procedure. To this end, sP rats were initially trained to lever-respond under a reinforcement requirement (RR) 55 (RR55) for alcohol. Achievement of RR55 resulted in the 20-min presentation of the alcohol (15%, v/v)-containing sipper bottle. Once stable levels of lever-responding and alcohol consumption were reached, rats were treated with 0, 25, 50, and 100 mg/kg GS39783 (i.g.) 60 min before the self-administration session. Rats displayed robust alcohol-seeking (as suggested by relatively short latencies to the first lever-response and high frequencies of lever-responding) and -taking (as suggested by alcohol intakes averaging approximately 1.5 g/kg) behaviors. Pretreatment with GS39783 inhibited both alcohol-seeking (the number of rats achieving RR55 and the mean RR value were virtually halved) and -taking (the amount of self-administered alcohol was reduced by approximately 60%). The results of the present study suggest the power of the “sipper” procedure in triggering high levels of alcohol-seeking and -taking behavior in sP rats. Further, these results extend to this additional procedure of alcohol self-administration the capacity of GS39783 to reduce the motivational properties of alcohol and alcohol consumption in sP rats. PMID:21423431

Recovery from ketamine-induced amnesia by blockade of GABA-A receptor in the medial prefrontal cortex of mice.

PubMed

Farahmandfar, Maryam; Akbarabadi, Ardeshir; Bakhtazad, Atefeh; Zarrindast, Mohammad-Reza

2017-03-06

Ketamine and other noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonists are known to induce deficits in learning and cognitive performance sensitive to prefrontal cortex (PFC) functions. The interaction of a glutamatergic and GABAergic systems is essential for many cognitive behaviors. In order to understand the effect of γ-aminobutyric acid (GABA)/glutamate interactions on learning and memory, we investigated the effects of intra medial prefrontal cortex (mPFC) injections of GABAergic agents on ketamine-induced amnesia using a one-trial passive avoidance task in mice. Pre-training systemic administration of ketamine (5, 10 and 15mg/kg, i.p.) dose-dependently decreased the memory acquisition of a one-trial passive avoidance task. Pre-training intra-mPFC injection of muscimol, GABAA receptor agonist (0.05, 0.1 and 0.2μg/mouse) and baclofen GABAB receptor agonist (0.05, 0.1, 0.5 and 1μg/mouse), impaired memory acquisition. However, co-pretreatment of different doses of muscimol and baclofen with a lower dose of ketamine (5mg/kg), which did not induce amnesia by itself, caused inhibition of memory formation. Our data showed that sole pre-training administration of bicuculline, GABA-A receptor antagonist and phaclofen GABA-B receptor antagonist into the mPFC, did not affect memory acquisition. In addition, the amnesia induced by pre-training ketamine (15mg/kg) was significantly decreased by the pretreatment of bicuculline (0.005, 0.1 and 0.5μg/mouse). It can be concluded that GABAergic system of the mPFC is involved in the ketamine-induced impairment of memory acquisition. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Promiscuous dimerization of the growth hormone secretagogue receptor (GHS-R1a) attenuates ghrelin-mediated signaling.

PubMed

Schellekens, Harriët; van Oeffelen, Wesley E P A; Dinan, Timothy G; Cryan, John F

2013-01-04

G protein-coupled receptors (GPCRs), such as the ghrelin receptor (GHS-R1a), the melanocortin 3 receptor (MC(3)), and the serotonin 2C receptor (5-HT(2C)), are well known for their key role in the homeostatic control of food intake and energy balance. Ghrelin is the only known gut peptide exerting an orexigenic effect and has thus received much attention as an anti-obesity drug target. In addition, recent data have revealed a critical role for ghrelin in dopaminergic mesolimbic circuits involved in food reward signaling. This study investigates the downstream signaling consequences and ligand-mediated co-internalization following heterodimerization of the GHS-R1a receptor with the dopamine 1 receptor, as well as that of the GHS-R1a-MC(3) heterodimer. In addition, a novel heterodimer between the GHS-R1a receptor and the 5-HT(2C) receptor was identified. Interestingly, dimerization of the GHS-R1a receptor with the unedited 5-HT(2C)-INI receptor, but not with the partially edited 5-HT(2C)-VSV isoform, significantly reduced GHS-R1a agonist-mediated calcium influx, which was completely restored following pharmacological blockade of the 5-HT(2C) receptor. These results combined suggest a potential novel mechanism for fine-tuning GHS-R1a receptor-mediated activity via promiscuous dimerization of the GHS-R1a receptor with other G protein-coupled receptors involved in appetite regulation and food reward. These findings may uncover novel mechanisms of significant relevance for the future pharmacological targeting of the GHS-R1a receptor in the homeostatic regulation of energy balance and in hedonic appetite signaling, both of which play a significant role in the development of obesity.

AMPA Receptors Mediate Acetylcholine Release from Starburst Amacrine Cells in the Rabbit Retina

PubMed Central

FIRTH, SALLY I.; LI, WEI; MASSEY, STEPHEN C.; MARSHAK, DAVID W.

2012-01-01

The light response of starburst amacrine cells is initiated by glutamate released from bipolar cells. To identify the receptors that mediate this response, we used a combination of anatomical and physiological techniques. An in vivo, rabbit eyecup was preloaded with [3H]-choline, and the [3H]-acetylcholine (ACh) released into the superfusate was monitored. A photopic, 3 Hz flashing light increased ACh release, and the selective AMPA receptor antagonist, GYKI 53655, blocked this light-evoked response. Nonselective AMPA/kainate agonists increased the release of ACh, but the specific kainate receptor agonist, SYM 2081, did not increase ACh release. Selective AMPA receptor antagonists, GYKI 53655 or GYKI 52466, also blocked the responses to agonists. We conclude that the predominant excitatory input to starburst amacrine cells is mediated by AMPA receptors. We also labeled lightly fixed rabbit retinas with antisera to choline acetyltransferase (ChAT), AMPA receptor subunits GluR1, GluR2/3, or GluR4, and kainate receptor subunits GluR6/7 and KA2. Labeled puncta were observed in the inner plexiform layer with each of these antisera to glutamate receptors, but only GluR2/3-IR puncta and GluR4-IR puncta were found on the ChAT-IR processes. The same was true of starburst cells injected intracellularly with Neurobiotin, and these AMPA receptor subunits were localized to two populations of puncta. The AMPA receptors are expected to desensitize rapidly, enhancing the sensitivity of starburst amacrine cells to moving or other rapidly changing stimuli. PMID:14515241

Stress-induced decrease of uterine blood flow in sheep is mediated by alpha 1-adrenergic receptors.

PubMed

Dreiling, Michelle; Bischoff, Sabine; Schiffner, Rene; Rupprecht, Sven; Kiehntopf, Michael; Schubert, Harald; Witte, Otto W; Nathanielsz, Peter W; Schwab, Matthias; Rakers, Florian

2016-09-01

Prenatal maternal stress can be transferred to the fetus via a catecholamine-dependent decrease of uterine blood flow (UBF). However, it is unclear which group of adrenergic receptors mediates this mechanism of maternal-fetal stress transfer. We hypothesized that in sheep, alpha 1-adrenergic receptors may play a key role in catecholamine mediated UBF decrease, as these receptors are mainly involved in peripheral vasoconstriction and are present in significant number in the uterine vasculature. After chronic instrumentation at 125 ± 1 days of gestation (dGA; term 150 dGA), nine pregnant sheep were exposed at 130 ± 1 dGA to acute isolation stress for one hour without visual, tactile, or auditory contact with their flockmates. UBF, blood pressure (BP), heart rate (HR), stress hormones, and blood gases were determined before and during this isolation challenge. Twenty-four hours later, experiments were repeated during alpha 1-adrenergic receptor blockage induced by a continuous intravenous infusion of urapidil. In both experiments, ewes reacted to isolation with an increase in serum norepinephrine, cortisol, BP, and HR as typical signs of activation of sympatho-adrenal and the hypothalamic-pituitary-adrenal axis. Stress-induced UBF decrease was prevented by alpha 1-adrenergic receptor blockage. We conclude that UBF decrease induced by maternal stress in sheep is mediated by alpha 1-adrenergic receptors. Future studies investigating prevention strategies of impact of prenatal maternal stress on fetal health should consider selective blockage of alpha 1-receptors to interrupt maternal-fetal stress transfer mediated by utero-placental malperfusion.

The Ionotropic Receptors IR21a and IR25a mediate cool sensing in Drosophila.

PubMed

Ni, Lina; Klein, Mason; Svec, Kathryn V; Budelli, Gonzalo; Chang, Elaine C; Ferrer, Anggie J; Benton, Richard; Samuel, Aravinthan Dt; Garrity, Paul A

2016-04-29

Animals rely on highly sensitive thermoreceptors to seek out optimal temperatures, but the molecular mechanisms of thermosensing are not well understood. The Dorsal Organ Cool Cells (DOCCs) of the Drosophila larva are a set of exceptionally thermosensitive neurons critical for larval cool avoidance. Here, we show that DOCC cool-sensing is mediated by Ionotropic Receptors (IRs), a family of sensory receptors widely studied in invertebrate chemical sensing. We find that two IRs, IR21a and IR25a, are required to mediate DOCC responses to cooling and are required for cool avoidance behavior. Furthermore, we find that ectopic expression of IR21a can confer cool-responsiveness in an Ir25a-dependent manner, suggesting an instructive role for IR21a in thermosensing. Together, these data show that IR family receptors can function together to mediate thermosensation of exquisite sensitivity.

RGS7/Gβ5/R7BP complex regulates synaptic plasticity and memory by modulating hippocampal GABABR-GIRK signaling

PubMed Central

Ostrovskaya, Olga; Xie, Keqiang; Masuho, Ikuo; Fajardo-Serrano, Ana; Lujan, Rafael; Wickman, Kevin; Martemyanov, Kirill A

2014-01-01

In the hippocampus, the inhibitory neurotransmitter GABA shapes the activity of the output pyramidal neurons and plays important role in cognition. Most of its inhibitory effects are mediated by signaling from GABAB receptor to the G protein-gated Inwardly-rectifying K+ (GIRK) channels. Here, we show that RGS7, in cooperation with its binding partner R7BP, regulates GABABR-GIRK signaling in hippocampal pyramidal neurons. Deletion of RGS7 in mice dramatically sensitizes GIRK responses to GABAB receptor stimulation and markedly slows channel deactivation kinetics. Enhanced activity of this signaling pathway leads to decreased neuronal excitability and selective disruption of inhibitory forms of synaptic plasticity. As a result, mice lacking RGS7 exhibit deficits in learning and memory. We further report that RGS7 is selectively modulated by its membrane anchoring subunit R7BP, which sets the dynamic range of GIRK responses. Together, these results demonstrate a novel role of RGS7 in hippocampal synaptic plasticity and memory formation. DOI: http://dx.doi.org/10.7554/eLife.02053.001 PMID:24755289

N-(4-Trifluoromethylphenyl)amide group of the synthetic histamine receptor agonist inhibits nicotinic acetylcholine receptor-mediated catecholamine secretion.

PubMed

Kim, Dong-Chan; Park, Yong-Soo; Jun, Dong-Jae; Hur, Eun-Mi; Kim, Sun-Hee; Choi, Bo-Hwa; Kim, Kyong-Tai

2006-02-28

The therapeutic targeting of nicotinic receptors requires the identification of drugs that selectively activate or inhibit a limited range of nicotine acetylcholine receptors (nAChRs). In this study, we identified N-(4-trifluoromethylphenyl)amide group of the synthetic histamine receptor ligands, histamine-trifluoromethyltoluide, that act as potent inhibitors of nAChRs in bovine adrenal chromaffin cells. Catecholamine secretion induced by the nAChRs agonist, 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), was significantly inhibited by histamine-trifluoromethyltoluide. Real time carbon-fiber amperometry confirmed the ability of histamine-trifluoromethyltoluide to inhibit DMPP-induced exocytosis in single chromaffin cells. We also found that histamine-trifluoromethyltoluide inhibited DMPP-induced [Ca(2+)](i) and [Na(+)](i) increases, as well as DMPP-induced inward currents in the absence of extracellular calcium. Histamine-trifluoromethyltoluide had no effect on [(3)H]nicotine binding or on calcium increases induced by high K(+), bradykinin, veratridine, histamine, and benzoylbenzoyl ATP. Among the synthetic histamine receptor ligands, clobenpropit exhibited similarity. In addition, 4'-nitroacetanilide also significantly attenuated nAChR-mediated catecholamine secretion. In conclusion, the N-(4-trifluoromethylphenyl)amide group of the histamine-trifluoromethyltoluide might be the critical moiety in the inhibition of nAChR-mediated CA secretion.

Estrous Cycle Regulation of Extrasynaptic δ-Containing GABAA Receptor-Mediated Tonic Inhibition and Limbic Epileptogenesis

PubMed Central

Wu, Xin; Gangisetty, Omkaram; Carver, Chase Matthew

2013-01-01

The ovarian cycle affects susceptibility to behavioral and neurologic conditions. The molecular mechanisms underlying these changes are poorly understood. Deficits in cyclical fluctuations in steroid hormones and receptor plasticity play a central role in physiologic and pathophysiologic menstrual conditions. It has been suggested that synaptic GABAA receptors mediate phasic inhibition in the hippocampus and extrasynaptic receptors mediate tonic inhibition in the dentate gyrus. Here we report a novel role of extrasynaptic δ-containing GABAA receptors as crucial mediators of the estrous cycle–related changes in neuronal excitability in mice, with hippocampus subfield specificity. In molecular and immunofluorescence studies, a significant increase occurred in δ-subunit, but not α4- and γ2-subunits, in the dentate gyrus during diestrus. However, δ-subunit upregulation was not evident in the CA1 region. The δ-subunit expression was undiminished by age and ovariectomy and in mice lacking progesterone receptors, but it was significantly reduced by finasteride, a neurosteroid synthesis inhibitor. Electrophysiologic studies confirmed greater potentiation of GABA currents by progesterone-derived neurosteroid allopregnanolone in dissociated dentate gyrus granule cells in diestrus than in CA1 pyramidal cells. The baseline conductance and allopregnanolone potentiation of tonic currents in dentate granule cells from hippocampal slices were higher than in CA1 pyramidal cells. In behavioral studies, susceptibility to hippocampus kindling epileptogenesis was lower in mice during diestrus. These results demonstrate the estrous cycle–related plasticity of neurosteroid-sensitive, δ-containing GABAA receptors that mediate tonic inhibition and seizure susceptibility. These findings may provide novel insight on molecular cascades of menstrual disorders like catamenial epilepsy, premenstrual syndrome, and migraine. PMID:23667248

EP2 receptors mediate airway relaxation to substance P, ATP, and PGE2.

PubMed

Fortner, C N; Breyer, R M; Paul, R J

2001-08-01

Substance P (SP) and ATP evoke transient, epithelium-dependent relaxation of constricted mouse tracheal smooth muscle. Relaxation to either SP or ATP is blocked by indomethacin, but the specific eicosanoid(s) involved have not been definitively identified. SP and ATP are reported to release PGE2 from airway epithelium in other species, suggesting PGE2 as a likely mediator in epithelium-dependent airway relaxation. Using mice homozygous for a gene-targeted deletion of the EP2 receptor [EP2(-/-)], one of the PGE2 receptors, we tested the hypothesis that PGE2 is the primary mediator of relaxation to SP or ATP. Relaxation in response to SP or ATP was significantly reduced in tracheas from EP2(-/-) mice. There were no differences between EP2(-/-) and wild-type tracheas in their physical dimensions, contraction to ACh, or relaxation to isoproterenol, thus ruling out any general alterations of smooth muscle function. There were also no differences between EP2(-/-) and wild-type tracheas in basal or stimulated PGE2 production. Exogenous PGE2 produced significantly less relaxation in EP2(-/-) tracheas compared with the wild type. Taken together, this experimental evidence supports the following two conclusions: EP2 receptors are of primary importance in airway relaxation to PGE2 and relaxation to SP or ATP is mediated through PGE2 acting on EP2 receptors.

Modulation of Pain Transmission by G Protein-Coupled Receptors

PubMed Central

Pan, Hui-Lin; Wu, Zi-Zhen; Zhou, Hong-Yi; Chen, Shao-Rui; Zhang, Hong-Mei; Li, De-Pei

2010-01-01

The heterotrimeric G protein-coupled receptors (GPCRs) represent the largest and most diverse family of cell surface receptors and proteins. GPCRs are widely distributed in the peripheral and central nervous systems and are one of the most important therapeutic targets in pain medicine. GPCRs are present on the plasma membrane of neurons and their terminals along the nociceptive pathways and are closely associated with the modulation of pain transmission. GPCRs that can produce analgesia upon activation include opioid, cannabinoid, α2-adrenergic, muscarinic acetylcholine, γ-aminobutyric acidB (GABAB), group II and III metabotropic glutamate, and somatostatin receptors. Recent studies have led to a better understanding of the role of these GPCRs in the regulation of pain transmission. Here, we review the current knowledge about the cellular and molecular mechanisms that underlie the analgesic actions of GPCR agonists, with a focus on their effects on ion channels expressed on nociceptive sensory neurons and on synaptic transmission at the spinal cord level. PMID:17959251

Adenovector GAD65 gene delivery into the rat trigeminal ganglion produces orofacial analgesia

PubMed Central

Vit, Jean-Philippe; Ohara, Peter T; Sundberg, Christopher; Rubi, Blanca; Maechler, Pierre; Liu, Chunyan; Puntel, Mariana; Lowenstein, Pedro; Castro, Maria; Jasmin, Luc

2009-01-01

Background Our goal is to use gene therapy to alleviate pain by targeting glial cells. In an animal model of facial pain we tested the effect of transfecting the glutamic acid decarboxylase (GAD) gene into satellite glial cells (SGCs) of the trigeminal ganglion by using a serotype 5 adenovector with high tropisms for glial cells. We postulated that GABA produced from the expression of GAD would reduce pain behavior by acting on GABA receptors on neurons within the ganglion. Results Injection of adenoviral vectors (AdGAD65) directly into the trigeminal ganglion leads to sustained expression of the GAD65 isoform over the 4 weeks observation period. Immunohistochemical analysis showed that adenovirus-mediated GAD65 expression and GABA synthesis were mainly in SGCs. GABAA and GABAB receptors were both seen in sensory neurons, yet only GABAA receptors decorated the neuronal surface. GABA receptors were not found on SGCs. Six days after injection of AdGAD65 into the trigeminal ganglion, there was a statistically significant decrease of pain behavior in the orofacial formalin test, a model of inflammatory pain. Rats injected with control virus (AdGFP or AdLacZ) had no reduction in their pain behavior. AdGAD65-dependent analgesia was blocked by bicuculline, a selective GABAA receptor antagonist, but not by CGP46381, a selective GABAB receptor antagonist. Conclusion Transfection of glial cells in the trigeminal ganglion with the GAD gene blocks pain behavior by acting on GABAA receptors on neuronal perikarya. PMID:19656360

Adenovector GAD65 gene delivery into the rat trigeminal ganglion produces orofacial analgesia.

PubMed

Vit, Jean-Philippe; Ohara, Peter T; Sundberg, Christopher; Rubi, Blanca; Maechler, Pierre; Liu, Chunyan; Puntel, Mariana; Lowenstein, Pedro; Castro, Maria; Jasmin, Luc

2009-08-05

Our goal is to use gene therapy to alleviate pain by targeting glial cells. In an animal model of facial pain we tested the effect of transfecting the glutamic acid decarboxylase (GAD) gene into satellite glial cells (SGCs) of the trigeminal ganglion by using a serotype 5 adenovector with high tropisms for glial cells. We postulated that GABA produced from the expression of GAD would reduce pain behavior by acting on GABA receptors on neurons within the ganglion. Injection of adenoviral vectors (AdGAD65) directly into the trigeminal ganglion leads to sustained expression of the GAD65 isoform over the 4 weeks observation period. Immunohistochemical analysis showed that adenovirus-mediated GAD65 expression and GABA synthesis were mainly in SGCs. GABAA and GABAB receptors were both seen in sensory neurons, yet only GABAA receptors decorated the neuronal surface. GABA receptors were not found on SGCs. Six days after injection of AdGAD65 into the trigeminal ganglion, there was a statistically significant decrease of pain behavior in the orofacial formalin test, a model of inflammatory pain. Rats injected with control virus (AdGFP or AdLacZ) had no reduction in their pain behavior. AdGAD65-dependent analgesia was blocked by bicuculline, a selective GABAA receptor antagonist, but not by CGP46381, a selective GABAB receptor antagonist. Transfection of glial cells in the trigeminal ganglion with the GAD gene blocks pain behavior by acting on GABAA receptors on neuronal perikarya.

DEPENDENCE OF PPAR LIGAND-INDUCED MAPK SIGNALING ON EPIDERMAL GROWTH FACTOR RECEPTOR TRANSACTIVATION HEPARIN-BINDING EGF CLEAVAGE MEDIATES ZINC-INDUCED EGF RECEPTOR PHOSPHORYLATION

EPA Science Inventory

Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that function as ligand-activated transcription factors regulating lipid metabolism and homeostasis. In addition to their ability to regulate PPAR-mediated gene transcription, PPARalpha and gamma li...

The miR-199-dynamin regulatory axis controls receptor-mediated endocytosis.

PubMed

Aranda, Juan F; Canfrán-Duque, Alberto; Goedeke, Leigh; Suárez, Yajaira; Fernández-Hernando, Carlos

2015-09-01

Small non-coding RNAs (microRNAs) are important regulators of gene expression that modulate many physiological processes; however, their role in regulating intracellular transport remains largely unknown. Intriguingly, we found that the dynamin (DNM) genes, a GTPase family of proteins responsible for endocytosis in eukaryotic cells, encode the conserved miR-199a and miR-199b family of miRNAs within their intronic sequences. Here, we demonstrate that miR-199a and miR-199b regulate endocytic transport by controlling the expression of important mediators of endocytosis such as clathrin heavy chain (CLTC), Rab5A, low-density lipoprotein receptor (LDLR) and caveolin-1 (Cav-1). Importantly, miR-199a-5p and miR-199b-5p overexpression markedly inhibits CLTC, Rab5A, LDLR and Cav-1 expression, thus preventing receptor-mediated endocytosis in human cell lines (Huh7 and HeLa). Of note, miR-199a-5p inhibition increases target gene expression and receptor-mediated endocytosis. Taken together, our work identifies a new mechanism by which microRNAs regulate intracellular trafficking. In particular, we demonstrate that the DNM, miR-199a-5p and miR-199b-5p genes act as a bifunctional locus that regulates endocytosis, thus adding an unexpected layer of complexity in the regulation of intracellular trafficking. © 2015. Published by The Company of Biologists Ltd.

The brain cytoplasmic RNA BC1 regulates dopamine D2 receptor-mediated transmission in the striatum.

PubMed

Centonze, Diego; Rossi, Silvia; Napoli, Ilaria; Mercaldo, Valentina; Lacoux, Caroline; Ferrari, Francesca; Ciotti, Maria Teresa; De Chiara, Valentina; Prosperetti, Chiara; Maccarrone, Mauro; Fezza, Filomena; Calabresi, Paolo; Bernardi, Giorgio; Bagni, Claudia

2007-08-15

Dopamine D(2) receptor (D(2)DR)-mediated transmission in the striatum is remarkably flexible, and changes in its efficacy have been heavily implicated in a variety of physiological and pathological conditions. Although receptor-associated proteins are clearly involved in specific forms of synaptic plasticity, the molecular mechanisms regulating the sensitivity of D(2) receptors in this brain area are essentially obscure. We have studied the physiological responses of the D(2)DR stimulations in mice lacking the brain cytoplasmic RNA BC1, a small noncoding dendritically localized RNA that is supposed to play a role in mRNA translation. We show that the efficiency of D(2)-mediated transmission regulating striatal GABA synapses is under the control of BC1 RNA, through a negative influence on D(2) receptor protein level affecting the functional pool of receptors. Ablation of the BC1 gene did not result in widespread dysregulation of synaptic transmission, because the sensitivity of cannabinoid CB(1) receptors was intact in the striatum of BC1 knock-out (KO) mice despite D(2) and CB(1) receptors mediated similar electrophysiological actions. Interestingly, the fragile X mental retardation protein FMRP, one of the multiple BC1 partners, is not involved in the BC1 effects on the D(2)-mediated transmission. Because D(2)DR mRNA is apparently equally translated in the BC1-KO and wild-type mice, whereas the protein level is higher in BC1-KO mice, we suggest that BC1 RNA controls D(2)DR indirectly, probably regulating translation of molecules involved in D(2)DR turnover and/or stability.

Glucocorticoid receptor-mediated induction of glutamine synthetase in skeletal muscle cells in vitro

NASA Technical Reports Server (NTRS)

Max, Stephen R.; Thomas, John W.; Banner, Carl; Vitkovic, Ljubisa; Konagaya, Masaaki

1987-01-01

The regulation by glucocorticoids of glutamine synthetase in L6 muscle cells in culture is studied. Glutamine synthetase activity was strikingly enhanced by dexamethasone. The dexamethasone-mediated induction of glutamine synthetase activity was blocked by RU38486, a glucocorticoid antagonist, indicating the involvement of intracellular glucocorticoid receptors in the induction process. RU38486 alone was without effect. Northern blot analysis revealed that dexamethasone-mediated enhancement of glutamine synthetase activity involves increased levels of glutamine synthetase mRNA. Glucocorticoids regulate the expression of glutamine synthetase mRNA in cultured muscle cells via interaction with intracellular receptors. Such regulation may be relevant to control of glutamine production by muscle.

Clinical, imaging, and follow-up observations of patients with anti-GABAB receptor encephalitis.

PubMed

Qiao, Song; Zhang, Yin-Xi; Zhang, Bi-Jun; Lu, Ru-Yi; Lai, Qi-Lun; Chen, Lin-Hui; Wu, Jiong

2017-05-01

Anti-gamma-aminobutyric acid B (anti-GABA B ) receptor encephalitis is a newly described type of autoimmune encephalitis. We report a case series of patients diagnosed with anti-GABA B receptor encephalitis in China, focusing on their presentations, laboratory and imaging results, and outcomes, as well as the treatment strategies which were employed. Data from patients diagnosed with anti-GABA B receptor encephalitis in the Second Affiliated Hospital, School of Medicine, Zhejiang University, from January 2014 to June 2015 were retrospectively collected and analyzed. Based on specific diagnostic criteria, seven cases were included. Six of the seven patients were males, and a median age at presentation of 56 years (range: 4-71 years). Seizures were the most common initial symptom, and all patients developed symptoms of typical limbic encephalitis during their disease course. Additional types of autoantibodies were identified in four patients. After presentation, three patients were found to have small cell lung cancer and one patient was eventually diagnosed with thymoma. All patients accepted first-line immune therapy, but only one chose tumor treatment. The three tumor-free patients had a good outcome, whereas those with tumors had a poor one. Finally, there were no relapses during follow-up. Anti-GABA B receptor encephalitis is a rare, unique autoimmune disease, and is often associated with tumors. It should be considered in the differential diagnosis for middle and senior-aged patients who present with predominantly limbic encephalitis symptoms. Importantly, earlier recognition of this potentially treatable condition could improve its overall prognosis.

Intra-accumbens baclofen, but not muscimol, mimics the effects of food withdrawal on feeding behaviour.

PubMed

Pulman, K G T; Somerville, E M; Clifton, P G

2010-11-01

Intra-accumbens stimulation of GABA receptors results in a robust increase in food intake. However the differential consequences of stimulating GABA(A) and GABA(B) receptors in the nucleus accumbens have not been extensively explored with respect to feeding behaviour. Here we compare the effects of the GABA(B) receptor agonist baclofen and GABA(A) receptor agonist muscimol, infused into the nucleus accumbens shell, on food intake and related behavior patterns. Baclofen (110-440 ρmol) dose dependently enhanced intake and delayed the onset of satiety within the test period as did the effects of 4-8h food withdrawal. Muscimol (220-660 ρmol) enhanced intake but also disrupted the sequence of associated behaviours at every dose tested. We conclude that GABA(B) receptors in the nucleus accumbens shell may play a role in relation to feeding motivation whereas GABA(A) receptors may, as previously suggested, have a more restricted role in relation to the motor components of approach to food and ingestion. Copyright © 2010 Elsevier Inc. All rights reserved.

Structural, signalling and regulatory properties of the group I metabotropic glutamate receptors: prototypic family C G-protein-coupled receptors.

PubMed Central

Hermans, E; Challiss, R A

2001-01-01

In 1991 a new type of G-protein-coupled receptor (GPCR) was cloned, the type 1a metabotropic glutamate (mGlu) receptor, which, despite possessing the defining seven-transmembrane topology of the GPCR superfamily, bore little resemblance to the growing number of other cloned GPCRs. Subsequent studies have shown that there are eight mammalian mGlu receptors that, together with the calcium-sensing receptor, the GABA(B) receptor (where GABA is gamma-aminobutyric acid) and a subset of pheromone, olfactory and taste receptors, make up GPCR family C. Currently available data suggest that family C GPCRs share a number of structural, biochemical and regulatory characteristics, which differ markedly from those of the other GPCR families, most notably the rhodopsin/family A GPCRs that have been most widely studied to date. This review will focus on the group I mGlu receptors (mGlu1 and mGlu5). This subgroup of receptors is widely and differentially expressed in neuronal and glial cells within the brain, and receptor activation has been implicated in the control of an array of key signalling events, including roles in the adaptative changes needed for long-term depression or potentiation of neuronal synaptic connectivity. In addition to playing critical physiological roles within the brain, the mGlu receptors are also currently the focus of considerable attention because of their potential as drug targets for the treatment of a variety of neurological and psychiatric disorders. PMID:11672421

Independent of 5-HT1A receptors, neurons in the paraventricular hypothalamus mediate ACTH responses from MDMA

PubMed Central

Zaretsky, Dmitry V.; Zaretskaia, Maria V.; DiMicco, Joseph A.; Durant, Pamela J.; Ross, Christian T.; Rusyniak, Daniel E.

2013-01-01

Acute and chronic complications from the substituted amphetamine 3,4-methylenedioxymethamphetamine (MDMA) are linked to activation of the hypothalamic-pituitary-adrenal (HPA) axis. How MDMA activates the HPA axis is not known. HPA responses to stress are known to be mediated through the paraventricular (PVH) hypothalamus and to involve serotonin-1a (5-HT1A) receptors. We sought to determine if the PVH and 5-HT1A receptors were also involved in mediating HPA responses to MDMA. Rats were pretreated with either saline or a 5-HT1A antagonist, WAY-100635 (WAY), followed by a systemic dose of MDMA (7.5 mg/kg i.v.). Animals pretreated with WAY had significantly lower plasma ACTH concentrations after MDMA. To determine if neurons in the PVH were involved, and if their involvement was mediated by 5-HT1A receptors, rats implanted with guide cannulas targeting the PVH were microinjected with the GABAA receptor agonist muscimol, aCSF, or WAY followed by MDMA. Compared to aCSF microinjections of muscimol significantly attenuated the MDMA-induced rise in plasma ACTH (126 vs. 588 pg/ml, P=<0.01). WAY had no effect. Our data demonstrates that neurons in the PVH, independent of 5-HT1A receptors, mediate ACTH responses to MDMA. PMID:23933156

G(o) transduces GABAB-receptor modulation of N-type calcium channels in cultured dorsal root ganglion neurons.

PubMed

Menon-Johansson, A S; Berrow, N; Dolphin, A C

1993-11-01

High-voltage-activated (HVA) calcium channel currents (IBa) were recorded from acutely replated cultured dorsal root ganglion (DRG) neurons. IBa was irreversibly inhibited by 56.9 +/- 2.7% by 1 microM omega-conotoxin-GVIA (omega-CTx-GVIA), whereas the 1,4-dihydropyridine antagonist nicardipine was ineffective. The selective gamma-aminobutyric acidB (GABAB) agonist, (-)-baclofen (50 microM), inhibited the HVA IBa by 30.7 +/- 5.4%. Prior application of omega-CTx-GVIA completely occluded inhibition of the HVA IBa by (-)-baclofen, indicating that in this preparation (-)-baclofen inhibits N-type current. To investigate which G protein subtype was involved, cells were replated in the presence of anti-G protein antisera. Under these conditions the antibodies were shown to enter the cells through transient pores created during the replating procedure. Replating DRGs in the presence of anti-G(o) antiserum, raised against the C-terminal decapeptide of the G alpha o subunit, reduced (-)-baclofen inhibition of the HVA IBa, whereas replating DRGs in the presence of the anti-Gi antiserum did not. Using anti-G alpha o antisera (1:2000) and confocal laser microscopy, G alpha o localisation was investigated in both unreplated and replated neurons. G alpha o immunoreactivity was observed at the plasma membrane, neurites, attachment plaques and perinuclear region, and was particularly pronounced at points of cell-to-cell contact. The plasma membrane G alpha o immunoreactivity was completely blocked by preincubation with the immunising G alpha o undecapeptide (1 microgram.ml-1) for 1 h at 37 degrees C. A similar treatment also blocked recognition of G alpha o in brain membranes on immunoblots.(ABSTRACT TRUNCATED AT 250 WORDS)

Unsaturated fatty acyl recognition by Frizzled receptors mediates dimerization upon Wnt ligand binding

PubMed Central

Nile, Aaron H.; Mukund, Susmith; Stanger, Karen; Wang, Weiru; Hannoush, Rami N.

2017-01-01

Frizzled (FZD) receptors mediate Wnt signaling in diverse processes ranging from bone growth to stem cell activity. Moreover, high FZD receptor expression at the cell surface contributes to overactive Wnt signaling in subsets of pancreatic, ovarian, gastric, and colorectal tumors. Despite the progress in biochemical understanding of Wnt–FZD receptor interactions, the molecular basis for recognition of Wnt cis-unsaturated fatty acyl groups by the cysteine-rich domain (CRD) of FZD receptors remains elusive. Here, we determined a crystal structure of human FZD7 CRD unexpectedly bound to a 24-carbon fatty acid. We also report a crystal structure of human FZD5 CRD bound to C16:1 cis-Δ9 unsaturated fatty acid. Both structures reveal a dimeric arrangement of the CRD. The lipid-binding groove exhibits flexibility and spans both monomers, adopting a U-shaped geometry that accommodates the fatty acid. Re-evaluation of the published mouse FZD8 CRD structure reveals that it also shares the same architecture as FZD5 and FZD7 CRDs. Our results define a common molecular mechanism for recognition of the cis-unsaturated fatty acyl group, a necessary posttranslational modification of Wnts, by multiple FZD receptors. The fatty acid bridges two CRD monomers, implying that Wnt binding mediates FZD receptor dimerization. Our data uncover possibilities for the arrangement of Wnt–FZD CRD complexes and shed structural insights that could aide in the identification of pharmacological strategies to modulate FZD receptor function. PMID:28377511

Chronic Exposure to Anabolic Androgenic Steroids Alters Neuronal Function in the Mammalian Forebrain via Androgen Receptor- and Estrogen Receptor-Mediated Mechanisms

PubMed Central

Penatti, Carlos A A; Porter, Donna M; Henderson, Leslie P

2009-01-01

Anabolic androgenic steroids (AAS) can promote detrimental effects on social behaviors for which γ-aminobutyric acid type A (GABAA) receptor-mediated circuits in the forebrain play a critical role. While all AAS bind to androgen receptors (AR), they may also be aromatized to estrogens and thus potentially impart effects via estrogen receptors (ER). Chronic exposure of wild type male mice to a combination of chemically distinct AAS increased action potential (AP) frequency, selective GABAA receptor subunit mRNAs, and GABAergic synaptic current decay in the medial preoptic area (mPOA). Experiments performed with pharmacological agents and in AR-deficient Tfm mutant mice suggest that the AAS-dependent enhancement of GABAergic transmission in wild type mice is AR-mediated. In AR-deficient mice, the AAS elicited dramatically different effects, decreasing AP frequency, sIPSC amplitude and frequency and the expression of selective GABAA receptor subunit mRNAs. Surprisingly, in the absence of AR signaling, the data indicate that the AAS do not act as ER agonists, but rather suggest a novel in vivo action in which the AAS inhibit aromatase and impair endogenous ER signaling. These results show that the AAS have the capacity to alter neuronal function in the forebrain via multiple steroid signaling mechanisms and suggest that effects of these steroids in the brain will depend not only on the balance of AR- vs. ER-mediated regulation for different target genes, but also on the ability of these drugs to alter steroid metabolism and thus the endogenous steroid milieu. PMID:19812324

Activation of the sigma receptor 1 modulates AMPA receptor-mediated light-evoked excitatory postsynaptic currents in rat retinal ganglion cells.

PubMed

Liu, Lei-Lei; Deng, Qin-Qin; Weng, Shi-Jun; Yang, Xiong-Li; Zhong, Yong-Mei

2016-09-22

Sigma receptor (σR), a unique receptor family, is classified into three subtypes: σR1, σR2 and σR3. It was previously shown that σR1 activation induced by 1μM SKF10047 (SKF) suppressed N-methyl-d-aspartate (NMDA) receptor-mediated responses of rat retinal ganglion cells (GCs) and the suppression was mediated by a distinct Ca(2+)-dependent phospholipase C (PLC)-protein kinase C (PKC) pathway. In the present work, using whole-cell patch-clamp techniques in rat retinal slice preparations, we further demonstrate that SKF of higher dosage (50μM) significantly suppressed AMPA receptor (AMPAR)-mediated light-evoked excitatory postsynaptic currents (L-EPSCs) of retinal ON-type GCs (ON GCs), and the effect was reversed by the σR1 antagonist BD1047, suggesting the involvement of σR1. The SKF (50μM) effect was unlikely due to a change in glutamate release from bipolar cells, as suggested by the unaltered paired-pulse ratio (PPR) of AMPAR-mediated EPSCs of ON GCs. SKF (50μM) did not change L-EPSCs of ON GCs when the G protein inhibitor GDP-β-S or the protein kinase G (PKG) inhibitor KT5823 was intracellularly infused. Calcium imaging further revealed that SKF (50μM) did not change intracellular calcium concentration in GCs and persisted to suppress L-EPSCs when intracellular calcium was chelated by BAPTA. The SKF (50μM) effect was intact when protein kinase A (PKA) and phosphatidylinostiol (PI)-PLC signaling pathways were both blocked. We conclude that the SKF (50μM) effect is Ca(2+)-independent, PKG-dependent, but not involving PKA, PI-PLC pathways. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

BK channel β1 subunits regulate airway contraction secondary to M2 muscarinic acetylcholine receptor mediated depolarization.

PubMed

Semenov, Iurii; Wang, Bin; Herlihy, Jeremiah T; Brenner, Robert

2011-04-01

The large conductance calcium- and voltage-activated potassium channel (BK channel) and its smooth muscle-specific β1 subunit regulate excitation–contraction coupling in many types of smooth muscle cells. However, the relative contribution of BK channels to control of M2- or M3-muscarinic acetylcholine receptor mediated airway smooth muscle contraction is poorly understood. Previously, we showed that knockout of the BK channel β1 subunit enhances cholinergic-evoked trachea contractions. Here, we demonstrate that the enhanced contraction of the BK β1 knockout can be ascribed to a defect in BK channel opposition of M2 receptor-mediated contractions. Indeed, the enhanced contraction of β1 knockout is eliminated by specific M2 receptor antagonism. The role of BK β1 to oppose M2 signalling is evidenced by a greater than fourfold increase in the contribution of L-type voltage-dependent calcium channels to contraction that otherwise does not occur with M2 antagonist or with β1 containing BK channels. The mechanism through which BK channels oppose M2-mediated recruitment of calcium channels is through a negative shift in resting voltage that offsets, rather than directly opposes, M2-mediated depolarization. The negative shift in resting voltage is reduced to similar extents by BK β1 knockout or by paxilline block of BK channels. Normalization of β1 knockout baseline voltage with low external potassium eliminated the enhanced M2-receptor mediated contraction. In summary, these findings indicate that an important function of BK/β1 channels is to oppose cholinergic M2 receptor-mediated depolarization and activation of calcium channels by restricting excitation–contraction coupling to more negative voltage ranges.

Inhibition of Na+−H+ exchange impairs receptor-mediated albumin endocytosis in renal proximal tubule-derived epithelial cells from opossum

PubMed Central

Gekle, Michael; Drumm, Karina; Mildenberger, Sigrid; Freudinger, Ruth; Gaßner, Birgit; Silbernagl, Stefan

1999-01-01

Receptor-mediated endocytosis is an important mechanism for transport of macromolecules and regulation of cell-surface receptor expression. In renal proximal tubules, receptor-mediated endocytosis mediates the reabsorption of filtered albumin. Acidification of the endocytic compartments is essential because it interferes with ligand-receptor dissociation, vesicle trafficking, fusion events and coat formation. Here we show that the activity of Na+−H+ exchanger isoform 3 (NHE3) is important for proper receptor-mediated endocytosis of albumin and endosomal pH homeostasis in a renal proximal tubular cell line (opossum kidney cells) which expresses NHE3 only. Depending on their inhibitory potency with respect to NHE3 and their lipophilicity, the NHE inhibitors EIPA, amiloride and HOE694 differentially reduced albumin endocytosis. The hydrophilic inhibitor HOE642 had no effect. Inhibition of NHE3 led to an alkalinization of early endosomes and to an acidification of the cytoplasm, indicating that Na+−H+ exchange contributes to the acidification of the early endosomal compartment due to the existence of a sufficient Na+ gradient across the endosomal membrane. Exclusive acidification of the cytoplasm with propionic acid or by removal of Na+ induced a significantly smaller reduction in endocytosis than that induced by inhibition of Na+−H+ exchange. Analysis of the inhibitory profiles indicates that in early endosomes and endocytic vesicles NHE3 is of major importance, whereas plasma membrane NHE3 plays a minor role. Thus, NHE3-mediated acidification along the first part of the endocytic pathway plays an important role in receptor-mediated endocytosis. Furthermore, the involvement of NHE3 offers new ways to explain the regulation of receptor-mediated endocytosis. PMID:10545138

The Ionotropic Receptors IR21a and IR25a mediate cool sensing in Drosophila

PubMed Central

Ni, Lina; Klein, Mason; Svec, Kathryn V; Budelli, Gonzalo; Chang, Elaine C; Ferrer, Anggie J; Benton, Richard; Samuel, Aravinthan DT; Garrity, Paul A

2016-01-01

Animals rely on highly sensitive thermoreceptors to seek out optimal temperatures, but the molecular mechanisms of thermosensing are not well understood. The Dorsal Organ Cool Cells (DOCCs) of the Drosophila larva are a set of exceptionally thermosensitive neurons critical for larval cool avoidance. Here, we show that DOCC cool-sensing is mediated by Ionotropic Receptors (IRs), a family of sensory receptors widely studied in invertebrate chemical sensing. We find that two IRs, IR21a and IR25a, are required to mediate DOCC responses to cooling and are required for cool avoidance behavior. Furthermore, we find that ectopic expression of IR21a can confer cool-responsiveness in an Ir25a-dependent manner, suggesting an instructive role for IR21a in thermosensing. Together, these data show that IR family receptors can function together to mediate thermosensation of exquisite sensitivity. DOI: http://dx.doi.org/10.7554/eLife.13254.001 PMID:27126188

Alterations in food intake elicited by GABA and opioid agonists and antagonists administered into the ventral tegmental area region of rats.

PubMed

Echo, Joyce A; Lamonte, Nicole; Ackerman, Tsippa F; Bodnar, Richard J

2002-05-01

Food intake is significantly increased following administration of mu-selective opioid agonists into the ventral tegmental area (VTA) region acting through multiple local opioid receptor subtypes. Since GABA receptor agonists in the VTA region are capable of eliciting feeding, the present study investigated whether feeding elicited by the mu-selective opioid agonist [D-Ala(2), NMe(4), Gly-ol(5)]-enkephalin (DAMGO) in the VTA region was altered by pretreatment into the same site with equimolar doses of either GABA(A) (bicuculline) or GABA(B) (saclofen) antagonists, and further, whether pretreatment with either general opioid or selective GABA receptor antagonists decreased feeding elicited by GABA(A) (muscimol) or GABA(B) (baclofen) agonists in the VTA region. DAMGO-induced feeding in the VTA region was dose-dependently decreased following pretreatment with either GABA(A) or GABA(B) antagonists in the absence of significant alterations in food intake by the antagonists per se. However, the presence of short-lived seizures following bicuculline in the VTA region suggests that this ingestive effect was caused by nonspecific actions. In contrast, GABA(B) receptors are involved in the full expression of mu-opioid agonist-induced feeding in this region since saclofen failed to elicit either seizure activity or a conditioned taste aversion. Pretreatment with naltrexone in the VTA region reduced intake elicited by baclofen, but not muscimol. Finally, baclofen-induced feeding was significantly reduced by saclofen, but not bicuculline, pretreatment in the VTA region. Therefore, possible coregulation between GABA(B) and opioid receptors in the VTA region, as suggested by immunocytochemical evidence, is supported by these behavioral effects upon ingestion.

Vortioxetine Treatment Reverses Subchronic PCP Treatment-Induced Cognitive Impairments: A Potential Role for Serotonin Receptor-Mediated Regulation of GABA Neurotransmission

PubMed Central

Pehrson, Alan L.; Pedersen, Christian S.; Tølbøl, Kirstine Sloth; Sanchez, Connie

2018-01-01

Major depressive disorder (MDD) is associated with cognitive impairments that may contribute to poor functional outcomes. Clinical data suggests that the multimodal antidepressant vortioxetine attenuates some cognitive impairments in MDD patients, but the mechanistic basis for these improvements is unclear. One theory suggests that vortioxetine improves cognition by suppressing γ-amino butyric acid (GABA)ergic neurotransmission, thereby increasing glutamatergic activation. Vortioxetine’s effects on cognition, GABA and glutamate neurotransmission have been supported in separate experiments, but no empirical work has directly connected vortioxetine’s cognitive effects to those on GABA and glutamate neurotransmission. In this paper, we attempt to bridge this gap by evaluating vortioxetine’s effects in the subchronic PCP (subPCP) model, which induces impaired cognitive function and altered GABA and glutamate neurotransmission. We demonstrate that acute or subchronic vortioxetine treatment attenuated subPCP-induced deficits in attentional set shifting (AST) performance, and that the selective 5-HT3 receptor antagonist ondansetron or the 5-HT reuptake inhibitor escitalopram could mimic this effect. Furthermore, acute vortioxetine treatment reversed subPCP-induced object recognition (OR) deficits in rats, while subchronic vortioxetine reversed subPCP-induced Object Recognition and object placement impairments in mice. Finally, subPCP treatment reduced GABAB receptor expression in a manner that was insensitive to vortioxetine treatment, and subchronic vortioxetine treatment alone, but not in combination with subPCP, significantly increased GABA’s affinity for the GABAA receptor. These data suggest that vortioxetine reverses cognitive impairments in a model associated with altered GABA and glutamate neurotransmission, further supporting the hypothesis that vortioxetine’s GABAergic and glutamatergic effects are relevant for cognitive function. PMID:29559911

Salicylate, an aspirin metabolite, specifically inhibits the current mediated by glycine receptors containing α1-subunits

PubMed Central

Lu, Y-G; Tang, Z-Q; Ye, Z-Y; Wang, H-T; Huang, Y-N; Zhou, K-Q; Zhang, M; Xu, T-L; Chen, L

2009-01-01

Background and purpose: Aspirin or its metabolite sodium salicylate is widely prescribed and has many side effects. Previous studies suggest that targeting neuronal receptors/ion channels is one of the pathways by which salicylate causes side effects in the nervous system. The present study aimed to investigate the functional action of salicylate on glycine receptors at a molecular level. Experimental approach: Whole-cell patch-clamp and site-directed mutagenesis were deployed to examine the effects of salicylate on the currents mediated by native glycine receptors in cultured neurones of rat inferior colliculus and by glycine receptors expressed in HEK293T cells. Key results: Salicylate effectively inhibited the maximal current mediated by native glycine receptors without altering the EC50 and the Hill coefficient, demonstrating a non-competitive action of salicylate. Only when applied simultaneously with glycine and extracellularly, could salicylate produce this antagonism. In HEK293T cells transfected with either α1-, α2-, α3-, α1β-, α2β- or α3β-glycine receptors, salicylate only inhibited the current mediated by those receptors that contained the α1-subunit. A single site mutation of I240V in the α1-subunit abolished inhibition by salicylate. Conclusions and implications: Salicylate is a non-competitive antagonist specifically on glycine receptors containing α1-subunits. This action critically involves the isoleucine-240 in the first transmembrane segment of the α1-subunit. Our findings may increase our understanding of the receptors involved in the side effects of salicylate on the central nervous system, such as seizures and tinnitus. PMID:19594751

Angiotensin II attenuates NMDA receptor-mediated neuronal cell death and prevents the associated reduction in Bcl-2 expression.

PubMed

Schelman, William R; Andres, Robert; Ferguson, Paul; Orr, Brent; Kang, Evan; Weyhenmeyer, James A

2004-09-10

While angiotensin II (Ang II) plays a major role in the regulation of blood pressure, fluid homeostasis and neuroendocrine function, recent studies have also implicated the peptide hormone in cell growth, differentiation and apoptosis. In support of this, we have previously demonstrated that Ang II attenuates N-methyl-D-aspartate (NMDA) receptor signaling [Molec. Brain Res. 48 (1997) 197]. To further examine the modulatory role of Ang II on NMDA receptor function, we investigated the effect of angiotensin receptor (AT) activation on NMDA-mediated cell death and the accompanying decrease in Bcl-2 expression. The viability of differentiated N1E-115 and NG108-15 neuronal cell lines was reduced following exposure to NMDA in a dose-dependent manner. MTT analysis (mitochondrial integrity) revealed a decrease in cell survival of 49.4+/-12.3% in NG108 cells and 79.9+/-6.8% in N1E cells following treatment with 10 mM NMDA for 20 h. Cytotoxicity in N1E cells was inhibited by the noncompetitive NMDA receptor antagonist, MK-801. Further, NMDA receptor-mediated cell death in NG108 cells was attenuated by treatment with Ang II. The Ang II effect was inhibited by both AT1 and AT2 receptor antagonists, losartan and PD123319, respectively, suggesting that both receptor subtypes may play a role in the survival effect of Ang II. Since it has been shown that activation of NMDA receptors alters the expression of Bcl-2 family proteins, Western blot analysis was performed in N1E cells to determine whether Ang II alters the NMDA-induced changes in Bcl-2 expression. A concentration-dependent decrease of intracellular Bcl-2 protein levels was observed following treatment with NMDA, and this reduction was inhibited by MK801. Addition of Ang II suppressed the NMDA receptor-mediated reduction in Bcl-2. The Ang II effect on NMDA-mediated changes in Bcl-2 levels was blocked by PD123319, but was not significantly changed by losartan, suggesting AT2 receptor specificity. Taken together, these

Asymmetrical Macromolecular Complex Formation of Lysophosphatidic Acid Receptor 2 (LPA2) Mediates Gradient Sensing in Fibroblasts*

PubMed Central

Ren, Aixia; Moon, Changsuk; Zhang, Weiqiang; Sinha, Chandrima; Yarlagadda, Sunitha; Arora, Kavisha; Wang, Xusheng; Yue, Junming; Parthasarathi, Kaushik; Heil-Chapdelaine, Rick; Tigyi, Gabor; Naren, Anjaparavanda P.

2014-01-01

Chemotactic migration of fibroblasts toward growth factors relies on their capacity to sense minute extracellular gradients and respond to spatially confined receptor-mediated signals. Currently, mechanisms underlying the gradient sensing of fibroblasts remain poorly understood. Using single-particle tracking methodology, we determined that a lysophosphatidic acid (LPA) gradient induces a spatiotemporally restricted decrease in the mobility of LPA receptor 2 (LPA2) on chemotactic fibroblasts. The onset of decreased LPA2 mobility correlates to the spatial recruitment and coupling to LPA2-interacting proteins that anchor the complex to the cytoskeleton. These localized PDZ motif-mediated macromolecular complexes of LPA2 trigger a Ca2+ puff gradient that governs gradient sensing and directional migration in response to LPA. Disruption of the PDZ motif-mediated assembly of the macromolecular complex of LPA2 disorganizes the gradient of Ca2+ puffs, disrupts gradient sensing, and reduces the directional migration of fibroblasts toward LPA. Our findings illustrate that the asymmetric macromolecular complex formation of chemoattractant receptors mediates gradient sensing and provides a new mechanistic basis for models to describe gradient sensing of fibroblasts. PMID:25542932

Dopamine D3 Receptors Mediate the Discriminative Stimulus Effects of Quinpirole in Free-Feeding Rats

PubMed Central

Baladi, Michelle G.; Newman, Amy H.

2010-01-01

The discriminative stimulus effects of dopamine (DA) D3/D2 receptor agonists are thought to be mediated by D2 receptors. To maintain responding, access to food is often restricted, which can alter neurochemical and behavioral effects of drugs acting on DA systems. This study established stimulus control with quinpirole in free-feeding rats and tested the ability of agonists to mimic and antagonists to attenuate the effects of quinpirole. The same antagonists were studied for their ability to attenuate quinpirole-induced yawning and hypothermia. DA receptor agonists apomorphine and lisuride, but not amphetamine and morphine, occasioned responding on the quinpirole lever. The discriminative stimulus effects of quinpirole were attenuated by the D3 receptor-selective antagonist N-{4-[4-(2,3-dichlorophenyl)-piperazin-1-yl]-trans-but-2-enyl}-4-pyridine-2-yl-benzamide HCl (PG01037) and the nonselective D3/D2 receptor antagonist raclopride, but not by the D2 receptor-selective antagonist 3-[4-(4-chlorophenyl)-4-hydroxypiperidin-1-yl]methyl-1H-indole (L-741,626); the potencies of PG01037 and raclopride to antagonize this effect of quinpirole paralleled their potencies to antagonize the ascending limb of the quinpirole yawning dose-response curve (thought to be mediated by D3 receptors). L-741,626 selectively antagonized the descending limb of the quinpirole yawning dose-response curve, and both L-741,626 and raclopride, but not PG01037, antagonized the hypothermic effects of quinpirole (thought to be mediated by D2 receptors). Food restriction (10 g/day/7 days) significantly decreased quinpirole-induced yawning without affecting the quinpirole discrimination. Many discrimination studies on DA receptor agonists use food-restricted rats; together with those studies, the current experiment using free-feeding rats suggests that feeding conditions affecting the behavioral effects of direct-acting DA receptor agonists might also have an impact on the effects of indirect

Dopamine D3 receptors mediate the discriminative stimulus effects of quinpirole in free-feeding rats.

PubMed

Baladi, Michelle G; Newman, Amy H; France, Charles P

2010-01-01

The discriminative stimulus effects of dopamine (DA) D3/D2 receptor agonists are thought to be mediated by D2 receptors. To maintain responding, access to food is often restricted, which can alter neurochemical and behavioral effects of drugs acting on DA systems. This study established stimulus control with quinpirole in free-feeding rats and tested the ability of agonists to mimic and antagonists to attenuate the effects of quinpirole. The same antagonists were studied for their ability to attenuate quinpirole-induced yawning and hypothermia. DA receptor agonists apomorphine and lisuride, but not amphetamine and morphine, occasioned responding on the quinpirole lever. The discriminative stimulus effects of quinpirole were attenuated by the D3 receptor-selective antagonist N-{4-[4-(2,3-dichlorophenyl)-piperazin-1-yl]-trans-but-2-enyl}-4-pyridine-2-yl-benzamide HCl (PG01037) and the nonselective D3/D2 receptor antagonist raclopride, but not by the D2 receptor-selective antagonist 3-[4-(4-chlorophenyl)-4-hydroxypiperidin-1-yl]methyl-1H-indole (L-741,626); the potencies of PG01037 and raclopride to antagonize this effect of quinpirole paralleled their potencies to antagonize the ascending limb of the quinpirole yawning dose-response curve (thought to be mediated by D3 receptors). L-741,626 selectively antagonized the descending limb of the quinpirole yawning dose-response curve, and both L-741,626 and raclopride, but not PG01037, antagonized the hypothermic effects of quinpirole (thought to be mediated by D2 receptors). Food restriction (10 g/day/7 days) significantly decreased quinpirole-induced yawning without affecting the quinpirole discrimination. Many discrimination studies on DA receptor agonists use food-restricted rats; together with those studies, the current experiment using free-feeding rats suggests that feeding conditions affecting the behavioral effects of direct-acting DA receptor agonists might also have an impact on the effects of indirect

Glutamate Receptors within the Mesolimbic Dopamine System Mediate Alcohol Relapse Behavior.

PubMed

Eisenhardt, Manuela; Leixner, Sarah; Luján, Rafael; Spanagel, Rainer; Bilbao, Ainhoa

2015-11-25

Glutamatergic input within the mesolimbic dopamine (DA) pathway plays a critical role in the development of addictive behavior. Although this is well established for some drugs of abuse, it is not known whether glutamate receptors within the mesolimbic system are involved in mediating the addictive properties of chronic alcohol use. Here we evaluated the contribution of mesolimbic NMDARs and AMPARs in mediating alcohol-seeking responses induced by environmental stimuli and relapse behavior using four inducible mutant mouse lines lacking the glutamate receptor genes Grin1 or Gria1 in either DA transporter (DAT) or D1R-expressing neurons. We first demonstrate the lack of GluN1 or GluA1 in either DAT- or D1R-expressing neurons in our mutant mouse lines by colocalization studies. We then show that GluN1 and GluA1 receptor subunits within these neuronal subpopulations mediate the alcohol deprivation effect, while having no impact on context- plus cue-induced reinstatement of alcohol-seeking behavior. We further validated these results pharmacologically by demonstrating similar reductions in the alcohol deprivation effect after infusion of the NMDAR antagonist memantine into the nucleus accumbens and ventral tegmental area of control mice, and a rescue of the mutant phenotype via pharmacological potentiation of AMPAR activity using aniracetam. In conclusion, dopamine neurons as well as D1R-expressing medium spiny neurons and their glutamatergic inputs via NMDARs and AMPARs act in concert to influence relapse responses. These results provide a neuroanatomical and molecular substrate for relapse behavior and emphasize the importance of glutamatergic drugs in modulating relapse behavior. Here we provide genetic and pharmacological evidence that glutamate receptors within the mesolimbic dopamine system play an essential role in alcohol relapse. Using various inducible and site-specific transgenic mouse models and pharmacological validation experiments, we show that critical

Influence of ER leak on resting cytoplasmic Ca2+ and receptor-mediated Ca2+ signalling in human macrophage.

PubMed

Layhadi, Janice A; Fountain, Samuel J

2017-06-03

Mechanisms controlling endoplasmic reticulum (ER) Ca 2+ homeostasis are important regulators of resting cytoplasmic Ca 2+ concentration ([Ca 2+ ] cyto ) and receptor-mediated Ca 2+ signalling. Here we investigate channels responsible for ER Ca 2+ leak in THP-1 macrophage and human primary macrophage. In the absence of extracellular Ca 2+ we employ ionomycin action at the plasma membrane to stimulate ER Ca 2+ leak. Under these conditions ionomycin elevates [Ca 2+ ] cyto revealing a Ca 2+ leak response which is abolished by thapsigargin. IP 3 receptors (Xestospongin C, 2-APB), ryanodine receptors (dantrolene), and translocon (anisomycin) inhibition facilitated ER Ca 2+ leak in model macrophage, with translocon inhibition also reducing resting [Ca 2+ ] cyto . In primary macrophage, translocon inhibition blocks Ca 2+ leak but does not influence resting [Ca 2+ ] cyto . We identify a role for translocon-mediated ER Ca 2+ leak in receptor-mediated Ca 2+ signalling in both model and primary human macrophage, whereby the Ca 2+ response to ADP (P2Y receptor agonist) is augmented following anisomycin treatment. In conclusion, we demonstrate a role of ER Ca 2+ leak via the translocon in controlling resting cytoplasmic Ca 2+ in model macrophage and receptor-mediated Ca 2+ signalling in model macrophage and primary macrophage. Copyright © 2017 Elsevier Inc. All rights reserved.

Steroid receptor coactivator-1 mediates estrogenic actions to prevent body weight gain in female mice

USDA-ARS?s Scientific Manuscript database

Estrogen receptor-alpha (ERalpha) expressed by hypothalamic proopiomelanocortin and steroidogenic factor-1 neurons largely mediates the antiobesity effects of estrogens in females. However, the critical molecular events that are coupled to ERalpha and mediate estrogenic effects on energy balance rem...

Regulated internalization of NMDA receptors drives PKD1-mediated suppression of the activity of residual cell-surface NMDA receptors.

PubMed

Fang, Xiao-Qian; Qiao, Haifa; Groveman, Bradley R; Feng, Shuang; Pflueger, Melissa; Xin, Wen-Kuan; Ali, Mohammad K; Lin, Shuang-Xiu; Xu, Jindong; Duclot, Florian; Kabbaj, Mohamed; Wang, Wei; Ding, Xin-Sheng; Santiago-Sim, Teresa; Jiang, Xing-Hong; Salter, Michael W; Yu, Xian-Min

2015-11-19

Constitutive and regulated internalization of cell surface proteins has been extensively investigated. The regulated internalization has been characterized as a principal mechanism for removing cell-surface receptors from the plasma membrane, and signaling to downstream targets of receptors. However, so far it is still not known whether the functional properties of remaining (non-internalized) receptor/channels may be regulated by internalization of the same class of receptor/channels. The N-methyl-D-aspartate receptor (NMDAR) is a principal subtype of glutamate-gated ion channel and plays key roles in neuronal plasticity and memory functions. NMDARs are well-known to undergo two types of regulated internalization - homologous and heterologous, which can be induced by high NMDA/glycine and DHPG, respectively. In the present work, we investigated effects of regulated NMDAR internalization on the activity of residual cell-surface NMDARs and neuronal functions. In electrophysiological experiments we discovered that the regulated internalization of NMDARs not only reduced the number of cell surface NMDARs but also caused an inhibition of the activity of remaining (non-internalized) surface NMDARs. In biochemical experiments we identified that this functional inhibition of remaining surface NMDARs was mediated by increased serine phosphorylation of surface NMDARs, resulting from the activation of protein kinase D1 (PKD1). Knockdown of PKD1 did not affect NMDAR internalization but prevented the phosphorylation and inhibition of remaining surface NMDARs and NMDAR-mediated synaptic functions. These data demonstrate a novel concept that regulated internalization of cell surface NMDARs not only reduces the number of NMDARs on the cell surface but also causes an inhibition of the activity of remaining surface NMDARs through intracellular signaling pathway(s). Furthermore, modulating the activity of remaining surface receptors may be an effective approach for treating receptor

BK channel β1 subunits regulate airway contraction secondary to M2 muscarinic acetylcholine receptor mediated depolarization

PubMed Central

Semenov, Iurii; Wang, Bin; Herlihy, Jeremiah T; Brenner, Robert

2011-01-01

Abstract The large conductance calcium- and voltage-activated potassium channel (BK channel) and its smooth muscle-specific β1 subunit regulate excitation–contraction coupling in many types of smooth muscle cells. However, the relative contribution of BK channels to control of M2- or M3-muscarinic acetylcholine receptor mediated airway smooth muscle contraction is poorly understood. Previously, we showed that knockout of the BK channel β1 subunit enhances cholinergic-evoked trachea contractions. Here, we demonstrate that the enhanced contraction of the BK β1 knockout can be ascribed to a defect in BK channel opposition of M2 receptor-mediated contractions. Indeed, the enhanced contraction of β1 knockout is eliminated by specific M2 receptor antagonism. The role of BK β1 to oppose M2 signalling is evidenced by a greater than fourfold increase in the contribution of L-type voltage-dependent calcium channels to contraction that otherwise does not occur with M2 antagonist or with β1 containing BK channels. The mechanism through which BK channels oppose M2-mediated recruitment of calcium channels is through a negative shift in resting voltage that offsets, rather than directly opposes, M2-mediated depolarization. The negative shift in resting voltage is reduced to similar extents by BK β1 knockout or by paxilline block of BK channels. Normalization of β1 knockout baseline voltage with low external potassium eliminated the enhanced M2-receptor mediated contraction. In summary, these findings indicate that an important function of BK/β1 channels is to oppose cholinergic M2 receptor-mediated depolarization and activation of calcium channels by restricting excitation–contraction coupling to more negative voltage ranges. PMID:21300746

Receptor-like cytoplasmic kinases are pivotal components in pattern recognition receptor-mediated signaling in plant immunity.

PubMed

Yamaguchi, Koji; Yamada, Kenta; Kawasaki, Tsutomu

2013-10-01

Innate immunity is generally initiated with recognition of conserved pathogen-associated molecular patterns (PAMPs). PAMPs are perceived by pattern recognition receptors (PRRs), leading to activation of a series of immune responses, including the expression of defense genes, ROS production and activation of MAP kinase. Recent progress has indicated that receptor-like cytoplasmic kinases (RLCKs) are directly activated by ligand-activated PRRs and initiate pattern-triggered immunity (PTI) in both Arabidopsis and rice. To suppress PTI, pathogens inhibit the RLCKs by many types of effectors, including AvrAC, AvrPphB and Xoo1488. In this review, we summarize recent advances in RLCK-mediated PTI in plants.

Med1 subunit of the mediator complex in nuclear receptor-regulated energy metabolism, liver regeneration, and hepatocarcinogenesis.

PubMed

Jia, Yuzhi; Viswakarma, Navin; Reddy, Janardan K

2014-01-01

Several nuclear receptors regulate diverse metabolic functions that impact on critical biological processes, such as development, differentiation, cellular regeneration, and neoplastic conversion. In the liver, some members of the nuclear receptor family, such as peroxisome proliferator-activated receptors (PPARs), constitutive androstane receptor (CAR), farnesoid X receptor (FXR), liver X receptor (LXR), pregnane X receptor (PXR), glucocorticoid receptor (GR), and others, regulate energy homeostasis, the formation and excretion of bile acids, and detoxification of xenobiotics. Excess energy burning resulting from increases in fatty acid oxidation systems in liver generates reactive oxygen species, and the resulting oxidative damage influences liver regeneration and liver tumor development. These nuclear receptors are important sensors of exogenous activators as well as receptor-specific endogenous ligands. In this regard, gene knockout mouse models revealed that some lipid-metabolizing enzymes generate PPARα-activating ligands, while others such as ACOX1 (fatty acyl-CoA oxidase1) inactivate these endogenous PPARα activators. In the absence of ACOX1, the unmetabolized ACOX1 substrates cause sustained activation of PPARα, and the resulting increase in energy burning leads to hepatocarcinogenesis. Ligand-activated nuclear receptors recruit the multisubunit Mediator complex for RNA polymerase II-dependent gene transcription. Evidence indicates that the Med1 subunit of the Mediator is essential for PPARα, PPARγ, CAR, and GR signaling in liver. Med1 null hepatocytes fail to respond to PPARα activators in that these cells do not show induction of peroxisome proliferation and increases in fatty acid oxidation enzymes. Med1-deficient hepatocytes show no increase in cell proliferation and do not give rise to liver tumors. Identification of nuclear receptor-specific coactivators and Mediator subunits should further our understanding of the complexities of metabolic

Receptor-Mediated Drug Delivery to Macrophages in Chemotherapy of Leishmaniasis

NASA Astrophysics Data System (ADS)

Mukhopadhyay, Amitabha; Chaudhuri, Gautam; Arora, Sunil K.; Sehgal, Shobha; Basu, Sandip K.

1989-05-01

Methotrexate coupled to maleylated bovine serum albumin was taken up efficiently through the ``scavenger'' receptors present on macrophages and led to selective killing of intracellular Leishmania mexicana amazonensis amastigotes in cultured hamster peritoneal macrophages. The drug conjugate was nearly 100 times as effective as free methotrexate in eliminating the intracellular parasites. Furthermore, in a model of experimental cutaneous leishmaniasis in hamsters, the drug conjugate brought about more than 90% reduction in the size of footpad lesions within 11 days. In contrast, the free drug at a similar concentration did not significantly affect lesion size. These studies demonstrate the potential of receptor-mediated drug delivery in the therapy of macrophage-associated diseases.

Selective dopamine receptor 4 activation mediates the hippocampal neuronal calcium response via IP3 and ryanodine receptors.

PubMed

Wang, Ya-Li; Wang, Jian-Gang; Guo, Fang-Li; Gao, Xia-Huan; Zhao, Dan-Dan; Zhang, Lin; Wang, Jian-Zhi; Lu, Cheng-Biao

2017-09-01

Intracellular calcium is a key factor in most cellular processes, including cell growth, differentiation, proliferation and neurotransmitter release. Dopamine (DA) mediates synaptic transmission by regulating the intracellular calcium content. It is not clear, however, which specific subunit of the DA receptor contributes to DA modulation of intracellular calcium content changes. Through the traditional technique of Fura-2 calcium imaging, this study demonstrated that the DA can induce transient calcium in cultured hippocampal neurons and that this response can be mimicked by a selective dopamine receptor 4 (DR4) agonist PD168077 (PD). PD-induced calcium transience can be blocked by a calcium chelator, such as BAPTA-AM, or by pre-treatment of neurons with thapsigargin, a IP 3 receptor antagonist, or a micromolar concentration of ryanodine, a ryanodine receptor (RyR) antagonist. However PD-induced calcium transience cannot be blocked by pre-treatment of neurons with a free-calcium medium or a cocktail of NMDA receptor, L-type calcium channel and alpha7 nicotinic acetylcholine receptor blockers. These results indicate that the calcium response induced by DR4 activation is mainly through activation of IP 3 receptor in internal stores, which is likely to contribute to the DA modulation of synaptic transmission and cognitive function. Copyright © 2017. Published by Elsevier B.V.

Effects of Chronic Alcohol Exposure on Kainate Receptor-Mediated Neurotransmission in the Hippocampus

DTIC Science & Technology

2004-09-01

hippocampus and hypothalamus. J Neurosci Res 63:200-208. Thorpe GH et al. (1989) Chemiluminescent enzyme immunoassay of alpha - fetoprotein based on an...cells. Proc Natl Acad Sci USA 83:6213-6215. Montpied Pet al. (1991) Prolonged ethanol inhalation decreases gamma-aminobutyric acidA receptor alpha ...potentials mediated via alpha -bungarotoxin-sensitive nico- tinic acetylcholine receptors in rat hippocampal interneurons. J Neurosci 18:8228- 8235

Mechanism of HSV infection through soluble adapter-mediated virus bridging to the EGF receptor

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

Nakano, Kenji, E-mail: kenakano@med.kyushu-u.ac.j; Kobayashi, Masatoshi; Nakamura, Kei-ichiro

2011-04-25

Herpes simplex virus entry into cells requires the binding of envelope glycoprotein D (gD) to an entry receptor. Depending on the cell, entry occurs by different mechanisms, including fusion at the cell surface or endocytosis. Here we examined the entry mechanism through a non-HSV receptor mediated by a soluble bi-specific adapter protein composed of recognition elements for gD and the EGF receptor (EGFR). Virus entered into endosomes using either EGF or an EGFR-specific single chain antibody (scFv) for receptor recognition. Infection was less efficient with the EGF adapter which could be attributed to its weaker binding to a viral gD.more » Infection mediated by the scFv adapter was pH sensitive, indicating that gD-EGFR bridging alone was insufficient for capsid release from endosomes. We also show that the scFv adapter enhanced infection of EGFR-expressing tumor tissue in vivo. Our results indicate that adapters may retarget HSV infection without drastically changing the entry mechanism.« less

Slow synaptic transmission mediated by TRPV1 channels in CA3 interneurons of the hippocampus.

PubMed

Eguchi, Noriomi; Hishimoto, Akitoyo; Sora, Ichiro; Mori, Masahiro

2016-03-11

Metabotropic glutamate receptors (mGluRs) modulate various neuronal functions in the central nervous system. Many studies reported that mGluRs have linkages to neuronal disorders such as schizophrenia and autism related disorders, indicating that mGluRs are involved in critical functions of the neuronal circuits. To study this possibility further, we recorded mGluR-induced synaptic responses in the interneurons of the CA3 stratum radiatum using rat hippocampal organotypic slice cultures. Electrical stimulation in the CA3 pyramidal cell layer evoked a slow inward current in the interneurons at a holding potential of -70mV in the presence of antagonists for AMPA/kainate receptors, NMDA receptors, GABAA receptors and GABAB receptors. The slow inward current was blocked in the absence of extracellular calcium, suggesting that this was a synaptic response. The slow excitatory postsynaptic current (EPSC) reversed near 0mV, reflecting an increase in a non-selective cationic conductance. The slow EPSC is mediated by group I mGluRs, as it was blocked by AP3, a group I mGluR antagonist. Neither a calcium chelator BAPTA nor a phospholipase C (PLC) inhibitor U73122 affected the slow EPSC. La(3+), a general TRP channel blocker or capsazepine, a selective TRPV1 channel antagonist significantly suppressed the slow EPSC. DHPG, a selective group I mGluRs agonist induced an inward current, which was suppressed by capsazepine. These results indicate that in the interneurons of the hippocampal CA3 stratum radiatum group I mGluRs activate TRPV1 channels independently of PLC and intracellular Ca(2+), resulting in the slow EPSC in the interneurons. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Activation of innate antiviral immune response via double-stranded RNA-dependent RLR receptor-mediated necroptosis

PubMed Central

Wang, Wei; Wang, Wei-Hua; Azadzoi, Kazem M.; Su, Ning; Dai, Peng; Sun, Jianbin; Wang, Qin; Liang, Ping; Zhang, Wentao; Lei, Xiaoying; Yan, Zhen; Yang, Jing-Hua

2016-01-01

Viruses induce double-stranded RNA (dsRNA) in the host cells. The mammalian system has developed dsRNA-dependent recognition receptors such as RLRs that recognize the long stretches of dsRNA as PAMPs to activate interferon-mediated antiviral pathways and apoptosis in severe infection. Here we report an efficient antiviral immune response through dsRNA-dependent RLR receptor-mediated necroptosis against infections from different classes of viruses. We demonstrated that virus-infected A549 cells were efficiently killed in the presence of a chimeric RLR receptor, dsCARE. It measurably suppressed the interferon antiviral pathway but promoted IL-1β production. Canonical cell death analysis by morphologic assessment, phosphatidylserine exposure, caspase cleavage and chemical inhibition excluded the involvement of apoptosis and consistently suggested RLR receptor-mediated necroptosis as the underlying mechanism of infected cell death. The necroptotic pathway was augmented by the formation of RIP1-RIP3 necrosome, recruitment of MLKL protein and the activation of cathepsin D. Contributing roles of RIP1 and RIP3 were confirmed by gene knockdown. Furthermore, the necroptosis inhibitor necrostatin-1 but not the pan-caspase inhibitor zVAD impeded dsCARE-dependent infected cell death. Our data provides compelling evidence that the chimeric RLR receptor shifts the common interferon antiviral responses of infected cells to necroptosis and leads to rapid death of the virus-infected cells. This mechanism could be targeted as an efficient antiviral strategy. PMID:26935990

M1 muscarinic receptor activation mediates cell death in M1-HEK293 cells.

PubMed

Graham, E Scott; Woo, Kerhan K; Aalderink, Miranda; Fry, Sandie; Greenwood, Jeffrey M; Glass, Michelle; Dragunow, Mike

2013-01-01

HEK293 cells have been used extensively to generate stable cell lines to study G protein-coupled receptors, such as muscarinic acetylcholine receptors (mAChRs). The activation of M1 mAChRs in various cell types in vitro has been shown to be protective. To further investigate M1 mAChR-mediated cell survival, we generated stable HEK293 cell-lines expressing the human M1 mAChR. M1 mAChRs were efficiently expressed at the cell surface and efficiently internalised within 1 h by carbachol. Carbachol also induced early signalling cascades similar to previous reports. Thus, ectopically expressed M1 receptors behaved in a similar fashion to the native receptor over short time periods of analysis. However, substantial cell death was observed in HEK293-M1 cells within 24 h after carbachol application. Death was only observed in HEK cells expressing M1 receptors and fully blocked by M1 antagonists. M1 mAChR-stimulation mediated prolonged activation of the MEK-ERK pathway and resulted in prolonged induction of the transcription factor EGR-1 (>24 h). Blockade of ERK signalling with U0126 did not reduce M1 mAChR-mediated cell-death significantly but inhibited the acute induction of EGR-1. We investigated the time-course of cell death using time-lapse microscopy and xCELLigence technology. Both revealed the M1 mAChR cytotoxicity occurs within several hours of M1 activation. The xCELLigence assay also confirmed that the ERK pathway was not involved in cell-death. Interestingly, the MEK blocker did reduce carbachol-mediated cleaved caspase 3 expression in HEK293-M1 cells. The HEK293 cell line is a widely used pharmacological tool for studying G-protein coupled receptors, including mAChRs. Our results highlight the importance of investigating the longer term fate of these cells in short term signalling studies. Identifying how and why activation of the M1 mAChR signals apoptosis in these cells may lead to a better understanding of how mAChRs regulate cell-fate decisions.

Photolysis of Caged Ca2+ But Not Receptor-Mediated Ca2+ Signaling Triggers Astrocytic Glutamate Release

PubMed Central

Smith, Nathan A.; Xu, Qiwu; Goldman, Siri; Peng, Weiguo; Huang, Jason H.; Takano, Takahiro; Nedergaard, Maiken

2013-01-01

Astrocytes in hippocampal slices can dynamically regulate synaptic transmission in a process mediated by increases in intracellular Ca2+. However, it is debated whether astrocytic Ca2+ signals result in release of glutamate. We here compared astrocytic Ca2+ signaling triggered by agonist exposure versus photolysis side by side. Using transgenic mice in which astrocytes selectively express the MrgA1 receptor, we found that receptor-mediated astrocytic Ca2+ signaling consistently triggered neuronal hyperpolarization and decreased the frequency of miniature excitatory postsynaptic currents (EPSCs). In contrast, photolysis of caged Ca2+ (o-nitrophenyl–EGTA) in astrocytes led to neuronal depolarization and increased the frequency of mEPSCs through a metabotropic glutamate receptor-mediated pathway. Analysis of transgenic mice in which astrocytic vesicular release is suppressed (dominant-negative SNARE mice) and pharmacological manipulations suggested that glutamate is primarily released by opening of anion channels rather than exocytosis. Combined, these studies show that photolysis but not by agonists induced astrocytic Ca2+ signaling triggers glutamate release. PMID:24174673

Mannose Receptor Mediates the Immune Response to Ganoderma atrum Polysaccharides in Macrophages.

PubMed

Li, Wen-Juan; Tang, Xiao-Fang; Shuai, Xiao-Xue; Jiang, Cheng-Jia; Liu, Xiang; Wang, Le-Feng; Yao, Yu-Fei; Nie, Shao-Ping; Xie, Ming-Yong

2017-01-18

The ability of mannose receptor (MR) to recognize the carbohydrate structures is well-established. Here, we reported that MR was crucial for the immune response to a Ganoderma atrum polysaccharide (PSG-1), as evidenced by elevation of MR in association with increase of phagocytosis and concentrations of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in normal macrophages. Elevation of MR triggered by PSG-1 also led to control lipopolysaccharide (LPS)-triggered inflammatory response via the increase of interleukin-10 (IL-10) and inhibition of phagocytosis and IL-1β. Anti-MR antibody partly attenuated PSG-1-mediated anti-inflammatory responses, while it could not affect TNF-α secretion, suggesting that another receptor was involved in PSG-1-triggered immunomodulatory effects. MR and toll-like receptor (TLR)4 coordinated the influences on the TLR4-mediated signaling cascade by the nuclear factor-κB (NF-κB) pathway in LPS-stimulated macrophages subjected to PSG-1. Collectively, immune response to PSG-1 required recognition by MR in macrophages. The NF-κB pathway served as a central role for the coordination of MR and TLR4 to elicit immune response to PSG-1.

Regulation of androgen receptor and histone deacetylase 1 by Mdm2-mediated ubiquitylation.

PubMed

Gaughan, Luke; Logan, Ian R; Neal, David E; Robson, Craig N

2005-01-01

The androgen receptor (AR) is a member of the nuclear hormone receptor family of transcription factors and plays a critical role in regulating the expression of genes involved in androgen-dependent and -independent tumour formation. Regulation of the AR is achieved by alternate binding of either histone acetyltransferase (HAT)-containing co-activator proteins, or histone deacetylase 1 (HDAC1). Factors that control AR stability may also constitute an important regulatory mechanism, a notion that has been confirmed with the finding that the AR is a direct target for Mdm2-mediated ubiquitylation and proteolysis. Using chromatin immunoprecipitation (ChIP) and re-ChIP analyses, we show that Mdm2 associates with AR and HDAC1 at the active androgen-responsive PSA promoter in LNCaP prostate cancer cells. Furthermore, we demonstrate that Mdm2-mediated modification of AR and HDAC1 catalyses protein destabilization and attenuates AR sactivity, suggesting that ubiquitylation of the AR and HDAC1 may constitute an additional mechanism for regulating AR function. We also show that HDAC1 and Mdm2 function co-operatively to reduce AR-mediated transcription that is attenuated by the HAT activity of the AR co-activator Tip60, suggesting interplay between acetylation status and receptor ubiquitylation in AR regulation. In all, our data indicates a novel role for Mdm2 in regulating components of the AR transcriptosome.

Multiple receptor subtypes mediate the effects of serotonin on rat subfornical organ neurons

NASA Technical Reports Server (NTRS)

Scrogin, K. E.; Johnson, A. K.; Schmid, H. A.

1998-01-01

The subfornical organ (SFO) receives significant serotonergic innervation. However, few reports have examined the functional effects of serotonin on SFO neurons. This study characterized the effects of serotonin on spontaneously firing SFO neurons in the rat brain slice. Of 31 neurons tested, 80% responded to serotonin (1-100 microM) with either an increase (n = 15) or decrease (n = 10) in spontaneous activity. Responses to serotonin were dose dependent and persisted after synaptic blockade. Excitatory responses could also be mimicked by the 5-hydroxytryptamine (5-HT)2A/2C receptor agonist 2,5-dimethoxy-4-iodoamphetamine (DOI; 1-10 microM) and could be blocked by the 5-HT2A/2C-receptor antagonist LY-53,857 (10 microM). LY-53,857 unmasked inhibitory responses to serotonin in 56% of serotonin-excited cells tested. Serotonin-inhibited cells were also inhibited by the 5-HT1A-receptor agonist 8-hydroxy-2(di-n-propylamino)tetralin (8-OH-DPAT; 1-10 microM; n = 7). The data indicate that SFO neurons are responsive to serotonin via postsynaptic activation of multiple receptor subtypes. The results suggest that excitatory responses to serotonin are mediated by 5-HT2A or 5-HT2C receptors and that inhibitory responses may be mediated by 5-HT1A receptors. In addition, similar percentages of serotonin-excited and -inhibited cells were also sensitive to ANG II. As such the functional relationship between serotonin and ANG II in the SFO remains unclear.

Multiple receptor subtypes mediate the effects of serotonin on rat subfornical organ neurons.

PubMed

Scrogin, K E; Johnson, A K; Schmid, H A

1998-12-01

The subfornical organ (SFO) receives significant serotonergic innervation. However, few reports have examined the functional effects of serotonin on SFO neurons. This study characterized the effects of serotonin on spontaneously firing SFO neurons in the rat brain slice. Of 31 neurons tested, 80% responded to serotonin (1-100 microM) with either an increase (n = 15) or decrease (n = 10) in spontaneous activity. Responses to serotonin were dose dependent and persisted after synaptic blockade. Excitatory responses could also be mimicked by the 5-hydroxytryptamine (5-HT)2A/2C receptor agonist 2,5-dimethoxy-4-iodoamphetamine (DOI; 1-10 microM) and could be blocked by the 5-HT2A/2C-receptor antagonist LY-53,857 (10 microM). LY-53,857 unmasked inhibitory responses to serotonin in 56% of serotonin-excited cells tested. Serotonin-inhibited cells were also inhibited by the 5-HT1A-receptor agonist 8-hydroxy-2(di-n-propylamino)tetralin (8-OH-DPAT; 1-10 microM; n = 7). The data indicate that SFO neurons are responsive to serotonin via postsynaptic activation of multiple receptor subtypes. The results suggest that excitatory responses to serotonin are mediated by 5-HT2A or 5-HT2C receptors and that inhibitory responses may be mediated by 5-HT1A receptors. In addition, similar percentages of serotonin-excited and -inhibited cells were also sensitive to ANG II. As such the functional relationship between serotonin and ANG II in the SFO remains unclear.

Copolymers of poly-L-lysine with serine and tryptophan form stable DNA vectors: implications for receptor-mediated gene transfer.

PubMed

Gómez-Valadés, A G; Molas, M; Vidal-Alabró, A; Bermúdez, J; Bartrons, R; Perales, J C

2005-01-20

Inefficient gene transfer and poor stability in physiological medium are important shortcomings for receptor-mediated gene transfer vectors. Here, we evaluate vectors formulated with random copolymers of L-lysine/L-serine (3:1) and L-lysine/L-tryptophan (4:1), focusing on both their biophysical and functional characterization. By means of dynamic light scattering (DLS) and transmission electron microscopy (TEM), we demonstrate that poly-L-lysine (pK), poly-L-lysine-L-tryptophan (pKW) and poly-L-lysine-L-serine (pKS) are able to form compacted, small particles when mixed with plasmid DNA in the absence of salt. Upon dilution in physiological medium, copolymers of both lys/ser and lys/trp do not aggregate, in contrast with poly-L-lysine DNA complexes as determined by scattering, DLS and TEM measurements. Tight packing, as demonstrated by resistance to heparin, SDS and trypsin treatments, is also featured in tryptophan-containing complexes. Successful receptor-mediated endocytosis gene transfer using galactosylated copolymers into cells expressing the asiagloglycoprotein receptor correlated with lack of aggregation. Particles obtained using galactosylated poly-L-lysine-L-tryptophan (Gal-pKW) copolymer demonstrated specific receptor-mediated gene transfer since reporter gene activity dropped in the presence of an excess ligand in the culture medium during transfection. Although copolymers of galactosylated poly-L-lysine-L-serine (Gal-pKS) do not aggregate in the presence of salt, they are not able to internalize in a specific receptor-mediated endocytosis fashion. The introduction of bulky aromatic/hydrophobic (tryptophan) or hydrophillic (serine) moieties into the positively charged vectors allows the compacted particles to disperse into salt-containing medium avoiding salt-induced aggregation. Moreover, tryptophan-containing particles are able to mediate specific gene transfer via receptor-mediated endocytosis.

Upregulation of genes related to bone formation by γ-amino butyric acid and γ-oryzanol in germinated brown rice is via the activation of GABAB-receptors and reduction of serum IL-6 in rats.

PubMed

Muhammad, Sani Ismaila; Maznah, Ismail; Mahmud, Rozi; Zuki, Abu Bakar Zakaria; Imam, Mustapha Umar

2013-01-01

GBR stimulates osteoblastogenesis by upregulation of bone formation genes, possibly via the activation of GABAB receptors and by inhibiting the activity of inflammatory cytokines and reactive oxygen species. Therefore, it could be used effectively in the management of osteoporosis.

Upregulation of genes related to bone formation by γ-amino butyric acid and γ-oryzanol in germinated brown rice is via the activation of GABAB-receptors and reduction of serum IL-6 in rats

PubMed Central

Muhammad, Sani Ismaila; Maznah, Ismail; Mahmud, Rozi; Zuki, Abu Bakar Zakaria; Imam, Mustapha Umar

2013-01-01

-treated groups. Conclusion GABA and ORZ from GBR stimulates osteoblastogenesis by upregulation of bone formation genes, possibly via the activation of GABAB receptors and by inhibiting the activity of inflammatory cytokines and reactive oxygen species. Therefore, it could be used effectively in the management of osteoporosis. PMID:24098073

Orphan nuclear receptor oestrogen-related receptor γ (ERRγ) plays a key role in hepatic cannabinoid receptor type 1-mediated induction of CYP7A1 gene expression

PubMed Central

Zhang, Yaochen; Kim, Don-Kyu; Lee, Ji-Min; Park, Seung Bum; Jeong, Won-IL; Kim, Seong Heon; Lee, In-Kyu; Lee, Chul-Ho; Chiang, John Y.L.; Choi, Hueng-Sik

2017-01-01

Bile acids are primarily synthesized from cholesterol in the liver and have important roles in dietary lipid absorption and cholesterol homoeostasis. Detailed roles of the orphan nuclear receptors regulating cholesterol 7α-hydroxylase (CYP7A1), the rate-limiting enzyme in bile acid synthesis, have not yet been fully elucidated. In the present study, we report that oestrogen-related receptor γ (ERRγ) is a novel transcriptional regulator of CYP7A1 expression. Activation of cannabinoid receptor type 1 (CB1 receptor) signalling induced ERRγ-mediated transcription of the CYP7A1 gene. Overexpression of ERRγ increased CYP7A1 expression in vitro and in vivo, whereas knockdown of ERRγ attenuated CYP7A1 expression. Deletion analysis of the CYP7A1 gene promoter and a ChIP assay revealed an ERRγ -binding site on the CYP7A1 gene promoter. Small heterodimer partner (SHP) inhibited the transcriptional activity of ERRγ and thus regulated CYP7A1 expression. Overexpression of ERRγ led to increased bile acid levels, whereas an inverse agonist of ERRγ, GSK5182, reduced CYP7A1 expression and bile acid synthesis. Finally, GSK5182 significantly reduced hepatic CB1 receptor-mediated induction of CYP7A1 expression and bile acid synthesis in alcohol-treated mice. These results provide the molecular mechanism linking ERRγ and bile acid metabolism. PMID:26348907

Cannabinoid CB1 receptor facilitation of substance P release in the rat spinal cord, measured as neurokinin 1 receptor internalization

PubMed Central

Zhang, Guohua; Chen, Wenling; Lao, Lijun; Marvizón, Juan Carlos G.

2010-01-01

The contribution of CB1 receptors in the spinal cord to cannabinoid analgesia is still unclear. The objective of this study was to investigate the effect of CB1 receptors on substance P release from primary afferent terminals in the spinal cord. Substance P release was measured as NK1 receptor internalization in lamina I neurons. It was induced in spinal cord slices by dorsal root stimulation and in live rats by a noxious stimulus. In spinal cord slices, the CB1 receptor antagonists AM251, AM281 and rimonabant partially but potently inhibited NK1 receptor internalization induced by electrical stimulation of the dorsal root. This was due to an inhibition of substance P release and not of NK1 receptor internalization itself, because AM251 and AM281 did not inhibit NK1 receptor internalization induced by exogenous substance P. The CB1 receptor agonist ACEA increased NK1 receptor internalization evoked by dorsal root stimulation. The effects of AM251 and ACEA cancelled each other. In vivo, AM251 injected intrathecally decreased NK1 receptor internalization in spinal segments L5 and L6 induced by noxious hind paw clamp. Intrathecal AM251 also produced analgesia to radiant heat stimulation of the paw. The inhibition by AM251 of NK1 receptor internalization was reversed by antagonists of μ-opioid and GABAB receptors. This indicates that CB1 receptors facilitate substance P release by inhibiting the release of GABA and opioids next to primary afferent terminals, producing disinhibition. This results in a pronociceptive effect of CB1 receptors in the spinal cord. PMID:20074214

NOP Receptor Mediates Anti-analgesia Induced by Agonist-Antagonist Opioids

PubMed Central

Gear, Robert W.; Bogen, Oliver; Ferrari, Luiz F.; Green, Paul G.; Levine, Jon D.

2014-01-01

Clinical studies have shown that agonist-antagonist opioid analgesics that produce their analgesic effect via action on the kappa-opioid receptor, produce a delayed-onset anti-analgesia in men but not women, an effect blocked by co-administration of a low dose of naloxone. We now report the same time-dependent anti-analgesia and its underlying mechanism in an animal model. Using the Randall-Selitto paw-withdrawal assay in male rats, we found that nalbuphine, pentazocine, and butorphanol each produced analgesia during the first hour followed by anti-analgesia starting at ~90 minutes after administration in males but not females, closely mimicking its clinical effects. As observed in humans, co-administration of nalbuphine with naloxone in a dose ratio of 12.5:1 blocked anti-analgesia but not analgesia. Administration of the highly selective kappa-opioid receptor agonist U69,593 produced analgesia without subsequent anti-analgesia, and confirmed by the failure of the selective kappa antagonist nor-binaltorphimine to block nalbuphine-induced anti-analgesia, indicating that anti-analgesia is not mediated by kappa-opioid receptors. We therefore tested the role of other receptors in nalbuphine anti-analgesia. Nociceptin/orphanin FQ (NOP) and sigma-1 and sigma-2 receptors were chosen on the basis of their known anti-analgesic effects and receptor binding studies. The selective NOP receptor antagonists, JTC801, and J113397, but not the sigma receptor antagonist, BD 1047, antagonized nalbuphine anti-analgesia. Furthermore, the NOP receptor agonist NNC 63-0532 produced anti-analgesia with the same delay in onset observed with the three agonist-antagonists, but without producing preceding analgesia and this anti-analgesia was also blocked by naloxone. These results strongly support the suggestion that clinically used agonist-antagonists act at the NOP receptor to produce anti-analgesia. PMID:24188792

The orphan receptor ALK7 and the Activin receptor ALK4 mediate signaling by Nodal proteins during vertebrate development.

PubMed

Reissmann, E; Jörnvall, H; Blokzijl, A; Andersson, O; Chang, C; Minchiotti, G; Persico, M G; Ibáñez, C F; Brivanlou, A H

2001-08-01

Nodal proteins have crucial roles in mesendoderm formation and left-right patterning during vertebrate development. The molecular mechanisms of signal transduction by Nodal and related ligands, however, are not fully understood. In this paper, we present biochemical and functional evidence that the orphan type I serine/threonine kinase receptor ALK7 acts as a receptor for mouse Nodal and Xenopus Nodal-related 1 (Xnr1). Receptor reconstitution experiments indicate that ALK7 collaborates with ActRIIB to confer responsiveness to Xnr1 and Nodal. Both receptors can independently bind Xnr1. In addition, Cripto, an extracellular protein genetically implicated in Nodal signaling, can independently interact with both Xnr1 and ALK7, and its expression greatly enhances the ability of ALK7 and ActRIIB to respond to Nodal ligands. The Activin receptor ALK4 is also able to mediate Nodal signaling but only in the presence of Cripto, with which it can also interact directly. A constitutively activated form of ALK7 mimics the mesendoderm-inducing activity of Xnr1 in Xenopus embryos, whereas a dominant-negative ALK7 specifically blocks the activities of Nodal and Xnr1 but has little effect on other related ligands. In contrast, a dominant-negative ALK4 blocks all mesoderm-inducing ligands tested, including Nodal, Xnr1, Xnr2, Xnr4, and Activin. In agreement with a role in Nodal signaling, ALK7 mRNA is localized to the ectodermal and organizer regions of Xenopus gastrula embryos and is expressed during early stages of mouse embryonic development. Therefore, our results indicate that both ALK4 and ALK7 can mediate signal transduction by Nodal proteins, although ALK7 appears to be a receptor more specifically dedicated to Nodal signaling.

The orphan receptor ALK7 and the Activin receptor ALK4 mediate signaling by Nodal proteins during vertebrate development

PubMed Central

Reissmann, Eva; Jörnvall, Henrik; Blokzijl, Andries; Andersson, Olov; Chang, Chenbei; Minchiotti, Gabriella; Persico, M. Graziella; Ibáñez, Carlos F.; Brivanlou, Ali H.

2001-01-01

Nodal proteins have crucial roles in mesendoderm formation and left–right patterning during vertebrate development. The molecular mechanisms of signal transduction by Nodal and related ligands, however, are not fully understood. In this paper, we present biochemical and functional evidence that the orphan type I serine/threonine kinase receptor ALK7 acts as a receptor for mouse Nodal and Xenopus Nodal-related 1 (Xnr1). Receptor reconstitution experiments indicate that ALK7 collaborates with ActRIIB to confer responsiveness to Xnr1 and Nodal. Both receptors can independently bind Xnr1. In addition, Cripto, an extracellular protein genetically implicated in Nodal signaling, can independently interact with both Xnr1 and ALK7, and its expression greatly enhances the ability of ALK7 and ActRIIB to respond to Nodal ligands. The Activin receptor ALK4 is also able to mediate Nodal signaling but only in the presence of Cripto, with which it can also interact directly. A constitutively activated form of ALK7 mimics the mesendoderm-inducing activity of Xnr1 in Xenopus embryos, whereas a dominant-negative ALK7 specifically blocks the activities of Nodal and Xnr1 but has little effect on other related ligands. In contrast, a dominant-negative ALK4 blocks all mesoderm-inducing ligands tested, including Nodal, Xnr1, Xnr2, Xnr4, and Activin. In agreement with a role in Nodal signaling, ALK7 mRNA is localized to the ectodermal and organizer regions of Xenopus gastrula embryos and is expressed during early stages of mouse embryonic development. Therefore, our results indicate that both ALK4 and ALK7 can mediate signal transduction by Nodal proteins, although ALK7 appears to be a receptor more specifically dedicated to Nodal signaling. PMID:11485994

VMAT2-mediated neurotransmission from midbrain leptin receptor neurons in feeding regulation

USDA-ARS?s Scientific Manuscript database

Leptin receptors (LepRs) expressed in the midbrain contribute to the action of leptin on feeding regulation. The midbrain neurons release a variety of neurotransmitters including dopamine (DA), glutamate and GABA. However, which neurotransmitter mediates midbrain leptin action on feeding remains unc...

Discrimination between NL1- and NL2-Mediated Nuclear Localization of the Glucocorticoid Receptor

PubMed Central

Savory, Joanne G. A.; Hsu, Brian; Laquian, Ian R.; Giffin, Ward; Reich, Terry; Haché, Robert J. G.; Lefebvre, Yvonne A.

1999-01-01

Glucocorticoid receptor (GR) cycles between a free liganded form that is localized to the nucleus and a heat shock protein (hsp)-immunophilin-complexed, unliganded form that is usually localized to the cytoplasm but that can also be nuclear. In addition, rapid nucleocytoplasmic exchange or shuttling of the receptor underlies its localization. Nuclear import of liganded GR is mediated through a well-characterized sequence, NL1, adjacent to the receptor DNA binding domain and a second, uncharacterized motif, NL2, that overlaps with the ligand binding domain. In this study we report that rapid nuclear import (half-life [t1/2] of 4 to 6 min) of agonist- and antagonist-treated GR and the localization of unliganded, hsp-associated GRs to the nucleus in G0 are mediated through NL1 and correlate with the binding of GR to pendulin/importin α. By contrast, NL2-mediated nuclear transfer of GR occurred more slowly (t1/2 = 45 min to 1 h), was agonist specific, and appeared to be independent of binding to importin α. Together, these results suggest that NL2 mediates the nuclear import of GR through an alternative nuclear import pathway. Nuclear export of GR was inhibited by leptomycin B, suggesting that the transfer of GR to the cytoplasm is mediated through the CRM1-dependent pathway. Inhibition of GR nuclear export by leptomycin B enhanced the nuclear localization of both unliganded, wild-type GR and hormone-treated NL1− GR. These results highlight that the subcellular localization of both liganded and unliganded GRs is determined, at least in part, by a flexible equilibrium between the rates of nuclear import and export. PMID:9891038

Coordinate regulation of estrogen-mediated fibronectin matrix assembly and epidermal growth factor receptor transactivation by the G protein-coupled receptor, GPR30.

PubMed

Quinn, Jeffrey A; Graeber, C Thomas; Frackelton, A Raymond; Kim, Minsoo; Schwarzbauer, Jean E; Filardo, Edward J

2009-07-01

Estrogen promotes changes in cytoskeletal architecture not easily attributed to the biological action of estrogen receptors, ERalpha and ERbeta. The Gs protein-coupled transmembrane receptor, GPR30, is linked to specific estrogen binding and rapid estrogen-mediated release of heparin-bound epidermal growth factor. Using marker rescue and dominant interfering mutant strategies, we show that estrogen action via GPR30 promotes fibronectin (FN) matrix assembly by human breast cancer cells. Stimulation with 17beta-estradiol or the ER antagonist, ICI 182, 780, results in the recruitment of FN-engaged integrin alpha5beta1 conformers to fibrillar adhesions and the synthesis of FN fibrils. Concurrent with this cellular response, GPR30 promotes the formation of Src-dependent, Shc-integrin alpha5beta1 complexes. Function-blocking antibodies directed against integrin alpha5beta1 or soluble Arg-Gly-Asp peptide fragments derived from FN specifically inhibited GPR30-mediated epidermal growth factor receptor transactivation. Estrogen-mediated FN matrix assembly and epidermal growth factor receptor transactivation were similarly disrupted in integrin beta1-deficient GE11 cells, whereas reintroduction of integrin beta1 into GE11 cells restored these responses. Mutant Shc (317Y/F) blocked GPR30-induced FN matrix assembly and tyrosyl phosphorylation of erbB1. Interestingly, relative to recombinant wild-type Shc, 317Y/F Shc was more readily retained in GPR30-induced integrin alpha5beta1 complexes, yet this mutant did not prevent endogenous Shc-integrin alpha5beta1 complex formation. Our results suggest that GPR30 coordinates estrogen-mediated FN matrix assembly and growth factor release in human breast cancer cells via a Shc-dependent signaling mechanism that activates integrin alpha5beta1.

Interleukin-1 receptor-associated kinase-1 plays an essential role for Toll-like receptor (TLR)7- and TLR9-mediated interferon-α induction

PubMed Central

Uematsu, Satoshi; Sato, Shintaro; Yamamoto, Masahiro; Hirotani, Tomonori; Kato, Hiroki; Takeshita, Fumihiko; Matsuda, Michiyuki; Coban, Cevayir; Ishii, Ken J.; Kawai, Taro; Takeuchi, Osamu; Akira, Shizuo

2005-01-01

Toll-like receptors (TLRs) recognize microbial pathogens and trigger innate immune responses. Among TLR family members, TLR7, TLR8, and TLR9 induce interferon (IFN)-α in plasmacytoid dendritic cells (pDCs). This induction requires the formation of a complex consisting of the adaptor MyD88, tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) and IFN regulatory factor (IRF) 7. Here we show an essential role of IL-1 receptor-associated kinase (IRAK)-1 in TLR7- and TLR9-mediated IRF7 signaling pathway. IRAK-1 directly bound and phosphorylated IRF7 in vitro. The kinase activity of IRAK-1 was necessary for transcriptional activation of IRF7. TLR7- and TLR9-mediated IFN-α production was abolished in Irak-1–deficient mice, whereas inflammatory cytokine production was not impaired. Despite normal activation of NF-κB and mitogen-activated protein kinases, IRF7 was not activated by a TLR9 ligand in Irak-1–deficient pDCs. These results indicated that IRAK-1 is a specific regulator for TLR7- and TLR9-mediated IFN-α induction in pDCs. PMID:15767370

Receptor-mediated protein kinase activation and the mechanism of transmembrane signaling in bacterial chemotaxis.

PubMed Central

Liu, Y; Levit, M; Lurz, R; Surette, M G; Stock, J B

1997-01-01

Chemotaxis responses of Escherichia coli and Salmonella are mediated by type I membrane receptors with N-terminal extracytoplasmic sensing domains connected by transmembrane helices to C-terminal signaling domains in the cytoplasm. Receptor signaling involves regulation of an associated protein kinase, CheA. Here we show that kinase activation by a soluble signaling domain construct involves the formation of a large complex, with approximately 14 receptor signaling domains per CheA dimer. Electron microscopic examination of these active complexes indicates a well defined bundle composed of numerous receptor filaments. Our findings suggest a mechanism for transmembrane signaling whereby stimulus-induced changes in lateral packing interactions within an array of receptor-sensing domains at the cell surface perturb an equilibrium between active and inactive receptor-kinase complexes within the cytoplasm. PMID:9405352

P2x7 Receptor-NADPH Oxidase-Axis Mediates Protein radical Formation And Kupffer Cell Activation in Carbon Tetrachloride-Mediated Steatohepatitis in Obese Mice

PubMed Central

Chatterjee, Saurabh; Rana, Ritu; Corbett, Jean; Kadiiska, Maria B.; Goldstein, Joyce; Mason, Ronald P.

2012-01-01

While some studies show that carbon tetrachloride-mediated metabolic oxidative stress exacerbates steatohepatitic-like lesions in obese mice, the redox mechanisms that trigger the innate immune system and accentuate the inflammatory cascade remain unclear. Here we have explored the role of the purinergic receptor P2X7-NADPH oxidase axis as a primary event in recognizing the heightened release of extracellular ATP from CCl4-treated hepatocytes and generating redoxmediated Kupffer cell activation in obese mice. We found that an underlying condition of obesity led to the formation of protein radicals and post-translational nitration, primarily in Kupffer cells, at 24 h post-CCl4 administration. The free radical-mediated oxidation of cellular macromolecules, which was NADPH oxidase- and P2X7 receptor-dependent, correlated well with the release of TNF- α and MCP-2 from Kupffer cells. The Kupffer cells in CCl4-treated mice exhibited increased expression of MHC Class II proteins and showed an activated phenotype. Increased expression of MHC Class II was inhibited by the NADPH oxidase inhibitor apocynin , P2X7 receptor antagonist A438709 hydrochloride, and genetic deletions of the NADPH oxidase p47 phox subunit or the P2X7 receptor. The P2X7 receptor acted upstream of NADPH oxidase activation by up-regulating the expression of the p47 phox subunit and p47 phox binding to the membrane subunit, gp91 phox. We conclude that the P2X7 receptor is a primary mediator of oxidative stress-induced exacerbation of inflammatory liver injury in obese mice via NADPH oxidase-dependent mechanisms. PMID:22343416

Molecular Mechanistic Insights into the Endothelial Receptor Mediated Cytoadherence of Plasmodium falciparum-Infected Erythrocytes

PubMed Central

Li, Ang; Lim, Tong Seng; Shi, Hui; Yin, Jing; Tan, Swee Jin; Li, Zhengjun; Low, Boon Chuan; Tan, Kevin Shyong Wei; Lim, Chwee Teck

2011-01-01

Cytoadherence or sequestration is essential for the pathogenesis of the most virulent human malaria species, Plasmodium falciparum (P. falciparum). Similar to leukocyte-endothelium interaction in response to inflammation, cytoadherence of P. falciparum infected red blood cells (IRBCs) to endothelium occurs under physiological shear stresses in blood vessels and involves an array of molecule complexes which cooperate to form stable binding. Here, we applied single-molecule force spectroscopy technique to quantify the dynamic force spectra and characterize the intrinsic kinetic parameters for specific ligand-receptor interactions involving two endothelial receptor proteins: thrombospondin (TSP) and CD36. It was shown that CD36 mediated interaction was much more stable than that mediated by TSP at single molecule level, although TSP-IRBC interaction appeared stronger than CD36-IRBC interaction in the high pulling rate regime. This suggests that TSP-mediated interaction may initiate cell adhesion by capturing the fast flowing IRBCs whereas CD36 functions as the ‘holder’ for providing stable binding. PMID:21437286

Cigarette Smoke–Induced CXCR3 Receptor Up-Regulation Mediates Endothelial Apoptosis

PubMed Central

Green, Linden A.; Petrusca, Daniela; Rajashekhar, Gangaraju; Gianaris, Tom; Schweitzer, Kelly S.; Wang, Liang; Justice, Matthew J.; Petrache, Irina

2012-01-01

Endothelial monocyte–activating polypeptide II (EMAP II) and interferon-inducible protein (IP)–10 are proinflammatory mediators, which in addition to their chemokine activities, selectively induce apoptosis in endothelial cells and are up-regulated in the lungs of cigarette smoke–exposed humans. Previously, we showed that EMAP II is an essential mediator of cigarette smoke–induced lung emphysema in mice linking endothelial cell apoptosis with inflammation. Here we addressed the role of the CXCR3 receptor in EMAP II–induced and IP-10–induced apoptosis in endothelial cells and its regulation by cigarette smoke. We found that both neutralizing antibodies and small inhibitory RNA to CXCR3 abrogated EMAP II–induced and IP-10–induced endothelial caspase-3 activation and DNA fragmentation. CXCR3 receptor surface expression in human lung microvascular endothelial cells and in lung tissue endothelium was up-regulated by exposure to cigarette smoke. In tissue culture conditions, EMAP II–induced and IP-10–induced apoptosis was enhanced by preincubation with cigarette smoke extract. Interestingly, serum starvation also induced CXCR3 up-regulation and enhanced EMAP II–induced endothelial apoptosis. Signal transduction via p38 mitogen-activated protein kinase activation was essential for CXCR3-induced cell death, but not for CXCR3 receptor up-regulation by cigarette smoke. In turn, protein nitration was required for CXCR3 receptor up-regulation by cigarette smoke and consequently for subsequent CXCR3-induced cell death. In conclusion, the concerted up-regulation of proinflammatory EMAP II, IP-10, and CXCR3 by cigarette smoke could sustain a cascade of cell death that may promote the alveolar tissue loss noted in human emphysema. PMID:22936405

Distinct combinations of variant ionotropic glutamate receptors mediate thermosensation and hygrosensation in Drosophila.

PubMed

Knecht, Zachary A; Silbering, Ana F; Ni, Lina; Klein, Mason; Budelli, Gonzalo; Bell, Rati; Abuin, Liliane; Ferrer, Anggie J; Samuel, Aravinthan Dt; Benton, Richard; Garrity, Paul A

2016-09-22

Ionotropic Receptors (IRs) are a large subfamily of variant ionotropic glutamate receptors present across Protostomia. While these receptors are most extensively studied for their roles in chemosensory detection, recent work has implicated two family members, IR21a and IR25a, in thermosensation in Drosophila . Here we characterize one of the most evolutionarily deeply conserved receptors, IR93a, and show that it is co-expressed and functions with IR21a and IR25a to mediate physiological and behavioral responses to cool temperatures. IR93a is also co-expressed with IR25a and a distinct receptor, IR40a, in a discrete population of sensory neurons in the sacculus, a multi-chambered pocket within the antenna. We demonstrate that this combination of receptors is required for neuronal responses to dry air and behavioral discrimination of humidity differences. Our results identify IR93a as a common component of molecularly and cellularly distinct IR pathways important for thermosensation and hygrosensation in insects.

Intra-accumbens baclofen, but not muscimol, increases second order instrumental responding for food reward in rats.

PubMed

Pulman, Kim G T; Somerville, Elizabeth M; Clifton, Peter G

2012-01-01

Stimulation of either GABA(A) or GABA(B) receptors within the nucleus accumbens shell strongly enhances food intake in rats. However the effects of subtype-selective stimulation of GABA receptors on instrumental responses for food reward are less well characterized. Here we contrast the effects of the GABA(A) receptor agonist muscimol and GABA(B) receptor agonist baclofen on instrumental responding for food using a second order reinforcement schedule. Bilateral intra-accumbens administration of baclofen (220-440 pmol) stimulated responding but a higher dose (660 pmol) induced stereotyped oral behaviour that interfered with responding. Baclofen (220-660 pmol) also stimulated intake of freely available chow. Muscimol (220-660 pmol) was without effect on responding for food on this schedule but did stimulate intake of freely available chow. Unilateral administration of either baclofen or muscimol (220 pmol) induced similar patterns of c-fos immunoreactivity in several hypothalamic sites but differed in its induction in the central nucleus of the amygdala. We conclude that stimulation of GABA(A) or GABA(B) receptors in the nucleus accumbens shell of rats produces clearly distinguishable effects on operant responding for food.

Modulation of AMPA receptor mediated current by nicotinic acetylcholine receptor in layer I neurons of rat prefrontal cortex

PubMed Central

Tang, Bo; Luo, Dong; Yang, Jie; Xu, Xiao-Yan; Zhu, Bing-Lin; Wang, Xue-Feng; Yan, Zhen; Chen, Guo-Jun

2015-01-01

Layer I neurons in the prefrontal cortex (PFC) exhibit extensive synaptic connections with deep layer neurons, implying their important role in the neural circuit. Study demonstrates that activation of nicotinic acetylcholine receptors (nAChRs) increases excitatory neurotransmission in this layer. Here we found that nicotine selectively increased the amplitude of AMPA receptor (AMPAR)-mediated current and AMPA/NMDA ratio, while without effect on NMDA receptor-mediated current. The augmentation of AMPAR current by nicotine was inhibited by a selective α7-nAChR antagonist methyllycaconitine (MLA) and intracellular calcium chelator BAPTA. In addition, nicotinic effect on mEPSC or paired-pulse ratio was also prevented by MLA. Moreover, an enhanced inward rectification of AMPAR current by nicotine suggested a functional role of calcium permeable and GluA1 containing AMPAR. Consistently, nicotine enhancement of AMPAR current was inhibited by a selective calcium-permeable AMPAR inhibitor IEM-1460. Finally, the intracellular inclusion of synthetic peptide designed to block GluA1 subunit of AMPAR at CAMKII, PKC or PKA phosphorylation site, as well as corresponding kinase inhibitor, blocked nicotinic augmentation of AMPA/NMDA ratio. These results have revealed that nicotine increases AMPAR current by modulating the phosphorylation state of GluA1 which is dependent on α7-nAChR and intracellular calcium. PMID:26370265

Differential effects of R-isovaline and the GABAB agonist, baclofen, in the guinea pig ileum.

PubMed

Fung, Timothy; Asseri, Khalid A; Asiri, Yahya I; Wall, Richard A; Schwarz, Stephan K W; Puil, Ernest; MacLeod, Bernard A

2016-11-15

R-isovaline is a non-proteinogenic amino acid which produces analgesia in a range of nociceptive assays. Mediation of this effect by metabotropic receptors for γ-aminobutyric acid (GABA) and glutamate, demonstrated by previous work, may depend on the type of tissue or receptor system. The objective of this study was to assess the activity of R-isovaline acting at GABA B and group II metabotropic glutamate receptors in guinea pig ileum, which is known to exhibit well-defined responses to GABA B agonists such as baclofen. The effects of bath-applied R-isovaline and RS-baclofen were examined on electrically evoked contractions of guinea pig ileum and during GABA B antagonism by CGP52432. In separate experiments, the group II metabotropic glutamate receptor agonist, LY354740 was applied to determine the functional presence of these receptors. R-isovaline (1-100mM) decreased the amplitude of ileal muscle contractions and increased tension. RS-baclofen reduced contraction amplitude, but decreased tension. CGP52432 did not prevent the effects of R-isovaline on contraction amplitude, but antagonized effects of RS-baclofen on contraction amplitude. The group II metabotropic glutamate receptor agonist, LY354740, produced no detectable effects on evoked contractions. R-isovaline differed significantly from RS-baclofen in its actions in the guinea pig ileum, indicated in particular by the finding that CGP52432 blocked only the effects of RS-baclofen. The ileal tissue did not respond to a group II metabotropic glutamate receptor agonist, previously shown to co-mediate R-isovaline analgesia. These findings raise the possibility of a novel therapeutic target at unknown receptors for R-isovaline-like compounds in the guinea pig ileum. Copyright © 2016 Elsevier B.V. All rights reserved.

Decoy receptor 3 suppresses TLR2-mediated B cell activation by targeting NF-κB.

PubMed

Huang, Zi-Ming; Kang, Jhi-Kai; Chen, Chih-Yu; Tseng, Tz-Hau; Chang, Chien-Wen; Chang, Yung-Chi; Tai, Shyh-Kuan; Hsieh, Shie-Liang; Leu, Chuen-Miin

2012-06-15

Decoy receptor 3 (DcR3) is a soluble protein in the TNFR superfamily. Its known ligands include Fas ligand, homologous to lymphotoxin, showing inducible expression, and competing with HSV glycoprotein D for herpes virus entry mediator, a receptor expressed by T lymphocytes, TNF-like molecule 1A, and heparan sulfate proteoglycans. DcR3 has been reported to modulate the functions of T cells, dendritic cells, and macrophages; however, its role in regulating B cell activation is largely unknown. In this study, we found that the DcR3.Fc fusion protein bound to human and mouse B cells and suppressed the activation of B cells. DcR3.Fc attenuated Staphylococcus aureus, IgM-, Pam(3)CSK(4)-, and LPS-mediated B cell proliferation but did not affect cytokine-induced B cell growth. In the presence of these mitogens, DcR3.Fc did not induce B cell apoptosis, suggesting that DcR3 may inhibit the signal(s) important for B cell activation. Because the combination of Fas.Fc, LT-βR.Fc (homologous to lymphotoxin, showing inducible expression, and competing with HSV glycoprotein D for herpes virus entry mediator, a receptor expressed by T lymphocytes receptor), and DR3.Fc (TNF-like molecule 1A receptor) did not suppress B cell proliferation and because the biological effect of DcR3.Fc on B cells was not blocked by heparin, we hypothesize that a novel ligand(s) of DcR3 mediates its inhibitory activity on B cells. Moreover, we found that TLR2-stimulated NF-κB p65 activation and NF-κB-driven luciferase activity were attenuated by DcR3.Fc. The TLR2-induced cytokine production by B cells was consistently reduced by DcR3. These results imply that DcR3 may regulate B cell activation by suppressing the activation of NF-κB.

Clozapine potentiation of GABA mediated cortical inhibition in treatment resistant schizophrenia.

PubMed

Kaster, Tyler S; de Jesus, Danilo; Radhu, Natasha; Farzan, Faranak; Blumberger, Daniel M; Rajji, Tarek K; Fitzgerald, Paul B; Daskalakis, Zafiris J

2015-07-01

Cortical inhibition (CI) deficits have been demonstrated in schizophrenia using transcranial magnetic stimulation (TMS). These CI deficits may be related to decreased GABA activity which may be involved in schizophrenia pathophysiology. Previous cross-sectional studies have also demonstrated greater CI in patients treated with clozapine than other typical/atypical antipsychotics. However, it is not clear if these differences in CI are a result of treatment-resistant illness which necessitates clozapine or are related to clozapine treatment. TMS measures of CI (i.e., cortical silent period (CSP) and short-interval cortical inhibition (SICI)) were measured over the motor cortex in 16 patients with schizophrenia before starting clozapine, then 6 weeks and 6 months after starting clozapine. CSP was significantly longer after 6 weeks of treatment with clozapine (p=0.014). From 6 weeks to 6 months, there was no significant difference in CSP (p>0.05). Short-interval cortical inhibition (SICI) was not significantly different at any time after treatment with clozapine (p>0.05). This prospective-longitudinal study demonstrates that treatment with clozapine is associated with an increase in GABAB mediated inhibitory neurotransmission. Potentiation of GABAB may be a novel neurotransmitter mechanism that is involved in the pathophysiology and treatment of schizophrenia. Copyright © 2015 Elsevier B.V. All rights reserved.

Neuropeptide Y restores non-receptor-mediated vasoconstrictive action in superior mesenteric arteries in portal hypertension.

PubMed

Hartl, Johannes; Dietrich, Peter; Moleda, Lukas; Müller-Schilling, Martina; Wiest, Reiner

2015-12-01

Vascular hyporeactivity to vasoconstrictors contributes to splanchnic arterial vasodilatation and hemodynamic dysregulation in portal hypertension. Neuropeptide Y (NPY), a sympathetic cotransmitter, has been shown to improve adrenergic vascular contractility in portal hypertensive rats and markedly attenuate hyperdynamic circulation. To further characterize the NPY-effects in portal hypertension, we investigated its role for non-receptor-mediated vasoconstriction in the superior mesenteric artery (SMA) of portal vein ligated (PVL) and sham-operated rats. Ex vivo SMA perfusion of PVL and sham rats was used to analyse the effects of NPY on pressure response to non-receptor-mediated vasoconstriction. Dose-response curves to KCl (30-300 mM) were used to bypass G protein-coupled receptor mechanisms. Potential involvement of the cyclooxygenase-pathway was tested by non-selective cyclooxygenase-inhibition using indomethacin. KCl-induced vascular contractility but not vascular sensitivity was significantly attenuated in PVL rats as compared with sham rats. Administration of NPY resulted in an augmentation of KCl-evoked vascular sensitivity being not different between study groups. However, KCl-induced vascular contractility was markedly more enhanced in PVL rats, thus, vascular response was no more significantly different between PVL and sham rats after addition of NPY. Administration of indomethacin abolished the NPY-induced enhancement of vasoconstriction. Receptor-independent vascular contractility is impaired in mesenteric arteries in portal hypertension. NPY improves non-receptor mediated mesenteric vasoconstriction more effective in portal hypertension than in healthy conditions correcting splanchnic vascular hyporesponsiveness. This beneficial vasoactive action of NPY adds to its well known more pronounced effects on adrenergic vasoconstriction in portal hypertension making it a promising therapeutic agent in portal hypertension. © 2015 John Wiley & Sons A

Tachykinin-induced contraction of the guinea-pig isolated oesophageal mucosa is mediated by NK2 receptors

PubMed Central

Kerr, Karen P; Thai, Binh; Coupar, Ian M

2000-01-01

The tachykinin receptor present in the guinea-pig oesophageal mucosa that mediates contractile responses of the muscularis mucosae has been characterized, using functional in vitro experiments. The NK1 receptor-selective agonist, [Sar9(O2)Met11]SP and the NK3 receptor-selective agonists, [MePhe7]-NKB and senktide, produced no response at submicromolar concentrations. The NK2 receptor-selective agonists, [Nle10]-NKA(4–10), and GR 64,349 produced concentration-dependent contractile effects with pD2 values of 8.20±0.16 and 8.30±0.15, respectively. The concentration-response curve to the non-selective agonist, NKA (pD2=8.13±0.04) was shifted significantly rightwards only by the NK2 receptor-selective antagonist, GR 159,897 and was unaffected by the NK1 receptor-selective antagonist, SR 140,333 and the NK3 receptor-selective antagonist, SB 222,200. The NK2 receptor-selective antagonist, GR 159,897, exhibited an apparent competitive antagonism against the NK2 receptor-selective agonist, GR 64,349 (apparent pKB value=9.29±0.16) and against the non-selective agonist, NKA (apparent pKB value=8.71±0.19). The NK2 receptor-selective antagonist, SR 48,968 exhibited a non-competitive antagonism against the NK2 receptor-selective agonist, [Nle10]-NKA(4–10). The pKB value was 10.84±0.19. It is concluded that the guinea-pig isolated oesophageal mucosa is a useful preparation for studying the effects of NK2 receptor-selective agonists and antagonists as the contractile responses to various tachykinins are mediated solely by NK2 receptors. PMID:11090121

Global phosphorylation analysis of beta-arrestin-mediated signaling downstream of a seven transmembrane receptor (7TMR).

PubMed

Xiao, Kunhong; Sun, Jinpeng; Kim, Jihee; Rajagopal, Sudarshan; Zhai, Bo; Villén, Judit; Haas, Wilhelm; Kovacs, Jeffrey J; Shukla, Arun K; Hara, Makoto R; Hernandez, Marylens; Lachmann, Alexander; Zhao, Shan; Lin, Yuan; Cheng, Yishan; Mizuno, Kensaku; Ma'ayan, Avi; Gygi, Steven P; Lefkowitz, Robert J

2010-08-24

beta-Arrestin-mediated signaling downstream of seven transmembrane receptors (7TMRs) is a relatively new paradigm for signaling by these receptors. We examined changes in protein phosphorylation occurring when HEK293 cells expressing the angiotensin II type 1A receptor (AT1aR) were stimulated with the beta-arrestin-biased ligand Sar(1), Ile(4), Ile(8)-angiotensin (SII), a ligand previously found to signal through beta-arrestin-dependent, G protein-independent mechanisms. Using a phospho-antibody array containing 46 antibodies against signaling molecules, we found that phosphorylation of 35 proteins increased upon SII stimulation. These SII-mediated phosphorylation events were abrogated after depletion of beta-arrestin 2 through siRNA-mediated knockdown. We also performed an MS-based quantitative phosphoproteome analysis after SII stimulation using a strategy of stable isotope labeling of amino acids in cell culture (SILAC). We identified 1,555 phosphoproteins (4,552 unique phosphopeptides), of which 171 proteins (222 phosphopeptides) showed increased phosphorylation, and 53 (66 phosphopeptides) showed decreased phosphorylation upon SII stimulation of the AT1aR. This study identified 38 protein kinases and three phosphatases whose phosphorylation status changed upon SII treatment. Using computational approaches, we performed system-based analyses examining the beta-arrestin-mediated phosphoproteome including construction of a kinase-substrate network for beta-arrestin-mediated AT1aR signaling. Our analysis demonstrates that beta-arrestin-dependent signaling processes are more diverse than previously appreciated. Notably, our analysis identifies an AT1aR-mediated cytoskeletal reorganization network whereby beta-arrestin regulates phosphorylation of several key proteins, including cofilin and slingshot. This study provides a system-based view of beta-arrestin-mediated phosphorylation events downstream of a 7TMR and opens avenues for research in a rapidly evolving area

Posttranslational Modification of HOIP Blocks Toll-Like Receptor 4-Mediated Linear-Ubiquitin-Chain Formation

PubMed Central

Bowman, James; Rodgers, Mary A.; Shi, Mude; Amatya, Rina; Hostager, Bruce; Iwai, Kazuhiro; Gao, Shou-Jiang

2015-01-01

ABSTRACT Linear ubiquitination is an atypical posttranslational modification catalyzed by the linear-ubiquitin-chain assembly complex (LUBAC), containing HOIP, HOIL-1L, and Sharpin. LUBAC facilitates NF-κB activation and inflammation upon receptor stimulation by ligating linear ubiquitin chains to critical signaling molecules. Indeed, linear-ubiquitination-dependent signaling is essential to prevent pyogenic bacterial infections that can lead to death. While linear ubiquitination is essential for intracellular receptor signaling upon microbial infection, this response must be measured and stopped to avoid tissue damage and autoimmunity. While LUBAC is activated upon bacterial stimulation, the mechanisms regulating LUBAC activity in response to bacterial stimuli have remained elusive. We demonstrate that LUBAC activity itself is downregulated through ubiquitination, specifically, ubiquitination of the catalytic subunit HOIP at the carboxyl-terminal lysine 1056. Ubiquitination of Lys1056 dynamically altered HOIP conformation, resulting in the suppression of its catalytic activity. Consequently, HOIP Lys1056-to-Arg mutation led not only to persistent LUBAC activity but also to prolonged NF-κB activation induced by bacterial lipopolysaccharide-mediated Toll-like receptor 4 (TLR4) stimulation, whereas it showed no effect on NF-κB activation induced by CD40 stimulation. This study describes a novel posttranslational regulation of LUBAC-mediated linear ubiquitination that is critical for specifically directing TLR4-mediated NF-κB activation. PMID:26578682

CD3ζ-based chimeric antigen receptors mediate T cell activation via cis- and trans-signalling mechanisms: implications for optimization of receptor structure for adoptive cell therapy

PubMed Central

Bridgeman, J S; Ladell, K; Sheard, V E; Miners, K; Hawkins, R E; Price, D A; Gilham, D E

2014-01-01

Chimeric antigen receptors (CARs) can mediate redirected lysis of tumour cells in a major histocompatibility complex (MHC)-independent manner, thereby enabling autologous adoptive T cell therapy for a variety of malignant neoplasms. Currently, most CARs incorporate the T cell receptor (TCR) CD3ζ signalling chain; however, the precise mechanisms responsible for CAR-mediated T cell activation are unclear. In this study, we used a series of immunoreceptor tyrosine-based activation motif (ITAM)-mutant and transmembrane-modified receptors to demonstrate that CARs activate T cells both directly via the antigen-ligated signalling chain and indirectly via associated chains within the TCR complex. These observations allowed us to generate new receptors capable of eliciting polyfunctional responses in primary human T cells. This work increases our understanding of CAR function and identifies new avenues for the optimization of CAR-based therapeutic interventions. PMID:24116999

Selective regulation of clathrin-mediated epidermal growth factor receptor signaling and endocytosis by phospholipase C and calcium

PubMed Central

Delos Santos, Ralph Christian; Bautista, Stephen; Lucarelli, Stefanie; Bone, Leslie N.; Dayam, Roya M.; Abousawan, John; Botelho, Roberto J.; Antonescu, Costin N.

2017-01-01

Clathrin-mediated endocytosis is a major regulator of cell-surface protein internalization. Clathrin and other proteins assemble into small invaginating structures at the plasma membrane termed clathrin-coated pits (CCPs) that mediate vesicle formation. In addition, epidermal growth factor receptor (EGFR) signaling is regulated by its accumulation within CCPs. Given the diversity of proteins regulated by clathrin-mediated endocytosis, how this process may distinctly regulate specific receptors is a key question. We examined the selective regulation of clathrin-dependent EGFR signaling and endocytosis. We find that perturbations of phospholipase Cγ1 (PLCγ1), Ca2+, or protein kinase C (PKC) impair clathrin-mediated endocytosis of EGFR, the formation of CCPs harboring EGFR, and EGFR signaling. Each of these manipulations was without effect on the clathrin-mediated endocytosis of transferrin receptor (TfR). EGFR and TfR were recruited to largely distinct clathrin structures. In addition to control of initiation and assembly of CCPs, EGF stimulation also elicited a Ca2+- and PKC-dependent reduction in synaptojanin1 recruitment to clathrin structures, indicating broad control of CCP assembly by Ca2+ signals. Hence EGFR elicits PLCγ1-calcium signals to facilitate formation of a subset of CCPs, thus modulating its own signaling and endocytosis. This provides evidence for the versatility of CCPs to control diverse cellular processes. PMID:28814502

[An analysis and literature review of two cases of autoimmune encephalitis with GABAB receptor antibodies].

PubMed

Zhang, M; Hao, H J; Liu, L P; Zhang, H H; Zhou, Y Y

2016-10-01

Autoimmune encephalitis with GABA B receptor antibodies has been rarely reported. Two cases of GABA B receptor antibodies encephalitis were presented here.Epilepsy was the onset symptom, followed by declined consciousness and frequent seizures. Fever was presented in the whole course of the disease. Myorhythmia of the two hands and pilomotor seizures were shown in the later course of the disease. No specificity was demonstrated in electroencephalograms and magnetic resonance imaging. Sensitive response was shown to the first-line immunotherapy.

ARF6 and GASP-1 are post-endocytic sorting proteins selectively involved in the intracellular trafficking of dopamine D2 receptors mediated by GRK and PKC in transfected cells

PubMed Central

Cho, DI; Zheng, M; Min, C; Kwon, KJ; Shin, CY; Choi, HK; Kim, KM

2013-01-01

Background and Purpose GPCRs undergo both homologous and heterologous regulatory processes in which receptor phosphorylation plays a critical role. The protein kinases responsible for each pathway are well established; however, other molecular details that characterize each pathway remain unclear. In this study, the molecular mechanisms that determine the differences in the functional roles and intracellular trafficking between homologous and PKC-mediated heterologous internalization pathways for the dopamine D2 receptor were investigated. Experimental Approach All of the S/T residues located within the intracellular loops of D2 receptor were mutated, and the residues responsible for GRK- and PKC-mediated internalization were determined in HEK-293 cells and SH-SY5Y cells. The functional role of receptor internalization and the cellular components that determine the post-endocytic fate of internalized D2 receptors were investigated in the transfected cells. Key Results T134, T225/S228/S229 and S325 were involved in PKC-mediated D2 receptor desensitization. S229 and adjacent S/T residues mediated the PKC-dependent internalization of D2 receptors, which induced down-regulation and desensitization. S/T residues within the second intracellular loop and T225 were the major residues involved in GRK-mediated internalization of D2 receptors, which induced receptor resensitization. ARF6 mediated the recycling of D2 receptors internalized in response to agonist stimulation. In contrast, GASP-1 mediated the down-regulation of D2 receptors internalized in a PKC-dependent manner. Conclusions and Implications GRK- and PKC-mediated internalizations of D2 receptors occur through different intracellular trafficking pathways and mediate distinct functional roles. Distinct S/T residues within D2 receptors and different sorting proteins are involved in the dissimilar regulation of D2 receptors by GRK2 and PKC. PMID:23082996

Receptor-Mediated Delivery of CRISPR-Cas9 Endonuclease for Cell-Type-Specific Gene Editing.

PubMed

Rouet, Romain; Thuma, Benjamin A; Roy, Marc D; Lintner, Nathanael G; Rubitski, David M; Finley, James E; Wisniewska, Hanna M; Mendonsa, Rima; Hirsh, Ariana; de Oñate, Lorena; Compte Barrón, Joan; McLellan, Thomas J; Bellenger, Justin; Feng, Xidong; Varghese, Alison; Chrunyk, Boris A; Borzilleri, Kris; Hesp, Kevin D; Zhou, Kaihong; Ma, Nannan; Tu, Meihua; Dullea, Robert; McClure, Kim F; Wilson, Ross C; Liras, Spiros; Mascitti, Vincent; Doudna, Jennifer A

2018-05-30

CRISPR-Cas RNA-guided endonucleases hold great promise for disrupting or correcting genomic sequences through site-specific DNA cleavage and repair. However, the lack of methods for cell- and tissue-selective delivery currently limits both research and clinical uses of these enzymes. We report the design and in vitro evaluation of S. pyogenes Cas9 proteins harboring asialoglycoprotein receptor ligands (ASGPrL). In particular, we demonstrate that the resulting ribonucleoproteins (Cas9-ASGPrL RNP) can be engineered to be preferentially internalized into cells expressing the corresponding receptor on their surface. Uptake of such fluorescently labeled proteins in liver-derived cell lines HEPG2 (ASGPr+) and SKHEP (control; diminished ASGPr) was studied by live cell imaging and demonstrates increased accumulation of Cas9-ASGPrL RNP in HEPG2 cells as a result of effective ASGPr-mediated endocytosis. When uptake occurred in the presence of a peptide with endosomolytic properties, we observed receptor-facilitated and cell-type specific gene editing that did not rely on electroporation or the use of transfection reagents. Overall, these in vitro results validate the receptor-mediated delivery of genome-editing enzymes as an approach for cell-selective gene editing and provide a framework for future potential applications to hepatoselective gene editing in vivo.

Cytotoxic-T-lymphocyte antigen 4 receptor signaling for lymphocyte adhesion is mediated by C3G and Rap1.

PubMed

Kloog, Yoel; Mor, Adam

2014-03-01

T-lymphocyte adhesion plays a critical role in both inflammatory and autoimmune responses. The small GTPase Rap1 is the key coordinator mediating T-cell adhesion to endothelial cells, antigen-presenting cells, and virus-infected cells. We describe a signaling pathway, downstream of the cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor, leading to Rap1-mediated adhesion. We identified a role for the Rap1 guanine nucleotide exchange factor C3G in the regulation of T-cell adhesion and showed that this factor is required for both T-cell receptor (TCR)-mediated and CTLA-4-mediated T-cell adhesion. Our data indicated that C3G translocates to the plasma membrane downstream of TCR signaling, where it regulates activation of Rap1. We also showed that CTLA-4 receptor signaling mediates tyrosine phosphorylation in the C3G protein, and that this is required for augmented activation of Rap1 and increased adhesion mediated by leukocyte function-associated antigen type 1 (LFA-1). Zap70 is required for C3G translocation to the plasma membrane, whereas the Src family member Hck facilitates C3G phosphorylation. These findings point to C3G and Hck as promising potential therapeutic targets for the treatment of T-cell-dependent autoimmune disorders.

Cytotoxic-T-Lymphocyte Antigen 4 Receptor Signaling for Lymphocyte Adhesion Is Mediated by C3G and Rap1

PubMed Central

Kloog, Yoel

2014-01-01

T-lymphocyte adhesion plays a critical role in both inflammatory and autoimmune responses. The small GTPase Rap1 is the key coordinator mediating T-cell adhesion to endothelial cells, antigen-presenting cells, and virus-infected cells. We describe a signaling pathway, downstream of the cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor, leading to Rap1-mediated adhesion. We identified a role for the Rap1 guanine nucleotide exchange factor C3G in the regulation of T-cell adhesion and showed that this factor is required for both T-cell receptor (TCR)-mediated and CTLA-4-mediated T-cell adhesion. Our data indicated that C3G translocates to the plasma membrane downstream of TCR signaling, where it regulates activation of Rap1. We also showed that CTLA-4 receptor signaling mediates tyrosine phosphorylation in the C3G protein, and that this is required for augmented activation of Rap1 and increased adhesion mediated by leukocyte function-associated antigen type 1 (LFA-1). Zap70 is required for C3G translocation to the plasma membrane, whereas the Src family member Hck facilitates C3G phosphorylation. These findings point to C3G and Hck as promising potential therapeutic targets for the treatment of T-cell-dependent autoimmune disorders. PMID:24396067

Cloning and expression profile of ionotropic receptors in the parasitoid wasp Microplitis mediator (Hymenoptera: Braconidae).

PubMed

Wang, Shan-Ning; Peng, Yong; Lu, Zi-Yun; Dhiloo, Khalid Hussain; Zheng, Yao; Shan, Shuang; Li, Rui-Jun; Zhang, Yong-Jun; Guo, Yu-Yuan

2016-07-01

Ionotropic receptors (IRs) mainly detect the acids and amines having great importance in many insect species, representing an ancient olfactory receptor family in insects. In the present work, we performed RNAseq of Microplitis mediator antennae and identified seventeen IRs. Full-length MmedIRs were cloned and sequenced. Phylogenetic analysis of the Hymenoptera IRs revealed that ten MmedIR genes encoded "antennal IRs" and seven encoded "divergent IRs". Among the IR25a orthologous groups, two genes, MmedIR25a.1 and MmedIR25a.2, were found in M. mediator. Gene structure analysis of MmedIR25a revealed a tandem duplication of IR25a in M. mediator. The tissue distribution and development specific expression of the MmedIR genes suggested that these genes showed a broad expression profile. Quantitative gene expression analysis showed that most of the genes are highly enriched in adult antennae, indicating the candidate chemosensory function of this family in parasitic wasps. Using immunocytochemistry, we confirmed that one co-receptor, MmedIR8a, was expressed in the olfactory sensory neurons. Our data will supply fundamental information for functional analysis of the IRs in parasitoid wasp chemoreception. Copyright © 2016 Elsevier Ltd. All rights reserved.

Ionotropic and metabotropic glutamate receptor mediation of glucocorticoid-induced apoptosis in hippocampal cells and the neuroprotective role of synaptic N-methyl-D-aspartate receptors.

PubMed

Lu, J; Goula, D; Sousa, N; Almeida, O F X

2003-01-01

Glutamate receptors have been proposed to mediate the apoptotic actions of glucocorticoids in hippocampal cells. To further analyze the role of glutamate receptors in this process, we pretreated primary hippocampal cells from neonatal (postnatal day 4) rats with antagonists of ionotropic glutamate receptor (iGluR) and metabotropic glutamate receptor (mGluR) antagonists before exposure to the specific glucocorticoid receptor agonist dexamethasone (DEX) at a dose of 1 microM. Dizocilpine (MK801; a general N-methyl-D-aspartic acid [NMDA] receptor antagonist, NMDAR antagonist) and ifenprodil (a specific ligand of the NMDAR 2B subunit, NR2B), were used to block iGluR; (RS)-alpha-ethyl-4-carboxyphenylglycine (E4CPG) and (RS)-alpha-cyclopropyl-4-phosphonophenyl-glycine (CPPG) were employed as I/II (E4CPG) and II/III (CPPG) mGluR antagonists. Blockade of iGluR resulted in a significant attenuation of DEX-induced cell death; the finding that ifenprodil exerted a similar potency to MK801 demonstrates the involvement of NR2B receptors in glucocorticoid-induced cell death. Apoptosis accounted for a significant amount of the cell loss observed, as detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling histochemistry for the in situ labeling of DNA breaks; apoptotic cells were distinguished from necrosis on the basis of morphological criteria, including chromatin condensation, membrane blebbing and presence of apoptotic bodies. Treatment with E4CPG and CPPG completely abolished the apoptotic response to DEX, thus showing the additional contribution of mGluR to the phenomenon. Further, dose-response studies with NMDA revealed that whereas high (10 microM) doses of NMDA themselves elicit cytotoxic responses, low (1-5 microM) concentrations of NMDA can effectively oppose DEX-induced cell death. Interestingly, the neuroprotective actions of low dose NMDA stimulation were abolished when either synaptic or extrasynaptic NMDA receptors were blocked with MK801

Dynamic regulation of glycinergic input to spinal dorsal horn neurones by muscarinic receptor subtypes in rats.

PubMed

Wang, Xiu-Li; Zhang, Hong-Mei; Li, De-Pei; Chen, Shao-Rui; Pan, Hui-Lin

2006-03-01

Activation of spinal muscarinic acetylcholine receptors (mAChRs) inhibits nociception. However, the cellular mechanisms of this action are not fully known. In this study, we determined the role of mAChR subtypes in regulation of synaptic glycine release in the spinal cord. Whole-cell voltage-clamp recordings were performed on lamina II neurones in the rat spinal cord slices. The mAChR agonist oxotremorine-M significantly increased the frequency of glycinergic sIPSCs but not mIPSCs. Surprisingly, the effect of oxotremorine-M on sIPSCs was largely attenuated at a higher concentration. On the other hand, 1-10 microm oxotremorine-M dose-dependently increased the frequency of sIPSCs in rats pretreated with intrathecal pertussis toxin. Furthermore, oxotremorine-M also dose-dependently increased the frequency of sIPSCs in the presence of himbacine (an M2/M4 mAChR antagonist) or AF-DX116 (an M2 mAChR antagonist). The M3 mAChR antagonist 4-DAMP abolished the stimulatory effect of oxotremorine-M on sIPSCs. Interestingly, the GABA(B) receptor antagonist CGP55845 potentiated the stimulatory effect of oxotremorine-M on sIPSCs. In the presence of CGP55845, both himbacine and AF-DX116 similarly reduced the potentiating effect of oxotremorine-M on sIPSCs. Collectively, these data suggest that the M3 subtype is present on the somatodendritic site of glycinergic neurones and is mainly responsible for muscarinic potentiation of glycinergic input to spinal dorsal horn neurones. Concurrent stimulation of mAChRs on adjacent GABAergic interneurones attenuates synaptic glycine release through presynaptic GABA(B) receptors on glycinergic interneurones. This study illustrates a complex dynamic interaction between GABAergic and glycinergic synapses in the spinal cord dorsal horn.

Distinct combinations of variant ionotropic glutamate receptors mediate thermosensation and hygrosensation in Drosophila

PubMed Central

Knecht, Zachary A; Silbering, Ana F; Ni, Lina; Klein, Mason; Budelli, Gonzalo; Bell, Rati; Abuin, Liliane; Ferrer, Anggie J; Samuel, Aravinthan DT; Benton, Richard; Garrity, Paul A

2016-01-01

Ionotropic Receptors (IRs) are a large subfamily of variant ionotropic glutamate receptors present across Protostomia. While these receptors are most extensively studied for their roles in chemosensory detection, recent work has implicated two family members, IR21a and IR25a, in thermosensation in Drosophila. Here we characterize one of the most evolutionarily deeply conserved receptors, IR93a, and show that it is co-expressed and functions with IR21a and IR25a to mediate physiological and behavioral responses to cool temperatures. IR93a is also co-expressed with IR25a and a distinct receptor, IR40a, in a discrete population of sensory neurons in the sacculus, a multi-chambered pocket within the antenna. We demonstrate that this combination of receptors is required for neuronal responses to dry air and behavioral discrimination of humidity differences. Our results identify IR93a as a common component of molecularly and cellularly distinct IR pathways important for thermosensation and hygrosensation in insects. DOI: http://dx.doi.org/10.7554/eLife.17879.001 PMID:27656904

Mechanism-Based Tumor-Targeting Drug Delivery System. Validation of Efficient Vitamin Receptor-Mediated Endocytosis and Drug Release

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

Chen, S.; Wong, S.; Zhao, X.

An efficient mechanism-based tumor-targeting drug delivery system, based on tumor-specific vitamin-receptor mediated endocytosis, has been developed. The tumor-targeting drug delivery system is a conjugate of a tumor-targeting molecule (biotin: vitamin H or vitamin B-7), a mechanism-based self-immolative linker and a second-generation taxoid (SB-T-1214) as the cytotoxic agent. This conjugate (1) is designed to be (i) specific to the vitamin receptors overexpressed on tumor cell surface and (ii) internalized efficiently through receptor-mediated endocytosis, followed by smooth drug release via glutathione-triggered self-immolation of the linker. In order to monitor and validate the sequence of events hypothesized, i.e., receptor-mediated endocytosis of the conjugate,more » drug release, and drug-binding to the target protein (microtubules), three fluorescent/fluorogenic molecular probes (2, 3, and 4) were designed and synthesized. The actual occurrence of these processes was unambiguously confirmed by means of confocal fluorescence microscopy (CFM) and flow cytometry using L1210FR leukemia cells, overexpressing biotin receptors. The molecular probe 4, bearing the taxoid linked to fluorescein, was also used to examine the cell specificity (i.e., efficacy of receptor-based cell targeting) for three cell lines, L1210FR (biotin receptors overexpressed), L1210 (biotin receptors not overexpressed), and WI38 (normal human lung fibroblast, biotin receptor negative). As anticipated, the molecular probe 4 exhibited high specificity only to L1210FR. To confirm the direct correlation between the cell-specific drug delivery and anticancer activity of the probe 4, its cytotoxicity against these three cell lines was also examined. The results clearly showed a good correlation between the two methods. In the same manner, excellent cell-specific cytotoxicity of the conjugate 1 (without fluorescein attachment to the taxoid) against the same three cell lines was confirmed. This

Endocrine Disrupting Chemicals Mediated through Binding Androgen Receptor Are Associated with Diabetes Mellitus

PubMed Central

Sakkiah, Sugunadevi; Wang, Tony; Zou, Wen; Wang, Yuping; Pan, Bohu; Tong, Weida; Hong, Huixiao

2017-01-01

Endocrine disrupting chemicals (EDCs) can mimic natural hormone to interact with receptors in the endocrine system and thus disrupt the functions of the endocrine system, raising concerns on the public health. In addition to disruption of the endocrine system, some EDCs have been found associated with many diseases such as breast cancer, prostate cancer, infertility, asthma, stroke, Alzheimer’s disease, obesity, and diabetes mellitus. EDCs that binding androgen receptor have been reported associated with diabetes mellitus in in vitro, animal, and clinical studies. In this review, we summarize the structural basis and interactions between androgen receptor and EDCs as well as the associations of various types of diabetes mellitus with the EDCs mediated through androgen receptor binding. We also discuss the perspective research for further understanding the impact and mechanisms of EDCs on the risk of diabetes mellitus. PMID:29295509

A dopamine D2 receptor mutant capable of G protein-mediated signaling but deficient in arrestin binding.

PubMed

Lan, Hongxiang; Liu, Yong; Bell, Michal I; Gurevich, Vsevolod V; Neve, Kim A

2009-01-01

Arrestins mediate G protein-coupled receptor desensitization, internalization, and signaling. Dopamine D(2) and D(3) receptors have similar structures but distinct characteristics of interaction with arrestins. The goals of this study were to compare arrestin-binding determinants in D(2) and D(3) receptors other than phosphorylation sites and to create a D(2) receptor that is deficient in arrestin binding. We first assessed the ability of purified arrestins to bind to glutathione transferase (GST) fusion proteins containing the receptor third intracellular loops (IC3). Arrestin3 bound to IC3 of both D(2) and D(3) receptors, with the affinity and localization of the binding site indistinguishable between the receptor subtypes. Mutagenesis of the GST-IC3 fusion proteins identified an important determinant of the binding of arrestin3 in the N-terminal region of IC3. Alanine mutations of this determinant (IYIV212-215) in the full-length D(2) receptor generated a signaling-biased receptor with intact ligand binding and G-protein coupling and activation, but deficient in receptor-mediated arrestin3 translocation to the membrane, agonist-induced receptor internalization, and agonist-induced desensitization in human embryonic kidney 293 cells. This mutation also decreased arrestin-dependent activation of extracellular signal-regulated kinases. The finding that nonphosphorylated D(2)-IC3 and D(3)-IC3 have similar affinity for arrestin is consistent with previous suggestions that the differential effects of D(2) and D(3) receptor activation on membrane translocation of arrestin and receptor internalization are due, at least in part, to differential phosphorylation of the receptors. In addition, these results imply that the sequence IYIV212-215 at the N terminus of IC3 of the D(2) receptor is a key element of the arrestin binding site.

Glutamate mediates the function of melanocortin receptor 4 on sim1 neurons in body weight regulation

USDA-ARS?s Scientific Manuscript database

The melanocortin receptor 4 (MC4R) is a well-established mediator of body weight homeostasis. However, the neurotransmitter(s) that mediate MC4R function remain largely unknown; as a result, little is known about the second-order neurons of the MC4R neural pathway. Single-minded 1 (Sim1)-expressing ...

A new meaning for “Gin & Tonic”

PubMed Central

Mody, Istvan; Glykys, Joseph; Wei, Weizheng

2007-01-01

Gamma-aminobutyric acid (GABA) is the main chemical inhibitory neurotransmitter in the brain. In the central nervous system (CNS) it acts on two distinct types of receptor: an ion channel, i.e., an “ionotropic” receptor permeable to Cl- and HCO3- (GABAA receptors) and a G-protein coupled “metabotropic” receptor that is linked to various effector mechanisms (GABAB receptors). This review will summarize novel developments in the physiology and pharmacology of GABAA receptors (GABAARs), specifically those found outside synapses. The focus will be on a particular combination of GABAAR subunits responsible for mediating tonic inhibition and sensitive to concentrations of ethanol legally considered to be sobriety impairing. Since the same receptors are also a preferred target for the metabolites of steroid hormones synthesized in the brain (neurosteroids), the ethanol-sensitive tonic inhibition may be a common pathway for interactions between the effects of alcohol and those of ovarian and stress-related neurosteroids. PMID:17521846

Receptor-mediated activation of nitric oxide synthesis by arginine in endothelial cells

PubMed Central

Joshi, Mahesh S.; Ferguson, T. Bruce; Johnson, Fruzsina K.; Johnson, Robert A.; Parthasarathy, Sampath; Lancaster, Jack R.

2007-01-01

Arginine contains the guanidinium group and thus has structural similarity to ligands of imidazoline and α-2 adrenoceptors (α-2 AR). Therefore, we investigated the possibility that exogenous arginine may act as a ligand for these receptors in human umbilical vein endothelial cells and activate intracellular nitric oxide (NO) synthesis. Idazoxan, a mixed antagonist of imidazoline and α-2 adrenoceptors, partly inhibited l-arginine-initiated NO formation as measured by a Griess reaction. Rauwolscine, a highly specific antagonist of α-2 AR, at very low concentrations completely inhibited NO formation. Like l-arginine, agmatine (decarboxylated arginine) also activated NO synthesis, however, at much lower concentrations. We found that dexmedetomidine, a specific agonist of α-2 AR was very potent in activating cellular NO, thus indicating a possible role for α-2 AR in l-arginine-mediated NO synthesis. d-arginine also activated NO production and could be inhibited by imidazoline and α-2 AR antagonists, thus indicating nonsubstrate actions of arginine. Pertussis toxin, an inhibitor of G proteins, attenuated l-arginine-mediated NO synthesis, thus indicating mediation via G proteins. l-type Ca2+ channel blocker nifedipine and phospholipase C inhibitor U73122 inhibited NO formation and thus implicated participation of a second messenger pathway. Finally, in isolated rat gracilis vessels, rauwolscine completely inhibited the l-arginine-initiated vessel relaxation. Taken together, these data provide evidence for binding of arginine to membrane receptor(s), leading to the activation of endothelial NO synthase (eNOS) NO production through a second messenger pathway. These findings provide a previously unrecognized mechanistic explanation for the beneficial effects of l-arginine in the cardiovascular system and thus provide new potential avenues for therapeutic development. PMID:17535904

Receptor-Mediated and Fluid-Phase Transcytosis of Horseradish Peroxidase across Rat Hepatocytes

PubMed Central

Ellinger, Isabella; Fuchs, Renate

2010-01-01

Horseradish peroxidase (HRP) is often used as a fluid-phase marker to characterize endocytic and transcytotic processes. Likewise, it has been applied to investigate the mechanisms of biliary secretion of fluid in rat liver hepatocytes. However, HRP contains mannose residues and thus binds to mannose receptors (MRs) on liver cells, including hepatocytes. To study the role of MR-mediated endocytosis of HRP transport in hepatocytes, we determined the influence of the oligosaccharid mannan on HRP biliary secretion in the isolated perfused rat liver. A 1-minute pulse of HRP was applied followed by marker-free perfusion. HRP appeared in bile with biphasic kinetics: a first peak at 7 minutes and a second peak at 15 minutes after labeling. Perfusion with 0.8 mg/mL HRP in the presence of a twofold excess of mannan reduced the first peak by 41% without effect on the second one. Together with recently published data on MR expression in rat hepatocytes this demonstrates two different mechanisms for HRP transcytosis: a rapid, receptor-mediated transport and a slower fluid-phase transport. PMID:20168981

Gefitinib enhances human colon cancer cells to TRAIL-induced apoptosis of via autophagy- and JNK-mediated death receptors upregulation.

PubMed

Chen, Lei; Meng, Yue; Guo, Xiaoqing; Sheng, Xiaotong; Tai, Guihua; Zhang, Fenglei; Cheng, Hairong; Zhou, Yifa

2016-11-01

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a potent cancer cell-specific apoptosis-inducing cytokine with little toxicity to most normal cells. Here, we report that gefitinib and TRAIL in combination produce a potent synergistic effect on TRAIL-sensitive human colon cancer HCT116 cells and an additive effect on TRAIL-resistant HT-29 cells. Interestingly, gefitinib increases the expression of cell surface receptors DR4 and DR5, possibly explaining the synergistic effect. Knockdown of DR4 and DR5 by siRNA significantly decreases gefitinib- and TRAIL-mediated cell apoptosis, supporting this idea. Because the inhibition of gefitinib-induced autophagy by 3-MA significantly decreases DR4 and DR5 upregulation, as well as reduces gefitinib- and TRAIL-induced apoptosis, we conclude that death receptor upregulation is autophagy mediated. Furthermore, our results indicate that death receptor expression may also be regulated by JNK activation, because pre-treatment of cells with JNK inhibitor SP600125 significantly decreases gefitinib-induced death receptor upregulation. Interestingly, SP600125 also inhibits the expression CHOP, yet CHOP has no impact on death receptor expressions. We also find here that phosphorylation of Akt and ERK might also be required for TRAIL sensitization. In summary, our results indicate that gefitinib effectively enhances TRAIL-induced apoptosis, likely via autophagy and JNK- mediated death receptor expression and phosphorylation of Akt and ERK.

Regulation of T cell receptor complex-mediated signaling by ubiquitin and ubiquitin-like modifications.

PubMed

Friend, Samantha F; Deason-Towne, Francina; Peterson, Lisa K; Berger, Allison J; Dragone, Leonard L

2014-01-01

Post-translational protein modifications are a dynamic method of regulating protein function in response to environmental signals. As with any cellular process, T cell receptor (TCR) complex-mediated signaling is highly regulated, since the strength and duration of TCR-generated signals governs T cell development and activation. While regulation of TCR complex-mediated signaling by phosphorylation has been well studied, regulation by ubiquitin and ubiquitin-like modifiers is still an emerging area of investigation. This review will examine how ubiquitin, E3 ubiquitin ligases, and other ubiquitin-like modifications such as SUMO and NEDD8 regulate TCR complex-mediated signaling.

Regulation of T cell receptor complex-mediated signaling by ubiquitin and ubiquitin-like modifications

PubMed Central

Friend, Samantha F; Deason-Towne, Francina; Peterson, Lisa K; Berger, Allison J; Dragone, Leonard L

2014-01-01

Post-translational protein modifications are a dynamic method of regulating protein function in response to environmental signals. As with any cellular process, T cell receptor (TCR) complex-mediated signaling is highly regulated, since the strength and duration of TCR-generated signals governs T cell development and activation. While regulation of TCR complex-mediated signaling by phosphorylation has been well studied, regulation by ubiquitin and ubiquitin-like modifiers is still an emerging area of investigation. This review will examine how ubiquitin, E3 ubiquitin ligases, and other ubiquitin-like modifications such as SUMO and NEDD8 regulate TCR complex-mediated signaling. PMID:25628960

Biotin-Tagged Polysaccharide Vesicular Nanocarriers for Receptor-Mediated Anticancer Drug Delivery in Cancer Cells.

PubMed

Deshpande, Nilesh Umakant; Jayakannan, Manickam

2018-06-15

Biotin-conjugated multi-stimuli-responsive polysaccharide vesicular nanocarriers are designed and developed, for the first time, to accomplish receptor-mediated endocytosis in cancer cells and to deliver anticancer drugs at the intracellular compartments. For this purpose, a new renewable hydrophobic unit was custom designed with redox-degradable disulphide and enzyme-biodegradable aliphatic ester chemical linkages and it was conjugated along with biotin on the dextran backbone. The dextran derivative self-assembled into nanovesicles of < 200 nm in size which were characterized by dynamic and static light scattering, electron and atomic force microscopes. Avidin-HABA assay established the high affinity of biotin-tagged dextran vesicles towards membrane-receptors up to 25 nM concentration. Doxorubicin-hydrochloride (DOX.HCl) loaded dextran vesicles exhibited stable formulation in PBS and FBS. Redox-degradation by glutathione (GSH) showed 60 % drug release whereas lysosomal esterase enzyme enabled > 98 % drug release in 12 h. Confocal microscope and flow cytometry assisted time-dependent cellular uptake studies revealed that the biotin-receptor over expressed cervical cancer cells (HeLa) exhibited larger drug accumulation through receptor-assisted endocytosis process. This process enabled the delivery of higher amount of DOX and significantly enhanced the killing in cancer cells (HeLa) compared to wild-type mouse embryonic fibroblast cells (WT-MEF, normal cells). Control experiments such as biotin pre-treatment in cancer cells and energy-suppressed cellular uptake at 4 C further supported the occurrence of receptor-mediated endocytosis by the biotin-tagged polymer vesicles. This report provides first insights into the targeted polysaccharide vesicle platform, and the proof-of-concept is successfully demonstrated in biotin receptors over expressed cervical cancer cells.

The anti-inflammatory effects of PGE2 on human lung macrophages are mediated by the EP4 receptor.

PubMed

Gill, Sharonjit K; Yao, Yiwen; Kay, Linda J; Bewley, Martin A; Marriott, Helen M; Peachell, Peter T

2016-11-01

PGE 2 inhibits cytokine generation from human lung macrophages. However, the EP receptor that mediates this beneficial anti-inflammatory effect of PGE 2 has not been defined. The aim of this study was to identify the EP receptor by which PGE 2 inhibits cytokine generation from human lung macrophages. This was determined by using recently developed EP receptor ligands. The effects of PGE 2 and EP-selective agonists on LPS-induced generation of TNF-α and IL-6 from macrophages were evaluated. The effects of EP 2 -selective (PF-04852946, PF-04418948) and EP 4 -selective (L-161,982, CJ-042794) receptor antagonists on PGE 2 responses were studied. The expression of EP receptor subtypes by human lung macrophages was determined by RT-PCR. PGE 2 inhibited LPS-induced and Streptococcus pneumoniae-induced cytokine generation from human lung macrophages. Analysis of mRNA levels indicated that macrophages expressed EP 2 and EP 4 receptors. L-902,688 (EP 4 receptor-selective agonist) was considerably more potent than butaprost (EP 2 receptor-selective agonist) as an inhibitor of TNF-α generation from macrophages. EP 2 receptor-selective antagonists had marginal effects on the PGE 2 inhibition of TNF-α generation, whereas EP 4 receptor-selective antagonists caused rightward shifts in the PGE 2 concentration-response curves. These studies demonstrate that the EP 4 receptor is the principal receptor that mediates the anti-inflammatory effects of PGE 2 on human lung macrophages. This suggests that EP 4 receptor agonists could be effective anti-inflammatory agents in human lung disease. © 2016 The British Pharmacological Society.

The CB1 Receptor as an Important Mediator of Hedonic Reward Processing

PubMed Central

Friemel, Chris M; Zimmer, Andreas; Schneider, Miriam

2014-01-01

The endocannabinoid (ECB) system has emerged recently as a key mediator for reward processing. It is well known that cannabinoids affect appetitive learning processes and can induce reinforcing and rewarding effects. However, the involvement of the ECB system in hedonic aspects of reward-related behavior is not completely understood. With the present study, we investigated the modulatory role of the ECB system on hedonic perception, measured by the pleasure attenuated startle (PAS) paradigm for a palatable food reward. Here, a conditioned odor is thought to induce a pleasant affective state that attenuates an aversive reflex—the acoustic startle response. Modulatory effects of the CB1 receptor antagonist/inverse agonist SR1411716 and the cannabinoid agonist WIN 55 212-2 on PAS were examined in rats. PAS was also measured in CB1 receptor knockout (KO) and wild-type (WT) mice. Pharmacological inhibition as well as the absence of CB1 receptors was found to reduce PAS, whereas WIN 55 212-2 administration increased PAS. Finally, presentation of a conditioned reward cue was found to induce striatal FosB/ΔFosB expression in WT mice, but not in KO mice, indicating a reduced stimulation of reward-related brain regions in conditioned KO mice by odor presentation. We here show that in addition to our previous studies in rats, PAS may also serve as a valuable and suitable measure to assess hedonic processing in mice. Our data further indicate that the ECB system, and in particular CB1 receptor signaling, appears to be highly important for the mediation of hedonic aspects of reward processing. PMID:24718372

Beta receptor-mediated modulation of the late positive potential in humans.

PubMed

de Rover, Mischa; Brown, Stephen B R E; Boot, Nathalie; Hajcak, Greg; van Noorden, Martijn S; van der Wee, Nic J A; Nieuwenhuis, Sander

2012-02-01

Electrophysiological studies have identified a scalp potential, the late positive potential (LPP), which is modulated by the emotional intensity of observed stimuli. Previous work has shown that the LPP reflects the modulation of activity in extrastriate visual cortical structures, but little is known about the source of that modulation. The present study investigated whether beta-adrenergic receptors are involved in the generation of the LPP. We used a genetic individual differences approach (experiment 1) and a pharmacological manipulation (experiment 2) to test the hypothesis that the LPP is modulated by the activation of β-adrenergic receptors. In experiment 1, we found that LPP amplitude depends on allelic variation in the β1-receptor gene polymorphism. In experiment 2, we found that LPP amplitude was modulated by the β-blocker propranolol in a direction dependent on subjects' level of trait anxiety: In participants with lower trait anxiety, propranolol led to a (nonsignificant) decrease in the LPP modulation; in participants with higher trait anxiety, propranolol increased the emotion-related LPP modulation. These results provide initial support for the hypothesis that the LPP reflects the downstream effects, in visual cortical areas, of β-receptor-mediated activation of the amygdala.

Estrogen Receptor-Related Receptor α Mediates Up-Regulation of Aromatase Expression by Prostaglandin E2 in Prostate Stromal Cells

PubMed Central

Miao, Lin; Shi, Jiandang; Wang, Chun-Yu; Zhu, Yan; Du, Xiaoling; Jiao, Hongli; Mo, Zengnan; Klocker, Helmut; Lee, Chung; Zhang, Ju

2010-01-01

Estrogen receptor-related receptor α (ERRα) is an orphan member of the nuclear receptor superfamily of transcription factors. ERRα is highly expressed in the prostate, especially in prostate stromal cells. However, little is known about the regulation and function of ERRα, which may contribute to the progression of prostatic diseases. We previously found that prostaglandin E2 (PGE2) up-regulated the expression of aromatase in prostate stromal cells. Here we show that PGE2 also up-regulates the expression of ERRα, which, as a transcription factor, further mediates the regulatory effects of PGE2 on the expression of aromatase. ERRα expression was up-regulated by PGE2 in prostate stromal cell line WPMY-1, which was mediated mainly through the protein kinase A signaling pathway by PGE2 receptor EP2. Suppression of ERRα activity by chlordane (an antagonist of ERRα) or small interfering RNA knockdown of ERRα blocked the increase of expression and promoter activity of aromatase induced by PGE2. Overexpression of ERRα significantly increased aromatase expression and promoter activity, which were further augmented by PGE2. Chromatin immunoprecipitation assay demonstrated that ERRα directly bound to the aromatase promoter in vivo, and PGE2 enhanced the recruitment of ERRα and promoted transcriptional regulatory effects on aromatase expression in WPMY-1. 17β-Estradiol concentration in WPMY-1 medium was up-regulated by ERRα expression, and that was further increased by PGE2. Our results provided evidence that ERRα contributed to local estrogen production by up-regulating aromatase expression in response to PGE2 and provided further insights into the potential role of ERRα in estrogen-related prostatic diseases. PMID:20351196

Toll Receptor-Mediated Hippo Signaling Controls Innate Immunity in Drosophila.

PubMed

Liu, Bo; Zheng, Yonggang; Yin, Feng; Yu, Jianzhong; Silverman, Neal; Pan, Duojia

2016-01-28

The Hippo signaling pathway functions through Yorkie to control tissue growth and homeostasis. How this pathway regulates non-developmental processes remains largely unexplored. Here, we report an essential role for Hippo signaling in innate immunity whereby Yorkie directly regulates the transcription of the Drosophila IκB homolog, Cactus, in Toll receptor-mediated antimicrobial response. Loss of Hippo pathway tumor suppressors or activation of Yorkie in fat bodies, the Drosophila immune organ, leads to elevated cactus mRNA levels, decreased expression of antimicrobial peptides, and vulnerability to infection by Gram-positive bacteria. Furthermore, Gram-positive bacteria acutely activate Hippo-Yorkie signaling in fat bodies via the Toll-Myd88-Pelle cascade through Pelle-mediated phosphorylation and degradation of the Cka subunit of the Hippo-inhibitory STRIPAK PP2A complex. Our studies elucidate a Toll-mediated Hippo signaling pathway in antimicrobial response, highlight the importance of regulating IκB/Cactus transcription in innate immunity, and identify Gram-positive bacteria as extracellular stimuli of Hippo signaling under physiological settings. Copyright © 2016 Elsevier Inc. All rights reserved.

Mu-Opioid (MOP) receptor mediated G-protein signaling is impaired in specific brain regions in a rat model of schizophrenia.

PubMed

Szűcs, Edina; Büki, Alexandra; Kékesi, Gabriella; Horváth, Gyöngyi; Benyhe, Sándor

2016-04-21

Schizophrenia is a complex mental health disorder. Clinical reports suggest that many patients with schizophrenia are less sensitive to pain than other individuals. Animal models do not interpret schizophrenia completely, but they can model a number of symptoms of the disease, including decreased pain sensitivities and increased pain thresholds of various modalities. Opioid receptors and endogenous opioid peptides have a substantial role in analgesia. In this biochemical study we investigated changes in the signaling properties of the mu-opioid (MOP) receptor in different brain regions, which are involved in the pain transmission, i.e., thalamus, olfactory bulb, prefrontal cortex and hippocampus. Our goal was to compare the transmembrane signaling mediated by MOP receptors in control rats and in a recently developed rat model of schizophrenia. Regulatory G-protein activation via MOP receptors were measured in [(35)S]GTPγS binding assays in the presence of a highly selective MOP receptor peptide agonist, DAMGO. It was found that the MOP receptor mediated activation of G-proteins was substantially lower in membranes prepared from the 'schizophrenic' model rats than in control animals. The potency of DAMGO to activate MOP receptor was also decreased in all brain regions studied. Taken together in our rat model of schizophrenia, MOP receptor mediated G-proteins have a reduced stimulatory activity compared to membrane preparations taken from control animals. The observed distinct changes of opioid receptor functions in different areas of the brain do not explain the augmented nociceptive threshold described in these animals. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Nuclear Membranes ETB Receptors Mediate ET-1-induced Increase of Nuclear Calcium in Human Left Ventricular Endocardial Endothelial Cells.

PubMed

Jules, Farah; Avedanian, Levon; Al-Khoury, Johny; Keita, Ramatoulaye; Normand, Alexandre; Bkaily, Ghassan; Jacques, Danielle

2015-07-01

In fetal human left ventricular endocardial endothelial cells (EECLs), both plasma membrane (PM) ET(A)R and ET(B)R were reported to mediate ET-1-induced increase of intracellular calcium [Ca](i); however, this effect was mediated by ET(A)R in right EECs (EECRs). In this study, we verified whether, as for the PM, nuclear membranes (NMs) ET-1 receptors activation in EECLs and EECRs induce an increase of nuclear calcium ([Ca](n)) and if this effect is mediated through the same receptor type as in PM. Using a plasmalemma-perforated technique and 3D confocal microscopy, our results showed that, as in PM intact cells, superfusion of nuclei of both cell types with cytosolic ET-1 induced a concentration-dependent sustained increase of [Ca](n). In EECRs, the ET(A)R antagonist prevented the effect of ET-1 on [Ca](n) without affecting EECLs. However, in both cell types, the effect of cytosolic ET-1 on [Ca](n) was prevented by the ETBR antagonist. In conclusion, both NMs' ET(A)R and ET(B)R mediated the effect of cytosolic ET-1 on [Ca](n) in EECRs. In contrast, only NMs' ET(B)R activation mediated the effect of cytosolic ET-1 in EECLs. Hence, the type of NMs' receptors mediating the effect of ET-1 on [Ca](n) are different from those of PM mediating the increase in [Ca](i).

Neurokinin-1 receptor mediates stress-exacerbated allergic airway inflammation and airway hyperresponsiveness in mice.

PubMed

Joachim, Ricarda A; Sagach, Viktoriya; Quarcoo, David; Dinh, Q Thai; Arck, Petra C; Klapp, Burghard F

2004-01-01

A wealth of clinical observation has suggested that stress and asthma morbidity are associated. We have previously established a mouse model of stress-exacerbated allergic airway inflammation, which reflects major clinical findings. The aim of the current study was to investigate the role of the neurokinin- (NK-)1 receptor in the mediation of stress effects in allergic airway inflammation. BALB/c mice were systemically sensitized with ovalbumin (OVA) on assay days 1, 14, and 21 and repeatedly challenged with OVA aerosol on days 26 and 27. Sound stress was applied to the animals for 24 hours, starting with the first airway challenge. Additionally, one group of stressed and one group of nonstressed mice received the highly specific NK-1 receptor antagonist RP 67580. Bronchoalveolar lavage fluid was obtained, and cell numbers and differentiation were determined. Airway hyperreactivity was measured in vitro by electrical field stimulation of tracheal smooth-muscle elements. Application of stress in sensitized and challenged animals resulted in a significant increase in leukocyte number in the bronchoalveolar lavage fluid. Furthermore, stressed animals showed enhanced airway reactivity. The increase of inflammatory cells and airway reactivity was blocked by treatment of animals with the NK-1 receptor antagonist. These data indicate that the NK-1 receptor plays an important role in mediating stress effects in allergen-induced airway inflammation.

Neuron-to-glia signaling mediated by excitatory amino acid receptors regulates ErbB receptor function in astroglial cells of the neuroendocrine brain.

PubMed

Dziedzic, Barbara; Prevot, Vincent; Lomniczi, Alejandro; Jung, Heike; Cornea, Anda; Ojeda, Sergio R

2003-02-01

Hypothalamic astroglial erbB tyrosine kinase receptors are required for the timely initiation of mammalian puberty. Ligand-dependent activation of these receptors sets in motion a glia-to-neuron signaling pathway that prompts the secretion of luteinizing hormone-releasing hormone (LHRH), the neuropeptide controlling sexual development, from hypothalamic neuroendocrine neurons. The neuronal systems that may regulate this growth factor-mediated back signaling to neuroendocrine neurons have not been identified. Here we demonstrate that hypothalamic astrocytes contain metabotropic receptors of the metabotropic glutamate receptor 5 subtype and the AMPA receptor subunits glutamate receptor 2 (GluR2) and GluR3. As in excitatory synapses, these receptors are in physical association with their respective interacting/clustering proteins Homer and PICK1. In addition, they are associated with erbB-1 and erbB-4 receptors. Concomitant activation of astroglial metabotropic and AMPA receptors results in the recruitment of erbB tyrosine kinase receptors and their respective ligands to the glial cell membrane, transactivation of erbB receptors via a mechanism requiring metalloproteinase activity, and increased erbB receptor gene expression. By facilitating erbB-dependent signaling and promoting erbB receptor gene expression in astrocytes, a neuron-to-glia glutamatergic pathway may represent a basic cell-cell communication mechanism used by the neuroendocrine brain to coordinate the facilitatory transsynaptic and astroglial input to LHRH neurons during sexual development.

LSD and DOB: interaction with 5-HT2A receptors to inhibit NMDA receptor-mediated transmission in the rat prefrontal cortex.

PubMed

Arvanov, V L; Liang, X; Russo, A; Wang, R Y

1999-09-01

Both the phenethylamine hallucinogen (-)-1-2, 5-dimethoxy-4-bromophenyl-2-aminopropane (DOB), a selective serotonin 5-HT2A,2C receptor agonist, and the indoleamine hallucinogen D-lysergic acid diethylamide (LSD, which binds to 5-HT1A, 1B, 1D, 1E, 1F, 2A, 2C, 5, 6, 7, dopamine D1 and D2, and alpha1 and alpha2 adrenergic receptors), but not their non-hallucinogenic congeners, inhibited N-methyl-D-aspartate (NMDA)-induced inward current and NMDA receptor-mediated synaptic responses evoked by electrical stimulation of the forceps minor in pyramidal cells of the prefrontal cortical slices. The inhibitory effect of hallucinogens was mimicked by 5-HT in the presence of selective 5-HT1A and 5-HT3 receptor antagonists. The inhibitory action of DOB, LSD and 5-HT on the NMDA transmission was blocked by the 5-HT2A receptor antagonists R-(+)-alpha-(2, 3-dimethoxyphenil)-1-[4-fluorophenylethyl]-4-piperidineme thanol (M100907) and ketanserin. However, at low concentrations, when both LSD and DOB by themselves only partially depressed the NMDA response, they blocked the inhibitory effect of 5-HT, suggesting a partial agonist action. Whereas N-(4-aminobutyl)-5-chloro-2-naphthalenesulphonamide (W-7, a calmodulin antagonist) and N-[2-[[[3-(4'-chlorophenyl)- 2-propenyl]methylamino]methyl]phenyl]-N-(2-hydroxyethyl)-4'-methoxy-b enzenesulphonamide phosphate (KN-93, a Ca2+/CaM-KII inhibitor), but not the negative control 2-[N-4'methoxybenzenesulphonyl]amino-N-(4'-chlorophenyl)-2-propeny l-N -methylbenzylamine phosphate (KN-92), blocked the inhibitory action of LSD and DOB, the selective protein kinase C inhibitor chelerythrine was without any effect. We conclude that phenethylamine and indoleamine hallucinogens may exert their hallucinogenic effect by interacting with 5-HT2A receptors via a Ca2+/CaM-KII-dependent signal transduction pathway as partial agonists and modulating the NMDA receptors-mediated sensory, perceptual, affective and cognitive processes.

Centralization of Noxious Stimulus-induced Analgesia (NSIA) is Related to Activity at Inhibitory Synapses in the Spinal Cord

PubMed Central

Tambeli, Claudia H.; Levine, Jon D.; Gear, Robert W.

2009-01-01

The duration of noxious stimulus-induced antinociception (NSIA) has been shown to outlast the pain stimulus that elicited it, however, the mechanism that determines the duration of analgesia is unknown. We evaluated the role of spinal excitatory and inhibitory receptors (NMDA, mGluR-5, mu-opioid, GABA-A, and GABA-B), previously implicated in NSIA initiation, in its maintenance. As in our previous studies, the supraspinal trigeminal jaw-opening reflex (JOR) in the rat was used for nociceptive testing because of its remoteness from the region of drug application, the lumbar spinal cord. NSIA was reversed by antagonists for two inhibitory receptors (GABA-B and mu-opioid) but not by antagonists for either of the two excitatory receptors (NMDA and mGluR-5), indicating that NSIA is maintained by ongoing activity at inhibitory synapses in the spinal cord. Furthermore, spinal administration of the GABA-B agonist baclofen mimicked NSIA in that it could be blocked by prior injection of the mu-opioid receptor antagonist H-D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) in nucleus accumbens. CTAP also blocked baclofen antinociception when administered in the spinal cord. We conclude that analgesia induced by noxious stimulation is maintained by activity in spinal inhibitory receptors. PMID:19375225

Characterization of the EP receptor types that mediate longitudinal smooth muscle contraction of human colon, mouse colon and mouse ileum.

PubMed

Fairbrother, S E; Smith, J E; Borman, R A; Cox, H M

2011-08-01

Prostaglandin E(2) (PGE(2) ) is an inflammatory mediator implicated in several gastrointestinal pathologies that affect normal intestinal transit. The aim was to establish the contribution of the four EP receptor types (EP(1-4) ), in human colon, that mediate PGE(2) -induced longitudinal smooth muscle contraction. Changes in isometric muscle tension of human colon, mouse colon and mouse ileum were measured in organ baths in response to receptor-specific agonists and antagonists. In addition, lidocaine was used to block neurogenic activity to investigate whether EP receptors were pre- or post-junctional. PGE(2) contracted longitudinal muscle from human and mouse colon and mouse ileum. These contractions were inhibited by the EP(1) receptor antagonist, EP(1) A in human colon, whereas a combination of EP(1) A and the EP(3) antagonist, L798106 inhibited agonist responses in both mouse preparations. The EP(3) agonist, sulprostone also increased muscle tension in both mouse tissues, and these responses were inhibited by lidocaine in the colon but not in the ileum. Although PGE(2) consistently contracted all three muscle preparations, butaprost decreased tension by activating smooth muscle EP(2) receptors in both colonic tissues. Alternatively, in mouse ileum, butaprost responses were lidocaine-sensitive, suggesting that it was activating prejunctional EP(2) receptors on inhibitory motor neurons. Conversely, EP(4) receptors were not functional in all the intestinal muscle preparations tested. PGE(2) -induced contraction of longitudinal smooth muscle is mediated by EP(1) receptors in human colon and by a combination of EP(1) and EP(3) receptors in mouse intestine, whereas EP(2) receptors modulate relaxation in all three preparations. © 2011 Blackwell Publishing Ltd.

Molecular basis for repression of liver X receptor-mediated gene transcription by receptor-interacting protein 140

PubMed Central

Jakobsson, Tomas; Osman, Waffa; Gustafsson, Jan-Åke; Zilliacus, Johanna; Wärnmark, Anette

2007-01-01

Similarities in physiological roles of LXR (liver X receptors) and co-repressor RIP140 (receptor-interacting protein 140) in regulating energy homoeostasis and lipid and glucose metabolism suggest that the effects of LXR could at least partly be mediated by recruitment of the co-repressor RIP140. In the present study, we have elucidated the molecular basis for regulation of LXR transcriptional activity by RIP140. LXR is evenly localized in the nucleus and neither the N-terminal domain nor the LBD (ligand-binding domain) is necessary for nuclear localization. Both LXR subtypes, LXRα and LXRβ, interact with RIP140 and co-localize in diffuse large nuclear domains. Interaction and co-localization are dependent on the LBD of the receptor. The C-terminal domain of RIP140 is sufficient for full repressive effect. None of the C-terminal NR (nuclear receptor)-boxes is required for the co-repressor activity, whereas the NR-box-like motif as well as additional elements in the C-terminal region are required for full repressive function. The C-terminal NR-box-like motif is necessary for interaction with LXRβ, whereas additional elements are needed for strong interaction with LXRα. In conclusion, our results suggest that co-repression of LXR activity by RIP140 involves an atypical binding mode of RIP140 and a repression element in the RIP140 C-terminus. PMID:17391100

Further evidence that tachykinin-induced contraction of human isolated bronchus is mediated only by NK2-receptors.

PubMed

Sheldrick, R L; Rabe, K F; Fischer, A; Magnussen, H; Coleman, R A

1995-11-01

The tachykinin-receptors mediating contraction of human bronchus have been characterized using both tachykinin-receptor selective agonists and blocking drugs under conditions where tachykinin metabolism by endogenous peptidases has been controlled, and true equilibrium conditions have been established. The findings that neurokinin A (EC50 = 2 nM) is the most potent agonist, and the NK2-receptor selective agonist, GR64349, is only 3-fold weaker, whereas agonists selective for NK1-receptors, substance P methyl ester, or NK3-receptors, senktide, are inactive, suggest that this effect is mediated exclusively by NK2-receptors. This is supported by observations that GR64349 is antagonised by the selective NK2-receptor blocking drugs, MEN10207 (pA2 = 6.7), R396 (pA2 = 6.1), (+/-)SR48968 (pA2 = 8.4) and GR159897 (pA2 = 8.6), but not by the NK1-receptor blocking drug, GR82334 (pA2 < 5). In approximately half of the preparations, the peptidase inhibitors, phosphoramidon (1 microM) and bestatin (100 microM), caused a marked and well-maintained contraction (approximately 20% of neurokinin A maximum), which may indicate a role for endogenous tachykinins in the regulation of tone in this preparation. This is supported by the finding that neurokinin A-immunoreactive nerve fibres are located around intrinsic neurones of local ganglia and within the smooth muscle layer of this preparation.

Both retinoic-acid-receptor- and retinoid-X-receptor-dependent signalling pathways mediate the induction of the brown-adipose-tissue-uncoupling-protein-1 gene by retinoids.

PubMed Central

Alvarez, R; Checa, M; Brun, S; Viñas, O; Mampel, T; Iglesias, R; Giralt, M; Villarroya, F

2000-01-01

The intracellular pathways and receptors mediating the effects of retinoic acid (RA) on the brown-fat-uncoupling-protein-1 gene (ucp-1) have been analysed. RA activates transcription of ucp-1 and the RA receptor (RAR) is known to be involved in this effect. However, co-transfection of an expression vector for retinoid-X receptor (RXR) increases the action of 9-cis RA but not the effects of all-trans RA on the ucp-1 promoter in brown adipocytes. Either RAR-specific ¿p-[(E)-2-(5,6,7,8,-tetrahydro-5,5,8, 8-tetramethyl-2-naphthalenyl)-1-propenyl]benzoic acid¿ or RXR-specific [isopropyl-(E,E)-(R,S)-11-methoxy-3,7, 11-trimethyldodeca-2,4-dienoate, or methoprene] synthetic compounds increase the expression of UCP-1 mRNA and the activity of chloramphenicol acetyltransferase expression vectors driven by the ucp-1 promoter. The RXR-mediated action of 9-cis RA requires the upstream enhancer region at -2469/-2318 in ucp-1. During brown-adipocyte differentiation RXRalpha and RXRgamma mRNA expression is induced in parallel with UCP-1 mRNA, whereas the mRNA for the three RAR subtypes, alpha, beta and gamma, decreases. Co-transfection of murine expression vectors for the different RAR and RXR subtypes indicates that RARalpha and RARbeta as well as RXRalpha are the major retinoid-receptor subtypes capable of mediating the responsiveness of ucp-1 to retinoids. It is concluded that the effects of retinoids on ucp-1 transcription involve both RAR- and RXR-dependent signalling pathways. The responsiveness of brown adipose tissue to retinoids in vivo relies on a complex combination of the capacity of RAR and RXR subtypes to mediate ucp-1 induction and their distinct expression in the differentiated brown adipocyte. PMID:10600643

Direct involvement of sigma-1 receptors in the dopamine D1 receptor-mediated effects of cocaine.

PubMed

Navarro, Gemma; Moreno, Estefanía; Aymerich, Marisol; Marcellino, Daniel; McCormick, Peter J; Mallol, Josefa; Cortés, Antoni; Casadó, Vicent; Canela, Enric I; Ortiz, Jordi; Fuxe, Kjell; Lluís, Carmen; Ferré, Sergi; Franco, Rafael

2010-10-26

It is well known that cocaine blocks the dopamine transporter. This mechanism should lead to a general increase in dopaminergic neurotransmission, and yet dopamine D(1) receptors (D(1)Rs) play a more significant role in the behavioral effects of cocaine than the other dopamine receptor subtypes. Cocaine also binds to σ-1 receptors, the physiological role of which is largely unknown. In the present study, D(1)R and σ(1)R were found to heteromerize in transfected cells, where cocaine robustly potentiated D(1)R-mediated adenylyl cyclase activation, induced MAPK activation per se and counteracted MAPK activation induced by D(1)R stimulation in a dopamine transporter-independent and σ(1)R-dependent manner. Some of these effects were also demonstrated in murine striatal slices and were absent in σ(1)R KO mice, providing evidence for the existence of σ(1)R-D(1)R heteromers in the brain. Therefore, these results provide a molecular explanation for which D(1)R plays a more significant role in the behavioral effects of cocaine, through σ(1)R-D(1)R heteromerization, and provide a unique perspective toward understanding the molecular basis of cocaine addiction.

Metabotropic glutamate receptor-mediated signaling dampens the HPA axis response to restraint stress

PubMed Central

Evanson, Nathan K.; Herman, James P.

2015-01-01

Glutamate is an important neurotransmitter in regulation of the neural portion of hypothalamus-pituitary-adrenal (HPA) axis activity, and signals through ionotropic and metabotropic receptors. In the current studies we investigated the role of hypothalamic paraventricular group I metabotropic glutamate receptors in regulation of the HPA axis response to restraint stress in rats. Direct injection of the group I metabotropic glutamate receptor agonist 3,5-dihydroxyphenylglycine (DHPG) into the PVN prior to restraint leads to blunting of the HPA axis response in awake animals. Consistent with this result, infusion of the group I receptor antagonist hexyl-homoibotenic acid (HIBO) potentiates the HPA axis response to restraint. The excitatory effect of blocking paraventricular group I metabotropic glutamate signaling is blocked by co-administration of dexamethasone into the PVN. However, the inhibitory effect of DHPG is not affected by co-administration of the cannabinoid CB1 receptor antagonist AM-251 into the PVN. Together, these results suggest that paraventricular group I metabotropic glutamate receptor signaling acts to dampen HPA axis reactivity. This effect appears to be similar to the rapid inhibitory effect of glucocorticoids at the PVN, but is not mediated by endocannabinoid signaling. PMID:25701594

P2Y6 receptor mediates colonic NaCl secretion via differential activation of cAMP-mediated transport

PubMed Central

Köttgen, Michael; Löffler, Thomas; Jacobi, Christoph; Nitschke, Roland; Pavenstädt, Hermann; Schreiber, Rainer; Frische, Sebastian; Nielsen, Søren; Leipziger, Jens

2003-01-01

Extracellular nucleotides are important regulators of epithelial ion transport. Here we investigated nucleotide-mediated effects on colonic NaCl secretion and the signal transduction mechanisms involved. Basolateral UDP induced a sustained activation of Cl– secretion, which was completely inhibited by 293B, a specific inhibitor of cAMP-stimulated basolateral KCNQ1/KCNE3 K+ channels. We therefore speculated that a basolateral P2Y6 receptor could increase cAMP. Indeed UDP elevated cAMP in isolated crypts. We identified an epithelial P2Y6 receptor using crypt [Ca2+]i measurements, RT-PCR, and immunohistochemistry. To investigate whether the rat P2Y6elevates cAMP, we coexpressed the P2Y1 or P2Y6 receptor together with the cAMP-regulated cystic fibrosis transmembrane conductance regulator (CFTR) Cl– channel in Xenopus oocytes. A two-electrode voltage clamp was used to monitor nucleotide-induced Cl– currents. In oocytes expressing the P2Y1 receptor, ATP transiently activated the endogenous Ca2+-activated Cl– current, but not CFTR. In contrast, in oocytes expressing the P2Y6receptor, UDP transiently activated the Ca2+-activated Cl– current and subsequently CFTR. CFTR Cl– currents were identified by their halide conductance sequence. In summary we find a basolateral P2Y6 receptor in colonic epithelial cells stimulating sustained NaCl secretion by way of a synergistic increase of [Ca2+]i and cAMP. In support of these data P2Y6 receptor stimulation differentially activates CFTR in Xenopus oocytes. PMID:12569163

Multivalent ligand-receptor-mediated interaction of small filled vesicles with a cellular membrane

NASA Astrophysics Data System (ADS)

Zhdanov, Vladimir P.

2017-07-01

The ligand-receptor-mediated contacts of small sub-100-nm-sized lipid vesicles (or nanoparticles) with the cellular membrane are of interest in the contexts of cell-to-cell communication, endocytosis of membrane-coated virions, and drug (RNA) delivery. In all these cases, the interior of vesicles is filled by biologically relevant content. Despite the diversity of such systems, the corresponding ligand-receptor interaction possesses universal features. One of them is that the vesicle-membrane contacts can be accompanied by the redistribution of ligands and receptors between the contact and contact-free regions. In particular, the concentrations of ligands and receptors may become appreciably higher in the contact regions and their composition may there be different compared to that in the suspended state in the solution. A statistical model presented herein describes the corresponding distribution of various ligands and receptors and allows one to calculate the related change of the free energy with variation of the vesicle-engulfment extent. The results obtained are used to clarify the necessary conditions for the vesicle-assisted pathway of drug delivery.

[Electrophysiological characteristics of hippocampal postnatal early development mediated by AMPA receptors in rats].

PubMed

Chen, Xue-Yi; Zhang, Ai-Feng; Zhao, Wen; Gao, Yu-Dan; Duan, Hong-Mei; Hao, Peng; Yang, Zhao-Yang; Li, Xiao-Guang

2018-04-25

The present study was aimed to investigate the electrophysiological characteristics of hippocampal postnatal early development mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors in rats. Forty-eight Wistar rats were divided into postnatal 0.5-, 1-, 2- and 3-month groups (n = 12). Spontaneous excitatory postsynaptic currents (sEPSCs) and field excitatory postsynaptic potentials (fEPSPs) mediated by AMPA receptors were recorded to evaluate the changes in the intrinsic membrane properties of hippocampal CA1 pyramidal neurons by using patch-clamp and MED64 planar microelectrode array technique respectively. The results showed that, during the period of postnatal 0.5-3 months, some of the intrinsic membrane properties of hippocampal CA1 pyramidal neurons, such as the membrane capacitance (Cm) and the resting membrane potential (RMP), showed no significant changes, while the membrane input resistance (Rin) and the time constant (τ) of the cells were decreased significantly. The amplitude, frequency and kinetics (both rise and decay times) of sEPSCs were significantly increased during the period of postnatal 0.5-1 month, but they were all decreased during the period of postnatal 1-3 months. In addition, the range of evoked fEPSPs in hippocamal CA1 region was significantly expanded, but the fEPSP amplitudes were decreased significantly during the period of postnatal 0.5-3 months. Furthermore, the evoked fEPSPs could be significantly inhibited by extracellular application of the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). These results suggest that AMPA receptor may act as a major type of excitatory receptor to regulate synaptic transmission and connections during the early stage of hippocampal postnatal development, which promotes the development and functional maturation of hippocampus in rats.

Control of glycinergic input to spinal dorsal horn neurons by distinct muscarinic receptor subtypes revealed using knockout mice.

PubMed

Zhang, Hong-Mei; Zhou, Hong-Yi; Chen, Shao-Rui; Gautam, Dinesh; Wess, Jürgen; Pan, Hui-Lin

2007-12-01

Muscarinic acetylcholine receptors (mAChRs) play an important role in the tonic regulation of nociceptive transmission in the spinal cord. However, how mAChR subtypes contribute to the regulation of synaptic glycine release is unknown. To determine their role, glycinergic spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded in lamina II neurons by using whole-cell recordings in spinal cord slices of wild-type (WT) and mAChR subtype knockout (KO) mice. In WT mice, the mAChR agonist oxotremorine-M dose-dependently decreased the frequency of sIPSCs in most neurons, but it had variable effects in other neurons. In contrast, in M3-KO mice, oxotremorine-M consistently decreased the glycinergic sIPSC frequency in all neurons tested, and in M2/M4 double-KO mice, it always increased the sIPSC frequency. In M2/M4 double-KO mice, the potentiating effect of oxotremorine-M was attenuated by higher concentrations in some neurons through activation of GABA(B) receptors. In pertussis toxin-treated WT mice, oxotremorine-M also consistently increased the sIPSC frequency. In M2-KO and M4-KO mice, the effect of oxotremorine-M on sIPSCs was divergent because of the opposing functions of the M3 subtype and the M2 and M4 subtypes. This study demonstrates that stimulation of the M2 and M4 subtypes inhibits glycinergic inputs to spinal dorsal horn neurons of mice, whereas stimulation of the M3 subtype potentiates synaptic glycine release. Furthermore, GABA(B) receptors are involved in the feedback regulation of glycinergic synaptic transmission in the spinal cord. This study revealed distinct functions of mAChR subtypes in controlling glycinergic input to spinal dorsal horn neurons.

Regulation of Androgen Receptor-Mediated Transcription by RPB5 Binding Protein URI/RMP ▿

PubMed Central

Mita, Paolo; Savas, Jeffrey N.; Djouder, Nabil; Yates, John R.; Ha, Susan; Ruoff, Rachel; Schafler, Eric D.; Nwachukwu, Jerome C.; Tanese, Naoko; Cowan, Nicholas J.; Zavadil, Jiri; Garabedian, Michael J.; Logan, Susan K.

2011-01-01

Androgen receptor (AR)-mediated transcription is modulated by interaction with coregulatory proteins. We demonstrate that the unconventional prefoldin RPB5 interactor (URI) is a new regulator of AR transcription and is critical for antagonist (bicalutamide) action. URI is phosphorylated upon androgen treatment, suggesting communication between the URI and AR signaling pathways. Whereas depletion of URI enhances AR-mediated gene transcription, overexpression of URI suppresses AR transcriptional activation and anchorage-independent prostate cancer cell growth. Repression of AR-mediated transcription is achieved, in part, by URI binding and regulation of androgen receptor trapped clone 27 (Art-27), a previously characterized AR corepressor. Consistent with this idea, genome-wide expression profiling in prostate cancer cells upon depletion of URI or Art-27 reveals substantially overlapping patterns of gene expression. Further, depletion of URI increases the expression of the AR target gene NKX-3.1, decreases the recruitment of Art-27, and increases AR occupancy at the NKX-3.1 promoter. While Art-27 can bind AR directly, URI is bound to chromatin prior to hormone-dependent recruitment of AR, suggesting a role for URI in modulating AR recruitment to target genes. PMID:21730289

Fetuin mediates hepatic uptake of negatively charged nanoparticles via scavenger receptor.

PubMed

Nagayama, Susumu; Ogawara, Ken-ichi; Minato, Keiko; Fukuoka, Yoshiko; Takakura, Yoshinobu; Hashida, Mitsuru; Higaki, Kazutaka; Kimura, Toshikiro

2007-02-01

We tried to evaluate the possible involvement of fetuin in the scavenger receptors (SRs)-mediated hepatic uptake of polystyrene nanospheres with the size of 50 nm (NS-50), which has surface negative charge (zeta potential=-21.8+/-2.3 mV). The liver perfusion studies in rats revealed that the hepatic uptake of NS-50 pre-coated with fetuin (NS-50-fetuin) was significantly inhibited by poly inosinic acid (poly I), a typical inhibitor of SRs, whereas that of plain NS-50 or NS-50 pre-coated with BSA (NS-50-BSA) was not. The uptake of NS-50-fetuin by cultured Kupffer cells was also significantly inhibited by poly I, and anti-class A scavenger receptors (SR-A) antibody, suggesting that fetuin on NS-50 mediated the recognition and internalization of NS-50 by Kupffer cells and at least SR-A would be responsible for the uptake. Taken that Western blot analysis confirmed that fetuin certainly adsorbed on the surface of NS-50 after the incubation of NS-50 with serum, the results obtained in the present study indicate that fetuin would be one of the serum proteins that were substantially involved in the hepatic uptake of NS-50 via SRs.

Mediation by neurotensin-receptors of effects of neurotensin on self-stimulation of the medial prefrontal cortex.

PubMed Central

Fernández, R.; Sabater, R.; Sáez, J. A.; Montes, R.; Alba, F.; Ferrer, J. M.

1996-01-01

1 Intracortical microinjections of neurotensin (NT) selectively decreased intracranial self-stimulation (ICSS) of the medial prefrontal cortex in the rat. 2 To elucidate whether this effect is mediated by NT receptors or by the formation of NT-dopamine complexes, we investigated the effects on ICSS of intracortical microinjections of neurotensin (1-11), an NT fragment that forms extracellular complexes with dopamine but does not bind to NT receptors. 3 We also studied the effects of the peripheral administration of SR 48692, a selective antagonist of NT receptors, on the inhibition of ICSS produced by the intracortical administration of NT. 4 Unilateral microinjections of neurotensin (1-11) at doses of 10, 20 and 40 nmol into the medial prefrontal cortex did not change the basal ICSS rate of this area. 5 The intraperitoneal administration of SR 48692 at doses of 0.08 and 0.16 mg kg-1 30 min before microinjection of 10 nmol of NT into the medial prefrontal cortex, antagonized the inhibition of ICSS produced by the neuropeptide. 6 These results demonstrate that the inhibitory effect of NT on ICSS is mediated by NT receptors. PMID:8886412

Microglia P2Y₆ receptors mediate nitric oxide release and astrocyte apoptosis.

PubMed

Quintas, Clara; Pinho, Diana; Pereira, Clara; Saraiva, Lucília; Gonçalves, Jorge; Queiroz, Glória

2014-09-03

During cerebral inflammation uracil nucleotides leak to the extracellular medium and activate glial pyrimidine receptors contributing to the development of a reactive phenotype. Chronically activated microglia acquire an anti-inflammatory phenotype that favors neuronal differentiation, but the impact of these microglia on astrogliosis is unknown. We investigated the contribution of pyrimidine receptors to microglia-astrocyte signaling in a chronic model of inflammation and its impact on astrogliosis. Co-cultures of astrocytes and microglia were chronically treated with lipopolysaccharide (LPS) and incubated with uracil nucleotides for 48 h. The effect of nucleotides was evaluated in methyl-[3H]-thymidine incorporation. Western blot and immunofluorescence was performed to detect the expression of P2Y6 receptors and the inducible form of nitric oxide synthase (iNOS). Nitric oxide (NO) release was quantified through Griess reaction. Cell death was also investigated by the LDH assay and by the TUNEL assay or Hoechst 33258 staining. UTP, UDP (0.001 to 1 mM) or PSB 0474 (0.01 to 10 μM) inhibited cell proliferation up to 43 ± 2% (n = 10, P <0.05), an effect prevented by the selective P2Y6 receptor antagonist MRS 2578 (1 μM). UTP was rapidly metabolized into UDP, which had a longer half-life. The inhibitory effect of UDP (1 mM) was abolished by phospholipase C (PLC), protein kinase C (PKC) and nitric oxide synthase (NOS) inhibitors. Both UDP (1 mM) and PSB 0474 (10 μM) increased NO release up to 199 ± 20% (n = 4, P <0.05), an effect dependent on P2Y6 receptors-PLC-PKC pathway activation, indicating that this pathway mediates NO release. Western blot and immunocytochemistry analysis indicated that P2Y6 receptors were expressed in the cultures being mainly localized in microglia. Moreover, the expression of iNOS was mainly observed in microglia and was upregulated by UDP (1 mM) or PSB 0474 (10 μM). UDP-mediated NO release induced apoptosis in astrocytes

Specific neurokinin receptors mediate plasma extravasation in the rat knee joint.

PubMed Central

Lam, F. Y.; Ferrell, W. R.

1991-01-01

1 Plasma extravasation in the rat knee joint was induced by intra-articular injection of neurokinins and specific neurokinin receptor agonists. 2 Pronounced plasma extravasation was produced by substance P (SP, 4-185 microM) and to a lesser extent by neurokinin-B (NKB, 83-413 microM), whereas neurokinin-A (NKA, 88-440 microM) and calcitonin gene-related peptide (CGRP, 26-130 microM) had no significant effect. 3 The specific neurokinin1 receptor agonist [Sar9, Met(O2)11]-substance P (NK1 agonist) in doses of 0.4-70 microM appeared to be more potent than SP in eliciting plasma extravasation. The neurokinin2 receptor agonist [Nle10]-neurokinin A4-10 (NK2 agonist) was not effective at 70 microM but produced a small and significant effect at 330 microM, whereas the neurokinin3 receptor agonist [MePhe7]-neurokinin B (NK3 agonist) was without effect at 40 microM or 400 microM. 4 Injections of SP or NKA into the synovial cavity of the rat knee were equally effective in producing marked plasma extravasation in remote sites such as the forelimb and hindlimb paws. 5 Co-administration experiments showed that the effects of SP were synergistic with NKA or the NK1 receptor agonist, but not with CGRP or the NK2 receptor agonist. 6 The rank order of potency was NK1 agonist greater than or equal to SP greater than NKB greater than NK2 agonist suggesting that NK1 receptors mediate plasma extravasation in the rat knee joint. PMID:1715229

Synergistic interaction between baclofen administration into the median raphe nucleus and inconsequential visual stimuli on investigatory behavior of rats

PubMed Central

Vollrath-Smith, Fiori R.; Shin, Rick

2011-01-01

Rationale Noncontingent administration of amphetamine into the ventral striatum or systemic nicotine increases responses rewarded by inconsequential visual stimuli. When these drugs are contingently administered, rats learn to self-administer them. We recently found that rats self-administer the GABAB receptor agonist baclofen into the median (MR) or dorsal (DR) raphe nuclei. Objectives We examined whether noncontingent administration of baclofen into the MR or DR increases rats’ investigatory behavior rewarded by a flash of light. Results Contingent presentations of a flash of light slightly increased lever presses. Whereas noncontingent administration of baclofen into the MR or DR did not reliably increase lever presses in the absence of visual stimulus reward, the same manipulation markedly increased lever presses rewarded by the visual stimulus. Heightened locomotor activity induced by intraperitoneal injections of amphetamine (3 mg/kg) failed to concur with increased lever pressing for the visual stimulus. These results indicate that the observed enhancement of visual stimulus seeking is distinct from an enhancement of general locomotor activity. Visual stimulus seeking decreased when baclofen was co-administered with the GABAB receptor antagonist, SCH 50911, confirming the involvement of local GABAB receptors. Seeking for visual stimulus also abated when baclofen administration was preceded by intraperitoneal injections of the dopamine antagonist, SCH 23390 (0.025 mg/kg), suggesting enhanced visual stimulus seeking depends on intact dopamine signals. Conclusions Baclofen administration into the MR or DR increased investigatory behavior induced by visual stimuli. Stimulation of GABAB receptors in the MR and DR appears to disinhibit the motivational process involving stimulus–approach responses. PMID:21904820

Ligand Receptor-Mediated Regulation of Growth in Plants.

PubMed

Haruta, Miyoshi; Sussman, Michael R

2017-01-01

Growth and development of multicellular organisms are coordinately regulated by various signaling pathways involving the communication of inter- and intracellular components. To form the appropriate body patterns, cellular growth and development are modulated by either stimulating or inhibiting these pathways. Hormones and second messengers help to mediate the initiation and/or interaction of the various signaling pathways in all complex multicellular eukaryotes. In plants, hormones include small organic molecules, as well as larger peptides and small proteins, which, as in animals, act as ligands and interact with receptor proteins to trigger rapid biochemical changes and induce the intracellular transcriptional and long-term physiological responses. During the past two decades, the availability of genetic and genomic resources in the model plant species, Arabidopsis thaliana, has greatly helped in the discovery of plant hormone receptors and the components of signal transduction pathways and mechanisms used by these immobile but highly complex organisms. Recently, it has been shown that two of the most important plant hormones, auxin and abscisic acid (ABA), act through signaling pathways that have not yet been recognized in animals. For example, auxins stimulate cell elongation by bringing negatively acting transcriptional repressor proteins to the proteasome to be degraded, thus unleashing the gene expression program required for increasing cell size. The "dormancy" inducing hormone, ABA, binds to soluble receptor proteins and inhibits a specific class of protein phosphatases (PP2C), which activates phosphorylation signaling leading to transcriptional changes needed for the desiccation of the seeds prior to entering dormancy. While these two hormone receptors have no known animal counterparts, there are also many similarities between animal and plant signaling pathways. For example, in plants, the largest single gene family in the genome is the protein kinase

The overexpressed human 46-kDa mannose 6-phosphate receptor mediates endocytosis and sorting of. beta. -glucuronidase

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

Watanabe, H.; Grubb, J.H.; Sly, W.S.

1990-10-01

The authors studied the function of the human small (46-kDa) mannose 6-phosphate receptor (SMPR) in transfected mouse L cells that do not express the larger insulin-like growth factor II/mannose 6-phosphate receptor. Cells overexpressing human SMPR were studied for enzyme binding to cell surface receptors, for binding to intracellular receptors in permeabilized cells, and for receptor-mediated endocytosis of recombinant human {beta}-glucuronidase. Specific binding to human SMPR in permeabilized cells showed a pH optimum between pH 6.0 and pH 6.5. Binding was significant in the present of EDTA but was enhanced by added divalent cations. Up to 2.3{percent} of the total functionalmore » receptor could be detected on the cell surface by enzyme binding. They present experiments showing that at very high levels of overexpression, and at pH 6.5, human SMPR mediated the endocytosis of {beta}-glucuronidase. At pH 7.5, the rate of endocytosis was only 14{percent} the rate seen at pH 6.5. Cells overexpressing human SMPR also showed reduced secretion of newly synthesized {beta}-glucuronidase when compared to cells transfected with vector only, suggesting that overexpressed human SMPR can participate in sorting of newly synthesized {beta}-glucuronidase and partially correct the sorting defect in mouse L cells that do not express the insulin-like growth factor II/mannose 6-phosphate receptor.« less

Oestrogen receptor-mediated expression of Olfactomedin 4 regulates the progression of endometrial adenocarcinoma

PubMed Central

Duan, Chao; Liu, Xubin; Liang, Shuang; Yang, Zheng; Xia, Meng; Wang, Liantang; Chen, Shangwu; Yu, Li

2014-01-01

Endometrial adenocarcinoma is the most common tumour of the female genital tract in developed countries, and oestrogen receptor (ER) signalling plays a pivotal role in its pathogenesis. When we used bioinformatics tools to search for the genes contributing to gynecological cancers, the expression of Olfactomedin 4 (OLFM4) was found by digital differential display to be associated with differentiation of endometrial adenocarcinoma. Aberrant expression of OLFM4 has been primarily reported in tumours of the digestive system. The mechanism of OLFM4 in tumuorigenesis is elusive. We investigated OLFM4 expression in endometrium, analysed the association of OLFM4 with ER signalling in endometrial adenocarcinoma, and examined the roles of OLFM4 in endometrial adenocarcinoma. Expression of OLFM4 was increased during endometrial carcinogenesis, linked to the differentiation of endometrioid adenocarcinoma, and positively related to the expression of oestrogen receptor-α (ERα) and progesterone receptor. Moreover, ERα-mediated signalling regulated expression of OLFM4, and knockdown of OLFM4 enhanced proliferation, migration and invasion of endometrial carcinoma cells. Down-regulation of OLFM4 was associated with decreased cumulative survival rate of patients with endometrioid adenocarcinoma. Our data suggested that impairment of ERα signal-mediated OLFM4 expression promoted the malignant progression of endometrioid adenocarcinoma, which may have significance for the therapy of this carcinoma. PMID:24495253

CXCR4 chemokine receptor signaling mediates pain in diabetic neuropathy

PubMed Central

2014-01-01

Background Painful Diabetic Neuropathy (PDN) is a debilitating syndrome present in a quarter of diabetic patients that has a substantial impact on their quality of life. Despite this significant prevalence and impact, current therapies for PDN are only partially effective. Moreover, the cellular mechanisms underlying PDN are not well understood. Neuropathic pain is caused by a variety of phenomena including sustained excitability in sensory neurons that reduces the pain threshold so that pain is produced in the absence of appropriate stimuli. Chemokine signaling has been implicated in the pathogenesis of neuropathic pain in a variety of animal models. We therefore tested the hypothesis that chemokine signaling mediates DRG neuronal hyperexcitability in association with PDN. Results We demonstrated that intraperitoneal administration of the specific CXCR4 antagonist AMD3100 reversed PDN in two animal models of type II diabetes. Furthermore DRG sensory neurons acutely isolated from diabetic mice displayed enhanced SDF-1 induced calcium responses. Moreover, we demonstrated that CXCR4 receptors are expressed by a subset of DRG sensory neurons. Finally, we observed numerous CXCR4 expressing inflammatory cells infiltrating into the DRG of diabetic mice. Conclusions These data suggest that CXCR4/SDF-1 signaling mediates enhanced calcium influx and excitability in DRG neurons responsible for PDN. Simultaneously, CXCR4/SDF-1 signaling may coordinate inflammation in diabetic DRG that could contribute to the development of pain in diabetes. Therefore, targeting CXCR4 chemokine receptors may represent a novel intervention for treating PDN. PMID:24961298

Astrocytes Regulate GLP-1 Receptor-Mediated Effects on Energy Balance

PubMed Central

Reiner, David J.; Mietlicki-Baase, Elizabeth G.; McGrath, Lauren E.; Zimmer, Derek J.; Bence, Kendra K.; Sousa, Gregory L.; Konanur, Vaibhav R.; Krawczyk, Joanna; Burk, David H.; Kanoski, Scott E.; Hermann, Gerlinda E.; Rogers, Richard C.

2016-01-01

Astrocytes are well established modulators of extracellular glutamate, but their direct influence on energy balance-relevant behaviors is largely understudied. As the anorectic effects of glucagon-like peptide-1 receptor (GLP-1R) agonists are partly mediated by central modulation of glutamatergic signaling, we tested the hypothesis that astrocytic GLP-1R signaling regulates energy balance in rats. Central or peripheral administration of a fluorophore-labeled GLP-1R agonist, exendin-4, localizes within astrocytes and neurons in the nucleus tractus solitarius (NTS), a hindbrain nucleus critical for energy balance control. This effect is mediated by GLP-1R, as the uptake of systemically administered fluorophore-tagged exendin-4 was blocked by central pretreatment with the competitive GLP-1R antagonist exendin-(9–39). Ex vivo analyses show prolonged exendin-4-induced activation (live cell calcium signaling) of NTS astrocytes and neurons; these effects are also attenuated by exendin-(9–39), indicating mediation by the GLP-1R. In vitro analyses show that the application of GLP-1R agonists increases cAMP levels in astrocytes. Immunohistochemical analyses reveal that endogenous GLP-1 axons form close synaptic apposition with NTS astrocytes. Finally, pharmacological inhibition of NTS astrocytes attenuates the anorectic and body weight-suppressive effects of intra-NTS GLP-1R activation. Collectively, data demonstrate a role for NTS astrocytic GLP-1R signaling in energy balance control. SIGNIFICANCE STATEMENT Glucagon-like peptide-1 receptor (GLP-1R) agonists reduce food intake and are approved by the Food and Drug Administration for the treatment of obesity, but the cellular mechanisms underlying the anorectic effects of GLP-1 require further investigation. Astrocytes represent a major cellular population in the CNS that regulates neurotransmission, yet the role of astrocytes in mediating energy balance is largely unstudied. The current data provide novel evidence that

Induction of osteoblast differentiation by selective activation of kinase-mediated actions of the estrogen receptor.

PubMed

Kousteni, Stavroula; Almeida, Maria; Han, Li; Bellido, Teresita; Jilka, Robert L; Manolagas, Stavros C

2007-02-01

Estrogens control gene transcription by cis or trans interactions of the estrogen receptor (ER) with target DNA or via the activation of cytoplasmic kinases. We report that selective activation of kinase-mediated actions of the ER with 4-estren-3alpha,17beta-diol (estren) or an estradiol-dendrimer conjugate, each a synthetic compound that stimulates kinase-mediated ER actions 1,000 to 10,000 times more potently than direct DNA interactions, induced osteoblastic differentiation in established cell lines of uncommitted osteoblast precursors and primary cultures of osteoblast progenitors by stimulating Wnt and BMP-2 signaling in a kinase-dependent manner. In sharp contrast, 17beta-estradiol (E(2)) suppressed BMP-2-induced osteoblast progenitor commitment and differentiation. Consistent with the in vitro findings, estren, but not E(2), stimulated Wnt/beta-catenin-mediated transcription in T-cell factor-lacZ transgenic mice. Moreover, E(2) stimulated BMP signaling in mice in which ERalpha lacks DNA binding activity and classical estrogen response element-mediated transcription (ERalpha(NERKI/-)) but not in wild-type controls. This evidence reveals for the first time the existence of a large signalosome in which inputs from the ER, kinases, bone morphogenetic proteins, and Wnt signaling converge to induce differentiation of osteoblast precursors. ER can either induce it or repress it, depending on whether the activating ligand (and presumably the resulting conformation of the receptor protein) precludes or accommodates ERE-mediated transcription.

Drug Management of Visceral Pain: Concepts from Basic Research

PubMed Central

Davis, Mellar P.

2012-01-01

Visceral pain is experienced by 40% of the population, and 28% of cancer patients suffer from pain arising from intra- abdominal metastasis or from treatment. Neuroanatomy of visceral nociception and neurotransmitters, receptors, and ion channels that modulate visceral pain are qualitatively or quantitatively different from those that modulate somatic and neuropathic pain. Visceral pain should be recognized as distinct pain phenotype. TRPV1, Na 1.8, and ASIC3 ion channels and peripheral kappa opioid receptors are important mediators of visceral pain. Mu agonists, gabapentinoids, and GABAB agonists reduce pain by binding to central receptors and channels. Combinations of analgesics and adjuvants in animal models have supra-additive antinociception and should be considered in clinical trials. This paper will discuss the neuroanatomy, receptors, ion channels, and neurotransmitters important to visceral pain and provide a basic science rationale for analgesic trials and management. PMID:22619712

Scavenger receptor mediates systemic RNA interference in ticks.

PubMed

Aung, Kyaw Min; Boldbaatar, Damdinsuren; Umemiya-Shirafuji, Rika; Liao, Min; Xuenan, Xuan; Suzuki, Hiroshi; Galay, Remil Linggatong; Tanaka, Tetsuya; Fujisaki, Kozo

2011-01-01

RNA interference is an efficient method to silence gene and protein expressions. Here, the class B scavenger receptor CD36 (SRB) mediated the uptake of exogenous dsRNAs in the induction of the RNAi responses in ticks. Unfed female Haemaphysalis longicornis ticks were injected with a single or a combination of H. longicornis SRB (HlSRB) dsRNA, vitellogenin-1 (HlVg-1) dsRNA, and vitellogenin receptor (HlVgR) dsRNA. We found that specific and systemic silencing of the HlSRB, HlVg-1, and HlVgR genes was achieved in ticks injected with a single dsRNA of HlSRB, HlVg-1, and HlVgR. In ticks injected first with HlVg-1 or HlVgR dsRNA followed 96 hours later with HlSRB dsRNA (HlVg-1/HlSRB or HlVgR/HlSRB), gene silencing of HlSRB was achieved in addition to first knockdown in HlVg-1 or HlVgR, and prominent phenotypic changes were observed in engorgement, mortality, and hatchability, indicating that a systemic and specific double knockdown of target genes had been simultaneously attained in these ticks. However, in ticks injected with HlSRB dsRNA followed 96 hours later with HlVg-1 or HlVgR dsRNAs, silencing of HlSRB was achieved, but no subsequent knockdown in HlVgR or HlVg-1 was observed. The Westernblot and immunohistochemical examinations revealed that the endogenous HlSRB protein was fully abolished in midguts of ticks injected with HlSRB/HlVg-1 dsRNAs but HlVg-1 was normally expressed in midguts, suggesting that HlVg-1 dsRNA-mediated RNAi was fully inhibited by the first knockdown of HlSRB. Similarly, the abolished localization of HlSRB protein was recognized in ovaries of ticks injected with HlSRB/HlVgR, while normal localization of HlVgR was observed in ovaries, suggesting that the failure to knock-down HlVgR could be attributed to the first knockdown of HlSRB. In summary, we demonstrated for the first time that SRB may not only mediate the effective knock-down of gene expression by RNAi but also play essential roles for systemic RNAi of ticks.

Scavenger Receptor Mediates Systemic RNA Interference in Ticks

PubMed Central

Aung, Kyaw Min; Boldbaatar, Damdinsuren; Umemiya-Shirafuji, Rika; Liao, Min; Xuenan, Xuan; Suzuki, Hiroshi; Linggatong Galay, Remil; Tanaka, Tetsuya; Fujisaki, Kozo

2011-01-01

RNA interference is an efficient method to silence gene and protein expressions. Here, the class B scavenger receptor CD36 (SRB) mediated the uptake of exogenous dsRNAs in the induction of the RNAi responses in ticks. Unfed female Haemaphysalis longicornis ticks were injected with a single or a combination of H. longicornis SRB (HlSRB) dsRNA, vitellogenin-1 (HlVg-1) dsRNA, and vitellogenin receptor (HlVgR) dsRNA. We found that specific and systemic silencing of the HlSRB, HlVg-1, and HlVgR genes was achieved in ticks injected with a single dsRNA of HlSRB, HlVg-1, and HlVgR. In ticks injected first with HlVg-1 or HlVgR dsRNA followed 96 hours later with HlSRB dsRNA (HlVg-1/HlSRB or HlVgR/HlSRB), gene silencing of HlSRB was achieved in addition to first knockdown in HlVg-1 or HlVgR, and prominent phenotypic changes were observed in engorgement, mortality, and hatchability, indicating that a systemic and specific double knockdown of target genes had been simultaneously attained in these ticks. However, in ticks injected with HlSRB dsRNA followed 96 hours later with HlVg-1 or HlVgR dsRNAs, silencing of HlSRB was achieved, but no subsequent knockdown in HlVgR or HlVg-1 was observed. The Westernblot and immunohistochemical examinations revealed that the endogenous HlSRB protein was fully abolished in midguts of ticks injected with HlSRB/HlVg-1 dsRNAs but HlVg-1 was normally expressed in midguts, suggesting that HlVg-1 dsRNA-mediated RNAi was fully inhibited by the first knockdown of HlSRB. Similarly, the abolished localization of HlSRB protein was recognized in ovaries of ticks injected with HlSRB/HlVgR, while normal localization of HlVgR was observed in ovaries, suggesting that the failure to knock-down HlVgR could be attributed to the first knockdown of HlSRB. In summary, we demonstrated for the first time that SRB may not only mediate the effective knock-down of gene expression by RNAi but also play essential roles for systemic RNAi of ticks. PMID:22145043

GABA(A) receptors mediate orexin-A induced stimulation of food intake.

PubMed

Kokare, Dadasaheb M; Patole, Angad M; Carta, Anna; Chopde, Chandrabhan T; Subhedar, Nishikant K

2006-01-01

Although the role of orexins in sleep/wake cycle and feeding behavior is well established, underlying mechanisms have not been fully understood. An attempt has been made to investigate the role of GABA(A) receptors and their benzodiazepine site on the orexin-A induced response to feeding. Different groups of rats were food deprived overnight and next day injected intracerebroventricularly (icv) with vehicle (artificial CSF; 5 microl/rat) or orexin-A (20-50 nM/rat) and the animals were given free access to food. Cumulative food intake was measured during light phase of light/dark cycle at 1-, 2-, 4- and 6-h post-injection time points. Orexin-A (30-50 nM/rat, icv) stimulated food intake at all the time points (P < 0.05). Prior administration of GABA(A) receptor agonists muscimol (25 ng/rat, icv) and diazepam (0.5 mg/kg, ip) at subeffective doses significantly potentiated the hyperphagic effect of orexin-A (30 nM/rat, icv). However, the effect was negated by the GABA(A) receptor antagonist bicuculline (1 mg/kg, ip). Interestingly, benzodiazepine receptor antagonist flumazenil (5 ng/rat, icv), augmented the orexin-A (30 nM/rat, icv) induced hyperphagia; the effect may be attributed to the intrinsic activity of the agent. The results suggest that the hyperphagic effect of orexin-A, at least in part, is mediated by enhanced GABA(A) receptor activity.

Metabotropic glutamate receptor-mediated signaling dampens the HPA axis response to restraint stress.

PubMed

Evanson, Nathan K; Herman, James P

2015-10-15

Glutamate is an important neurotransmitter in the regulation of the neural portion of hypothalamus-pituitary-adrenal (HPA) axis activity, and signals through ionotropic and metabotropic receptors. In the current studies we investigated the role of hypothalamic paraventricular group I metabotropic glutamate receptors in the regulation of the HPA axis response to restraint stress in rats. Direct injection of the group I metabotropic glutamate receptor agonist 3,5-dihydroxyphenylglycine (DHPG) into the PVN prior to restraint leads to blunting of the HPA axis response in awake animals. Consistent with this result, infusion of the group I receptor antagonist hexyl-homoibotenic acid (HIBO) potentiates the HPA axis response to restraint. The excitatory effect of blocking paraventricular group I metabotropic glutamate signaling is blocked by co-administration of dexamethasone into the PVN. However, the inhibitory effect of DHPG is not affected by co-administration of the cannabinoid CB1 receptor antagonist AM-251 into the PVN. Together, these results suggest that paraventricular group I metabotropic glutamate receptor signaling acts to dampen HPA axis reactivity. This effect appears to be similar to the rapid inhibitory effect of glucocorticoids at the PVN, but is not mediated by endocannabinoid signaling. Copyright © 2015 Elsevier Inc. All rights reserved.

Interaction of medullary P2 and glutamate receptors mediates the vasodilation in the hindlimb of rat.

PubMed

Korim, Willian Seiji; Ferreira-Neto, Marcos L; Pedrino, Gustavo R; Pilowsky, Paul M; Cravo, Sergio L

2012-12-01

In the nucleus tractus solitarii (NTS) of rats, blockade of extracellular ATP breakdown to adenosine reduces arterial blood pressure (AP) increases that follow stimulation of the hypothalamic defense area (HDA). The effects of ATP on NTS P2 receptors, during stimulation of the HDA, are still unclear. The aim of this study was to determine whether activation of P2 receptors in the NTS mediates cardiovascular responses to HDA stimulation. Further investigation was taken to establish if changes in hindlimb vascular conductance (HVC) elicited by electrical stimulation of the HDA, or activation of P2 receptors in the NTS, are relayed in the rostral ventrolateral medulla (RVLM); and if those responses depend on glutamate release by ATP acting on presynaptic terminals. In anesthetized and paralyzed rats, electrical stimulation of the HDA increased AP and HVC. Blockade of P2 or glutamate receptors in the NTS, with bilateral microinjections of suramin (10 mM) or kynurenate (50 mM) reduced only the evoked increase in HVC by 75 % or more. Similar results were obtained with the blockade combining both antagonists. Blockade of P2 and glutamate receptors in the RVLM also reduced the increases in HVC to stimulation of the HDA by up to 75 %. Bilateral microinjections of kynurenate in the RVLM abolished changes in AP and HVC to injections of the P2 receptor agonist α,β-methylene ATP (20 mM) into the NTS. The findings suggest that HDA-NTS-RVLM pathways in control of HVC are mediated by activation of P2 and glutamate receptors in the brainstem in alerting-defense reactions.

GABAa and GABAc receptor-mediated modulation of responses to color stimuli: electroretinographic study in the turtle Emys orbicularis.

PubMed

Kupenova, Petia; Vitanova, Lily; Popova, Elka

2010-04-01

GABAergic transmission is involved in color coding in the retina. The specific contribution of different GABA receptors to spectral sensitivity of the retinal responses is not well characterized. We studied GABAa and GABAc receptor-mediated effects on the intensity-response functions of the electroretinographic ON (b-wave) and OFF (d-wave) responses to color stimuli. For this purpose, we compared the effects of GABAa receptor blockade by bicuculline with the effects of GABAa + GABAc receptor blockade by picrotoxin. The blockade of both GABAa and GABAc receptors caused an amplitude increase of the electroretinographic responses, but the effects of the two blockades depended in a specific manner on stimulus intensity and wavelength. The effects of GABAa receptor blockade showed distinct color ON/OFF asymmetry. The absolute and relative sensitivities of the ON responses to blue stimuli and OFF responses to red stimuli were increased to the greatest degree while the sensitivity of the ON responses to red stimuli and OFF responses to blue stimuli was least increased. In contrast, color ON/OFF asymmetry was not typical of the effects of GABAc receptor blockade. The most prominent GABAc effect was the sensitivity increase of the ON and OFF responses to blue stimuli and, to some lesser extent, to green stimuli. The results of this study indicate a specific role of GABAa and GABAc receptor-mediated influences in processing of chromatic information in the distal retina.

Cholera Toxin Inhibits the T-Cell Antigen Receptor-Mediated Increases in Inositol Trisphosphate and Cytoplasmic Free Calcium

NASA Astrophysics Data System (ADS)

Imboden, John B.; Shoback, Dolores M.; Pattison, Gregory; Stobo, John D.

1986-08-01

The addition of monoclonal antibodies to the antigen receptor complex on the malignant human T-cell line Jurkat generates increases in inositol trisphosphate and in the concentration of cytoplasmic free calcium. Exposure of Jurkat cells to cholera toxin for 3 hr inhibited these receptor-mediated events and led to a selective, partial loss of the antigen receptor complex from the cellular surface. None of the effects of cholera toxin on the antigen receptor complex were mimicked by the B subunit of cholera toxin or by increasing intracellular cAMP levels with either forskolin or 8-bromo cAMP. These results suggest that a cholera toxin substrate can regulate signal transduction by the T-cell antigen receptor.

Discoidin Domain Receptor-1 Deficiency Attenuates Atherosclerotic Calcification and Smooth Muscle Cell-Mediated Mineralization

PubMed Central

Ahmad, Pamela J.; Trcka, Daniel; Xue, Siming; Franco, Christopher; Speer, Mei Y.; Giachelli, Cecilia M.; Bendeck, Michelle P.

2009-01-01

Intimal calcification is a feature of advanced atherosclerotic disease that predicts a two- to eightfold increase in the risk of coronary events. Type I collagen promotes vascular smooth muscle cell-mediated calcification, although the mechanism by which this occurs is unknown. The discoidin domain receptor 1 (DDR1) is a collagen receptor that is emerging as a critical mediator of atherosclerosis. To determine whether DDR1 is involved in intimal calcification, we fed male Ddr1−/−;Ldlr−/− and Ddr1+/+;Ldlr−/− mice an atherogenic diet for 6, 12, or 24 weeks. DDR1 deficiency significantly reduced the calcium content of the aortic arch, and microcomputed tomography demonstrated a significant decrease in hydroxyapatite deposition after 24 weeks of atherogenic diet. Reduced calcification was correlated with decreases in macrophage accumulation and tumor necrosis factor α staining, suggesting that the reduction in calcification was in part due to decreased inflammation. The chondrogenic markers type II collagen, type X collagen, and Sox-9 were expressed within the mineralized foci. An in vitro assay performed with vascular smooth muscle cells revealed that DDR1 was required for cell-mediated calcification of the matrix, and Ddr1+/+ smooth muscle cells expressed more alkaline phosphatase activity, whereas Ddr1−/− smooth muscle cells expressed elevated levels of mRNA for nucleotide pyrophosphatase phosphodiesterase 1, an inhibitor of tissue mineralization. Taken together, our results demonstrate that DDR1 mediates an important mechanism for atherosclerotic calcification. PMID:19893047

Discoidin domain receptor-1 deficiency attenuates atherosclerotic calcification and smooth muscle cell-mediated mineralization.

PubMed

Ahmad, Pamela J; Trcka, Daniel; Xue, Siming; Franco, Christopher; Speer, Mei Y; Giachelli, Cecilia M; Bendeck, Michelle P

2009-12-01

Intimal calcification is a feature of advanced atherosclerotic disease that predicts a two- to eightfold increase in the risk of coronary events. Type I collagen promotes vascular smooth muscle cell-mediated calcification, although the mechanism by which this occurs is unknown. The discoidin domain receptor 1 (DDR1) is a collagen receptor that is emerging as a critical mediator of atherosclerosis. To determine whether DDR1 is involved in intimal calcification, we fed male Ddr1(-/-);Ldlr(-/-) and Ddr1(+/+);Ldlr(-/-) mice an atherogenic diet for 6, 12, or 24 weeks. DDR1 deficiency significantly reduced the calcium content of the aortic arch, and microcomputed tomography demonstrated a significant decrease in hydroxyapatite deposition after 24 weeks of atherogenic diet. Reduced calcification was correlated with decreases in macrophage accumulation and tumor necrosis factor alpha staining, suggesting that the reduction in calcification was in part due to decreased inflammation. The chondrogenic markers type II collagen, type X collagen, and Sox-9 were expressed within the mineralized foci. An in vitro assay performed with vascular smooth muscle cells revealed that DDR1 was required for cell-mediated calcification of the matrix, and Ddr1(+/+) smooth muscle cells expressed more alkaline phosphatase activity, whereas Ddr1(-/-) smooth muscle cells expressed elevated levels of mRNA for nucleotide pyrophosphatase phosphodiesterase 1, an inhibitor of tissue mineralization. Taken together, our results demonstrate that DDR1 mediates an important mechanism for atherosclerotic calcification.

Functional evidence for the rapid desensitization of 5-HT(3) receptors on vagal afferents mediating the Bezold-Jarisch reflex

NASA Technical Reports Server (NTRS)

Whalen, E. J.; Johnson, A. K.; Lewis, S. J.

2000-01-01

The aim of this study was to determine whether 5-hydroxytryptamine (5-HT)(3) receptors on cardiopulmonary afferents mediating the Bezold-Jarisch reflex (BJR) desensitize upon repeated exposure to selective agonists. BJR-mediated falls in heart rate, diastolic arterial blood pressure and cardiac output elicited by the 5-HT(3)-receptor agonists, phenylbiguanide (100 microg/kg, i.v.) or 2-methyl-5-HT (100 microg/kg, i.v.), progressively diminished upon repeated injection in conscious rats. The BJR responses elicited by 5-HT (40 microg/kg, i.v.) were markedly reduced in rats which had received the above injections of phenylbiguanide or 2-methyl-5-HT whereas the BJR responses elicited by L-S-nitrosocysteine (10 micromol/kg, i.v.) were similar before and after the injections of the 5-HT(3) receptor agonists. These findings suggest that tachyphylaxis to 5-HT(3) receptor agonists may be due to the desensitization of 5-HT(3) receptors on cardiopulmonary afferents rather than the impairment of the central or peripheral processing of the BJR.

Selective regulation of clathrin-mediated epidermal growth factor receptor signaling and endocytosis by phospholipase C and calcium.

PubMed

Delos Santos, Ralph Christian; Bautista, Stephen; Lucarelli, Stefanie; Bone, Leslie N; Dayam, Roya M; Abousawan, John; Botelho, Roberto J; Antonescu, Costin N

2017-10-15

Clathrin-mediated endocytosis is a major regulator of cell-surface protein internalization. Clathrin and other proteins assemble into small invaginating structures at the plasma membrane termed clathrin-coated pits (CCPs) that mediate vesicle formation. In addition, epidermal growth factor receptor (EGFR) signaling is regulated by its accumulation within CCPs. Given the diversity of proteins regulated by clathrin-mediated endocytosis, how this process may distinctly regulate specific receptors is a key question. We examined the selective regulation of clathrin-dependent EGFR signaling and endocytosis. We find that perturbations of phospholipase Cγ1 (PLCγ1), Ca 2+ , or protein kinase C (PKC) impair clathrin-mediated endocytosis of EGFR, the formation of CCPs harboring EGFR, and EGFR signaling. Each of these manipulations was without effect on the clathrin-mediated endocytosis of transferrin receptor (TfR). EGFR and TfR were recruited to largely distinct clathrin structures. In addition to control of initiation and assembly of CCPs, EGF stimulation also elicited a Ca 2+ - and PKC-dependent reduction in synaptojanin1 recruitment to clathrin structures, indicating broad control of CCP assembly by Ca 2+ signals. Hence EGFR elicits PLCγ1-calcium signals to facilitate formation of a subset of CCPs, thus modulating its own signaling and endocytosis. This provides evidence for the versatility of CCPs to control diverse cellular processes. © 2017 Delos Santos et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Modulation of receptor-mediated gonadotropin action in rat testes by dietary fat.

PubMed

Sebokova, E; Garg, M L; Clandinin, M T

1988-06-01

The effect of feeding diets enriched with 18:2 omega 6, 18:3 omega 3, or saturated fatty acids on lipid composition and receptor-mediated action of luteinizing hormone/human chorionic gonadotropin (LH/hCG) in rat testicular plasma membranes was investigated. Linoleic and alpha-linolenic acid treatments reduced total phospholipid and cholesterol content of the testicular plasma membrane and altered membrane phospholipid composition. Change in phospholipid and cholesterol content after feeding the polyunsaturated fats decreased cholesterol to phospholipid ratios and binding capacity of the LH/hCG receptor in the testicular plasma membrane. LH-stimulated adenylate cyclase activity was decreased in animals fed the linolenic acid-rich diet. NaF-stimulated adenylate cyclase activity was decreased in animals fed diets high in either polyunsaturated fatty acid. Decreased plasma membrane LH/hCG receptor content was associated with decreased testosterone production in Leydig cells in response to LH in the linolenic acid-fed group. It is suggested that change in cholesterol-to-phospholipid ratios alters the physical properties of testicular plasma membranes in a manner that influences accessibility of LH/hCG receptors in testicular tissue.

The role of GABA in NMDA-dependent long term depression (LTD) of rat medial vestibular nuclei.

PubMed

Grassi, S; Della Torre, G; Capocchi, G; Zampolini, M; Pettorossi, V E

1995-11-20

The role of GABA in NMDA-dependent long term depression (LTD) in the medial vestibular nuclei (MVN) was studied on rat brainstem slices. High frequency stimulation (HFS) of the primary vestibular afferents induces a long lasting reduction of the polysynaptic (N2) component of the field potentials recorded in the dorsal portion of the MVN. The induction but not the maintenance of this depression was abolished by AP5, a specific blocking agent for glutamate NMDA receptors. The involvement of GABA in mediating the depression was checked by applying the GABAA and GABAB receptor antagonists, bicuculline and saclofen, before and after HFS. Under bicuculline and saclofen perfusion, HFS provoked a slight potentiation of the N2 wave, while the N2 depression clearly emerged after drug wash-out. This indicates that GABA is not involved in inducing the long term effect, but it is necessary for its expression. Similarly, the LTD reversed and a slight potentiation appeared when both drugs were administered after its induction. Most of these effects were due to the bicuculline, suggesting that GABAA receptors contribute to LTD more than GABAB do. According to our results, it is unlikely that the long lasting vestibular depression is the result of a homosynaptic LTD. On the contrary, our findings suggest that the depression is due to an enhancement of the GABA inhibitory effect, caused by an HFS dependent increase in gabaergic interneuron activity, which resets vestibular neuron excitability at a lower level.

Ethanol increases GABAergic transmission at both pre- and postsynaptic sites in rat central amygdala neurons

PubMed Central

Roberto, Marisa; Madamba, Samuel G.; Moore, Scott D.; Tallent, Melanie K.; Siggins, George R.

2003-01-01

We examined the interaction of ethanol with the γ-aminobutyric acid (GABA)ergic system in neurons of slices of the rat central amygdala nucleus (CeA), a brain region thought to be critical for the reinforcing effects of ethanol. Brief superfusion of 11–66 mM ethanol significantly increased GABA type A (GABAA) receptor-mediated inhibitory postsynaptic potentials (IPSPs) and currents (IPSCs) in most CeA neurons, with a low apparent EC50 of 20 mM. Acute superfusion of 44 mM ethanol increased the amplitude of evoked GABAA IPSPs and IPSCs in 70% of CeA neurons. The ethanol enhancement of IPSPs and IPSCs occurred to a similar extent in the presence of the GABA type B (GABAB) receptor antagonist CGP 55845A, suggesting that this receptor is not involved in the ethanol effect on CeA neurons. Ethanol superfusion also decreased paired-pulse facilitation of evoked GABAA IPSPs and IPSCs and always increased the frequency and sometimes the amplitude of spontaneous miniature GABAA IPSCs as well as responses to local GABA application, indicating both presynaptic and postsynaptic sites of action for ethanol. Thus, the CeA is the first brain region to reveal, without conditional treatments such as GABAB antagonists, consistent, low-dose ethanol enhancement of GABAergic transmission at both pre- and postsynaptic sites. These findings add further support to the contention that the ethanol–GABA interaction in CeA plays an important role in the reinforcing effects of ethanol. PMID:12566570

Inositol trisphosphate receptor mediated spatiotemporal calcium signalling.

PubMed

Miyazaki, S

1995-04-01

Spatiotemporal Ca2+ signalling in the cytoplasm is currently understood as an excitation phenomenon by analogy with electrical excitation in the plasma membrane. In many cell types, Ca2+ waves and Ca2+ oscillations are mediated by inositol 1,4,5-trisphosphate (IP3) receptor/Ca2+ channels in the endoplasmic reticulum membrane, with positive feedback between cytosolic Ca2+ and IP3-induced Ca2+ release creating a regenerative process. Remarkable advances have been made in the past year in the analysis of subcellular Ca2+ microdomains using confocal microscopy and of Ca2+ influx pathways that are functionally coupled to IP3-induced Ca2+ release. Ca2+ signals can be conveyed into the nucleus and mitochondria. Ca2+ entry from outside the cell allows repetitive Ca2+ release by providing Ca2+ to refill the endoplasmic reticulum stores, thus giving rise to frequency-encoded Ca2+ signals.

A-kinase anchoring protein 150 mediates transient receptor potential family V type 1 sensitivity to phosphatidylinositol-4,5-bisphosphate.

PubMed

Jeske, Nathaniel A; Por, Elaine D; Belugin, Sergei; Chaudhury, Sraboni; Berg, Kelly A; Akopian, Armen N; Henry, Michael A; Gomez, Ruben

2011-06-08

A-kinase anchoring protein 150 (AKAP150) is a scaffolding protein that controls protein kinase A- and C-mediated phosphorylation of the transient receptor potential family V type 1 (TRPV1), dictating receptor response to nociceptive stimuli. The phospholipid phosphatidylinositol-4,5-bisphosphate (PIP(2)) anchors AKAP150 to the plasma membrane in naive conditions and also affects TRPV1 activity. In the present study, we sought to determine whether the effects of PIP(2) on TRPV1 are mediated through AKAP150. In trigeminal neurons and CHO cells, the manipulation of cellular PIP(2) led to significant changes in the association of AKAP150 and TRPV1. Following PIP(2) degradation, increased TRPV1:AKAP150 coimmunoprecipitation was observed, resulting in increased receptor response to capsaicin treatment. Phospholipase C activation in neurons isolated from AKAP150(-/-) animals indicated that PIP(2)-mediated inhibition of TRPV1 in the whole-cell environment requires expression of the scaffolding protein. Furthermore, the addition of PIP(2) to neurons isolated from AKAP150 wild-type mice reduced PKA sensitization of TRPV1 compared with isolated neurons from AKAP150(-/-) mice. These findings suggest that PIP(2) degradation increases AKAP150 association with TRPV1 in the whole-cell environment, leading to sensitization of the receptor to nociceptive stimuli.

Toll-like Receptor-mediated Down-regulation of the Deubiquitinase Cylindromatosis (CYLD) Protects Macrophages from Necroptosis in Wild-derived Mice*

PubMed Central

Schworer, Stephen A.; Smirnova, Irina I.; Kurbatova, Irina; Bagina, Uliana; Churova, Maria; Fowler, Trent; Roy, Ananda L.; Degterev, Alexei; Poltorak, Alexander

2014-01-01

Pathogen recognition by the innate immune system initiates the production of proinflammatory cytokines but can also lead to programmed host cell death. Necroptosis, a caspase-independent cell death pathway, can contribute to the host defense against pathogens or cause damage to host tissues. Receptor-interacting protein (RIP1) is a serine/threonine kinase that integrates inflammatory and necroptotic responses. To investigate the mechanisms of RIP1-mediated activation of immune cells, we established a genetic screen on the basis of RIP1-mediated necroptosis in wild-derived MOLF/EiJ mice, which diverged from classical laboratory mice over a million years ago. When compared with C57BL/6, MOLF/EiJ macrophages were resistant to RIP1-mediated necroptosis induced by Toll-like receptors. Using a forward genetic approach in a backcross panel of mice, we identified cylindromatosis (CYLD), a deubiquitinase known to act directly on RIP1 and promote necroptosis in TNF receptor signaling, as the gene conferring the trait. We demonstrate that CYLD is required for Toll-like receptor-induced necroptosis and describe a novel mechanism by which CYLD is down-regulated at the transcriptional level in MOLF/EiJ macrophages to confer protection from necroptosis. PMID:24706750

Phytomelatonin receptor PMTR1-mediated signaling regulates stomatal closure in Arabidopsis thaliana.

PubMed

Wei, Jian; Li, Dong-Xu; Zhang, Jia-Rong; Shan, Chi; Rengel, Zed; Song, Zhong-Bang; Chen, Qi

2018-04-27

Melatonin has been detected in plants in 1995; however, the function and signaling pathway of this putative phytohormone are largely undetermined due to a lack of knowledge about its receptor. Here, we discovered the first phytomelatonin receptor (CAND2/PMTR1) in Arabidopsis thaliana and found that melatonin governs the receptor-dependent stomatal closure. The application of melatonin induced stomatal closure through the heterotrimeric G protein α subunit-regulated H 2 O 2 and Ca 2+ signals. The Arabidopsis mutant lines lacking AtCand2 that encodes a candidate G protein-coupled receptor were insensitive to melatonin-induced stomatal closure. Accordingly, the melatonin-induced H 2 O 2 production and Ca 2+ influx were completely abolished in cand2. CAND2 is a membrane protein that interacts with GPA1 and the expression of AtCand2 was tightly regulated by melatonin in various organs and guard cells. CAND2 showed saturable and specific 125 I-melatonin binding, with apparent K d (dissociation constant) of 0.73 ± 0.10 nmol/L (r 2  = .99), demonstrating this protein is a phytomelatonin receptor (PMTR1). Our results suggest that the phytomelatonin regulation of stomatal closure is dependent on its receptor CAND2/PMTR1-mediated H 2 O 2 and Ca 2+ signaling transduction cascade. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A Novel Ecdysone Receptor Mediates Steroid-Regulated Developmental Events during the Mid-Third Instar of Drosophila

PubMed Central

Costantino, Benjamin F. B.; Bricker, Daniel K.; Alexandre, Kelly; Shen, Kate; Merriam, John R.; Antoniewski, Christophe; Callender, Jenna L.; Henrich, Vincent C.; Presente, Asaf; Andres, Andrew J.

2008-01-01

The larval salivary gland of Drosophila melanogaster synthesizes and secretes glue glycoproteins that cement developing animals to a solid surface during metamorphosis. The steroid hormone 20-hydroxyecdysone (20E) is an essential signaling molecule that modulates most of the physiological functions of the larval gland. At the end of larval development, it is known that 20E—signaling through a nuclear receptor heterodimer consisting of EcR and USP—induces the early and late puffing cascade of the polytene chromosomes and causes the exocytosis of stored glue granules into the lumen of the gland. It has also been reported that an earlier pulse of hormone induces the temporally and spatially specific transcriptional activation of the glue genes; however, the receptor responsible for triggering this response has not been characterized. Here we show that the coordinated expression of the glue genes midway through the third instar is mediated by 20E acting to induce genes of the Broad Complex (BRC) through a receptor that is not an EcR/USP heterodimer. This result is novel because it demonstrates for the first time that at least some 20E-mediated, mid-larval, developmental responses are controlled by an uncharacterized receptor that does not contain an RXR-like component. PMID:18566664

Susceptibility to T cell-mediated liver injury is enhanced in asialoglycoprotein receptor-deficient mice.

PubMed

McVicker, Benita L; Thiele, Geoffrey M; Casey, Carol A; Osna, Natalia A; Tuma, Dean J

2013-05-01

T cell activation and associated pro-inflammatory cytokine production is a pathological feature of inflammatory liver disease. It is also known that liver injury is associated with marked impairments in the function of many hepatic proteins including a hepatocyte-specific binding protein, the asialoglycoprotein receptor (ASGPR). Recently, it has been suggested that hepatic ASGPRs may play an important role in the physiological regulation of T lymphocytes, leading to our hypothesis that ASGPR defects correlate with inflammatory-mediated events in liver diseases. Therefore, in this study we investigated whether changes in hepatocellular ASGPR expression were related to the dysregulation of intrahepatic T lymphocytes and correlate with the development of T-cell mediated hepatitis. Mice lacking functional ASGPRs (receptor-deficient, RD), and wild-type (WT) controls were intravenously injected with T-cell mitogens, Concanavalin A (Con A) or anti-CD3 antibody. As a result of T cell mitogen treatment, RD mice lacking hepatic ASGPRs displayed enhancements in liver pathology, transaminase activities, proinflammatory cytokine expression, and caspase activation compared to that observed in normal WT mice. Furthermore, FACS analysis demonstrated that T-cell mitogen administration resulted in a significant rise in the percentage of CD8+ lymphocytes present in the livers of RD animals versus WT mice. Since these two mouse strains differ only in whether they express the hepatic ASGPR, it can be concluded that proper ASGPR function exerts a protective effect against T cell mediated hepatitis and that impairments to this hepatic receptor could be related to the accumulation of cytotoxic T cells that are observed in inflammatory liver diseases. Published by Elsevier B.V.

Ror2 Receptor Mediates Wnt11 Ligand Signaling and Affects Convergence and Extension Movements in Zebrafish*

PubMed Central

Bai, Yan; Tan, Xungang; Zhang, Haifeng; Liu, Chengdong; Zhao, Beibei; Li, Yun; Lu, Ling; Liu, Yunzhang; Zhou, Jianfeng

2014-01-01

The receptor-tyrosine kinase Ror2 acts as an alternative receptor or co-receptor for Wnt5a and mediates Wnt5a-induced convergent extension movements during embryogenesis in mice and Xenopus as well as the polarity and migration of several cell types during development. However, little is known about whether Ror2 function is conserved in other vertebrates or is involved in other non-canonical Wnt ligands in vivo. In this study we demonstrated that overexpression of dominant-negative ror2 (ror2-TM) mRNA in zebrafish embryos resulted in convergence and extension defects and incompletely separated eyes, which is consistent with observations from slb/wnt11 mutants or wnt11 knockdown morphants. Moreover, the co-injection of ror2-TM mRNA and a wnt11 morpholino or the coexpression of ror2 and wnt11 in zebrafish embryos synergetically induced more severe convergence and extension defects. Transplantation studies further demonstrated that the Ror2 receptor responded to the Wnt11 ligand and regulated cell migration and cell morphology during gastrulation. DnRor2 inhibited the action of Wnt11, which was revealed by a decreased percentage of Wnt11-induced convergence and extension defects. Ror2 physically interacts with Wnt11. The intracellular Tyr-647 and Ser-863 sites of Ror2 are essential for mediating the action of Wnt11. Dishevelled and RhoA act downstream of Wnt11-Ror2 to regulate convergence and extension movements. Overall, our data suggest an important role of Ror2 in mediating Wnt11 signaling and in regulating convergence and extension movements in zebrafish. PMID:24928507

Tachykinin-mediated respiratory effects in conscious guinea pigs: modulation by NK1 and NK2 receptor antagonists.

PubMed

Kudlacz, E M; Logan, D E; Shatzer, S A; Farrell, A M; Baugh, L E

1993-09-07

Tachykinins, in particular neurokinin A and substance P, produce a number of airway effects which may contribute to respiratory diseases such as asthma. We examined the ability of aerosolized substance P, neurokinin A or capsaicin to produce respiratory alterations in conscious guinea pigs using modified whole body plethysmography. Substance P-mediated dyspnea and significant respiratory events were inhibited by the NK1 receptor antagonist, CP-96,345. Neurokinin A-mediated respiratory effects were ablated by the NK2 receptor antagonists: MEN 10207, MDL 29,913 and SR 48,968, the latter being the most potent. The peptide-based antagonist, MEN 10207, produced respiratory effects itself suggesting partial agonist activity. The cyclic hexapeptide, MDL 29,913, relaxed airway smooth muscle via mechanisms other than tachykinin antagonism. NK2 but not NK1 receptor antagonists were able to delay the onset of capsaicin-induced dyspnea, although alone they did not usually (in approximately 10% of the animals) eliminate the response. However, when NK2 receptor antagonists were combined with CP-96,345, the incidence of dyspnea induced by capsaicin decreased significantly (40%) suggesting that both tachykinins contribute to dyspnea in this system.

Muscarinic receptors mediate cold stress-induced detrusor overactivity in type 2 diabetes mellitus rats.

PubMed

Imamura, Tetsuya; Ishizuka, Osamu; Ogawa, Teruyuki; Yamagishi, Takahiro; Yokoyama, Hitoshi; Minagawa, Tomonori; Nakazawa, Masaki; Gautam, Sudha Silwal; Nishizawa, Osamu

2014-10-01

This study determined if muscarinic receptors could mediate the cold stress-induced detrusor overactivity induced in type 2 diabetes mellitus rats. Ten-week-old female Goto-Kakizaki diabetic rats (n = 12) and Wister Kyoto non-diabetic rats (n = 12) were maintained on a high-fat diet for 4 weeks. Cystometric investigations of the unanesthetized rats were carried out at room temperature (27 ± 2°C) for 20 min. They were intravenously administered imidafenacin (0.3 mg/kg, n = 6) or vehicle (n = 6). After 5 min, the rats were transferred to a low temperature (4 ± 2°C) for 40 min where the cystometry was continued. The rats were then returned to room temperature for the final cystometric measurements. Afterwards, expressions of bladder muscarinic receptor M3 and M2 messenger ribonucleic acids and proteins were assessed by reverse transcription polymerase chain reaction and immunohistochemistry. In non-diabetic Wister Kyoto rats, imidafenacin did not reduce cold stress-induced detrusor overactivity. In diabetic Goto-Kakizaki rats, just after transfer to a low temperature, the cold stress-induced detrusor overactivity in imidafenacin-treated rats was reduced compared with vehicle-treated rats. Within the urinary bladders, the ratio of M3 to M2 receptor messenger ribonucleic acid in the diabetic Goto-Kakizaki rats was significantly higher than that of the non-diabetic Wister Kyoto rats. The proportion of muscarinic M3 receptor-positive area within the detrusor in diabetic Goto-Kakizaki rats was also significantly higher than that in non-diabetic Wister Kyoto rats. Imidafenacin partially inhibits cold stress-induced detrusor overactivity in diabetic Goto-Kakizaki rats. In this animal model, muscarinic M3 receptors partially mediate cold stress-induced detrusor overactivity. © 2014 The Japanese Urological Association.

An endogenous 55 kDa TNF receptor mediates cell death in a neural cell line.

PubMed

Sipe, K J; Srisawasdi, D; Dantzer, R; Kelley, K W; Weyhenmeyer, J A

1996-06-01

Tumor necrosis factor-alpha (TNF) is associated with developmental and injury-related events in the central nervous system (CNS). In the present study, we have examined the role of TNF on neurons using the clonal murine neuroblastoma line, N1E-115 (N1E). N1E cells represent a well-defined model for studying neuronal development since they can be maintained as either undifferentiated, mitotically active neuroblasts or as differentiated, mature neurons. Northern and reverse transcription-polymerase chain reaction (RT-PCR) analyses revealed that both undifferentiated and differentiated N1Es express transcripts for the 55 kDa TNF receptor (TNFR), but not the 75 kDa TNFR. The biological activity of the expressed TNF receptor was demonstrated by a dose dependent cytotoxicity to either recombinant murine or human TNF when the cells were incubated with the transcriptional inhibitor actinomycin D. The lack of the 75 kDa receptor mRNA expression and the dose dependent response to rHuTNF, an agonist specific for the murine 55 kDa receptor, suggest that the TNF induced cytotoxicity is mediated through the 55 kDa receptor in both the undifferentiated and differentiated N1Es. Light microscopic observations, flow cytometric analysis of hypodiploid DNA, and electrophoretic analysis of nucleosomal DNA fragmentation of N1Es treated with actinomycin D and TNF revealed features characteristic of both necrotic and apoptotic cell death. These findings demonstrate that blast and mature N1E cells express the 55 kDa TNF receptor which is responsible for inducing both necrotic and apoptotic death in these cells. The observation that actinomycin D renders N1E cells susceptible to the cytotoxic effects of TNF indicates that a sensitization step, such as removal of an endogenous protective factor or viral-mediated inhibition of transcription, may be necessary for TNF cytotoxicity in neurons.

Α2 GABAA receptor sub-units in the ventral hippocampus and α5 GABAA receptor sub-units in the dorsal hippocampus mediate anxiety and fear memory.

PubMed

McEown, K; Treit, D

2013-11-12

Temporary neuronal inactivation of the ventral hippocampus with the GABAA agonist muscimol suppresses unconditioned fear behavior (anxiety) but inactivation of the dorsal hippocampus does not. On the other hand, inactivating the dorsal hippocampus disrupts fear memory, while inactivating the ventral hippocampus does not. Here we investigate the roles of hippocampal GABAA receptor sub-units in mediating these anxiolytic and amnesic effects of GABAA receptor agonists. We microinfused TPA023 (α2 agonist) or TB-21007 (inverse α5 agonist) into the dorsal or ventral hippocampus prior to testing rats in two animal models of anxiety: the elevated plus-maze and shock-probe burying test. Twenty-four hours later rats were re-tested in the shock-probe chamber with a non-electrified probe to assess their memory of the initial shock-probe experience (i.e., fear memory). We found that TPA023 was anxiolytic in the plus-maze and shock-probe burying tests when microinfused into the ventral hippocampus. However, TPA023 did not affect anxiety-related behavior when infused into the dorsal hippocampus. Conversely, we found that the α5 sub-unit inverse agonist TB-21007 impaired rats' memory of the initial shock-probe experience when infused into the dorsal hippocampus, but not when infused into the ventral hippocampus. This double dissociation suggests that α2 GABAA receptor sub-units in the ventral hippocampus mediate unconditioned fear or anxiety, while α5 GABAA receptor sub-units in the dorsal hippocampus mediate conditioned fear memory. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

Protease-activated receptor 2-mediated protection of myocardial ischemia-reperfusion injury: role of transient receptor potential vanilloid receptors

PubMed Central

Zhong, Beihua

2009-01-01

Activation of the protease-activated receptor 2 (PAR2) or the transient receptor potential vanilloid type 1 (TRPV1) channels expressed in cardiac sensory afferents containing calcitonin gene-related peptide (CGRP) and/or substance P (SP) has been proposed to play a protective role in myocardial ischemia-reperfusion (I/R) injury. However, the interaction between PAR2 and TRPV1 is largely unknown. Using gene-targeted TRPV1-null mutant (TRPV1−/−) or wild-type (WT) mice, we test the hypothesis that TRPV1 contributes to PAR2-mediated cardiac protection via increasing the release of CGRP and SP. Immunofluorescence labeling showed that TRPV1 coexpressed with PAR2, PKC-ε, or PKAc in cardiomyocytes, cardiac blood vessels, and perivascular nerves in WT but not TRPV1−/− hearts. WT or TRPV1−/− hearts were Langendorff perfused with the selective PAR2 agonist, SLIGRL, in the presence or absence of various antagonists, followed by 35 min of global ischemia and 40 min of reperfusion (I/R). The recovery rate of coronary flow, the maximum rate of left ventricular pressure development, left ventricular end-diastolic pressure, and left ventricular developed pressure were evaluated after I/R. SLIGRL improved the recovery of hemodynamic parameters, decreased lactate dehydrogenase release, and reduced the infarct size in both WT and TRPV1−/− hearts (P < 0.05). The protection of SLIGRL was significantly surpassed for WT compared with TRPV1−/− hearts (P < 0.05). CGRP8–37, a selective CGRP receptor antagonist, RP67580, a selective neurokinin-1 receptor antagonist, PKC-ε V1–2, a selective PKC-ε inhibitor, or H-89, a selective PKA inhibitor, abolished SLIGRL protection by inhibiting the recovery of the rate of coronary flow, maximum rate of left ventricular pressure development, and left ventricular developed pressure, and increasing left ventricular end-diastolic pressure in WT but not TRPV1−/− hearts. Radioimmunoassay showed that SLIGRL increased the release

Inhibitory Effects of Polyacetylene Compounds from Panax ginseng on Neurotrophin Receptor-Mediated Hair Growth.

PubMed

Suzuki, Aoi; Matsuura, Daisuke; Kanatani, Hirotoshi; Yano, Shingo; Tsunakawa, Mitsuo; Matsuyama, Shigeru; Shigemori, Hideyuki

2017-01-01

Neurotrophins play an important role in the control of the hair growth cycle. Therefore, neurotrophin receptor antagonists have therapeutic potential for the treatment of hair growth disorders. In this study, we investigated the inhibitory effect of Panax ginseng, a medicinal plant commonly used to treat alopecia, on the binding of neurotrophins to their receptors. In addition, we isolated and characterized the bioactive compounds of P. ginseng extracts. P. ginseng hexane extracts strongly inhibited brain-derived neurotrophic factor (BDNF)-TrkB and β-nerve growth factor (β-NGF)-p75 neurotrophin receptor (p75NTR) binding. Furthermore, we identified the following 6 polyacetylene compounds as the bioactive components in P. ginseng hexane extract: panaxynol (1), panaxydol (2), panaxydol chlorohydrin (3), 1,8-heptadecadiene-4,6-diyne-3,10-diol (4), panaxytriol (5), and dihydropanaxacol (6). In particular, compounds 4, 5, and 6 significantly inhibited BDNF-TrkB binding in a dose-dependent manner. To identify the structural component mediating the inhibitory effect, we investigated the effects of the hydroxyl moiety in these compounds. We found that the inhibitory effect of panaxytriol (5) was strong, whereas the inhibitory effect of Ac-panaxytriol (7) was relatively weak. Our findings suggest that P. ginseng-derived polyacetylenes with a hydroxyl moiety might provide therapeutic benefits to patients with hair growth disorders such as alopecia by inhibiting the binding of neurotrophins to their receptors. Although saponins have been proposed to be the primary mediators of the effects of P. ginseng on hair growth, this study revealed that polyacetylene compounds exert similar effects.

Grp94 Protein Delivers γ-Aminobutyric Acid Type A (GABAA) Receptors to Hrd1 Protein-mediated Endoplasmic Reticulum-associated Degradation.

PubMed

Di, Xiao-Jing; Wang, Ya-Juan; Han, Dong-Yun; Fu, Yan-Lin; Duerfeldt, Adam S; Blagg, Brian S J; Mu, Ting-Wei

2016-04-29

Proteostasis maintenance of γ-aminobutyric acid type A (GABAA) receptors dictates their function in controlling neuronal inhibition in mammalian central nervous systems. However, as a multisubunit, multispan, integral membrane protein, even wild type subunits of GABAA receptors fold and assemble inefficiently in the endoplasmic reticulum (ER). Unassembled and misfolded subunits undergo ER-associated degradation (ERAD), but this degradation process remains poorly understood for GABAA receptors. Here, using the α1 subunits of GABAA receptors as a model substrate, we demonstrated that Grp94, a metazoan-specific Hsp90 in the ER lumen, uses its middle domain to interact with the α1 subunits and positively regulates their ERAD. OS-9, an ER-resident lectin, acts downstream of Grp94 to further recognize misfolded α1 subunits in a glycan-dependent manner. This delivers misfolded α1 subunits to the Hrd1-mediated ubiquitination and the valosin-containing protein-mediated extraction pathway. Repressing the initial ERAD recognition step by inhibiting Grp94 enhances the functional surface expression of misfolding-prone α1(A322D) subunits, which causes autosomal dominant juvenile myoclonic epilepsy. This study clarifies a Grp94-mediated ERAD pathway for GABAA receptors, which provides a novel way to finely tune their function in physiological and pathophysiological conditions. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

FcgammaRIIB signals inhibit BLyS signaling and BCR-mediated BLyS receptor up-regulation.

PubMed

Crowley, Jenni E; Stadanlick, Jason E; Cambier, John C; Cancro, Michael P

2009-02-12

These studies investigate how interactions between the BCR and FcgammaRIIB affect B lymphocyte stimulator (BLyS) recep-tor expression and signaling. Previous studies showed that BCR ligation up-regulates BLyS binding capacity in mature B cells, reflecting increased BLyS receptor levels. Here we show that FcgammaRIIB coaggregation dampens BCR-induced BLyS receptor up-regulation. This cross-regulation requires BCR and FcgammaRIIB coligation, and optimal action relies on the Src-homology-2 (SH2)-containing inositol 5 phosphase-1 (SHIP1). Subsequent to FcgammaRIIB/BCR coaggregation, the survival promoting actions of BLyS are attenuated, reflecting reduced BLyS receptor signaling capacity in terms of Pim 2 maintenance, noncanonical NF-kappaB activation, and Bcl-xL levels. These findings link the negative regulatory functions of FcgammaRIIB with BLyS-mediated B-cell survival.

Human Milk Components Modulate Toll-Like Receptor-Mediated Inflammation.

PubMed

He, YingYing; Lawlor, Nathan T; Newburg, David S

2016-01-01

Toll-like receptor (TLR) signaling is central to innate immunity. Aberrant expression of TLRs is found in neonatal inflammatory diseases. Several bioactive components of human milk modulate TLR expression and signaling pathways, including soluble toll-like receptors (sTLRs), soluble cluster of differentiation (sCD) 14, glycoproteins, small peptides, and oligosaccharides. Some milk components, such as sialyl (α2,3) lactose and lacto-N-fucopentaose III, are reported to increase TLR signaling; under some circumstances this might contribute toward immunologic balance. Human milk on the whole is strongly anti-inflammatory, and contains abundant components that depress TLR signaling pathways: sTLR2 and sCD14 inhibit TLR2 signaling; sCD14, lactadherin, lactoferrin, and 2'-fucosyllactose attenuate TLR4 signaling; 3'-galactosyllactose inhibits TLR3 signaling, and β-defensin 2 inhibits TLR7 signaling. Feeding human milk to neonates decreases their risk of sepsis and necrotizing enterocolitis. Thus, the TLR regulatory components found in human milk hold promise as benign oral prophylactic and therapeutic treatments for the many gastrointestinal inflammatory disorders mediated by abnormal TLR signaling. © 2016 American Society for Nutrition.

IL-6 Overexpression in ERG-Positive Prostate Cancer Is Mediated by Prostaglandin Receptor EP2.

PubMed

Merz, Constanze; von Mässenhausen, Anne; Queisser, Angela; Vogel, Wenzel; Andrén, Ove; Kirfel, Jutta; Duensing, Stefan; Perner, Sven; Nowak, Michael

2016-04-01

Prostate cancer is the most diagnosed cancer in men and multiple risk factors and genetic alterations have been described. The TMPRSS2-ERG fusion event and the overexpression of the transcription factor ERG are present in approximately 50% of all prostate cancer patients, however, the clinical outcome is still controversial. Prostate tumors produce various soluble factors, including the pleiotropic cytokine IL-6, regulating cellular processes such as proliferation and metastatic segregation. Here, we used prostatectomy samples in a tissue microarray format and analyzed the co-expression and the clinicopathologic data of ERG and IL-6 using immunohistochemical double staining and correlated the read-out with clinicopathologic data. Expression of ERG and IL-6 correlated strongly in prostate tissue samples. Forced expression of ERG in prostate tumor cell lines resulted in significantly increased secretion of IL-6, whereas the down-regulation of ERG decreased IL-6 secretion. By dissecting the underlying mechanism in prostate tumor cell lines we show the ERG-mediated up-regulation of the prostanoid receptors EP2 and EP3. The prostanoid receptor EP2 was overexpressed in human prostate cancer tissue. Furthermore, the proliferation rate and IL-6 secretion in DU145 cells was reduced after treatment with EP2-receptor antagonist. Collectively, our study shows that the expression of ERG in prostate cancer is linked to the expression of IL-6 mediated by the prostanoid receptor EP2. Copyright © 2016 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Interleukin 1 amplifies receptor-mediated activation of phospholipase A2 in 3T3 fibroblasts.

PubMed Central

Burch, R M; Connor, J R; Axelrod, J

1988-01-01

Human recombinant interleukin 1 alpha (IL-1 alpha) and IL-1 beta stimulated prostaglandin E2 synthesis in 3T3 fibroblasts in a time- and concentration-dependent manner. Enhanced prostaglandin E2 synthesis after IL-1 treatment was apparent by 1 hr and continued to increase for at least 2 days. Half-maximal stimulation occurred at 0.5 pM IL-1 alpha or IL-1 beta, and both interleukins were equally effective, with maximal stimulation occurring in response to 5-10 pM IL-1. In contrast to IL-1, bradykinin stimulation of prostaglandin E2 synthesis is rapid; its effect is maximal by 5 min. In cells that had been pretreated with IL-1 for 24 hr, prostaglandin E2 synthesis in response to bradykinin was amplified more than 10-fold. IL-1 also amplified the receptor-mediated formation of prostaglandin E2 by bombesin and thrombin. The lymphokine did not affect bradykinin receptor number or affinity. IL-1 treatment induced phospholipase A2 and cyclooxygenase but not phospholipase C or prostaglandin E isomerase. It also enhanced bradykinin-stimulated GTPase activity, suggesting possible induction of the GTP-binding regulatory protein coupled to the bradykinin receptor. Thus, IL-1 enhanced receptor-mediated release of prostaglandin E2 in response to bradykinin, bombesin, and thrombin by increasing the cellular levels of phospholipase A2, cyclooxygenase, and GTP-binding regulatory protein(s). PMID:2901097

Epithelial estrogen receptor 1 intrinsically mediates squamous differentiation in the mouse vagina.

PubMed

Miyagawa, Shinichi; Iguchi, Taisen

2015-10-20

Estrogen-mediated actions in female reproductive organs are tightly regulated, mainly through estrogen receptor 1 (ESR1). The mouse vaginal epithelium cyclically exhibits cell proliferation and differentiation in response to estrogen and provides a unique model for analyzing the homeostasis of stratified squamous epithelia. To address the role of ESR1-mediated tissue events during homeostasis, we analyzed mice with a vaginal epithelium-specific knockout of Esr1 driven by keratin 5-Cre (K5-Esr1KO). We show here that loss of epithelial ESR1 in the vagina resulted in aberrant epithelial cell proliferation in the suprabasal cell layers and led to failure of keratinized differentiation. Gene expression analysis showed that several known estrogen target genes, including erbB growth factor ligands, were not induced by estrogen in the K5-Esr1KO mouse vagina. Organ culture experiments revealed that the addition of erbB growth factor ligands, such as amphiregulin, could activate keratinized differentiation in the absence of epithelial ESR1. Thus, epithelial ESR1 integrates estrogen and growth factor signaling to mediate regulation of cell proliferation in squamous differentiation, and our results provide new insights into estrogen-mediated homeostasis in female reproductive organs.

Airborne fine particulate matter causes murine bronchial hyperreactivity via MAPK pathway-mediated M3 muscarinic receptor upregulation.

PubMed

Wang, Rong; Xiao, Xue; Shen, Zhenxing; Cao, Lei; Cao, Yongxiao

2017-02-01

Regarding the human health effects, airborne fine particulate matter 2.5 (PM 2.5 ) is an important environmental risk factor. However, the underlying molecular mechanisms are largely unknown. The present study examined the hypothesis that PM 2.5 causes bronchial hyperreactivity by upregulated muscarinic receptors via the mitogen-activated protein kinase (MAPK) pathway. The isolated rat bronchi segments were cultured with different concentration of PM 2.5 for different time. The contractile response of the bronchi segments were recorded by a sensitive myograph. The mRNA and protein expression levels of M 3 muscarinic receptors were studied by quantitative real-time PCR and immunohistochemistry, respectively. The muscarinic receptors agonist, carbachol induced a remarkable contractile response on fresh and DMSO cultured bronchial segments. Compared with the fresh or DMSO culture groups, 1.0 µg/mL of PM 2.5 cultured for 24 h significantly enhanced muscarinic receptor-mediated contractile responses in bronchi with a markedly increased maximal contraction. In addition, the expression levels of mRNA and protein for M 3 muscarinic receptors in bronchi of PM 2.5 group were higher than that of fresh or DMSO culture groups. SB203580 (p38 inhibitor) and U0126 (MEK1/2 inhibitor) significantly inhibited the PM 2.5 -induced enhanced contraction and increased mRNA and protein expression of muscarinic receptors. However, JNK inhibitor SP600125 had no effect on PM 2.5 -induced muscarinic receptor upregulation and bronchial hyperreactivity. In conclusion, airborne PM 2.5 upregulates muscarinic receptors, which causes subsequently bronchial hyperreactivity shown as enhanced contractility in bronchi. This process may be mediated by p38 and MEK1/2 MAPK pathways. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 371-381, 2017. © 2016 Wiley Periodicals, Inc.

An XA21-Associated Kinase (OsSERK2) Regulates Immunity Mediated by the XA21 and XA3 Immune Receptors

PubMed Central

Chen, Xuewei; Zuo, Shimin; Schwessinger, Benjamin; Chern, Mawsheng; Canlas, Patrick E.; Ruan, Deling; Zhou, Xiaogang; Wang, Jing; Daudi, Arsalan; Petzold, Christopher J.; Heazlewood, Joshua L.; Ronald, Pamela C.

2014-01-01

The rice XA21 immune receptor kinase and the structurally related XA3 receptor confer immunity to Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial leaf blight. Here we report the isolation of OsSERK2 (rice somatic embryogenesis receptor kinase 2) and demonstrate that OsSERK2 positively regulates immunity mediated by XA21 and XA3 as well as the rice immune receptor FLS2 (OsFLS2). Rice plants silenced for OsSerk2 display altered morphology and reduced sensitivity to the hormone brassinolide. OsSERK2 interacts with the intracellular domains of each immune receptor in the yeast two-hybrid system in a kinase activity-dependent manner. OsSERK2 undergoes bidirectional transphosphorylation with XA21 in vitro and forms a constitutive complex with XA21 in vivo. These results demonstrate an essential role for OsSERK2 in the function of three rice immune receptors and suggest that direct interaction with the rice immune receptors is critical for their function. Taken together, our findings suggest that the mechanism of OsSERK2-meditated regulation of rice XA21, XA3, and FLS2 differs from that of AtSERK3/BAK1-mediated regulation of Arabidopsis FLS2 and EFR. PMID:24482436

Pleiotropic AT1 receptor signaling pathways mediating physiological and pathogenic actions of angiotensin II.

PubMed

Hunyady, László; Catt, Kevin J

2006-05-01

Angiotensin II (Ang II) activates a wide spectrum of signaling responses via the AT1 receptor (AT1R) that mediate its physiological control of blood pressure, thirst, and sodium balance and its diverse pathological actions in cardiovascular, renal, and other cell types. Ang II-induced AT1R activation via Gq/11 stimulates phospholipases A2, C, and D, and activates inositol trisphosphate/Ca2+ signaling, protein kinase C isoforms, and MAPKs, as well as several tyrosine kinases (Pyk2, Src, Tyk2, FAK), scaffold proteins (G protein-coupled receptor kinase-interacting protein 1, p130Cas, paxillin, vinculin), receptor tyrosine kinases, and the nuclear factor-kappaB pathway. The AT1R also signals via Gi/o and G11/12 and stimulates G protein-independent signaling pathways, such as beta-arrestin-mediated MAPK activation and the Jak/STAT. Alterations in homo- or heterodimerization of the AT1R may also contribute to its pathophysiological roles. Many of the deleterious actions of AT1R activation are initiated by locally generated, rather than circulating, Ang II and are concomitant with the harmful effects of aldosterone in the cardiovascular system. AT1R-mediated overproduction of reactive oxygen species has potent growth-promoting, proinflammatory, and profibrotic actions by exerting positive feedback effects that amplify its signaling in cardiovascular cells, leukocytes, and monocytes. In addition to its roles in cardiovascular and renal disease, agonist-induced activation of the AT1R also participates in the development of metabolic diseases and promotes tumor progression and metastasis through its growth-promoting and proangiogenic activities. The recognition of Ang II's pathogenic actions is leading to novel clinical applications of angiotensin-converting enzyme inhibitors and AT1R antagonists, in addition to their established therapeutic actions in essential hypertension.

Kainate receptors mediate signaling in both transient and sustained OFF bipolar cell pathways in mouse retina.

PubMed

Borghuis, Bart G; Looger, Loren L; Tomita, Susumu; Demb, Jonathan B

2014-04-30

A fundamental question in sensory neuroscience is how parallel processing is implemented at the level of molecular and circuit mechanisms. In the retina, it has been proposed that distinct OFF cone bipolar cell types generate fast/transient and slow/sustained pathways by the differential expression of AMPA- and kainate-type glutamate receptors, respectively. However, the functional significance of these receptors in the intact circuit during light stimulation remains unclear. Here, we measured glutamate release from mouse bipolar cells by two-photon imaging of a glutamate sensor (iGluSnFR) expressed on postsynaptic amacrine and ganglion cell dendrites. In both transient and sustained OFF layers, cone-driven glutamate release from bipolar cells was blocked by antagonists to kainate receptors but not AMPA receptors. Electrophysiological recordings from bipolar and ganglion cells confirmed the essential role of kainate receptors for signaling in both transient and sustained OFF pathways. Kainate receptors mediated responses to contrast modulation up to 20 Hz. Light-evoked responses in all mouse OFF bipolar pathways depend on kainate, not AMPA, receptors.

Ionotropic Chemosensory Receptors Mediate the Taste and Smell of Polyamines.

PubMed

Hussain, Ashiq; Zhang, Mo; Üçpunar, Habibe K; Svensson, Thomas; Quillery, Elsa; Gompel, Nicolas; Ignell, Rickard; Grunwald Kadow, Ilona C

2016-05-01

The ability to find and consume nutrient-rich diets for successful reproduction and survival is fundamental to animal life. Among the nutrients important for all animals are polyamines, a class of pungent smelling compounds required in numerous cellular and organismic processes. Polyamine deficiency or excess has detrimental effects on health, cognitive function, reproduction, and lifespan. Here, we show that a diet high in polyamine is beneficial and increases reproductive success of flies, and we unravel the sensory mechanisms that attract Drosophila to polyamine-rich food and egg-laying substrates. Using a combination of behavioral genetics and in vivo calcium imaging, we demonstrate that Drosophila uses multisensory detection to find and evaluate polyamines present in overripe and fermenting fruit, their favored feeding and egg-laying substrate. In the olfactory system, two coexpressed ionotropic receptors (IRs), IR76b and IR41a, mediate the long-range attraction to the odor. In the gustatory system, multimodal taste sensation by IR76b receptor and GR66a bitter receptor neurons is used to evaluate quality and valence of the polyamine providing a mechanism for the fly's high attraction to polyamine-rich and sweet decaying fruit. Given their universal and highly conserved biological roles, we propose that the ability to evaluate food for polyamine content may impact health and reproductive success also of other animals including humans.

Ionotropic Chemosensory Receptors Mediate the Taste and Smell of Polyamines

PubMed Central

Üçpunar, Habibe K.; Svensson, Thomas; Quillery, Elsa; Gompel, Nicolas; Ignell, Rickard; Grunwald Kadow, Ilona C.

2016-01-01

The ability to find and consume nutrient-rich diets for successful reproduction and survival is fundamental to animal life. Among the nutrients important for all animals are polyamines, a class of pungent smelling compounds required in numerous cellular and organismic processes. Polyamine deficiency or excess has detrimental effects on health, cognitive function, reproduction, and lifespan. Here, we show that a diet high in polyamine is beneficial and increases reproductive success of flies, and we unravel the sensory mechanisms that attract Drosophila to polyamine-rich food and egg-laying substrates. Using a combination of behavioral genetics and in vivo calcium imaging, we demonstrate that Drosophila uses multisensory detection to find and evaluate polyamines present in overripe and fermenting fruit, their favored feeding and egg-laying substrate. In the olfactory system, two coexpressed ionotropic receptors (IRs), IR76b and IR41a, mediate the long-range attraction to the odor. In the gustatory system, multimodal taste sensation by IR76b receptor and GR66a bitter receptor neurons is used to evaluate quality and valence of the polyamine providing a mechanism for the fly’s high attraction to polyamine-rich and sweet decaying fruit. Given their universal and highly conserved biological roles, we propose that the ability to evaluate food for polyamine content may impact health and reproductive success also of other animals including humans. PMID:27145030

Tachykinin NK2 receptors predominantly mediate tachykinin-induced contractions in ovine trachea.

PubMed

Reynolds, A M; Reynolds, P; Holmes, M; Scicchitano, R

1998-01-12

In vitro studies were conducted to characterize the contractile effects of tachykinins in normal ovine trachea with a view in the future to compare tachykinin contractile responses in allergic tissue. Tracheal smooth muscle strips were prepared for in vitro studies of isometric contraction in response to cumulative addition of carbachol, acetylcholine, histamine, neuropeptide gamma, substance P, neurokinin A, neurokinin B, [Sar9, Met(O2)11]substance P, [Nle10]neurokinin A-(4-10), and [Succinyl-Asp6, Me-Phe8]substance P-(6-11) (senktide). The rank order of potency was neuropeptide gamma > carbachol > neurokinin A > or = [Nle10]neurokinin A-(4-10) > acetylcholine > or = histamine. Phosphoramidon enhanced the contractile response to neurokinin A and substance P, but not to neuropeptide gamma, [Sar9, Met(O2)11]substance P or senktide. Repeated cumulative concentration responses for acetylcholine, substance P, neurokinin A, [Sar9, Met(O2)11]substance P and histamine were also conducted to test for tachyphylaxis. No tachyphylaxis to acetylcholine, substance P, or neurokinin A was observed, however, [Sar9, Met(O2)11]substance P and histamine did exhibit tachyphylaxis. Atropine had no effect on tracheal contractions to neurokinin A and substance P, while [Sar9, Met(O2)11]substance P contractions were atropine sensitive. Pyrilamine did not affect substance P-induced tracheal smooth muscle contractions, indicating that the response to substance P was not mediated by histamine release. These results show that, in vitro, natural tachykinins induce tracheal smooth muscle contraction predominantly by a direct effect mediated by tachykinin NK2 receptors, and a small tachykinin NK1 receptor mediated cholinergic mechanism.

Selectivity of commonly used inhibitors of clathrin-mediated and caveolae-dependent endocytosis of G protein-coupled receptors.

PubMed

Guo, Shuohan; Zhang, Xiaohan; Zheng, Mei; Zhang, Xiaowei; Min, Chengchun; Wang, Zengtao; Cheon, Seung Hoon; Oak, Min-Ho; Nah, Seung-Yeol; Kim, Kyeong-Man

2015-10-01

Among the multiple G protein-coupled receptor (GPCR) endocytic pathways, clathrin-mediated endocytosis (CME) and caveolar endocytosis are more extensively characterized than other endocytic pathways. A number of endocytic inhibitors have been used to block CME; however, systemic studies to determine the selectivity of these inhibitors are needed. Clathrin heavy chain or caveolin1-knockdown cells have been employed to determine the specificity of various chemical and molecular biological tools for CME and caveolar endocytosis. Sucrose, concanavalin A, and dominant negative mutants of dynamin blocked other endocytic pathways, in addition to CME. In particular, concanavalin A nonspecifically interfered with the signaling of several GPCRs tested in the study. Decreased pH, monodansylcadaverine, and dominant negative mutants of epsin were more specific for CME than other treatments were. A recently introduced CME inhibitor, Pitstop2™, showed only marginal selectivity for CME and interfered with receptor expression on the cell surface. Blockade of receptor endocytosis by epsin mutants and knockdown of the clathrin heavy chain enhanced the β2AR-mediated ERK activation. Overall, our studies show that previous experimental results should be interpreted with discretion if they included the use of endocytic inhibitors that were previously thought to be CME-selective. In addition, our study shows that endocytosis of β2 adrenoceptor through clathrin-mediated pathway has negative effects on ERK activation. Copyright © 2015 Elsevier B.V. All rights reserved.

Selective enhancement of NMDA receptor-mediated locomotor hyperactivity by male sex hormones in mice.

PubMed

van den Buuse, Maarten; Low, Jac Kee; Kwek, Perrin; Martin, Sally; Gogos, Andrea

2017-09-01

Altered glutamate NMDA receptor function is implicated in schizophrenia, and gender differences have been demonstrated in this illness. This study aimed to investigate the interaction of gonadal hormones with NMDA receptor-mediated locomotor hyperactivity and PPI disruption in mice. The effect of 0.25 mg/kg of MK-801 on locomotor activity was greater in male mice than in female mice. Gonadectomy (by surgical castration) significantly reduced MK-801-induced hyperlocomotion in male mice, but no effect of gonadectomy was seen in female mice or on amphetamine-induced locomotor hyperactivity. The effect of MK-801 on prepulse inhibition of startle (PPI) was similar in intact and castrated male mice and in ovariectomized (OVX) female mice. In contrast, there was no effect of MK-801 on PPI in intact female mice. Forebrain NMDA receptor density, as measured with [ 3 H]MK-801 autoradiography, was significantly higher in male than in female mice but was not significantly altered by either castration or OVX. These results suggest that male sex hormones enhance the effect of NMDA receptor blockade on psychosis-like behaviour. This interaction was not seen in female mice and was independent of NMDA receptor density in the forebrain. Male sex hormones may be involved in psychosis by an interaction with NMDA receptor hypofunction.

Different Classes of Glutamate Receptors Mediate Distinct Behaviors in a Single Brainstem Nucleus

NASA Astrophysics Data System (ADS)

Dye, John; Heiligenberg, Walter; Keller, Clifford H.; Kawasaki, Masashi

1989-11-01

We have taken advantage of the increasing understanding of glutamate neuropharmacology to probe mechanisms of well-defined vertebrate behaviors. Here we report a set of experiments that suggests distinct roles for two major classes of glutamate receptors in a discrete premotor nucleus of the brainstem. The medullary pacemaker nucleus of weakly electric fish is an endogenous oscillator that controls the electric organ discharge (EOD). Its regular frequency of firing is modulated during several distinct behaviors. The pacemaker nucleus continues firing regularly when isolated in vitro, and modulatory behaviors can be reproduced by stimulating the descending input pathway. Glutamate agonists applied to the pacemaker in vitro produced increases in frequency, while glutamate antagonists selectively blocked stimulus-induced modulations. Experiments with glutamate antagonists in the intact animal resulted in specific effects on two well-characterized behaviors. Our data indicate that these behaviors are separately mediated in the pacemaker by receptors displaying characteristics of the kainate/quisqualate and N-methyl-D-aspartate subtypes of glutamate receptor, respectively.

P2X7 ionotropic receptor is functionally expressed in rabbit articular chondrocytes and mediates extracellular ATP cytotoxicity.

PubMed

Tanigawa, Hitoshi; Toyoda, Futoshi; Kumagai, Kosuke; Okumura, Noriaki; Maeda, Tsutomu; Matsuura, Hiroshi; Imai, Shinji

2018-05-29

Extracellular ATP regulates various cellular functions by engaging multiple subtypes of P2 purinergic receptors. In many cell types, the ionotropic P2X7 receptor mediates pathological events such as inflammation and cell death. However, the importance of this receptor in chondrocytes remains largely unexplored. Here, we report the functional identification of P2X7 receptor in articular chondrocytes and investigate the involvement of P2X7 receptors in ATP-induced cytotoxicity. Chondrocytes were isolated from rabbit articular cartilage, and P2X7 receptor currents were examined using the whole-cell patch-clamp technique. ATP-induced cytotoxicity was evaluated by measuring caspase-3/7 activity, lactate dehydrogenase (LDH) leakage, and prostagrandin E 2 (PGE 2 ) release using microscopic and fluorimetric/colorimetric evaluation. Extracellular ATP readily evoked a cationic current without obvious desensitization. This ATP-activated current was dose related, but required millimolar concentrations. A more potent P2X7 receptor agonist, BzATP, also activated this current but at 100-fold lower concentrations. ATP-induced currents were largely abolished by selective P2X7 antagonists, suggesting a predominant role for the P2X7 receptor. RT-PCR confirmed the presence of P2X7 in chondrocytes. Heterologous expression of a rabbit P2X7 clone successfully reproduced the ATP-induced current. Exposure of chondrocytes to ATP increased caspase-3/7 activities, an effect that was totally abrogated by P2X7 receptor antagonists. Extracellular ATP also enhanced LDH release, which was partially attenuated by the P2X7 inhibitor. The P2X7 receptor-mediated elevation in apoptotic caspase signaling was accompanied by increased PGE 2 release and was attenuated by inhibition of either phospholipase A 2 or cyclooxygenase-2. This study provides direct evidence for the presence of functional P2X7 receptors in articular chondrocytes. Our results suggest that the P2X7 receptor is a potential therapeutic

A role for inflammatory mediators in heterologous desensitization of CysLT1 receptor in human monocytes

PubMed Central

Capra, Valérie; Accomazzo, Maria Rosa; Gardoni, Fabrizio; Barbieri, Silvia; Rovati, G. Enrico

2010-01-01

Cysteinyl-leukotrienes (cysteinyl-LT) are rapidly generated at sites of inflammation and, in addition to their role in asthma, rhinitis, and other immune disorders, are increasingly regarded as significant inflammatory factors in cancer, gastrointestinal, cardiovascular diseases. We recently demonstrated that in monocyte/macrophage–like U937 cells, extracellular nucleotides heterologously desensitize CysLT1 receptor (CysLT1R)-induced Ca2+ transients. Given that monocytes express a number of other inflammatory and chemoattractant receptors, this study was aimed at characterizing transregulation between these different stimuli. We demonstrate that in U937 cells and in primary human monocytes, a series of inflammatory mediators activating Gi-coupled receptor (FPR1, BLT1) desensitize CysLT1R-induced Ca2+ response unidirectionally through activation of PKC. Conversely, PAF-R, exclusively coupled to Gq, cross-desensitizes CysLT1R without the apparent involvement of any kinase. Interestingly, Gs-coupled receptors (β2AR, H1/2R, EP2/4R) are also able to desensitize CysLT1R response through activation of PKA. Heterologous desensitization seems to affect mostly the Gi-mediated signaling of the CysLT1R. The hierarchy of desensitization among agonists may be important for leukocyte signal processing at the site of inflammation. Considering that monocytes/macrophages are likely to be the major source of cysteinyl-LT in many immunological and inflammatory processes, shedding light on how their receptors are regulated will certainly help to better understand the role of these cells in orchestrating this complex network of integrated signals. PMID:19965602

Maternal high-salt diet altered PKC/MLC20 pathway and increased ANG II receptor-mediated vasoconstriction in adult male rat offspring.

PubMed

Li, Weisheng; Lv, Juanxiu; Wu, Jue; Zhou, Xiuwen; Jiang, Lin; Zhu, Xiaolin; Tu, Qing; Tang, Jiaqi; Liu, Yanping; He, Axin; Zhong, Yuan; Xu, Zhice

2016-07-01

High-salt diet (HSD) is associated with cardiovascular diseases. This study aims at ascertaining the influence of maternal HSD on offspring's angiotensin II (ANG II)-mediated vasoconstriction and the underlying mechanisms. In comparison to a normal-salt diet, HSD used in pregnancy in rats changed the ultrastructures of the coronary artery (CA) in 5-month-old male offspring, and increased ANG II-mediated CA contractility. Measurement of [Ca(2+) ]i in CA using fluorescent fura-2, a Ca(2+) indicator, showed that ANG II-mediated increases in [Ca(2+) ]i were the same between HSD and normal-salt diet groups, but the ratio of diameter change/[Ca(2+) ]i induced by ANG II were significantly higher in HSD groups. Angiotensin II receptor type 1, not angiotensin II receptor type 2, caused ANG II-mediated vasoconstriction. Protein kinase C (PKC) inhibitor GF109203X attenuated the ANG II-mediated vasoconstriction, PKC agonist phorbol12,13-dibutyrate produced a greater contraction. There was an increase in PKCβ mRNA and the corresponding protein abundance in the offspring, whereas other PKC subunits PKCα, PKCδ, and PKCε did not change. Moreover, 20 kDa myosin light chain phosphorylation levels were increased in HSD group. Maternal HSD affected the developmental programing for the offspring CA, with increased ANG II-mediated vasoconstrictions. The angiotensin II receptor type 1-PKC-20 kDa myosin light chain phosphorylation pathway was the possible mediated cellular mechanism. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Characterization of muscarinic receptors mediating relaxation and contraction in the rat iris dilator muscle.

PubMed Central

Masuda, Y; Yamahara, N S; Tanaka, M; Ryang, S; Kawai, T; Imaizumi, Y; Watanabe, M

1995-01-01

1. The characteristics of muscarinic receptors mediating relaxation and/or contraction in the rat iris dilator muscle were examined. 2. Relaxation was induced in a dilator muscle by application of acetylcholine (ACh) at low doses (3 microM or less) and contraction was induced by high doses. Methacholine and carbachol also showed biphasic effects similar to those of ACh; in contrast, bethanechol, arecoline, pilocarpine and McN-A-343 induced mainly relaxation but no substantial contraction. 3. After parasympathetic denervation by ciliary ganglionectomy, the relaxant response to muscarinic agonists disappeared upon nerve stimulation. Application of McN-A-343 and pilocarpine induced only small contractions in denervated dilator muscles, indicating that these are partial agonists for contraction. 4. pA2 values of pirenzepine, methoctramine, AF-DX 116, himbacine, and 4-DAMP for antagonism to pilocarpine-induced relaxation in normal dilator muscles and those for antagonism to ACh-induced contraction in denervated dilator muscles were determined. The pA2 values for antagonism to relaxation of all these antagonists were most similar to those for M3-type muscarinic receptors. 5. Although pA2 values for contraction of these antagonists, except for methoctramine, were very close to those for relaxation, contraction was not significantly antagonized by methoctramine. Contraction might be mediated by M3-like receptors which have a very low affinity for methoctramine. 6. In conclusion, ACh-induced biphasic responses in rat iris dilator muscles were clearly distinguished from each other by specific muscarinic agonists and parasympathetic denervation, whereas muscarinic receptors could not be subclassified according to the pA2 values of 5 specific antagonists only. PMID:7539696

Pharmacological profile of the receptors that mediate external carotid vasoconstriction by 5-HT in vagosympathectomized dogs.

PubMed Central

Villalón, C. M.; Ramírez-San Juan, E.; Castillo, C.; Castillo, E.; López-Muñoz, F. J.; Terrón, J. A.

1995-01-01

1. 5-Hydroxytryptamine (5-HT) can produce vasodilatation or vasoconstriction of the canine external carotid bed depending upon the degree of carotid sympathetic tone. Hence, external carotid vasodilatation to 5-HT in dogs with intact sympathetic tone is primarily mediated by prejunctional 5-HT1-like receptors similar to the 5-HT1D subtype, which inhibit the carotid sympathetic outflow. The present investigation is devoted to the pharmacological analysis of the receptors mediating external carotid vasoconstriction by 5-HT in vagosympathectomized dogs. 2. Intracarotid (i.c.) infusions for 1 min of 5-HT (0.3, 1, 3, 10, 30 and 100 micrograms) resulted in dose-dependent decreases in both external carotid blood flow and the corresponding conductance; both mean arterial blood pressure and heart rate remained unchanged during the infusions of 5-HT. These responses to 5-HT were resistant to blockade by antagonists at 5-HT2 (ritanserin) and 5-HT3/5-HT4 (tropisetron) receptors, but were partly blocked by the 5-HT1-like and 5-HT2 receptor antagonist, methiothepin (0.3 mg kg-1); higher doses of methiothepin (1 and 3 mg kg-1) caused little, if any, further blockade. These methiothepin (3 mg kg-1)-resistant responses to 5-HT were not significantly antagonized by MDL 72222 (0.3 mg kg-1) or tropisetron (3 mg kg-1). 3. The external carotid vasoconstrictor effects of 5-HT were mimicked by the selective 5-HT1-like receptor agonist, sumatriptan (3, 10, 30 and 100 micrograms during 1 min, i.c.), which produced dose-dependent decreases in external carotid blood flow and the corresponding conductance; these effects of sumatriptan were dose-dependently antagonized by methiothepin (0.3, 1 and 3 mg kg-1), but not by 5-HT1D-like receptor blocking doses of metergoline (0.1 mg kg-1). 4. The above vasoconstrictor effects of 5-HT remained unaltered after administration of phentolamine, propranolol, atropine, hexamethonium, brompheniramine, cimetidine and haloperidol, thus excluding the

Chicken adenovirus (CELO virus) particles augment receptor-mediated DNA delivery to mammalian cells and yield exceptional levels of stable transformants.

PubMed Central

Cotten, M; Wagner, E; Zatloukal, K; Birnstiel, M L

1993-01-01

Delivery of genes via receptor-mediated endocytosis is severely limited by the poor exit of endocytosed DNA from the endosome. A large enhancement in delivery efficiency has been obtained by including human adenovirus particles in the delivery system. This enhancement is probably a function of the natural adenovirus entry mechanism, which must include passage through or disruption of the endosomal membrane. In an effort to identify safer virus particles useful in this application, we have tested the chicken adenovirus CELO virus for its ability to augment receptor-mediated gene delivery. We report here that CELO virus possesses pH-dependent, liposome disruption activity similar to that of human adenovirus type 5. Furthermore, the chicken adenovirus can be used to augment receptor-mediated gene delivery to levels comparable to those found for the human adenovirus when it is physically linked to polylysine ligand-condensed DNA particles. The chicken adenovirus has the advantage of being produced inexpensively in embryonated eggs, and the virus is naturally replication defective in mammalian cells, even in the presence of wild-type human adenovirus. Images PMID:8099627

Ran GTPase promotes cancer progression via Met receptor-mediated downstream signaling

PubMed Central

Yuen, Hiu-Fung; Chan, Ka-Kui; Platt-Higgins, Angela; Dakir, El-Habib; Matchett, Kyle B.; Haggag, Yusuf Ahmed; Jithesh, Puthen V.; Habib, Tanwir; Faheem, Ahmed; Dean, Fennell A.; Morgan, Richard; Rudland, Philip S.; El-Tanani, Mohamed

2016-01-01

It has been shown previously that cancer cells with an activated oncogenic pathway, including Met activation, require Ran for growth and survival. Here, we show that knockdown of Ran leads to a reduction of Met receptor expression in several breast and lung cancer cell lines. This, in turn suppressed HGF expression and the Met-mediated activation of the Akt pathway, as well as cell adhesion, migration, and invasion. In a cell line model where Met amplification has previously been shown to contribute to gefitinib resistance, Ran knockdown sensitized cells to gefitinib-mediated inhibition of Akt and ERK1/2 phosphorylation and consequently reduced cell proliferation. We further demonstrate that Met reduction-mediated by knockdown of Ran, occurs at the post-transcriptional level, probably via a matrix metalloproteinase. Moreover, the level of immunoreactive Ran and Met are positively associated in human breast cancer specimens, suggesting that a high level of Ran may be a pre-requisite for Met overexpression. Interestingly, a high level of immunoreactive Ran dictates the prognostic significance of Met, indicating that the co-overexpression of Met and Ran may be associated with cancer progression and could be used in combination as a prognostic indicator. PMID:27716616

Gonadotrophin-releasing activity of neurohypophysial hormones: II. The pituitary oxytocin receptor mediating gonadotrophin release differs from that of corticotrophs.

PubMed

Evans, J J; Catt, K J

1989-07-01

Neurohypophysial hormones stimulate gonadotrophin release from dispersed rat anterior pituitary cells in vitro, acting through receptors distinct from those which mediate the secretory response to gonadotrophin-releasing hormone (GnRH). The LH response to oxytocin was not affected by the presence of the phosphodiesterase inhibitor, methyl isobutylxanthine, but was diminished in the absence of extracellular calcium and was progressively increased as the calcium concentration in the medium was raised to normal. In addition, the calcium channel antagonist, nifedipine, suppressed oxytocin-stimulated secretion of LH. It is likely that the mechanisms of LH release induced by GnRH and neurohypophysial hormones are similar, although stimulation of gonadotrophin secretion is mediated by separate receptor systems. Oxytocin was more active than vasopressin in releasing LH, but less active in releasing ACTH. The highly selective oxytocin agonist, [Thr4,Gly7]oxytocin, elicited concentration-dependent secretion of LH but had little effect on corticotrophin secretion. The neurohypophysial hormone antagonist analogues, [d(CH2)5Tyr(Me)2]vasopressin, [d(CH2)5Tyr(Me)2,Orn8]vasotocin and [d(CH2)5D-Tyr(Et)2Val4,Cit8]vasopressin, inhibited the LH response to both oxytocin and vasopressin. However, [d(CH2)5Tyr(Me)2]vasopressin was much less effective in inhibiting the ACTH response to the neurohypophysial hormones, and [d(CH2)5Tyr-(Me)2,Orn8]vasotocin and [d(CH2)5D-Tyr(Et)2,Val4,Cit8]vasopressin exhibited no inhibitory activity against ACTH release. Thus, agonist and antagonist analogues of neurohypophysial hormones display divergent activities with regard to LH and ACTH responses, and the neuropeptide receptor mediating gonadotroph activation is clearly different from that on the corticotroph. Whereas the corticotroph receptor is a vasopressin-type receptor an oxytocin-type receptor is responsible for gonadotrophin release by neurohypophysial hormones.

Activation of Brain Somatostatin Signaling Suppresses CRF Receptor-Mediated Stress Response.

PubMed

Stengel, Andreas; Taché, Yvette F

2017-01-01

Corticotropin-releasing factor (CRF) is the hallmark brain peptide triggering the response to stress and mediates-in addition to the stimulation of the hypothalamus-pituitary-adrenal (HPA) axis-other hormonal, behavioral, autonomic and visceral components. Earlier reports indicate that somatostatin-28 injected intracerebroventricularly counteracts the acute stress-induced ACTH and catecholamine release. Mounting evidence now supports that activation of brain somatostatin signaling exerts a broader anti-stress effect by blunting the endocrine, autonomic, behavioral (with a focus on food intake) and visceral gastrointestinal motor responses through the involvement of distinct somatostatin receptor subtypes.

Lipid-Mediated Regulation of Embedded Receptor Kinases via Parallel Allosteric Relays.

PubMed

Ghosh, Madhubrata; Wang, Loo Chien; Ramesh, Ranita; Morgan, Leslie K; Kenney, Linda J; Anand, Ganesh S

2017-02-28

Membrane-anchored receptors are essential cellular signaling elements for stimulus sensing, propagation, and transmission inside cells. However, the contributions of lipid interactions to the function and dynamics of embedded receptor kinases have not been described in detail. In this study, we used amide hydrogen/deuterium exchange mass spectrometry, a sensitive biophysical approach, to probe the dynamics of a membrane-embedded receptor kinase, EnvZ, together with functional assays to describe the role of lipids in receptor kinase function. Our results reveal that lipids play an important role in regulating receptor function through interactions with transmembrane segments, as well as through peripheral interactions with nonembedded domains. Specifically, the lipid membrane allosterically modulates the activity of the embedded kinase by altering the dynamics of a glycine-rich motif that is critical for phosphotransfer from ATP. This allostery in EnvZ is independent of membrane composition and involves direct interactions with transmembrane and periplasmic segments, as well as peripheral interactions with nonembedded domains of the protein. In the absence of the membrane-spanning regions, lipid allostery is propagated entirely through peripheral interactions. Whereas lipid allostery impacts the phosphotransferase function of the kinase, extracellular stimulus recognition is mediated via a four-helix bundle subdomain located in the cytoplasm, which functions as the osmosensing core through osmolality-dependent helical stabilization. Our findings emphasize the functional modularity in a membrane-embedded kinase, separated into membrane association, phosphotransferase function, and stimulus recognition. These components are integrated through long-range communication relays, with lipids playing an essential role in regulation. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Postnatal aniracetam treatment improves prenatal ethanol induced attenuation of AMPA receptor-mediated synaptic transmission.

PubMed

Wijayawardhane, Nayana; Shonesy, Brian C; Vaglenova, Julia; Vaithianathan, Thirumalini; Carpenter, Mark; Breese, Charles R; Dityatev, Alexander; Suppiramaniam, Vishnu

2007-06-01

Aniracetam is a nootropic compound and an allosteric modulator of AMPA receptors (AMPARs) which mediate synaptic mechanisms of learning and memory. Here we analyzed impairments in AMPAR-mediated synaptic transmission caused by moderate prenatal ethanol exposure and investigated the effects of postnatal aniracetam treatment on these abnormalities. Pregnant Sprague-Dawley rats were gavaged with ethanol or isocaloric sucrose throughout pregnancy, and subsequently the offspring were treated with aniracetam on postnatal days (PND) 18 to 27. Hippocampal slices prepared from these pups on PND 28 to 34 were used for the whole-cell patch-clamp recordings of AMPAR-mediated spontaneous and miniature excitatory postsynaptic currents in CA1 pyramidal cells. Our results indicate that moderate ethanol exposure during pregnancy results in impaired hippocampal AMPAR-mediated neurotransmission, and critically timed aniracetam treatment can abrogate this deficiency. These results highlight the possibility that aniracetam treatment can restore synaptic transmission and ameliorate cognitive deficits associated with the fetal alcohol syndrome.

The SRC homology 2 domain of Rin1 mediates its binding to the epidermal growth factor receptor and regulates receptor endocytosis.

PubMed

Barbieri, M Alejandro; Kong, Chen; Chen, Pin-I; Horazdovsky, Bruce F; Stahl, Philip D

2003-08-22