Efficient syntheses of polyamine and polyamine amide voltage-sensitive calcium channel blockers: FTX-3.3 and sFTX-3.3.

PubMed

Moya, E; Blagbrough, I S

1996-02-01

Efficient syntheses of FTX-3.3 and sFTX-3.3, voltage-sensitive calcium channel blockers are described. These modified polyamines were prepared from selectively protected polyamines and purified on a practical scale.

Three types of neuronal calcium channel with different calcium agonist sensitivity.

PubMed

Nowycky, M C; Fox, A P; Tsien, R W

How many types of calcium channels exist in neurones? This question is fundamental to understanding how calcium entry contributes to diverse neuronal functions such as transmitter release, neurite extension, spike initiation and rhythmic firing. There is considerable evidence for the presence of more than one type of Ca conductance in neurones and other cells. However, little is known about single-channel properties of diverse neuronal Ca channels, or their responsiveness to dihydropyridines, compounds widely used as labels in Ca channel purification. Here we report evidence for the coexistence of three types of Ca channel in sensory neurones of the chick dorsal root ganglion. In addition to a large conductance channel that contributes long-lasting current at strong depolarizations (L), and a relatively tiny conductance that underlies a transient current activated at weak depolarizations (T), we find a third type of unitary activity (N) that is neither T nor L. N-type Ca channels require strongly negative potentials for complete removal of inactivation (unlike L) and strong depolarizations for activation (unlike T). The dihydropyridine Ca agonist Bay K 8644 strongly increases the opening probability of L-, but not T- or N-type channels.

L-type Voltage-Gated Calcium Channels in Conditioned Fear: A Genetic and Pharmacological Analysis

ERIC Educational Resources Information Center

McKinney, Brandon C.; Sze, Wilson; White, Jessica A.; Murphy, Geoffrey G.

2008-01-01

Using pharmacological approaches, others have suggested that L-type voltage-gated calcium channels (L-VGCCs) mediate both consolidation and extinction of conditioned fear. In the absence of L-VGCC isoform-specific antagonists, we have begun to investigate the subtype-specific role of LVGCCs in consolidation and extinction of conditioned fear…

L-Type Calcium Channels Modulation by Estradiol.

PubMed

Vega-Vela, Nelson E; Osorio, Daniel; Avila-Rodriguez, Marco; Gonzalez, Janneth; García-Segura, Luis Miguel; Echeverria, Valentina; Barreto, George E

2017-09-01

Voltage-gated calcium channels are key regulators of brain function, and their dysfunction has been associated with multiple conditions and neurodegenerative diseases because they couple membrane depolarization to the influx of calcium-and other processes such as gene expression-in excitable cells. L-type calcium channels, one of the three major classes and probably the best characterized of the voltage-gated calcium channels, act as an essential calcium binding proteins with a significant biological relevance. It is well known that estradiol can activate rapidly brain signaling pathways and modulatory/regulatory proteins through non-genomic (or non-transcriptional) mechanisms, which lead to an increase of intracellular calcium that activate multiple kinases and signaling cascades, in the same way as L-type calcium channels responses. In this context, estrogens-L-type calcium channels signaling raises intracellular calcium levels and activates the same signaling cascades in the brain probably through estrogen receptor-independent modulatory mechanisms. In this review, we discuss the available literature on this area, which seems to suggest that estradiol exerts dual effects/modulation on these channels in a concentration-dependent manner (as a potentiator of these channels in pM concentrations and as an inhibitor in nM concentrations). Indeed, estradiol may orchestrate multiple neurotrophic responses, which open a new avenue for the development of novel estrogen-based therapies to alleviate different neuropathologies. We also highlight that it is essential to determine through computational and/or experimental approaches the interaction between estradiol and L-type calcium channels to assist these developments, which is an interesting area of research that deserves a closer look in future biomedical research.

Differential facilitation of N- and P/Q-type calcium channels during trains of action potential-like waveforms

PubMed Central

Currie, Kevin P M; Fox, Aaron P

2002-01-01

Inhibition of presynaptic voltage-gated calcium channels by direct G-protein βγ subunit binding is a widespread mechanism that regulates neurotransmitter release. Voltage-dependent relief of this inhibition (facilitation), most likely to be due to dissociation of the G-protein from the channel, may occur during bursts of action potentials. In this paper we compare the facilitation of N- and P/Q-type Ca2+ channels during short trains of action potential-like waveforms (APWs) using both native channels in adrenal chromaffin cells and heterologously expressed channels in tsA201 cells. While both N- and P/Q-type Ca2+ channels exhibit facilitation that is dependent on the frequency of the APW train, there are important quantitative differences. Approximately 20 % of the voltage-dependent inhibition of N-type ICa was reversed during a train while greater than 40 % of the inhibition of P/Q-type ICa was relieved. Changing the duration or amplitude of the APW dramatically affected the facilitation of N-type channels but had little effect on the facilitation of P/Q-type channels. Since the ratio of N-type to P/Q-type Ca2+ channels varies widely between synapses, differential facilitation may contribute to the fine tuning of synaptic transmission, thereby increasing the computational repertoire of neurons. PMID:11882675

Molecular Interactions between Tarantula Toxins and Low-Voltage-Activated Calcium Channels

PubMed Central

Salari, Autoosa; Vega, Benjamin S.; Milescu, Lorin S.; Milescu, Mirela

2016-01-01

Few gating-modifier toxins have been reported to target low-voltage-activated (LVA) calcium channels, and the structural basis of toxin sensitivity remains incompletely understood. Studies of voltage-gated potassium (Kv) channels have identified the S3b–S4 “paddle motif,” which moves at the protein-lipid interface to drive channel opening, as the target for these amphipathic neurotoxins. Voltage-gated calcium (Cav) channels contain four homologous voltage sensor domains, suggesting multiple toxin binding sites. We show here that the S3–S4 segments within Cav3.1 can be transplanted into Kv2.1 to examine their individual contributions to voltage sensing and pharmacology. With these results, we now have a more complete picture of the conserved nature of the paddle motif in all three major voltage-gated ion channel types (Kv, Nav, and Cav). When screened with tarantula toxins, the four paddle sequences display distinct toxin binding properties, demonstrating that gating-modifier toxins can bind to Cav channels in a domain specific fashion. Domain III was the most commonly and strongly targeted, and mutagenesis revealed an acidic residue that is important for toxin binding. We also measured the lipid partitioning strength of all toxins tested and observed a positive correlation with their inhibition of Cav3.1, suggesting a key role for membrane partitioning. PMID:27045173

6-OHDA induced calcium influx through N-type calcium channel alters membrane properties via PKA pathway in substantia nigra pars compacta dopaminergic neurons.

PubMed

Qu, Liang; Wang, Yuan; Zhang, Hai-Tao; Li, Nan; Wang, Qiang; Yang, Qian; Gao, Guo-Dong; Wang, Xue-Lian

2014-07-11

Voltage gated calcium channels (VGCC) are sensitive to oxidative stress, and their activation or inactivation can impact cell death. Although these channels have been extensively studied in expression systems, their role in the brain, particularly in the substantia nigra pars compacta (SNc), remain controversial. In this study, we assessed 6-hydroxydopamine (6-OHDA) induced transformation of firing pattern and functional changes of calcium channels in SNc dopaminergic neurons. Application of 6-OHDA (0.5-2mM) evoked a dose-dependent, desensitizing inward current and intracellular free calcium concentration ([Ca(2+)]i) rise. In voltage clamp, ω-conotoxin-sensitive Ca(2+) current modulation mediated by 6-OHDA reflected an altered sensitivity. Furthermore, we found that 6-OHDA modulated Ca(2+) currents through PKA pathway. These results provided evidence for the potential role of VGCCs and PKA involved in oxidative stress in degeneration of SNc neurons in Parkinson's disease (PD). Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

A Critical Neurodevelopmental Role for L-Type Voltage-Gated Calcium Channels in Neurite Extension and Radial Migration.

PubMed

Kamijo, Satoshi; Ishii, Yuichiro; Horigane, Shin-Ichiro; Suzuki, Kanzo; Ohkura, Masamichi; Nakai, Junichi; Fujii, Hajime; Takemoto-Kimura, Sayaka; Bito, Haruhiko

2018-06-13

Despite many association studies linking gene polymorphisms and mutations of L-type voltage-gated Ca 2+ channels (VGCCs) in neurodevelopmental disorders such as autism and schizophrenia, the roles of specific L-type VGCC during brain development remain unclear. Calcium signaling has been shown to be essential for neurodevelopmental processes such as sculpting of neurites, functional wiring, and fine tuning of growing networks. To investigate this relationship, we performed submembraneous calcium imaging using a membrane-tethered genetically encoded calcium indicator (GECI) Lck-G-CaMP7. We successfully recorded s pontaneous regenerative calcium transients (SRCaTs) in developing mouse excitatory cortical neurons prepared from both sexes before synapse formation. SRCaTs originated locally in immature neurites independently of somatic calcium rises and were significantly more elevated in the axons than in dendrites. SRCaTs were not blocked by tetrodoxin, a Na + channel blocker, but were strongly inhibited by hyperpolarization, suggesting a voltage-dependent source. Pharmacological and genetic manipulations revealed the critical importance of the Ca v 1.2 (CACNA1C) pore-forming subunit of L-type VGCCs, which were indeed expressed in immature mouse brains. Consistently, knocking out Ca v 1.2 resulted in significant alterations of neurite outgrowth. Furthermore, expression of a gain-of-function Ca v 1.2 mutant found in Timothy syndrome, an autosomal dominant multisystem disorder exhibiting syndromic autism, resulted in impaired radial migration of layer 2/3 excitatory neurons, whereas postnatal abrogation of Ca v 1.2 enhancement could rescue cortical malformation. Together, these lines of evidence suggest a critical role for spontaneous opening of L-type VGCCs in neural development and corticogenesis and indicate that L-type VGCCs might constitute a perinatal therapeutic target for neuropsychiatric calciochannelopathies. SIGNIFICANCE STATEMENT Despite many association

The Physiology, Pathology, and Pharmacology of Voltage-Gated Calcium Channels and Their Future Therapeutic Potential

PubMed Central

Zamponi, Gerald W.; Striessnig, Joerg; Koschak, Alexandra

2015-01-01

Voltage-gated calcium channels are required for many key functions in the body. In this review, the different subtypes of voltage-gated calcium channels are described and their physiologic roles and pharmacology are outlined. We describe the current uses of drugs interacting with the different calcium channel subtypes and subunits, as well as specific areas in which there is strong potential for future drug development. Current therapeutic agents include drugs targeting L-type CaV1.2 calcium channels, particularly 1,4-dihydropyridines, which are widely used in the treatment of hypertension. T-type (CaV3) channels are a target of ethosuximide, widely used in absence epilepsy. The auxiliary subunit α2δ-1 is the therapeutic target of the gabapentinoid drugs, which are of value in certain epilepsies and chronic neuropathic pain. The limited use of intrathecal ziconotide, a peptide blocker of N-type (CaV2.2) calcium channels, as a treatment of intractable pain, gives an indication that these channels represent excellent drug targets for various pain conditions. We describe how selectivity for different subtypes of calcium channels (e.g., CaV1.2 and CaV1.3 L-type channels) may be achieved in the future by exploiting differences between channel isoforms in terms of sequence and biophysical properties, variation in splicing in different target tissues, and differences in the properties of the target tissues themselves in terms of membrane potential or firing frequency. Thus, use-dependent blockers of the different isoforms could selectively block calcium channels in particular pathologies, such as nociceptive neurons in pain states or in epileptic brain circuits. Of important future potential are selective CaV1.3 blockers for neuropsychiatric diseases, neuroprotection in Parkinson’s disease, and resistant hypertension. In addition, selective or nonselective T-type channel blockers are considered potential therapeutic targets in epilepsy, pain, obesity, sleep, and

Effects of osmotic swelling on voltage-gated calcium channel currents in rat anterior pituitary cells.

PubMed

Ben-Tabou De-Leon, Shlomo; Blotnick, Edna; Nussinovitch, Itzhak

2003-10-01

Decrease in extracellular osmolarity ([Os]e) results in stimulation of hormone secretion from pituitary cells. Different mechanisms can account for this stimulation of hormone secretion. In this study we examined the possibility that hyposmolarity directly modulates voltage-gated calcium influx in pituitary cells. The effects of hyposmolarity on L-type (IL) and T-type (IT) calcium currents in pituitary cells were investigated by using two hyposmotic stimuli, moderate (18-22% decrease in [Os]e) and strong (31-32% decrease in [Os]e). Exposure to moderate hyposmotic stimuli resulted in three response types in IL (a decrease, a biphasic effect, and an increase in IL) and in increase in IT. Exposure to strong hyposmotic stimuli resulted only in increases in both IL and IT. Similarly, in intact pituitary cells (perforated patch method), exposure to either moderate or strong hyposmotic stimuli resulted only in increases in both IL and IT. Thus it appears that the main effect of decrease in [Os]e is increase in calcium channel currents. This increase was differential (IL were more sensitive than IT) and voltage independent. In addition, we show that these hyposmotic effects cannot be explained by activation of an anionic conductance or by an increase in cell membrane surface area. In conclusion, this study shows that hyposmotic swelling of pituitary cells can directly modulate voltage-gated calcium influx. This hyposmotic modulation of IL and IT may contribute to the previously reported hyposmotic stimulation of hormone secretion. The mechanisms underlying these hyposmotic effects and their possible physiological relevance are discussed.

Voltage-Gated Calcium Channels

NASA Astrophysics Data System (ADS)

Zamponi, Gerald Werner

Voltage Gated Calcium Channels is the first comprehensive book in the calcium channel field, encompassing over thirty years of progress towards our understanding of calcium channel structure, function, regulation, physiology, pharmacology, and genetics. This book balances contributions from many of the leading authorities in the calcium channel field with fresh perspectives from risings stars in the area, taking into account the most recent literature and concepts. This is the only all-encompassing calcium channel book currently available, and is an essential resource for academic researchers at all levels in the areas neuroscience, biophysics, and cardiovascular sciences, as well as to researchers in the drug discovery area.

Effects of inositol trisphosphate on calcium mobilization in high-voltage and saponin-permeabilized platelets

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

Gear, A.R.L.; Hallam, T.J.

1986-03-01

Interest in phosphatidylinositol metabolism has been greatly stimulated by the findings that diglyceride and inositol phosphates may serve as second messengers in modulating cellular function. Formation of 1,4,5-inositol trisphosphate (IP/sub 3/), in particular, has been linked to mobilization of intracellular calcium in a number of cell types. The authors have examined the ability of IP/sub 3/ to mobilize calcium in human platelets permeabilized by either saponin or high-voltage discharge. Saponin at 15 ..mu..g/ml effectively permeabilized platelets to exogenous inositol 1,4,5-trisphosphate which released bound (/sup 45/Ca) within 1 min and with a Ka of 7.4 +/- 4.1 ..mu..M. A small (25%)more » azide-sensitive pool was also responsive to inositol trisphosphate. The calcium pools were completely discharged by A-23187 and the ATP-dependent uptake was prevented by dinitrophenol. In contrast to the result with saponin, platelets accessed by high-voltage discharge were insensitive to challenge by inositol 1,4,5-trisphosphate. The data suggest that while inositol 1,4,5-trisphosphate can rapidly mobilize platelet calcium, the ability to demonstrate this depends on the method of permeabilization.« less

EFFECTS OF PYRETHROIDS ON VOLTAGE-SENSITIVE CALCIUM CHANNELS: A CRITICAL EVALUATION OF STRENGTHS, WEAKNESSES, DATA NEEDS, AND RELATIONSHIP TO ASSESSMENT OF CUMULATIVE NEUROTOXICITY.

EPA Science Inventory

A recently published review (Soderlund et al., 2002, Toxicology 171, 3-59.) of the mechanisms of acute neurotoxicity of pyrethroid compounds postulated that voltage-sensitive calcium channels (VSCC) may be a target of some pyrethroid compounds and that effects on VSCC may contrib...

Regulation of Spinal Substance P Release by Intrathecal Calcium Channel Blockade

PubMed Central

Takasusuki, Toshifumi; Yaksh, Tony L.

2012-01-01

Background We investigated the role of different voltage sensitive calcium channels expressed at presynaptic afferent terminals in substance P release and on nociceptive behavior evoked by intraplantar formalin by examining the effects of intrathecally delivered N- (ziconotide), T- (mibefradil) and L-type voltage sensitive calcium channels blockers (diltiazem and verapamil). Methods Rats received intrathecal pretreatment with saline or doses of morphine, ziconotide, mibefradil, diltiazem or verapamil. The effect of these injections upon flinching evoked by intraplantar formalin (5%, 50μl) was quantified. To assess substance P release, the incidence of neurokinin 1 receptor internalization in the ipsilateral and contralateral lamina I was determined in immunofluorescent stained tissues. Results Intrathecal morphine (20μg), ziconotide (0.3, 0.6 and 1μg), mibefradil (100μg, but not 50μg), diltiazem (500μg, but not 300μg) and verapamil (200μg, but not 50 and 100μg) reduced paw flinching in phase 2 as compared to vehicle control (P < 0.05), with no effect upon phase 1. Ziconotide (0.3, 0.6 and 1μg) and morphine (20μg) significantly inhibited neurokinin 1 receptor internalization (P < 0.05), but mibefradil, diltiazem and verapamil at the highest doses had no effect. Conclusion These results emphasize the role in vivo of N-, but not T- and L-type voltage sensitive calcium channels in mediating the stimulus evoked substance P release from small primary afferents and suggest that T- and L-type voltage sensitive calcium channels blockers exert antihyperalgesic effects by an action on other populations of afferents or mechanisms involving post synaptic excitability. PMID:21577088

Nitric oxide augments voltage-activated calcium currents of crustacea (Idotea baltica) skeletal muscle.

PubMed

Erxleben, C; Hermann, A

2001-03-16

Invertebrate skeletal muscle contraction is regulated by calcium influx through voltage-dependent calcium channels in the sarcolemmal membrane. In present study we investigated the effects of nitric oxide (NO) donors on calcium currents of single skeletal muscle fibres from the marine isopod, Idotea baltica, using two-electrode voltage clamp recording techniques. The NO donors, S-nitrosocysteine, S-nitroso-N-acetyl-penicillamine or hydroxylamine reversibly increased calcium inward currents in a time dependent manner. The increase of the current was prevented by methylene blue. Our experiments suggest that NO increases calcium inward currents. NO, by acting on calcium ion channels in the sarcolemmal membrane, therefore, may directly be involved in the modulation of muscle contraction.

Retinoschisin, a New Binding Partner for L-type Voltage-gated Calcium Channels in the Retina*

PubMed Central

Shi, Liheng; Jian, Kuihuan; Ko, Michael L.; Trump, Dorothy; Ko, Gladys Y.-P.

2009-01-01

The L-type voltage-gated calcium channels (L-VGCCs) are activated under high depolarization voltages. They are vital for diverse biological events, including cell excitability, differentiation, and synaptic transmission. In retinal photoreceptors, L-VGCCs are responsible for neurotransmitter release and are under circadian influences. However, the mechanism of L-VGCC regulation in photoreceptors is not fully understood. Here, we show that retinoschisin, a highly conserved extracellular protein, interacts with the L-VGCCα1D subunit and regulates its activities in a circadian manner. Mutations in the gene encoding retinoschisin (RS1) cause retinal disorganization that leads to early onset of macular degeneration. Since ion channel activities can be modulated through interactions with extracellular proteins, disruption of these interactions can alter physiology and be the root cause of disease states. Co-immunoprecipitation and mammalian two-hybrid assays showed that retinoschisin and the N-terminal fragment of the L-VGCCα1 subunit physically interacted with one another. The expression and secretion of retinoschisin are under circadian regulation with a peak at night and nadir during the day. Inhibition of L-type VGCCs decreased membrane-bound retinoschisin at night. Overexpression of a missense RS1 mutant gene, R141G, into chicken cone photoreceptors caused a decrease of L-type VGCC currents at night. Our findings demonstrate a novel bidirectional relationship between an ion channel and an extracellular protein; L-type VGCCs regulate the circadian rhythm of retinoschisin secretion, whereas secreted retinoschisin feeds back to regulate L-type VGCCs. Therefore, physical interactions between L-VGCCα1 subunits and retinoschisin play an important role in the membrane retention of L-VGCCα1 subunits and photoreceptor-bipolar synaptic transmission. PMID:19074145

Distribution of L-type calcium channels in rat thalamic neurones.

PubMed

Budde, T; Munsch, T; Pape, H C

1998-02-01

One major pathway for calcium entry into neurones is through voltage-activated calcium channels. The distribution of calcium channels over the membrane surface is important for their contribution to neuronal function. Electrophysiological recordings from thalamic cells in situ and after acute isolation demonstrated the presence of high-voltage activated calcium currents. The use of specific L-type calcium channel agonists and antagonists of the dihydropyridine type revealed an about 40% contribution of L-type channels to the total high-voltage-activated calcium current. In order to localize L-type calcium channels in thalamic neurones, fluorescent dihydropyridines were used. They were combined with the fluorescent dye RH414, which allowed the use of a ratio technique and thereby the determination of channel density. The distribution of L-type channels was analysed in the three main thalamic cell types: thalamocortical relay cells, local interneurones and reticular thalamic neurones. While channel density was highest in the soma and decreased significantly in the dendritic region, channels appeared to be clustered differentially in the three types of cells. In thalamocortical cells, L-type channels were clustered in high density around the base of dendrites, while they were more evenly distributed on the soma of interneurones. Reticular thalamic neurones exhibited high density of L-type channels in more central somatic regions. The differential localization of L-type calcium channels found in this study implies their predominate involvement in the regulation of somatic and proximal dendritic calcium-dependent processes, which may be of importance for specific thalamic functions, such as those mediating the transition from rhythmic burst activity during sleep to single spike activity during wakefulness or regulating the relay of visual information.

Fast activation of dihydropyridine-sensitive calcium channels of skeletal muscle. Multiple pathways of channel gating

PubMed Central

1996-01-01

Dihydropyridine (DHP) receptors of the transverse tubule membrane play two roles in excitation-contraction coupling in skeletal muscle: (a) they function as the voltage sensor which undergoes fast transition to control release of calcium from sarcoplasmic reticulum, and (b) they provide the conducting unit of a slowly activating L-type calcium channel. To understand this dual function of the DHP receptor, we studied the effect of depolarizing conditioning pulse on the activation kinetics of the skeletal muscle DHP-sensitive calcium channels reconstituted into lipid bilayer membranes. Activation of the incorporated calcium channel was imposed by depolarizing test pulses from a holding potential of -80 mV. The gating kinetics of the channel was studied with ensemble averages of repeated episodes. Based on a first latency analysis, two distinct classes of channel openings occurred after depolarization: most had delayed latencies, distributed with a mode of 70 ms (slow gating); a small number of openings had short first latencies, < 12 ms (fast gating). A depolarizing conditioning pulse to +20 mV placed 200 ms before the test pulse (-10 mV), led to a significant increase in the activation rate of the ensemble averaged-current; the time constant of activation went from tau m = 110 ms (reference) to tau m = 45 ms after conditioning. This enhanced activation by the conditioning pulse was due to the increase in frequency of fast open events, which was a steep function of the intermediate voltage and the interval between the conditioning pulse and the test pulse. Additional analysis demonstrated that fast gating is the property of the same individual channels that normally gate slowly and that the channels adopt this property after a sojourn in the open state. The rapid secondary activation seen after depolarizing prepulses is not compatible with a linear activation model for the calcium channel, but is highly consistent with a cyclical model. A six- state cyclical model is

Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

NASA Technical Reports Server (NTRS)

Jorgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne; Civitelli, Roberto; Sorensen, Ole Helmer; Steinberg, Thomas H.

2003-01-01

The propagation of mechanically induced intercellular calcium waves (ICW) among osteoblastic cells occurs both by activation of P2Y (purinergic) receptors by extracellular nucleotides, resulting in "fast" ICW, and by gap junctional communication in cells that express connexin43 (Cx43), resulting in "slow" ICW. Human osteoblastic cells transmit intercellular calcium signals by both of these mechanisms. In the current studies we have examined the mechanism of slow gap junction-dependent ICW in osteoblastic cells. In ROS rat osteoblastic cells, gap junction-dependent ICW were inhibited by removal of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx43 (UMR/Cx43) we confirmed that nifedipine sensitivity of ICW required Cx43 expression. In human osteoblastic cells, gap junction-dependent ICW also required activation of L-type calcium channels and influx of extracellular calcium.

Apo calmodulin binding to the L-type voltage-gated calcium channel Ca{sub v}1.2 IQ peptide

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

Lian Luyun; Myatt, Daniel; Kitmitto, Ashraf

2007-02-16

The influx of calcium through the L-type voltage-gated calcium channels (LTCCs) is the trigger for the process of calcium-induced calcium release (CICR) from the sarcoplasmic recticulum, an essential step for cardiac contraction. There are two feedback mechanisms that regulate LTCC activity: calcium-dependent inactivation (CDI) and calcium-dependent facilitation (CDF), both of which are mediated by calmodulin (CaM) binding. The IQ domain (aa 1645-1668) housed within the cytoplasmic domain of the LTCC Ca{sub v}1.2 subunit has been shown to bind both calcium-loaded (Ca{sup 2+}CaM ) and calcium-free CaM (apoCaM). Here, we provide new data for the structural basis for the interaction ofmore » apoCaM with the IQ peptide using NMR, revealing that the apoCaM C-lobe residues are most significantly perturbed upon complex formation. In addition, we have employed transmission electron microscopy of purified LTCC complexes which shows that both apoCaM and Ca{sup 2+}CaM can bind to the intact channel.« less

Spinal antinociception of synthetic omega-conotoxin SO-3, a selective N-type neuronal voltage-sensitive calcium channel blocker, and its effects on morphine analgesia in chemical stimulus tests in rodent.

PubMed

Yan, Ling-Di; Liu, Yan-Li; Zhang, Lei; Dong, Hua-Jin; Zhou, Pei-Lan; Su, Rui-Bin; Gong, Ze-Hui; Huang, Pei-Tang

2010-06-25

SO-3, a novel Omega-superfamily conotoxin derived from Conus striatus, selectively inhibits N-type neuronal voltage-sensitive calcium channels. In current study, antinociception of SO-3 compared with MVIIA or morphine and its effects on morphine analgesia were investigated in rodent chemical stimulus tests after acute or repeated intrathecal administration. In mice acetic acid writhing test, similar to MVIIA, SO-3 caused dose- and time-dependent spinal antinociception with ED(50) of 0.25 microg/kg and t(1/2) of 4h, which was more potent and longer-acting than morphine. In rat formalin test after intrathecal bolus injection, SO-3 produced dose- and time-dependent antinociception by suppressing acute (ED(50), 1.79 microg/kg) and tonic phases (ED(50), 0.41 microg/kg), which was similar to MVIIA and approximately 10-fold potency and twice longer-acting of morphine in blocking tonic phase responses. After repeated intrathecal injections twice daily for 5 consecutive days, SO-3 produced analgesia without loss of potency whereas morphine produced analgesia tolerance in rat formalin test; further, SO-3 still produced potent analgesia in morphine-tolerant rats. SO-3 co-administered with morphine left-shift the dose-response curve of morphine in mice acetic acid writhing test and significantly potentiated morphine analgesia in rat formalin test. No changes in motor function were seen in mice or rats receiving antinociceptive doses of SO-3 whereas MVIIA caused motor dysfunction at doses of 1.0-2.0 microg/kg in rats. This study showed that (1) novel SO-3 produced potent and long-acting spinal antinociception without observable motor dysfunction, (2) SO-3 significantly potentiated morphine analgesia, (3) After repeated intrathecal administration, SO-3 produced neither tolerance nor cross-tolerance to morphine analgesia. (c) 2010 Elsevier B.V. All rights reserved.

Spinal blockage of P/Q- or N-type voltage-gated calcium channels modulates functional and symptomatic changes related to haemorrhagic cystitis in mice

PubMed Central

Silva, R B M; Sperotto, N D M; Andrade, E L; Pereira, T C B; Leite, C E; de Souza, A H; Bogo, M R; Morrone, F B; Gomez, M V; Campos, M M

2015-01-01

Background and Purpose Spinal voltage-gated calcium channels (VGCCs) are pivotal regulators of painful and inflammatory alterations, representing attractive therapeutic targets. We examined the effects of epidural administration of the P/Q- and N-type VGCC blockers Tx3-3 and Phα1β, respectively, isolated from the spider Phoneutria nigriventer, on symptomatic, inflammatory and functional changes allied to mouse cyclophosphamide (CPA)-induced haemorrhagic cystitis (HC). The effects of P. nigriventer-derived toxins were compared with those displayed by MVIIC and MVIIA, extracted from the cone snail Conus magus. Experimental Approach HC was induced by a single i.p. injection of CPA (300 mg·kg–1). Dose- and time-related effects of spinally administered P/Q and N-type VGCC blockers were assessed on nociceptive behaviour and macroscopic inflammation elicited by CPA. The effects of toxins were also evaluated on cell migration, cytokine production, oxidative stress, functional cystometry alterations and TRPV1, TRPA1 and NK1 receptor mRNA expression. Key Results The spinal blockage of P/Q-type VGCC by Tx3-3 and MVIIC or N-type VGCC by Phα1β attenuated nociceptive and inflammatory events associated with HC, including bladder oxidative stress and cytokine production. CPA produced a slight increase in bladder TRPV1 and TRPA1 mRNA expression, which was reversed by all the toxins tested. Noteworthy, Phα1β strongly prevented bladder neutrophil migration, besides HC-related functional alterations, and its effects were potentiated by co-injecting the selective NK1 receptor antagonist CP-96345. Conclusions and Implications Our results shed new light on the role of spinal P/Q and N-type VGCC in bladder dysfunctions, pointing out Phα1β as a promising alternative for treating complications associated with CPA-induced HC. PMID:25298144

P-type calcium channels in rat neocortical neurones.

PubMed Central

Brown, A M; Sayer, R J; Schwindt, P C; Crill, W E

1994-01-01

1. The high threshold, voltage-activated (HVA) calcium current was recorded from acutely isolated rat neocortical pyramidal neurones using the whole-cell patch technique to examine the effect of agents that block P-type calcium channels and to compare their effects to those of omega-conotoxin GVIA (omega-CgTX) and nifedipine. 2. When applied at a saturating concentration (100 nM) the peptide toxins omega-Aga-IVA and synthetic omega-Aga-IVA blocked 31.5 and 33.0% of the HVA current respectively. 3. A saturating concentration of nifedipine (10 microM) inhibited 48.2% of the omega-Aga-IVA-sensitive current, whereas saturating concentrations of both omega-Aga-IVA (100 nM) and omega-CgTX (10 microM) blocked separate specific components of the HVA current. 4. Partially purified funnel web spider toxin (FTX) at a dilution of 1:1000 blocked 81.4% of the HVA current and occluded the inhibitory effect of omega-Aga-IVA. Synthetic FTX 3.3 arginine polyamine (sFTX) at a concentration of 1 mM blocked 61.2% of the HVA current rapidly and reversibly. The effects of sFTX were partially occluded by pre-application of omega-Aga-IVA. We conclude that neither FTX nor sFTX blocked a specific component of the HVA current in these cells. 5. In view of the specificity of omega-Aga-IVA for P-type calcium channels in other preparations and for a specific component of the HVA current in dissociated neocortical neurones we conclude that about 30% of the HVA current in these neurones flow through P-channels. PMID:7517449

P-type calcium channels in rat neocortical neurones.

PubMed

Brown, A M; Sayer, R J; Schwindt, P C; Crill, W E

1994-03-01

1. The high threshold, voltage-activated (HVA) calcium current was recorded from acutely isolated rat neocortical pyramidal neurones using the whole-cell patch technique to examine the effect of agents that block P-type calcium channels and to compare their effects to those of omega-conotoxin GVIA (omega-CgTX) and nifedipine. 2. When applied at a saturating concentration (100 nM) the peptide toxins omega-Aga-IVA and synthetic omega-Aga-IVA blocked 31.5 and 33.0% of the HVA current respectively. 3. A saturating concentration of nifedipine (10 microM) inhibited 48.2% of the omega-Aga-IVA-sensitive current, whereas saturating concentrations of both omega-Aga-IVA (100 nM) and omega-CgTX (10 microM) blocked separate specific components of the HVA current. 4. Partially purified funnel web spider toxin (FTX) at a dilution of 1:1000 blocked 81.4% of the HVA current and occluded the inhibitory effect of omega-Aga-IVA. Synthetic FTX 3.3 arginine polyamine (sFTX) at a concentration of 1 mM blocked 61.2% of the HVA current rapidly and reversibly. The effects of sFTX were partially occluded by pre-application of omega-Aga-IVA. We conclude that neither FTX nor sFTX blocked a specific component of the HVA current in these cells. 5. In view of the specificity of omega-Aga-IVA for P-type calcium channels in other preparations and for a specific component of the HVA current in dissociated neocortical neurones we conclude that about 30% of the HVA current in these neurones flow through P-channels.

Power conditioning using dynamic voltage restorers under different voltage sag types.

PubMed

Saeed, Ahmed M; Abdel Aleem, Shady H E; Ibrahim, Ahmed M; Balci, Murat E; El-Zahab, Essam E A

2016-01-01

Voltage sags can be symmetrical or unsymmetrical depending on the causes of the sag. At the present time, one of the most common procedures for mitigating voltage sags is by the use of dynamic voltage restorers (DVRs). By definition, a DVR is a controlled voltage source inserted between the network and a sensitive load through a booster transformer injecting voltage into the network in order to correct any disturbance affecting a sensitive load voltage. In this paper, modelling of DVR for voltage correction using MatLab software is presented. The performance of the device under different voltage sag types is described, where the voltage sag types are introduced using the different types of short-circuit faults included in the environment of the MatLab/Simulink package. The robustness of the proposed device is evaluated using the common voltage sag indices, while taking into account voltage and current unbalance percentages, where maintaining the total harmonic distortion percentage of the load voltage within a specified range is desired. Finally, several simulation results are shown in order to highlight that the DVR is capable of effective correction of the voltage sag while minimizing the grid voltage unbalance and distortion, regardless of the fault type.

Power conditioning using dynamic voltage restorers under different voltage sag types

PubMed Central

Saeed, Ahmed M.; Abdel Aleem, Shady H.E.; Ibrahim, Ahmed M.; Balci, Murat E.; El-Zahab, Essam E.A.

2015-01-01

Voltage sags can be symmetrical or unsymmetrical depending on the causes of the sag. At the present time, one of the most common procedures for mitigating voltage sags is by the use of dynamic voltage restorers (DVRs). By definition, a DVR is a controlled voltage source inserted between the network and a sensitive load through a booster transformer injecting voltage into the network in order to correct any disturbance affecting a sensitive load voltage. In this paper, modelling of DVR for voltage correction using MatLab software is presented. The performance of the device under different voltage sag types is described, where the voltage sag types are introduced using the different types of short-circuit faults included in the environment of the MatLab/Simulink package. The robustness of the proposed device is evaluated using the common voltage sag indices, while taking into account voltage and current unbalance percentages, where maintaining the total harmonic distortion percentage of the load voltage within a specified range is desired. Finally, several simulation results are shown in order to highlight that the DVR is capable of effective correction of the voltage sag while minimizing the grid voltage unbalance and distortion, regardless of the fault type. PMID:26843975

Simultaneous mapping of membrane voltage and calcium in zebrafish heart in vivo reveals chamber-specific developmental transitions in ionic currents

PubMed Central

Hou, Jennifer H.; Kralj, Joel M.; Douglass, Adam D.; Engert, Florian; Cohen, Adam E.

2014-01-01

The cardiac action potential (AP) and the consequent cytosolic Ca2+ transient are key indicators of cardiac function. Natural developmental processes, as well as many drugs and pathologies change the waveform, propagation, or variability (between cells or over time) of these parameters. Here we apply a genetically encoded dual-function calcium and voltage reporter (CaViar) to study the development of the zebrafish heart in vivo between 1.5 and 4 days post fertilization (dpf). We developed a high-sensitivity spinning disk confocal microscope and associated software for simultaneous three-dimensional optical mapping of voltage and calcium. We produced a transgenic zebrafish line expressing CaViar under control of the heart-specific cmlc2 promoter, and applied ion channel blockers at a series of developmental stages to map the maturation of the action potential in vivo. Early in development, the AP initiated via a calcium current through L-type calcium channels. Between 90 and 102 h post fertilization (hpf), the ventricular AP switched to a sodium-driven upswing, while the atrial AP remained calcium driven. In the adult zebrafish heart, a sodium current drives the AP in both the atrium and ventricle. Simultaneous voltage and calcium imaging with genetically encoded reporters provides a new approach for monitoring cardiac development, and the effects of drugs on cardiac function. PMID:25309445

Simultaneous mapping of membrane voltage and calcium in zebrafish heart in vivo reveals chamber-specific developmental transitions in ionic currents.

PubMed

Hou, Jennifer H; Kralj, Joel M; Douglass, Adam D; Engert, Florian; Cohen, Adam E

2014-01-01

The cardiac action potential (AP) and the consequent cytosolic Ca(2+) transient are key indicators of cardiac function. Natural developmental processes, as well as many drugs and pathologies change the waveform, propagation, or variability (between cells or over time) of these parameters. Here we apply a genetically encoded dual-function calcium and voltage reporter (CaViar) to study the development of the zebrafish heart in vivo between 1.5 and 4 days post fertilization (dpf). We developed a high-sensitivity spinning disk confocal microscope and associated software for simultaneous three-dimensional optical mapping of voltage and calcium. We produced a transgenic zebrafish line expressing CaViar under control of the heart-specific cmlc2 promoter, and applied ion channel blockers at a series of developmental stages to map the maturation of the action potential in vivo. Early in development, the AP initiated via a calcium current through L-type calcium channels. Between 90 and 102 h post fertilization (hpf), the ventricular AP switched to a sodium-driven upswing, while the atrial AP remained calcium driven. In the adult zebrafish heart, a sodium current drives the AP in both the atrium and ventricle. Simultaneous voltage and calcium imaging with genetically encoded reporters provides a new approach for monitoring cardiac development, and the effects of drugs on cardiac function.

Omega-conotoxin- and nifedipine-insensitive voltage-operated calcium channels mediate K(+)-induced release of pro-thyrotropin-releasing hormone-connecting peptides Ps4 and Ps5 from perifused rat hypothalamic slices.

PubMed

Valentijn, K; Tranchand Bunel, D; Vaudry, H

1992-07-01

4 and Ps5, are released by K(+)-induced depolarization through activation of voltage-sensitive calcium channels. The calcium channels appear to have a pharmacological profile different from that of L- and N-type channels. Although, their insensitivity to low Cd2+ concentrations and sensitivity to Ni2+ ions would support the involvement of T-type calcium channels, the lack of effect of amiloride suggests that they belong to a yet undefined class of calcium channels.

Molecular and functional expression of voltage-operated calcium channels during osteogenic differentiation of human mesenchymal stem cells.

PubMed

Zahanich, Ihor; Graf, Eva M; Heubach, Jürgen F; Hempel, Ute; Boxberger, Sabine; Ravens, Ursula

2005-09-01

We used the patch-clamp technique and RT-PCR to study the molecular and functional expression of VOCCs in undifferentiated hMSCs and in cells undergoing osteogenic differentiation. L-type Ca2+ channel blocker nifedipine did not influence alkaline phosphatase activity, calcium, and phosphate accumulation of hMSCs during osteogenic differentiation. This study suggests that osteogenic differentiation of hMSCs does not require L-type Ca2+ channel function. During osteogenic differentiation, mesenchymal stem cells from human bone marrow (hMSCs) must adopt the calcium handling of terminally differentiated osteoblasts. There is evidence that voltage-operated calcium channels (VOCCs), including L-type calcium channels, are involved in regulation of osteoblast function. We therefore studied whether VOCCs play a critical role during osteogenic differentiation of hMSCs. Osteogenic differentiation was induced in hMSCs cultured in maintenance medium (MM) by addition of ascorbate, beta-glycerophosphate, and dexamethasone (ODM) and was assessed by measuring alkaline phosphatase activity, expression of osteopontin, osteoprotegerin, RANKL, and mineralization. Expression of Ca2+ channel alpha1 subunits was shown by semiquantitative or single cell RT-PCR. Voltage-activated calcium currents of hMSCs were measured with the whole cell voltage-clamp technique. mRNA for the pore-forming alpha1C and alpha1G subunits of the L-type and T-type Ca2+ channels, respectively, was found in comparable amounts in cells cultured in MM or ODM. The limitation of L-type Ca2+ currents to a subpopulation of hMSCs was confirmed by single cell RT-PCR, where mRNA for the alpha1C subunits was detectable in only 50% of the cells cultured in MM. Dihydropyridine-sensitive L-type Ca2+ currents were found in 13% of cells cultured in MM and in 12% of the cells cultured in ODM. Under MM and ODM culture conditions, the cells positive for L-type Ca2+ currents were significantly larger than cells without Ca2+ currents

D1 receptors physically interact with N-type calcium channels to regulate channel distribution and dendritic calcium entry.

PubMed

Kisilevsky, Alexandra E; Mulligan, Sean J; Altier, Christophe; Iftinca, Mircea C; Varela, Diego; Tai, Chao; Chen, Lina; Hameed, Shahid; Hamid, Jawed; Macvicar, Brian A; Zamponi, Gerald W

2008-05-22

Dopamine signaling through D1 receptors in the prefrontal cortex (PFC) plays a critical role in the maintenance of higher cognitive functions, such as working memory. At the cellular level, these functions are predicated to involve alterations in neuronal calcium levels. The dendrites of PFC neurons express D1 receptors and N-type calcium channels, yet little information exists regarding their coupling. Here, we show that D1 receptors potently inhibit N-type channels in dendrites of rat PFC neurons. Using coimmunoprecipitation, we demonstrate the existence of a D1 receptor-N-type channel signaling complex in this region, and we provide evidence for a direct receptor-channel interaction. Finally, we demonstrate the importance of this complex to receptor-channel colocalization in heterologous systems and in PFC neurons. Our data indicate that the N-type calcium channel is an important physiological target of D1 receptors and reveal a mechanism for D1 receptor-mediated regulation of cognitive function in the PFC.

The effects of piracetam and its novel peptide analogue GVS-111 on neuronal voltage-gated calcium and potassium channels.

PubMed

Solntseva, E I; Bukanova, J V; Ostrovskaya, R U; Gudasheva, T A; Voronina, T A; Skrebitsky, V G

1997-07-01

1. With the use of the two-microelectrode voltage-clamp method, three types of voltage-activated ionic currents were examined in isolated neurons of the snail Helix pomatia: high-threshold Ca2+ current (ICa), high-threshold Ca(2+)-dependent K+ current (IK(Ca)) and high-threshold K+ current independent of Ca2+ (IK(V)). 2. The effect of bath application of the nootropics piracetam and a novel piracetam peptide analog, ethyl ester of N-phenyl-acetyl-L-prolyl-glycine (GVS-111), on these three types of voltage-activated ionic currents was studied. 3. In more than half of the tested cells, ICa was resistant to both piracetam and GVS-111. In the rest of the cells, ICa decreased 19 +/- 7% with 2 mM of piracetam and 39 +/- 14% with 2 microM of GVS-111. 4. IK(V) in almost all cells tested was resistant to piracetam at concentrations up to 2 mM. However, IK(V) in two-thirds of the cells was sensitive to GVS-111, being suppressed 49 +/- 18% with 1 microM GVS-111. 5. IK(Ca) appeared to be the most sensitive current of those studied to both piracetam and GVS-111. Piracetam at 1 mM and GVS-111 at 0.1 microM decreased the amplitude of IK(Ca) in most of the cells examined by 49 +/- 19% and 69 +/- 24%, respectively. 6. The results suggest that piracetam and GVS-111 suppression of voltage-activated calcium and potassium currents of the neuronal membrane may regulate (both up and down) Ca2+ influx into neurons.

Calcium/calmodulin-dependent serine protein kinase CASK modulates the L-type calcium current.

PubMed

Nafzger, Sabine; Rougier, Jean-Sebastien

2017-01-01

The L-type voltage-gated calcium channel Ca v 1.2 mediates the calcium influx into cells upon membrane depolarization. The list of cardiopathies associated to Ca v 1.2 dysfunctions highlights the importance of this channel in cardiac physiology. Calcium/calmodulin-dependent serine protein kinase (CASK), expressed in cardiac cells, has been identified as a regulator of Ca v 2.2 channels in neurons, but no experiments have been performed to investigate its role in Ca v 1.2 regulation. Full length or the distal C-terminal truncated of the pore-forming Ca v 1.2 channel (Ca v 1.2α1c), both present in cardiac cells, were expressed in TsA-201 cells. In addition, a shRNA silencer, or scramble as negative control, of CASK was co-transfected in order to silence CASK endogenously expressed. Three days post-transfection, the barium current was increased only for the truncated form without alteration of the steady state activation and inactivation biophysical properties. The calcium current, however, was increased after CASK silencing with both types of Ca v 1.2α1c subunits suggesting that, in absence of calcium, the distal C-terminal counteracts the CASK effect. Biochemistry experiments did not reveals neither an alteration of Ca v 1.2 channel protein expression after CASK silencing nor an interaction between Ca v 1.2α1c subunits and CASK. Nevertheless, after CASK silencing, single calcium channel recordings have shown an increase of the voltage-gated calcium channel Ca v 1.2 open probability explaining the increase of the whole-cell current. This study suggests CASK as a novel regulator of Ca v 1.2 via a modulation of the voltage-gated calcium channel Ca v 1.2 open probability. Copyright © 2016 Elsevier Ltd. All rights reserved.

High-voltage-activated calcium current subtypes in mouse DRG neurons adapt in a subpopulation-specific manner after nerve injury.

PubMed

Murali, Swetha S; Napier, Ian A; Mohammadi, Sarasa A; Alewood, Paul F; Lewis, Richard J; Christie, MacDonald J

2015-03-01

Changes in ion channel function and expression are characteristic of neuropathic pain. Voltage-gated calcium channels (VGCCs) are integral for neurotransmission and membrane excitability, but relatively little is known about changes in their expression after nerve injury. In this study, we investigate whether peripheral nerve ligation is followed by changes in the density and proportion of high-voltage-activated (HVA) VGCC current subtypes in dorsal root ganglion (DRG) neurons, the contribution of presynaptic N-type calcium channels in evoked excitatory postsynaptic currents (EPSCs) recorded from dorsal horn neurons in the spinal cord, and the changes in expression of mRNA encoding VGCC subunits in DRG neurons. Using C57BL/6 mice [8- to 11-wk-old males (n = 91)] for partial sciatic nerve ligation or sham surgery, we performed whole cell patch-clamp recordings on isolated DRG neurons and dorsal horn neurons and measured the expression of all VGCC subunits with RT-PCR in DRG neurons. After nerve injury, the density of P/Q-type current was reduced overall in DRG neurons. There was an increase in the percentage of N-type and a decrease in that of P/Q-type current in medium- to large-diameter neurons. No changes were found in the contribution of presynaptic N-type calcium channels in evoked EPSCs recorded from dorsal horn neurons. The α2δ-1 subunit was upregulated by 1.7-fold and γ-3, γ-2, and β-4 subunits were all downregulated 1.7-fold in injured neurons compared with sham-operated neurons. This comprehensive characterization of HVA VGCC subtypes in mouse DRG neurons after nerve injury revealed changes in N- and P/Q-type current proportions only in medium- to large-diameter neurons. Copyright © 2015 the American Physiological Society.

Methamphetamine acutely inhibits voltage-gated calcium channels but chronically up-regulates L-type channels.

PubMed

Andres, Marilou A; Cooke, Ian M; Bellinger, Frederick P; Berry, Marla J; Zaporteza, Maribel M; Rueli, Rachel H; Barayuga, Stephanie M; Chang, Linda

2015-07-01

In neurons, calcium (Ca(2+) ) channels regulate a wide variety of functions ranging from synaptic transmission to gene expression. They also induce neuroplastic changes that alter gene expression following psychostimulant administration. Ca(2+) channel blockers have been considered as potential therapeutic agents for the treatment of methamphetamine (METH) dependence because of their ability to reduce drug craving among METH users. Here, we studied the effects of METH exposure on voltage-gated Ca(2+) channels using SH-SY5Y cells as a model of dopaminergic neurons. We found that METH has different short- and long-term effects. A short-term effect involves immediate (< 5 min) direct inhibition of Ca(2+) ion movements through Ca(2+) channels. Longer exposure to METH (20 min or 48 h) selectively up-regulates the expression of only the CACNA1C gene, thus increasing the number of L-type Ca(2+) channels. This up-regulation of CACNA1C is associated with the expression of the cAMP-responsive element-binding protein (CREB), a known regulator of CACNA1C gene expression, and the MYC gene, which encodes a transcription factor that putatively binds to a site proximal to the CACNA1C gene transcription initiation site. The short-term inhibition of Ca(2+) ion movement and later, the up-regulation of Ca(2+) channel gene expression together suggest the operation of cAMP-responsive element-binding protein- and C-MYC-mediated mechanisms to compensate for Ca(2+) channel inhibition by METH. Increased Ca(2+) current density and subsequent increased intracellular Ca(2+) may contribute to the neurodegeneration accompanying chronic METH abuse. Methamphetamine (METH) exposure has both short- and long-term effects. Acutely, methamphetamine directly inhibits voltage-gated calcium channels. Chronically, neurons compensate by up-regulating the L-type Ca(2+) channel gene, CACNA1C. This compensatory mechanism is mediated by transcription factors C-MYC and CREB, in which CREB is linked to the

Inhibition of N-Type Calcium Channels by Fluorophenoxyanilide Derivatives

PubMed Central

Gleeson, Ellen C.; Graham, Janease E.; Spiller, Sandro; Vetter, Irina; Lewis, Richard J.; Duggan, Peter J.; Tuck, Kellie L.

2015-01-01

A set of fluorophenoxyanilides, designed to be simplified analogues of previously reported ω-conotoxin GVIA mimetics, were prepared and tested for N-type calcium channel inhibition in a SH-SY5Y neuroblastoma FLIPR assay. N-type or Cav2.2 channel is a validated target for the treatment of refractory chronic pain. Despite being significantly less complex than the originally designed mimetics, up to a seven-fold improvement in activity was observed. PMID:25871286

Role of N-type calcium channels in autonomic neurotransmission in guineapig isolated left atria

PubMed Central

Serone, Adrian P; Angus, James A

1999-01-01

Calcium entry via neuronal calcium channels is essential for the process of neurotransmission. We investigated the calcium channel subtypes involved in the operation of cardiac autonomic neurotransmission by examining the effects of selective calcium channel blockers on the inotropic responses to electrical field stimulation (EFS) of driven (4 Hz) guineapig isolated left atria. In this tissue, a previous report (Hong & Chang, 1995) found no evidence for N-type channels involved in the vagal negative inotropic response and only weak involvement in sympathetic responses. The effects of cumulative concentrations of the selective N-type calcium channel blocker, ω-conotoxin GVIA (GVIA; 0.1–10 nM) and the nonselective N-, P/Q-type calcium channel blocker, ω-conotoxin MVIIC (MVIIC; 0.01–10 nM) were examined on the positive (with atropine, 1 μM present) and negative (with propranolol, 1 μM and clonidine, 1 μM present) inotropic responses to EFS (eight trains, each train four pulses per punctate stimulus). GVIA caused complete inhibition of both cardiac vagal and sympathetic inotropic responses to EFS. GVIA was equipotent at inhibiting positive (pIC50 9.29±0.08) and negative (pIC50 9.13±0.17) inotropic responses. MVIIC also mediated complete inhibition of inotropic responses to EFS and was 160 and 85 fold less potent than GVIA at inhibiting positive (pIC50 7.08±0.10) and negative (pIC50 7.20±0.14) inotropic responses, respectively. MVIIC was also equipotent at inhibiting both sympathetic and vagal responses. Our data demonstrates that N-type calcium channels account for all the calcium current required for cardiac autonomic neurotransmission in the guinea-pig isolated left atrium. PMID:10433500

Beta-Estradiol Regulates Voltage-Gated Calcium Channels and Estrogen Receptors in Telocytes from Human Myometrium.

PubMed

Banciu, Adela; Banciu, Daniel Dumitru; Mustaciosu, Cosmin Catalin; Radu, Mihai; Cretoiu, Dragos; Xiao, Junjie; Cretoiu, Sanda Maria; Suciu, Nicolae; Radu, Beatrice Mihaela

2018-05-09

Voltage-gated calcium channels and estrogen receptors are essential players in uterine physiology, and their association with different calcium signaling pathways contributes to healthy and pathological conditions of the uterine myometrium. Among the properties of the various cell subtypes present in human uterine myometrium, there is increasing evidence that calcium oscillations in telocytes (TCs) contribute to contractile activity and pregnancy. Our study aimed to evaluate the effects of beta-estradiol on voltage-gated calcium channels and estrogen receptors in TCs from human uterine myometrium and to understand their role in pregnancy. For this purpose, we employed patch-clamp recordings, ratiometric Fura-2-based calcium imaging analysis, and qRT-PCR techniques for the analysis of cultured human myometrial TCs derived from pregnant and non-pregnant uterine samples. In human myometrial TCs from both non-pregnant and pregnant uterus, we evidenced by qRT-PCR the presence of genes encoding for voltage-gated calcium channels (Cav3.1, Ca3.2, Cav3.3, Cav2.1), estrogen receptors (ESR1, ESR2, GPR30), and nuclear receptor coactivator 3 (NCOA3). Pregnancy significantly upregulated Cav3.1 and downregulated Cav3.2, Cav3.3, ESR1, ESR2, and NCOA3, compared to the non-pregnant condition. Beta-estradiol treatment (24 h, 10, 100, 1000 nM) downregulated Cav3.2, Cav3.3, Cav1.2, ESR1, ESR2, GRP30, and NCOA3 in TCs from human pregnant uterine myometrium. We also confirmed the functional expression of voltage-gated calcium channels by patch-clamp recordings and calcium imaging analysis of TCs from pregnant human myometrium by perfusing with BAY K8644, which induced calcium influx through these channels. Additionally, we demonstrated that beta-estradiol (1000 nM) antagonized the effect of BAY K8644 (2.5 or 5 µM) in the same preparations. In conclusion, we evidenced the presence of voltage-gated calcium channels and estrogen receptors in TCs from non-pregnant and pregnant human

[Human calcium channelopathies. Voltage-gated Ca(2+) channels in etiology, pathogenesis, and pharmacotherapy of neurologic disorders].

PubMed

Weiergräber, M; Hescheler, J; Schneider, T

2008-04-01

Voltage-gated calcium channels are key components in a variety of physiological processes. Within the last decade an increasing number of voltage-gated Ca(2+) channelopathies in both humans and animal models has been described, most of which are related to the neurologic and muscular system. In humans, mutations were found in L-type Ca(v)1.2 and Ca(v)1.4 Ca(2+) channels as well as the non-L-type Ca(v)2.1 and T-type Ca(v)3.2 channels, resulting in altered electrophysiologic properties. Based on their widespread distribution within the CNS, voltage-gated calcium channels are of particular importance in the etiology and pathogenesis of various forms of epilepsy and neuropsychiatric disorders. In this review we characterise the different human Ca(2+) channelopathies known so far, further illuminating basic pathophysiologic mechanisms and clinical aspects.

Voltage-Gated Calcium Influx Modifies Cholinergic Inhibition of Inner Hair Cells in the Immature Rat Cochlea.

PubMed

Zachary, Stephen; Nowak, Nathaniel; Vyas, Pankhuri; Bonanni, Luke; Fuchs, Paul Albert

2018-06-20

Until postnatal day (P) 12, inner hair cells of the rat cochlea are invested with both afferent and efferent synaptic connections. With the onset of hearing at P12, the efferent synapses disappear, and afferent (ribbon) synapses operate with greater efficiency. This change coincides with increased expression of voltage-gated potassium channels, the loss of calcium-dependent electrogenesis, and the onset of graded receptor potentials driven by sound. The transient efferent synapses include near-membrane postsynaptic cisterns thought to regulate calcium influx through the hair cell's α9-containing and α10-containing nicotinic acetylcholine receptors. This influx activates small-conductance Ca 2+ -activated K + (SK) channels. Serial-section electron microscopy of inner hair cells from two 9-d-old (male) rat pups revealed many postsynaptic efferent cisterns and presynaptic afferent ribbons whose average minimal separation in five cells ranged from 1.1 to 1.7 μm. Efferent synaptic function was studied in rat pups (age, 7-9 d) of either sex. The duration of these SK channel-mediated IPSCs was increased by enhanced calcium influx through L-type voltage-gated channels, combined with ryanodine-sensitive release from internal stores-presumably the near-membrane postsynaptic cistern. These data support the possibility that inner hair cell calcium electrogenesis modulates the efficacy of efferent inhibition during the maturation of inner hair cell synapses. SIGNIFICANCE STATEMENT Strict calcium buffering is essential for cellular function. This problem is especially acute for compact hair cells where increasing cytoplasmic calcium promotes the opposing functions of closely adjoining afferent and efferent synapses. The near-membrane postsynaptic cistern at efferent synapses segregates synaptic calcium signals by acting as a dynamic calcium store. The hair cell serves as an informative model for synapses with postsynaptic cisterns (C synapses) found in central neurons

Role of T-type calcium channels in myogenic tone of skeletal muscle resistance arteries.

PubMed

VanBavel, Ed; Sorop, Oana; Andreasen, Ditte; Pfaffendorf, Martin; Jensen, Boye L

2002-12-01

T-type calcium channels may be involved in the maintenance of myogenic tone. We tested their role in isolated rat cremaster arterioles obtained after CO(2) anesthesia and decapitation. Total RNA was analyzed by RT-PCR and Southern blotting for calcium channel expression. We observed expression of voltage-operated calcium (Ca(V)) channels Ca(V)3.1 (T-type), Ca(V)3.2 (T-type), and Ca(V)1.2 (L-type) in cremaster arterioles (n = 3 rats). Amplification products were observed only in the presence of reverse transcriptase and cDNA. Concentration-response curves of the relatively specific L-type blocker verapamil and the relatively specific T-type blockers mibefradil and nickel were made on cannulated vessels with either myogenic tone (75 mmHg) or a similar level of constriction induced by 30 mM K(+) at 35 mmHg. Mibefradil and nickel were, respectively, 162-fold and 300-fold more potent in inhibiting myogenic tone compared with K(+)-induced constriction [log(IC(50), M): mibefradil, basal -7.3 +/- 0.2 (n = 9) and K(+) -5.1 +/- 0.1 (n = 5); nickel, basal -4.1 +/- 0.2 (n = 5) and K(+) -1.6 +/- 0.5 (n = 5); means +/- SE]. Verapamil had a 17-fold more potent effect [log(IC(50), M): basal -6.6 +/- 0.1 (n = 5); K(+) -5.4 +/- 0.3 (n = 4); all log(IC(50)) P < 0.05, basal vs. K(+)]. These data suggest that T-type calcium channels are expressed and involved in maintenance of myogenic tone in rat cremaster muscle arterioles.

The L-Type Voltage-Gated Calcium Channel Ca [subscript V] 1.2 Mediates Fear Extinction and Modulates Synaptic Tone in the Lateral Amygdala

ERIC Educational Resources Information Center

Temme, Stephanie J.; Murphy, Geoffrey G.

2017-01-01

L-type voltage-gated calcium channels (LVGCCs) have been implicated in both the formation and the reduction of fear through Pavlovian fear conditioning and extinction. Despite the implication of LVGCCs in fear learning and extinction, studies of the individual LVGCC subtypes, Ca[subscript V]1.2 and Ca[subscript V] 1.3, using transgenic mice have…

Evaluation of the inhibitory effect of dihydropyridines on N-type calcium channel by virtual three-dimensional pharmacophore modeling.

PubMed

Ogihara, Takuo; Kano, Takashi; Kakinuma, Chihaya

2009-01-01

Currently, a new type of calcium channel blockers, which can inhibit not only L-type calcium channels abundantly expressed in vascular smooth muscles, but also N-type calcium channels that abound in the sympathetic nerve endings, have been developed. In this study, analysis on a like-for-like basis of the L- and N-type calcium channel-inhibitory activity of typical dihydropyridine-type calcium-channel blockers (DHPs) was performed. Moreover, to understand the differences of N-type calcium channel inhibition among DHPs, the binding of DHPs to the channel was investigated by means of hypothetical three-dimensional pharmacophore modeling using multiple calculated low-energy conformers of the DHPs. All of the tested compounds, i.e. cilnidipine (CAS 132203-70-4), efonidipine (CAS 111011-76-8), amlodipine (CAS 111470-99-6), benidipine (CAS 85387-35-5), azelnidipine (CAS 123524-52-7) and nifedipine (CAS 21829-25-4), potently inhibited the L-type calcium channel, whereas only cilnidipine inhibited the N-type calcium channel (IC50 value: 51.2 nM). A virtual three-dimensional structure of the N-type calcium channel was generated by using the structure of the peptide omega-conotoxin GVIA, a standard inhibitor of the channel, and cilnidipine was found to fit well into this pharmacophore model. Lipophilic potential maps of omega-conotoxin GVIA and cilnidipine supported this finding. Conformational overlay of cilnidipine and the other DHPs indicated that amlodipine and nifedipine were not compatible with the pharmacophore model because they did not contain an aromatic ring that was functionally equivalent to Tyr13 of omega-conotoxin GVIA. Azelnidipine, benidipine, and efonidipine, which have this type of aromatic ring, were not positively identified due to intrusions into the excluded volume. Estimation of virtual three-dimensional structures of proteins, such as ion channels, by using standard substrates and/or inhibitors may be a useful method to explore the mechanisms of

Fragile X mental retardation protein controls synaptic vesicle exocytosis by modulating N-type calcium channel density

NASA Astrophysics Data System (ADS)

Ferron, Laurent; Nieto-Rostro, Manuela; Cassidy, John S.; Dolphin, Annette C.

2014-04-01

Fragile X syndrome (FXS), the most common heritable form of mental retardation, is characterized by synaptic dysfunction. Synaptic transmission depends critically on presynaptic calcium entry via voltage-gated calcium (CaV) channels. Here we show that the functional expression of neuronal N-type CaV channels (CaV2.2) is regulated by fragile X mental retardation protein (FMRP). We find that FMRP knockdown in dorsal root ganglion neurons increases CaV channel density in somata and in presynaptic terminals. We then show that FMRP controls CaV2.2 surface expression by targeting the channels to the proteasome for degradation. The interaction between FMRP and CaV2.2 occurs between the carboxy-terminal domain of FMRP and domains of CaV2.2 known to interact with the neurotransmitter release machinery. Finally, we show that FMRP controls synaptic exocytosis via CaV2.2 channels. Our data indicate that FMRP is a potent regulator of presynaptic activity, and its loss is likely to contribute to synaptic dysfunction in FXS.

Voltage imaging to understand connections and functions of neuronal circuits.

PubMed

Antic, Srdjan D; Empson, Ruth M; Knöpfel, Thomas

2016-07-01

Understanding of the cellular mechanisms underlying brain functions such as cognition and emotions requires monitoring of membrane voltage at the cellular, circuit, and system levels. Seminal voltage-sensitive dye and calcium-sensitive dye imaging studies have demonstrated parallel detection of electrical activity across populations of interconnected neurons in a variety of preparations. A game-changing advance made in recent years has been the conceptualization and development of optogenetic tools, including genetically encoded indicators of voltage (GEVIs) or calcium (GECIs) and genetically encoded light-gated ion channels (actuators, e.g., channelrhodopsin2). Compared with low-molecular-weight calcium and voltage indicators (dyes), the optogenetic imaging approaches are 1) cell type specific, 2) less invasive, 3) able to relate activity and anatomy, and 4) facilitate long-term recordings of individual cells' activities over weeks, thereby allowing direct monitoring of the emergence of learned behaviors and underlying circuit mechanisms. We highlight the potential of novel approaches based on GEVIs and compare those to calcium imaging approaches. We also discuss how novel approaches based on GEVIs (and GECIs) coupled with genetically encoded actuators will promote progress in our knowledge of brain circuits and systems. Copyright © 2016 the American Physiological Society.

Voltage imaging to understand connections and functions of neuronal circuits

PubMed Central

Antic, Srdjan D.; Empson, Ruth M.

2016-01-01

Understanding of the cellular mechanisms underlying brain functions such as cognition and emotions requires monitoring of membrane voltage at the cellular, circuit, and system levels. Seminal voltage-sensitive dye and calcium-sensitive dye imaging studies have demonstrated parallel detection of electrical activity across populations of interconnected neurons in a variety of preparations. A game-changing advance made in recent years has been the conceptualization and development of optogenetic tools, including genetically encoded indicators of voltage (GEVIs) or calcium (GECIs) and genetically encoded light-gated ion channels (actuators, e.g., channelrhodopsin2). Compared with low-molecular-weight calcium and voltage indicators (dyes), the optogenetic imaging approaches are 1) cell type specific, 2) less invasive, 3) able to relate activity and anatomy, and 4) facilitate long-term recordings of individual cells' activities over weeks, thereby allowing direct monitoring of the emergence of learned behaviors and underlying circuit mechanisms. We highlight the potential of novel approaches based on GEVIs and compare those to calcium imaging approaches. We also discuss how novel approaches based on GEVIs (and GECIs) coupled with genetically encoded actuators will promote progress in our knowledge of brain circuits and systems. PMID:27075539

Voltage-sensitive rhodol with enhanced two-photon brightness.

PubMed

Kulkarni, Rishikesh U; Kramer, Daniel J; Pourmandi, Narges; Karbasi, Kaveh; Bateup, Helen S; Miller, Evan W

2017-03-14

We have designed, synthesized, and applied a rhodol-based chromophore to a molecular wire-based platform for voltage sensing to achieve fast, sensitive, and bright voltage sensing using two-photon (2P) illumination. Rhodol VoltageFluor-5 (RVF5) is a voltage-sensitive dye with improved 2P cross-section for use in thick tissue or brain samples. RVF5 features a dichlororhodol core with pyrrolidyl substitution at the nitrogen center. In mammalian cells under one-photon (1P) illumination, RVF5 demonstrates high voltage sensitivity (28% ΔF/F per 100 mV) and improved photostability relative to first-generation voltage sensors. This photostability enables multisite optical recordings from neurons lacking tuberous sclerosis complex 1, Tsc1, in a mouse model of genetic epilepsy. Using RVF5, we show that Tsc1 KO neurons exhibit increased activity relative to wild-type neurons and additionally show that the proportion of active neurons in the network increases with the loss of Tsc1. The high photostability and voltage sensitivity of RVF5 is recapitulated under 2P illumination. Finally, the ability to chemically tune the 2P absorption profile through the use of rhodol scaffolds affords the unique opportunity to image neuronal voltage changes in acutely prepared mouse brain slices using 2P illumination. Stimulation of the mouse hippocampus evoked spiking activity that was readily discerned with bath-applied RVF5, demonstrating the utility of RVF5 and molecular wire-based voltage sensors with 2P-optimized fluorophores for imaging voltage in intact brain tissue.

Voltage-sensitive rhodol with enhanced two-photon brightness

PubMed Central

Kulkarni, Rishikesh U.; Kramer, Daniel J.; Pourmandi, Narges; Karbasi, Kaveh; Bateup, Helen S.

2017-01-01

We have designed, synthesized, and applied a rhodol-based chromophore to a molecular wire-based platform for voltage sensing to achieve fast, sensitive, and bright voltage sensing using two-photon (2P) illumination. Rhodol VoltageFluor-5 (RVF5) is a voltage-sensitive dye with improved 2P cross-section for use in thick tissue or brain samples. RVF5 features a dichlororhodol core with pyrrolidyl substitution at the nitrogen center. In mammalian cells under one-photon (1P) illumination, RVF5 demonstrates high voltage sensitivity (28% ΔF/F per 100 mV) and improved photostability relative to first-generation voltage sensors. This photostability enables multisite optical recordings from neurons lacking tuberous sclerosis complex 1, Tsc1, in a mouse model of genetic epilepsy. Using RVF5, we show that Tsc1 KO neurons exhibit increased activity relative to wild-type neurons and additionally show that the proportion of active neurons in the network increases with the loss of Tsc1. The high photostability and voltage sensitivity of RVF5 is recapitulated under 2P illumination. Finally, the ability to chemically tune the 2P absorption profile through the use of rhodol scaffolds affords the unique opportunity to image neuronal voltage changes in acutely prepared mouse brain slices using 2P illumination. Stimulation of the mouse hippocampus evoked spiking activity that was readily discerned with bath-applied RVF5, demonstrating the utility of RVF5 and molecular wire-based voltage sensors with 2P-optimized fluorophores for imaging voltage in intact brain tissue. PMID:28242676

Calcium channel blockers and transmitter release at the normal human neuromuscular junction.

PubMed

Protti, D A; Reisin, R; Mackinley, T A; Uchitel, O D

1996-05-01

Transmitter release evoked by nerve stimulation is highly dependent on Ca2+ entry through voltage-activated plasma membrane channels. Calcium influx may be modified in some neuromuscular diseases like Lambert-Eaton syndrome and amyotrophic lateral sclerosis. We studied the pharmacologic sensitivity of the transmitter release process to different calcium channel blockers in normal human muscles and found that funnel web toxin and omega-Agatoxin-IVA, both P-type calcium channel blockers, blocked nerve-elicited muscle action potentials and inhibited evoked synaptic transmission. The transmitter release was not affected either by nitrendipine, an L-type channel blocker, or omega-Conotoxin-GVIA, an N-type channel blocker. The pharmacologic profile of neuromuscular transmission observed in normal human muscles indicates that P-like channels mediate transmitter release at the motor nerve terminals.

Micromolar-Affinity Benzodiazepine Receptors Regulate Voltage-Sensitive Calcium Channels in Nerve Terminal Preparations

NASA Astrophysics Data System (ADS)

Taft, William C.; Delorenzo, Robert J.

1984-05-01

Benzodiazepines in micromolar concentrations significantly inhibit depolarization-sensitive Ca2+ uptake in intact nerve-terminal preparations. Benzodiazepine inhibition of Ca2+ uptake is concentration dependent and stereospecific. Micromolar-affinity benzodiazepine receptors have been identified and characterized in brain membrane and shown to be distinct from nanomolar-affinity benzodiazepine receptors. Evidence is presented that micromolar, and not nanomolar, benzodiazepine binding sites mediate benzodiazepine inhibition of Ca2+ uptake. Irreversible binding to micromolar benzodiazepine binding sites also irreversibly blocked depolarization-dependent Ca2+ uptake in synaptosomes, indicating that these compounds may represent a useful marker for identifying the molecular components of Ca2+ channels in brain. Characterization of benzodiazepine inhibition of Ca2+ uptake demonstrates that these drugs function as Ca2+ channel antagonists, because benzodiazepines effectively blocked voltage-sensitive Ca2+ uptake inhibited by Mn2+, Co2+, verapamil, nitrendipine, and nimodipine. These results indicate that micromolar benzodiazepine binding sites regulate voltage-sensitive Ca2+ channels in brain membrane and suggest that some of the neuronal stabilizing effects of micromolar benzodiazepine receptors may be mediated by the regulation of Ca2+ conductance.

Micromolar-affinity benzodiazepine receptors regulate voltage-sensitive calcium channels in nerve terminal preparations.

PubMed Central

Taft, W C; DeLorenzo, R J

1984-01-01

Benzodiazepines in micromolar concentrations significantly inhibit depolarization-sensitive Ca2+ uptake in intact nerve-terminal preparations. Benzodiazepine inhibition of Ca2+ uptake is concentration dependent and stereospecific. Micromolar-affinity benzodiazepine receptors have been identified and characterized in brain membrane and shown to be distinct from nanomolar-affinity benzodiazepine receptors. Evidence is presented that micromolar, and not nanomolar, benzodiazepine binding sites mediate benzodiazepine inhibition of Ca2+ uptake. Irreversible binding to micromolar benzodiazepine binding sites also irreversibly blocked depolarization-dependent Ca2+ uptake in synaptosomes, indicating that these compounds may represent a useful marker for identifying the molecular components of Ca2+ channels in brain. Characterization of benzodiazepine inhibition of Ca2+ uptake demonstrates that these drugs function as Ca2+ channel antagonists, because benzodiazepines effectively blocked voltage-sensitive Ca2+ uptake inhibited by Mn2+, Co2+, verapamil, nitrendipine, and nimodipine. These results indicate that micromolar benzodiazepine binding sites regulate voltage-sensitive Ca2+ channels in brain membrane and suggest that some of the neuronal stabilizing effects of micromolar benzodiazepine receptors may be mediated by the regulation of Ca2+ conductance. PMID:6328498

Lack of voltage-dependent calcium channel opening during the calcium influx induced by progesterone in human sperm. Effect of calcium channel deactivation and inactivation.

PubMed

Guzmán-Grenfell, Alberto Martín; González-Martínez, Marco T

2004-01-01

Progesterone induces calcium influx and acrosomal exocytosis in human sperm. Pharmacologic evidence suggests that voltage-dependent calcium channels (VDCCs) are involved. In this study, membrane potential (Vm) and intracellular calcium concentration ([Ca(2+)](i)) were monitored simultaneously to assess the effect of VDCC gating on the calcium influx triggered by progesterone. Holding the Vm to values that maintained VDCCs in a deactivated (-71 mV) closed state inhibited the calcium influx induced by progesterone by approximately 40%. At this Vm, the acrosomal reaction induced by progesterone, but not by A23187, was inhibited. However, when the Vm was held at -15 mV (which maintains VDCCs in an inactivated closed state), the progesterone-induced calcium influx was stimulated. Furthermore, the progesterone and voltage-dependent calcium influxes were additive. These findings indicate that progesterone does not produce VDCC gating in human sperm.

Palette of fluorinated voltage-sensitive hemicyanine dyes

PubMed Central

Yan, Ping; Acker, Corey D.; Zhou, Wen-Liang; Lee, Peter; Bollensdorff, Christian; Negrean, Adrian; Lotti, Jacopo; Sacconi, Leonardo; Antic, Srdjan D.; Kohl, Peter; Mansvelder, Huibert D.; Pavone, Francesco S.; Loew, Leslie M.

2012-01-01

Optical recording of membrane potential permits spatially resolved measurement of electrical activity in subcellular regions of single cells, which would be inaccessible to electrodes, and imaging of spatiotemporal patterns of action potential propagation in excitable tissues, such as the brain or heart. However, the available voltage-sensitive dyes (VSDs) are not always spectrally compatible with newly available optical technologies for sensing or manipulating the physiological state of a system. Here, we describe a series of 19 fluorinated VSDs based on the hemicyanine class of chromophores. Strategic placement of the fluorine atoms on the chromophores can result in either blue or red shifts in the absorbance and emission spectra. The range of one-photon excitation wavelengths afforded by these new VSDs spans 440–670 nm; the two-photon excitation range is 900–1,340 nm. The emission of each VSD is shifted by at least 100 nm to the red of its one-photon excitation spectrum. The set of VSDs, thus, affords an extended toolkit for optical recording to match a broad range of experimental requirements. We show the sensitivity to voltage and the photostability of the new VSDs in a series of experimental preparations ranging in scale from single dendritic spines to whole heart. Among the advances shown in these applications are simultaneous recording of voltage and calcium in single dendritic spines and optical electrophysiology recordings using two-photon excitation above 1,100 nm. PMID:23169660

Calcium current in type I hair cells isolated from the semicircular canal crista ampullaris of the rat.

PubMed

Almanza, Angélica; Vega, Rosario; Soto, Enrique

2003-12-24

The low voltage gain in type I hair cells implies that neurotransmitter release at their afferent synapse should be mediated by low voltage activated calcium channels, or that some peculiar mechanism should be operating in this synapse. With the patch clamp technique, we studied the characteristics of the Ca(2+) current in type I hair cells enzymatically dissociated from rat semicircular canal crista ampullaris. Calcium current in type I hair cells exhibited a slow inactivation (during 2-s depolarizing steps), was sensitive to nimodipine and was blocked by Cd(2+) and Ni(2+). This current was activated at potentials above -60 mV, had a mean half maximal activation of -36 mV, and exhibited no steady-state inactivation at holding potentials between -100 and -60 mV. This data led us to conclude that hair cell Ca(2+) current is most likely of the L type. Thus, other mechanisms participating in neurotransmitter release such as K(+) accumulation in the synaptic cleft, modulation of K(+) currents by nitric oxide, participation of a Na(+) current and possible metabotropic cascades activated by depolarization should be considered.

Expression of voltage-activated calcium channels in the early zebrafish embryo.

PubMed

Sanhueza, Dayán; Montoya, Andro; Sierralta, Jimena; Kukuljan, Manuel

2009-05-01

Increases in cytosolic calcium concentrations regulate many cellular processes, including aspects of early development. Calcium release from intracellular stores and calcium entry through non-voltage-gated channels account for signalling in non-excitable cells, whereas voltage-gated calcium channels (CaV) are important in excitable cells. We report the expression of multiple transcripts of CaV, identified by its homology to other species, in the early embryo of the zebrafish, Danio rerio, at stages prior to the differentiation of excitable cells. CaV mRNAs and proteins were detected as early as the 2-cell stages, which indicate that they arise from both maternal and zygotic transcription. Exposure of embryos to pharmacological blockers of CaV does not perturb early development significantly, although late effects are appreciable. These results suggest that CaV may have a role in calcium homeostasis and control of cellular process during early embryonic development.

Cardiac voltage gated calcium channels and their regulation by β-adrenergic signaling.

PubMed

Kumari, Neema; Gaur, Himanshu; Bhargava, Anamika

2018-02-01

Voltage-gated calcium channels (VGCCs) are the predominant source of calcium influx in the heart leading to calcium-induced calcium release and ultimately excitation-contraction coupling. In the heart, VGCCs are modulated by the β-adrenergic signaling. Signaling through β-adrenergic receptors (βARs) and modulation of VGCCs by β-adrenergic signaling in the heart are critical signaling and changes to these have been significantly implicated in heart failure. However, data related to calcium channel dysfunction in heart failure is divergent and contradictory ranging from reduced function to no change in the calcium current. Many recent studies have highlighted the importance of functional and spatial microdomains in the heart and that may be the key to answer several puzzling questions. In this review, we have briefly discussed the types of VGCCs found in heart tissues, their structure, and significance in the normal and pathological condition of the heart. More importantly, we have reviewed the modulation of VGCCs by βARs in normal and pathological conditions incorporating functional and structural aspects. There are different types of βARs, each having their own significance in the functioning of the heart. Finally, we emphasize the importance of location of proteins as it relates to their function and modulation by co-signaling molecules. Its implication on the studies of heart failure is speculated. Copyright © 2017 Elsevier Inc. All rights reserved.

Calcium-sensitive and insensitive transient outward current in rabbit ventricular myocytes.

PubMed Central

Hiraoka, M; Kawano, S

1989-01-01

1. A suction pipette whole-cell voltage-clamp technique was used to record membrane currents and potentials of isolated ventricular myocytes from rabbit hearts. 2. Transient outward current (Ito) was activated by voltage steps positive to -20 mV, increasing in amplitude with further depolarization to reach a maximum around +70 mV. The current attained its peak within 10 ms and then it inactivated for 100-200 ms. 3. A large portion of Ito still remained after the calcium current (ICa) was blocked when depolarizing pulses were applied at a frequency of 0.1 Hz or less. Therefore, this current component is referred to as calcium-insensitive Ito or It. 4. It showed voltage- and time-dependent inactivation similar to that observed in Purkinje fibres and other cardiac preparations. 5. The reversal potential of It depended on external K+ concentration, [K+]o, with a slope of 32 mV per 10-fold change in the presence of a normal [Na+]o (143 mM), while the slope was 48 mV per 10-fold change in low [Na+]o (1.0 mM). 6. It was completely inhibited by 2-4 mM-4-aminopyridine. Ito in the presence of ICa was also partially blocked by 4-aminopyridine and the remainder was abolished by 5 mM-caffeine. 7. The calcium-insensitive and caffeine-sensitive Ito differed in their decay rates as well as in their recovery time courses. The former was predominantly available at a slow pulsing rate, while the latter increased its amplitude with high-frequency depolarization. 8. The caffeine-sensitive Ito was inhibited by a blockade of ICa, by replacing Ca2+ with Sr2+, by external application of ryanodine and by internal application of EGTA. This indicates that the current is calcium-sensitive and is dependent on increased myoplasmic Ca2+ through Ca2+ influx via the sarcolemma and Ca2+ release from the sarcoplasmic reticulum. The current is therefore designated as IK, Ca. 9. The physiological functions of IK, Ca and It are indicated by their contribution to ventricular repolarization at fast and

Forskolin Regulates L-Type Calcium Channel through Interaction between Actinin 4 and β3 Subunit in Osteoblasts.

PubMed

Zhang, Xuemei; Li, Fangping; Guo, Lin; Hei, Hongya; Tian, Lulu; Peng, Wen; Cai, Hui

2015-01-01

Voltage-dependent L-type calcium channels that permit cellular calcium influx are essential in calcium-mediated modulation of cellular signaling. Although the regulation of voltage-dependent L-type calcium channels is linked to many factors including cAMP-dependent protein kinase A (PKA) activity and actin cytoskeleton, little is known about the detailed mechanisms underlying the regulation in osteoblasts. Our present study investigated the modulation of L-type calcium channel activities through the effects of forskolin on actin reorganization and on its functional interaction with actin binding protein actinin 4. The results showed that forskolin did not significantly affect the trafficking of pore forming α1c subunit and its interaction with actin binding protein actinin 4, whereas it significantly increased the expression of β3 subunit and its interaction with actinin 4 in osteoblast cells as assessed by co-immunoprecipitation, pull-down assay, and immunostaining. Further mapping showed that the ABD and EF domains of actinin 4 were interaction sites. This interaction is independent of PKA phosphorylation. Knockdown of actinin 4 significantly decreased the activities of L-type calcium channels. Our study revealed a new aspect of the mechanisms by which the forskolin activation of adenylyl cyclase - cAMP cascade regulates the L-type calcium channel in osteoblast cells, besides the PKA mediated phosphorylation of the channel subunits. These data provide insight into the important role of interconnection among adenylyl cyclase, cAMP, PKA, the actin cytoskeleton, and the channel proteins in the regulation of voltage-dependent L-type calcium channels in osteoblast cells.

Forskolin Regulates L-Type Calcium Channel through Interaction between Actinin 4 and β3 Subunit in Osteoblasts

PubMed Central

Guo, Lin; Hei, Hongya; Tian, Lulu; Peng, Wen; Cai, Hui

2015-01-01

Voltage-dependent L-type calcium channels that permit cellular calcium influx are essential in calcium-mediated modulation of cellular signaling. Although the regulation of voltage-dependent L-type calcium channels is linked to many factors including cAMP-dependent protein kinase A (PKA) activity and actin cytoskeleton, little is known about the detailed mechanisms underlying the regulation in osteoblasts. Our present study investigated the modulation of L-type calcium channel activities through the effects of forskolin on actin reorganization and on its functional interaction with actin binding protein actinin 4. The results showed that forskolin did not significantly affect the trafficking of pore forming α1c subunit and its interaction with actin binding protein actinin 4, whereas it significantly increased the expression of β3 subunit and its interaction with actinin 4 in osteoblast cells as assessed by co-immunoprecipitation, pull-down assay, and immunostaining. Further mapping showed that the ABD and EF domains of actinin 4 were interaction sites. This interaction is independent of PKA phosphorylation. Knockdown of actinin 4 significantly decreased the activities of L-type calcium channels. Our study revealed a new aspect of the mechanisms by which the forskolin activation of adenylyl cyclase - cAMP cascade regulates the L-type calcium channel in osteoblast cells, besides the PKA mediated phosphorylation of the channel subunits. These data provide insight into the important role of interconnection among adenylyl cyclase, cAMP, PKA, the actin cytoskeleton, and the channel proteins in the regulation of voltage-dependent L-type calcium channels in osteoblast cells. PMID:25902045

N-acetylcysteine-induced vasodilation involves voltage-gated potassium channels in rat aorta.

PubMed

Han, Wei-Qing; Zhu, Ding-Liang; Wu, Ling-Yun; Chen, Qi-Zhi; Guo, Shu-Jie; Gao, Ping-Jin

2009-05-22

N-acetylcysteine (NAC) has a protective effect against vascular dysfunction by decreasing the level of reactive oxygen species (ROS) in experimental and human hypertension. This study was designed to examine whether NAC would relax vascular rings in vitro via nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway, extracellular Ca2+ and/or K+ channels. Rat aortic arteries were mounted in an organ bath, contracted with 0.1, 0.5 or 1 micromol/L phenylephrine to plateau, and the vasodilatory effect of NAC was examined in the absence or presence of ROS scavengers, inhibitors of NO-cGMP pathway or K+ channels. Vascular smooth muscle cells (VSMCs) were loaded with a calcium sensitive fluorescent dye fluo-3 AM, and [Ca2+](i) was determined with laser-scanning confocal microscopy. NAC (0.1-4 mmol/L) dose-dependently relaxed rat aorta pre-contracted with phenylephrine. Endothelium removal, endothelial nitric oxide synthase inhibitor N(omega)-Nitro-l-arginine (L-NNA) (100 micromol/L) or soluble guanylyl cyclase (sGC) inhibitor (ODQ) (10 micromol/L) did not affect NAC-induced vasodilation. In contrast, NAC-induced vasodilation was blunted after extracellular calcium was removed and calcium imaging showed that 4 mmol/L NAC quickly decreased [Ca2+](i) in fluo-3 AM loaded VSMCs. NAC-induced vasodilation was significantly reduced in the presence of voltage-gated K+ channels (Kv) inhibitor 4-aminopyridine (4-AP). The vasodilatory effect of NAC may be explained at least partly by activation of voltage-gated K+ channels.

P-type voltage-dependent calcium channel mediates presynaptic calcium influx and transmitter release in mammalian synapses.

PubMed Central

Uchitel, O D; Protti, D A; Sanchez, V; Cherksey, B D; Sugimori, M; Llinás, R

1992-01-01

We have studied the effect of the purified toxin from the funnel-web spider venom (FTX) and its synthetic analog (sFTX) on transmitter release and presynaptic currents at the mouse neuromuscular junction. FTX specifically blocks the omega-conotoxin- and dihydropyridine-insensitive P-type voltage-dependent Ca2+ channel (VDCC) in cerebellar Purkinje cells. Mammalian neuromuscular transmission, which is insensitive to N- or L-type Ca2+ channel blockers, was effectively abolished by FTX and sFTX. These substances blocked the muscle contraction and the neurotransmitter release evoked by nerve stimulation. Moreover, presynaptic Ca2+ currents recorded extracellularly from the interior of the perineural sheaths of nerves innervating the mouse levator auris muscle were specifically blocked by both natural toxin and synthetic analogue. In a parallel set of experiments, K(+)-induced Ca45 uptake by brain synaptosomes was also shown to be blocked or greatly diminished by FTX and sFTX. These results indicate that the predominant VDCC in the motor nerve terminals, and possibly in a significant percentage of brain synapses, is the P-type channel. Images PMID:1348859

P-type voltage-dependent calcium channel mediates presynaptic calcium influx and transmitter release in mammalian synapses.

PubMed

Uchitel, O D; Protti, D A; Sanchez, V; Cherksey, B D; Sugimori, M; Llinás, R

1992-04-15

We have studied the effect of the purified toxin from the funnel-web spider venom (FTX) and its synthetic analog (sFTX) on transmitter release and presynaptic currents at the mouse neuromuscular junction. FTX specifically blocks the omega-conotoxin- and dihydropyridine-insensitive P-type voltage-dependent Ca2+ channel (VDCC) in cerebellar Purkinje cells. Mammalian neuromuscular transmission, which is insensitive to N- or L-type Ca2+ channel blockers, was effectively abolished by FTX and sFTX. These substances blocked the muscle contraction and the neurotransmitter release evoked by nerve stimulation. Moreover, presynaptic Ca2+ currents recorded extracellularly from the interior of the perineural sheaths of nerves innervating the mouse levator auris muscle were specifically blocked by both natural toxin and synthetic analogue. In a parallel set of experiments, K(+)-induced Ca45 uptake by brain synaptosomes was also shown to be blocked or greatly diminished by FTX and sFTX. These results indicate that the predominant VDCC in the motor nerve terminals, and possibly in a significant percentage of brain synapses, is the P-type channel.

Modulation of voltage- and Ca2+-dependent gating of CaV1.3 L-type calcium channels by alternative splicing of a C-terminal regulatory domain.

PubMed

Singh, Anamika; Gebhart, Mathias; Fritsch, Reinhard; Sinnegger-Brauns, Martina J; Poggiani, Chiara; Hoda, Jean-Charles; Engel, Jutta; Romanin, Christoph; Striessnig, Jörg; Koschak, Alexandra

2008-07-25

Low voltage activation of Ca(V)1.3 L-type Ca(2+) channels controls excitability in sensory cells and central neurons as well as sinoatrial node pacemaking. Ca(V)1.3-mediated pacemaking determines neuronal vulnerability of dopaminergic striatal neurons affected in Parkinson disease. We have previously found that in Ca(V)1.4 L-type Ca(2+) channels, activation, voltage, and calcium-dependent inactivation are controlled by an intrinsic distal C-terminal modulator. Because alternative splicing in the Ca(V)1.3 alpha1 subunit C terminus gives rise to a long (Ca(V)1.3(42)) and a short form (Ca(V)1.3(42A)), we investigated if a C-terminal modulatory mechanism also controls Ca(V)1.3 gating. The biophysical properties of both splice variants were compared after heterologous expression together with beta3 and alpha2delta1 subunits in HEK-293 cells. Activation of calcium current through Ca(V)1.3(42A) channels was more pronounced at negative voltages, and inactivation was faster because of enhanced calcium-dependent inactivation. By investigating several Ca(V)1.3 channel truncations, we restricted the modulator activity to the last 116 amino acids of the C terminus. The resulting Ca(V)1.3(DeltaC116) channels showed gating properties similar to Ca(V)1.3(42A) that were reverted by co-expression of the corresponding C-terminal peptide C(116). Fluorescence resonance energy transfer experiments confirmed an intramolecular protein interaction in the C terminus of Ca(V)1.3 channels that also modulates calmodulin binding. These experiments revealed a novel mechanism of channel modulation enabling cells to tightly control Ca(V)1.3 channel activity by alternative splicing. The absence of the C-terminal modulator in short splice forms facilitates Ca(V)1.3 channel activation at lower voltages expected to favor Ca(V)1.3 activity at threshold voltages as required for modulation of neuronal firing behavior and sinoatrial node pacemaking.

Disruption of the IS6-AID linker affects voltage-gated calcium channel inactivation and facilitation.

PubMed

Findeisen, Felix; Minor, Daniel L

2009-03-01

Two processes dominate voltage-gated calcium channel (Ca(V)) inactivation: voltage-dependent inactivation (VDI) and calcium-dependent inactivation (CDI). The Ca(V)beta/Ca(V)alpha(1)-I-II loop and Ca(2+)/calmodulin (CaM)/Ca(V)alpha(1)-C-terminal tail complexes have been shown to modulate each, respectively. Nevertheless, how each complex couples to the pore and whether each affects inactivation independently have remained unresolved. Here, we demonstrate that the IS6-alpha-interaction domain (AID) linker provides a rigid connection between the pore and Ca(V)beta/I-II loop complex by showing that IS6-AID linker polyglycine mutations accelerate Ca(V)1.2 (L-type) and Ca(V)2.1 (P/Q-type) VDI. Remarkably, mutations that either break the rigid IS6-AID linker connection or disrupt Ca(V)beta/I-II association sharply decelerate CDI and reduce a second Ca(2+)/CaM/Ca(V)alpha(1)-C-terminal-mediated process known as calcium-dependent facilitation. Collectively, the data strongly suggest that components traditionally associated solely with VDI, Ca(V)beta and the IS6-AID linker, are essential for calcium-dependent modulation, and that both Ca(V)beta-dependent and CaM-dependent components couple to the pore by a common mechanism requiring Ca(V)beta and an intact IS6-AID linker.

Disruption of the IS6-AID Linker Affects Voltage-gated Calcium Channel Inactivation and Facilitation

PubMed Central

Findeisen, Felix

2009-01-01

Two processes dominate voltage-gated calcium channel (CaV) inactivation: voltage-dependent inactivation (VDI) and calcium-dependent inactivation (CDI). The CaVβ/CaVα1-I-II loop and Ca2+/calmodulin (CaM)/CaVα1–C-terminal tail complexes have been shown to modulate each, respectively. Nevertheless, how each complex couples to the pore and whether each affects inactivation independently have remained unresolved. Here, we demonstrate that the IS6–α-interaction domain (AID) linker provides a rigid connection between the pore and CaVβ/I-II loop complex by showing that IS6-AID linker polyglycine mutations accelerate CaV1.2 (L-type) and CaV2.1 (P/Q-type) VDI. Remarkably, mutations that either break the rigid IS6-AID linker connection or disrupt CaVβ/I-II association sharply decelerate CDI and reduce a second Ca2+/CaM/CaVα1–C-terminal–mediated process known as calcium-dependent facilitation. Collectively, the data strongly suggest that components traditionally associated solely with VDI, CaVβ and the IS6-AID linker, are essential for calcium-dependent modulation, and that both CaVβ-dependent and CaM-dependent components couple to the pore by a common mechanism requiring CaVβ and an intact IS6-AID linker. PMID:19237593

Molecular mechanism underlying β1 regulation in voltage- and calcium-activated potassium (BK) channels

PubMed Central

Castillo, Karen; Contreras, Gustavo F.; Pupo, Amaury; Torres, Yolima P.; Neely, Alan; González, Carlos; Latorre, Ramon

2015-01-01

Being activated by depolarizing voltages and increases in cytoplasmic Ca2+, voltage- and calcium-activated potassium (BK) channels and their modulatory β-subunits are able to dampen or stop excitatory stimuli in a wide range of cellular types, including both neuronal and nonneuronal tissues. Minimal alterations in BK channel function may contribute to the pathophysiology of several diseases, including hypertension, asthma, cancer, epilepsy, and diabetes. Several gating processes, allosterically coupled to each other, control BK channel activity and are potential targets for regulation by auxiliary β-subunits that are expressed together with the α (BK)-subunit in almost every tissue type where they are found. By measuring gating currents in BK channels coexpressed with chimeras between β1 and β3 or β2 auxiliary subunits, we were able to identify that the cytoplasmic regions of β1 are responsible for the modulation of the voltage sensors. In addition, we narrowed down the structural determinants to the N terminus of β1, which contains two lysine residues (i.e., K3 and K4), which upon substitution virtually abolished the effects of β1 on charge movement. The mechanism by which K3 and K4 stabilize the voltage sensor is not electrostatic but specific, and the α (BK)-residues involved remain to be identified. This is the first report, to our knowledge, where the regulatory effects of the β1-subunit have been clearly assigned to a particular segment, with two pivotal amino acids being responsible for this modulation. PMID:25825713

A 700 V narrow channel nJFET with low pinch-off voltage and suppressed drain-induced barrier lowering effect

NASA Astrophysics Data System (ADS)

Mao, Kun; Qiao, Ming; Zhang, WenTong; Zhang, Bo; Li, Zhaoji

2014-11-01

This paper proposes a 700 V narrow channel region triple-RESURF (reduced surface field) n-type junction field-effect transistor (NCT-nJFET). Compared to traditional structures, low pinch-off voltage (VP) with unobvious drain-induced barrier lowering (DIBL) effect and large saturated current (IDsat) are achieved. This is because p-type buried layer (Pbury) and PWELL are introduced to shape narrow n-type channel in JFET channel region. DIBL sensitivity (SDIBL) is firstly introduced in this paper to analyze the DIBL effect of high-voltage long-channel JFET. Ultra-high breakdown voltage is obtained by triple RESURF technology. Experimental results show that proposed NCT-nJFET achieves 24-V VP, 3.5% SDIBL, 2.3-mA IDsat, 800-V OFF-state breakdown voltage (OFF-BV) and 650-V ON-state breakdown voltage when VGS equals 0 V (ON-BV).

Functional and pharmacological consequences of the distribution of voltage-gated calcium channels in the renal blood vessels.

PubMed

Hansen, P B L

2013-04-01

Calcium channel blockers are widely used to treat hypertension because they inhibit voltage-gated calcium channels that mediate transmembrane calcium influx in, for example, vascular smooth muscle and cardiomyocytes. The calcium channel family consists of several subfamilies, of which the L-type is usually associated with vascular contractility. However, the L-, T- and P-/Q-types of calcium channels are present in the renal vasculature and are differentially involved in controlling vascular contractility, thereby contributing to regulation of kidney function and blood pressure. In the preglomerular vascular bed, all the three channel families are present. However, the T-type channel is the only channel in cortical efferent arterioles which is in contrast to the juxtamedullary efferent arteriole, and that leads to diverse functional effects of L- and T-type channel inhibition. Furthermore, by different mechanisms, T-type channels may contribute to both constriction and dilation of the arterioles. Finally, P-/Q-type channels are involved in the regulation of human intrarenal arterial contractility. The calcium blockers used in the clinic affect not only L-type but also P-/Q- and T-type channels. Therefore, the distinct effect obtained by inhibiting a given subtype or set of channels under experimental settings should be considered when choosing a calcium blocker for treatment. T-type channels seem to be crucial for regulating the GFR and the filtration fraction. Use of blockers is expected to lead to preferential efferent vasodilation, reduction of glomerular pressure and proteinuria. Therefore, renovascular T-type channels might provide novel therapeutic targets, and may have superior renoprotective effects compared to conventional calcium blockers. Acta Physiologica © 2013 Scandinavian Physiological Society.

Wide-field in vivo neocortical calcium dye imaging using a convection-enhanced loading technique combined with simultaneous multiwavelength imaging of voltage-sensitive dyes and hemodynamic signals

PubMed Central

Ma, Hongtao; Harris, Samuel; Rahmani, Redi; Lacefield, Clay O.; Zhao, Mingrui; Daniel, Andy G. S.; Zhou, Zhiping; Bruno, Randy M.; Berwick, Jason; Schwartz, Theodore H.

2014-01-01

Abstract. In vivo calcium imaging is an incredibly powerful technique that provides simultaneous information on fast neuronal events, such as action potentials and subthreshold synaptic activity, as well as slower events that occur in the glia and surrounding neuropil. Bulk-loading methods that involve multiple injections can be used for single-cell as well as wide-field imaging studies. However, multiple injections result in inhomogeneous loading as well as multiple sites of potential cortical injury. We used convection-enhanced delivery to create smooth, continuous loading of a large area of the cortical surface through a solitary injection site and demonstrated the efficacy of the technique using confocal microscopy imaging of single cells and physiological responses to single-trial events of spontaneous activity, somatosensory-evoked potentials, and epileptiform events. Combinations of calcium imaging with voltage-sensitive dye and intrinsic signal imaging demonstrate the utility of this technique in neurovascular coupling investigations. Convection-enhanced loading of calcium dyes may be a useful technique to advance the study of cortical processing when widespread loading of a wide-field imaging is required. PMID:25525611

Wide-field in vivo neocortical calcium dye imaging using a convection-enhanced loading technique combined with simultaneous multiwavelength imaging of voltage-sensitive dyes and hemodynamic signals.

PubMed

Ma, Hongtao; Harris, Samuel; Rahmani, Redi; Lacefield, Clay O; Zhao, Mingrui; Daniel, Andy G S; Zhou, Zhiping; Bruno, Randy M; Berwick, Jason; Schwartz, Theodore H

2014-07-24

In vivo calcium imaging is an incredibly powerful technique that provides simultaneous information on fast neuronal events, such as action potentials and subthreshold synaptic activity, as well as slower events that occur in the glia and surrounding neuropil. Bulk-loading methods that involve multiple injections can be used for single-cell as well as wide-field imaging studies. However, multiple injections result in inhomogeneous loading as well as multiple sites of potential cortical injury. We used convection-enhanced delivery to create smooth, continuous loading of a large area of the cortical surface through a solitary injection site and demonstrated the efficacy of the technique using confocal microscopy imaging of single cells and physiological responses to single-trial events of spontaneous activity, somatosensory-evoked potentials, and epileptiform events. Combinations of calcium imaging with voltage-sensitive dye and intrinsic signal imaging demonstrate the utility of this technique in neurovascular coupling investigations. Convection-enhanced loading of calcium dyes may be a useful technique to advance the study of cortical processing when widespread loading of a wide-field imaging is required.

Circadian phase-dependent effect of nitric oxide on L-type voltage-gated calcium channels in avian cone photoreceptors

PubMed Central

Ko, Michael L.; Shi, Liheng; Huang, Cathy Chia-Yu; Grushin, Kirill; Park, So-Young; Ko, Gladys Y.-P.

2014-01-01

Nitric oxide (NO) plays an important role in phase-shifting of circadian neuronal activities in the suprachiasmatic nucleus and circadian behavior activity rhythms. In the retina, NO production is increased in a light-dependent manner. While endogenous circadian oscillators in retinal photoreceptors regulate their physiological states, it is not clear whether NO also participates in the circadian regulation of photoreceptors. In the present study, we demonstrate that NO is involved in the circadian phase-dependent regulation of L-type voltage-gated calcium channels (L-VGCCs). In chick cone photoreceptors, the L-VGCCα1 subunit expression and the maximal L-VGCC currents are higher at night, and both Ras-MAPK (mitogen-activated protein kinase)-Erk (extracellular-signal-regulated kinase) and Ras-phosphatidylinositol 3 kinase (PI3K)-protein kinase B (Akt) are part of the circadian output pathways regulating L-VGCCs. The NO-cGMP-protein kinase G (PKG) pathway decreases L-VGCCα1 subunit expression and L-VGCC currents at night, but not during the day, and exogenous NO donor or cGMP decreases the phosphorylation of Erk and Akt at night. The protein expression of neural NO synthase (nNOS) is also under circadian control, with both nNOS and NO production being higher during the day. Taken together, NO/cGMP/PKG signaling is involved as part of the circadian output pathway to regulate L-VGCCs in cone photoreceptors. PMID:23895452

Dopamine Induces LTP Differentially in Apical and Basal Dendrites through BDNF and Voltage-Dependent Calcium Channels

ERIC Educational Resources Information Center

Navakkode, Sheeja; Sajikumar, Sreedharan; Korte, Martin; Soong, Tuck Wah

2012-01-01

The dopaminergic modulation of long-term potentiation (LTP) has been studied well, but the mechanism by which dopamine induces LTP (DA-LTP) in CA1 pyramidal neurons is unknown. Here, we report that DA-LTP in basal dendrites is dependent while in apical dendrites it is independent of activation of L-type voltage-gated calcium channels (VDCC).…

Antibodies to voltage-gated potassium and calcium channels in epilepsy.

PubMed

Majoie, H J Marian; de Baets, Mark; Renier, Willy; Lang, Bethan; Vincent, Angela

2006-10-01

To determine the prevalence of antibodies to ion channels in patients with long standing epilepsy. Although the CNS is thought to be protected from circulating antibodies by the blood brain barrier, glutamate receptor antibodies have been reported in Rasmussen's encephalitis, glutamic acid decarboxylase (GAD) antibodies have been found in a few patients with epilepsy, and antibodies to voltage-gated potassium channels (VGKC) have been found in a non-paraneoplastic form of limbic encephalitis (with amnesia and seizures) that responds to immunosuppressive therapy. We retrospectively screened sera from female epilepsy patients (n=106) for autoantibodies to VGKC (Kv 1.1, 1.2 or 1.6), voltage-gated calcium channels (VGCC) (P/Q-type), and GAD. All positive results, based on the values of control data [McKnight, K., Jiang, Y., et al. (2005). Serum antibodies in epilepsy and seizure-associated disorders. Neurology 65, 1730-1735], were retested at lower serum concentrations, and results compared with previously published control data. Demographics, medical history, and epilepsy related information was gathered. The studied group consisted predominantly of patients with long standing drug resistant epilepsy. VGKC antibodies were raised (>100 pM) in six patients. VGCC antibodies (>45 pM) were slightly raised in only one patient. GAD antibodies were <3 U/ml in all patients. The clinical features of the patients with VGKC antibodies differed from previously described patients with limbic encephalitis-like syndrome, and were not different with respect to seizure type, age at first seizure, duration of epilepsy, or use of anti-epileptic drugs from the VGKC antibody negative patients. The results demonstrate that antibodies to VGKC are present in 6% of patients with typical long-standing epilepsy, but whether these antibodies are pathogenic or secondary to the primary disease process needs to be determined.

P/Q-type calcium channel modulators

PubMed Central

Nimmrich, V; Gross, G

2012-01-01

P/Q-type calcium channels are high-voltage-gated calcium channels contributing to vesicle release at synaptic terminals. A number of neurological diseases have been attributed to malfunctioning of P/Q channels, including ataxia, migraine and Alzheimer's disease. To date, only two specific P/Q-type blockers are known: both are peptides deriving from the spider venom of Agelenopsis aperta, ω-agatoxins. Other peptidic calcium channel blockers with activity at P/Q channels are available, albeit with less selectivity. A number of low molecular weight compounds modulate P/Q-type currents with different characteristics, and some exhibit a peculiar bidirectional pattern of modulation. Interestingly, there are a number of therapeutics in clinical use, which also show P/Q channel activity. Because selectivity as well as the exact mode of action is different between all P/Q-type channel modulators, the interpretation of clinical and experimental data is complicated and needs a comprehensive understanding of their target profile. The situation is further complicated by the fact that information on potency varies vastly in the literature, which may be the result of different experimental systems, conditions or the splice variants of the P/Q channel. This review attempts to provide a comprehensive overview of the compounds available that affect the P/Q-type channel and should help with the interpretation of results of in vitro experiments and animal models. It also aims to explain some clinical observations by implementing current knowledge about P/Q channel modulation of therapeutically used non-selective drugs. Chances and challenges of the development of P/Q channel-selective molecules are discussed. PMID:22670568

Development of a sensitive setup for laser spectroscopy studies of very exotic calcium isotopes

NASA Astrophysics Data System (ADS)

Garcia Ruiz, R. F.; Gorges, C.; Bissell, M.; Blaum, K.; Gins, W.; Heylen, H.; Koenig, K.; Kaufmann, S.; Kowalska, M.; Krämer, J.; Lievens, P.; Malbrunot-Ettenauer, S.; Neugart, R.; Neyens, G.; Nörtershäuser, W.; Yordanov, D. T.; Yang, X. F.

2017-04-01

An experimental setup for sensitive high-resolution measurements of hyperfine structure spectra of exotic calcium isotopes has been developed and commissioned at the COLLAPS beam line at ISOLDE, CERN. The technique is based on the radioactive detection of decaying isotopes after optical pumping and state selective neutralization (ROC) (Vermeeren et al 1992 Phys. Rev. Lett. 68 1679). The improvements and developments necessary to extend the applicability of the experimental technique to calcium isotopes produced at rates as low as few ions s-1 are discussed. Numerical calculations of laser-ion interaction and ion-beam simulations were explored to obtain the optimum performance of the experimental setup. Among the implemented features are a multi-step optical pumping region for sensitive measurements of isotopes with hyperfine splitting, a high-voltage platform for adequate control of low-energy ion beams and simultaneous β-detection of neutralized and remaining ions. The commissioning of the experimental setup, and the first online results on neutron-rich calcium isotopes are presented.

L-Histidine sensing by calcium sensing receptor inhibits voltage-dependent calcium channel activity and insulin secretion in β-cells

PubMed Central

Parkash, Jai; Asotra, Kamlesh

2011-01-01

Aims Our goal was to test the hypothesis that the histidine-induced activation of calcium sensing receptor (CaR) can regulate calcium channel activity of L-type voltage dependent calcium channel (VDCC) due to increased spatial interaction between CaR and VDCC in β-cells and thus modulate glucose-induced insulin secretion. Main methods Rat insulinoma (RINr1046-38) insulin-producing β-cells were cultured in RPMI-1640 medium on 25 mm diameter glass coverslips in six-well culture plates in a 5% CO2 incubator at 37°C. The intracellular calcium concentration, [Ca2+]i, was determined by ratio fluorescence microscopy using Fura-2AM. The spatial interactions between CaR and L-type VDCC in β-cells were measured by immunofluorescence confocal microscopy using a Nikon C1 laser scanning confocal microscope. The insulin release was determined by enzyme-linked immunosorbent assay (ELISA). Key findings The additions of increasing concentrations of L-histidine along with 10 mM glucose resulted in 57% decrease in [Ca2+]i. The confocal fluorescence imaging data showed 5.59 to 8.62-fold increase in colocalization correlation coefficient between CaR and VDCC in β-cells exposed to L-histidine thereby indicating increased membrane delimited spatial interactions between these two membrane proteins. The insulin ELISA data showed 54% decrease in 1st phase of glucose-induced insulin secretion in β-cells exposed to increasing concentrations of L-histidine. Significance L-histidine-induced increased spatial interaction of CaR with VDCC can inhibit calcium channel activity of VDCC and consequently regulate glucose-induced insulin secretion by β-cells. The L-type VDCC could therefore be potential therapeutic target in diabetes. PMID:21219913

Voltage-gated currents in identified rat olfactory receptor neurons.

PubMed

Trombley, P Q; Westbrook, G L

1991-02-01

Whole-cell recording techniques were used to characterize voltage-gated membrane currents in neonatal rat olfactory receptor neurons (ORNs) in cell culture. Mature ORNs were identified in culture by their characteristic bipolar morphology, by retrograde labeling techniques, and by olfactory marker protein (OMP) immunoreactivity. ORNs did not have spontaneous activity, but fired action potentials to depolarizing current pulses. Action potentials were blocked by tetrodotoxin (TTX), which contrasts with the TTX-resistant action potentials in salamander olfactory receptor cells (e.g., Firestein and Werblin, 1987). Prolonged, suprathreshold current pulses evoked only a single action potential; however, repetitive firing up to 35 Hz could be elicited by a series of brief depolarizing pulses. Under voltage clamp, the TTX-sensitive sodium current had activation and inactivation properties similar to other excitable cells. In TTX and 20 mM barium, sustained inward current were evoked by voltage steps positive to -30 mV. This current was blocked by Cd (100 microM) and by nifedipine (IC50 = 368 nM) consistent with L-type calcium channels in other neurons. No T-type calcium current was observed. Voltage steps positive to -20 mV also evoked an outward current that did not inactivate during 100-msec depolarizations. Tail current analysis of this current was consistent with a selective potassium conductance. The outward current was blocked by external tetraethylammonium but was unaffected by Cd or 4-aminopyridine (4-AP) or by removal of external calcium. A transient outward current was not observed. The 3 voltage-dependent conductances in cultured rat ORNs appear to be sufficient for 2 essential functions: action potential generation and transmitter release. As a single odorant-activated channel can trigger an action potential (e.g., Lynch and Barry, 1989), the repetitive firing seen with brief depolarizing pulses suggests that ORNs do not integrate sensory input, but rather act

Resveratrol-induced antinociception is involved in calcium channels and calcium/caffeine-sensitive pools.

PubMed

Pan, Xiaoyu; Chen, Jiechun; Wang, Weijie; Chen, Ling; Wang, Lin; Ma, Quan; Zhang, Jianbo; Chen, Lichao; Wang, Gang; Zhang, Meixi; Wu, Hao; Cheng, Ruochuan

2017-02-07

Resveratrol has been widely investigated for its potential health properties, although little is known about its mechanism in vivo. Previous studies have indicated that resveratrol produces antinociceptive effects in mice. Calcium channels and calcium/caffeine-sensitive pools are reported to be associated with analgesic effect. The present study was to explore the involvement of Ca2+ channel and calcium/caffeine-sensitive pools in the antinociceptive response of resveratrol. Tail-flick test was used to assess antinociception in mice treated with resveratrol or the combinations of resveratrol with MK 801, nimodipine, CaCl2, ryanodine and ethylene glycol tetraacetic acid (EGTA), respectively. The Ca2+/calmodulin-dependent protein kinase II (CaMKII) and brain-derived neurotrophic factor (BDNF) levels in the spinal cord were also investigated when treated with the above drugs. The results showed that resveratrol increased the tail flick latency in the tail-flick test, in dose-dependent manner. N-methyl-D-aspartate (NMDA) glutamate receptor antagonist MK 801 potentiated the antinociceptive effects of sub-threshold dose of resveratrol at 10 mg/kg. Ca2+ channel blocker, however, abolished the antinociceptive effects of resveratrol. In contrast to these results, EGTA or ryanodine treatment (i.c.v.) potentiated resveratrol-induced antinociception. There was a significant decrease in p-CaMKII and an increase in BDNF expression in the spinal cord when combined with MK 801, nimodipine, ryanodine and EGTA. While an increase in p-CaMKII level and a decrease in BDNF expression were observed when high dose of resveratrol combined with CaCl2. These findings suggest that resveratrol exhibits the antinociceptive effects by inhibition of calcium channels and calcium/caffeine-sensitive pools.

Restricting calcium currents is required for correct fiber type specification in skeletal muscle

PubMed Central

Sultana, Nasreen; Dienes, Beatrix; Benedetti, Ariane; Tuluc, Petronel; Szentesi, Peter; Sztretye, Monika; Rainer, Johannes; Hess, Michael W.; Schwarzer, Christoph; Obermair, Gerald J.; Csernoch, Laszlo

2016-01-01

ABSTRACT Skeletal muscle excitation-contraction (EC) coupling is independent of calcium influx. In fact, alternative splicing of the voltage-gated calcium channel CaV1.1 actively suppresses calcium currents in mature muscle. Whether this is necessary for normal development and function of muscle is not known. However, splicing defects that cause aberrant expression of the calcium-conducting developmental CaV1.1e splice variant correlate with muscle weakness in myotonic dystrophy. Here, we deleted CaV1.1 (Cacna1s) exon 29 in mice. These mice displayed normal overall motor performance, although grip force and voluntary running were reduced. Continued expression of the developmental CaV1.1e splice variant in adult mice caused increased calcium influx during EC coupling, altered calcium homeostasis, and spontaneous calcium sparklets in isolated muscle fibers. Contractile force was reduced and endurance enhanced. Key regulators of fiber type specification were dysregulated and the fiber type composition was shifted toward slower fibers. However, oxidative enzyme activity and mitochondrial content declined. These findings indicate that limiting calcium influx during skeletal muscle EC coupling is important for the secondary function of the calcium signal in the activity-dependent regulation of fiber type composition and to prevent muscle disease. PMID:26965373

Calcium-dependent inactivation of calcium channels in cochlear hair cells of the chicken.

PubMed

Lee, Seunghwan; Briklin, Olga; Hiel, Hakim; Fuchs, Paul

2007-09-15

Voltage-gated calcium channels support both spontaneous and sound-evoked neurotransmitter release from ribbon synapses of cochlear hair cells. A variety of regulatory mechanisms must cooperate to ensure the appropriate level of activity in the restricted pool of synaptic calcium channels ( approximately 100) available to each synaptic ribbon. One potential feedback mechanism, calcium-dependent inactivation (CDI) of voltage-gated, L-type calcium channels, can be modulated by calmodulin-like calcium-binding proteins. CDI of voltage-gated calcium current was studied in hair cells of the chicken's basilar papilla (analogous to the mammalian cochlea) after blocking the predominant potassium conductances. For inactivating currents produced by 2.5 s steps to the peak of the current-voltage relation (1 mm EGTA internal calcium buffer), single exponential fits yielded an average decay time constant of 1.92 +/- 0.18 s (mean +/- s.e.m., n = 12) at 20-22 degrees C, while recovery occurred with a half-time of approximately 10 s. Inactivation produced no change in reversal potential, arguing that the observed relaxation did not result from alternative processes such as calcium accumulation or activation of residual potassium currents. Substitution of external calcium with barium greatly reduced inactivation, while inhibition of endoplasmic calcium pumps with t-benzohydroquinone (BHQ) or thapsigargin made inactivation occur faster and to a greater extent. Raising external calcium 10-fold (from 2 to 20 mm) increased peak current 3-fold, but did not alter the extent or time course of CDI. However, increasing levels of internal calcium buffer consistently reduced the rate and extent of inactivation. With 1 mm EGTA buffering and in 2 mm external calcium, the available pool of calcium channels was half-inactivated near the resting membrane potential (-50 mV). CDI may be further regulated by calmodulin-like calcium-binding proteins (CaBPs). mRNAs for several CaBPs are expressed in

Bio-inspired voltage-dependent calcium channel blockers.

PubMed

Yang, Tingting; He, Lin-Ling; Chen, Ming; Fang, Kun; Colecraft, Henry M

2013-01-01

Ca(2+) influx via voltage-dependent CaV1/CaV2 channels couples electrical signals to biological responses in excitable cells. CaV1/CaV2 channel blockers have broad biotechnological and therapeutic applications. Here we report a general method for developing novel genetically encoded calcium channel blockers inspired by Rem, a small G-protein that constitutively inhibits CaV1/CaV2 channels. We show that diverse cytosolic proteins (CaVβ, 14-3-3, calmodulin and CaMKII) that bind pore-forming α1-subunits can be converted into calcium channel blockers with tunable selectivity, kinetics and potency, simply by anchoring them to the plasma membrane. We term this method 'channel inactivation induced by membrane-tethering of an associated protein' (ChIMP). ChIMP is potentially extendable to small-molecule drug discovery, as engineering FK506-binding protein into intracellular sites within CaV1.2-α1C permits heterodimerization-initiated channel inhibition with rapamycin. The results reveal a universal method for developing novel calcium channel blockers that may be extended to develop probes for a broad cohort of unrelated ion channels.

Reduced voltage sensitivity in a K+-channel voltage sensor by electric field remodeling

PubMed Central

González-Pérez, Vivian; Stack, Katherine; Boric, Katica; Naranjo, David

2010-01-01

Propagation of the nerve impulse relies on the extreme voltage sensitivity of Na+ and K+ channels. The transmembrane movement of four arginine residues, located at the fourth transmembrane segment (S4), in each of their four voltage-sensing domains is mostly responsible for the translocation of 12 to 13 eo across the transmembrane electric field. Inserting additional positively charged residues between the voltage-sensing arginines in S4 would, in principle, increase voltage sensitivity. Here we show that either positively or negatively charged residues added between the two most external sensing arginines of S4 decreased voltage sensitivity of a Shaker voltage-gated K+-channel by up to ≈50%. The replacement of Val363 with a charged residue displaced inwardly the external boundaries of the electric field by at least 6 Å, leaving the most external arginine of S4 constitutively exposed to the extracellular space and permanently excluded from the electric field. Both the physical trajectory of S4 and its electromechanical coupling to open the pore gate seemed unchanged. We propose that the separation between the first two sensing charges at resting is comparable to the thickness of the low dielectric transmembrane barrier they must cross. Thus, at most a single sensing arginine side chain could be found within the field. The conserved hydrophobic nature of the residues located between the voltage-sensing arginines in S4 may shape the electric field geometry for optimal voltage sensitivity in voltage-gated ion channels. PMID:20194763

Point mutation of a conserved aspartate, D69, in the muscarinic M2 receptor does not modify voltage-sensitive agonist potency.

PubMed

Ågren, Richard; Sahlholm, Kristoffer; Nilsson, Johanna; Århem, Peter

2018-01-29

The muscarinic M 2 receptor (M 2 R) has been shown to display voltage-sensitive agonist binding, based on G protein-activated inward rectifier potassium channel (GIRK) opening and radioligand binding at different membrane voltages. A conserved aspartate in transmembrane segment (TM) II of M 2 R, D69, has been proposed as the voltage sensor. While a recent paper instead presented evidence of tyrosines in TMs III, VI, and VII acting as voltage sensors, these authors were not able to record GIRK channel activation by a D69N mutant M 2 R. In the present study, we succeeded in recording ACh-induced GIRK channel activation by this mutant at -80 and 0 mV. The acetylcholine EC 50 was about 2.5-fold higher at 0 mV, a potency shift very similar to that observed at wild-type M 2 R, indicating that voltage sensitivity persists at the D69N mutant. Thus, our present observations corroborate the notion that D69 is not responsible for voltage sensitivity of the M 2 R. Copyright © 2018 Elsevier Inc. All rights reserved.

The Positive Inotropic Effect of Pyruvate Involves an Increase in Myofilament Calcium Sensitivity

PubMed Central

Torres, Carlos A. A.; Varian, Kenneth D.; Canan, Cynthia H.; Davis, Jonathan P.; Janssen, Paul M. L.

2013-01-01

Pyruvate is a metabolic fuel that is a potent inotropic agent. Despite its unique inotropic and antioxidant properties, the molecular mechanism of its inotropic mechanism is still largely unknown. To examine the inotropic effect of pyruvate in parallel with intracellular calcium handling under near physiological conditions, we measured pH, myofilament calcium sensitivity, developed force, and calcium transients in ultra thin rabbit heart trabeculae at 37 °C loaded iontophoretically with the calcium indicator bis-fura-2. By contrasting conditions of control versus sarcoplasmic reticulum block (with either cyclopiazonic acid and ryanodine or with thapsigargin) we were able to characterize and isolate the effects of pyruvate on sarcoplasmic reticulum calcium handling and developed force. A potassium contracture technique was subsequently utilized to assess the force-calcium relationship and thus the myofilament calcium sensitivity. Pyruvate consistently increased developed force whether or not the sarcoplasmic reticulum was blocked (16.8±3.5 to 24.5±5.1 vs. 6.9±2.6 to 12.5±4.4 mN/mm2, non-blocked vs. blocked sarcoplasmic reticulum respectively, p<0.001, n = 9). Furthermore, the sensitizing effect of pyruvate on the myofilaments was demonstrated by potassium contractures (EC50 at baseline versus 20 minutes of pyruvate infusion (peak force development) was 701±94 vs. 445±65 nM, p<0.01, n = 6). This study is the first to demonstrate that a leftward shift in myofilament calcium sensitivity is an important mediator of the inotropic effect of pyruvate. This finding can have important implications for future development of therapeutic strategies in the management of heart failure. PMID:23691074

Divergent biophysical properties, gating mechanisms, and possible functions of the two skeletal muscle Ca(V)1.1 calcium channel splice variants.

PubMed

Tuluc, Petronel; Flucher, Bernhard E

2011-12-01

Voltage-gated calcium channels are multi-subunit protein complexes that specifically allow calcium ions to enter the cell in response to membrane depolarization. But, for many years it seemed that the skeletal muscle calcium channel Ca(V)1.1 is the exception. The classical splice variant Ca(V)1.1a activates slowly, has a very small current amplitude and poor voltage sensitivity. In fact adult muscle fibers work perfectly well even in the absence of calcium influx. Recently a new splice variant of the skeletal muscle calcium channel Ca(V)1.1e has been characterized. The lack of the 19 amino acid exon 29 in this splice variant results in a rapidly activating calcium channel with high current amplitude and good voltage sensitivity. Ca(V)1.1e is the dominant channel in embryonic muscle, where the expression of this high calcium-conducting Ca(V)1.1 isoform readily explains developmental processes depending on L-type calcium currents. Moreover, the availability of these two structurally similar but functionally distinct channel variants facilitates the analysis of the molecular mechanisms underlying the unique current properties of the classical Ca(V)1.1a channel.

Differential calcium sensitivity in NaV 1.5 mixed syndrome mutants.

PubMed

Abdelsayed, Mena; Baruteau, Alban-Elouen; Gibbs, Karen; Sanatani, Shubhayan; Krahn, Andrew D; Probst, Vincent; Ruben, Peter C

2017-09-15

SCN5a mutations may express gain-of-function (Long QT Syndrome-3), loss-of-function (Brugada Syndrome 1) or both (mixed syndromes), depending on the mutation and environmental triggers. One such trigger may be an increase in cytosolic calcium, accompanying exercise. Many mixed syndromes mutants, including ∆KPQ, E1784K, 1795insD and Q1909R, are found in calcium-sensitive regions. Elevated cytosolic calcium attenuates gain-of-function properties in ∆KPQ, 1795insD and Q1909R, but not in E1784K. By contrast, elevated cytosolic calcium further exacerbates gain-of-function in E1784K by destabilizing slow inactivation. Action potential modelling, using a modified O'Hara Rudy model, suggests that elevated heart rate rescues action potential duration in ∆KPQ, 1795insD and Q1909R, but not in E1784K. Action potential simulations suggest that E1784K carriers have an increased intracellular sodium-to-calcium ratio under bradycardia and tachycardia conditions. Elevated cytosolic calcium, which is common during high heart rates, ameliorates or exacerbates the mixed syndrome phenotype depending on the genetic signature. Inherited arrhythmias may arise from mutations in the gene for SCN5a, which encodes the cardiac voltage-gated sodium channel, Na V 1.5. Mutants in Na V 1.5 result in Brugada Syndrome (BrS1), Long-QT Syndrome (LQT3) or mixed syndromes (an overlap of BrS1/LQT3). Exercise is a potential arrhythmogenic trigger in mixed syndromes. We aimed to determine the effects of elevated cytosolic calcium, which is common during exercise, in mixed syndrome Na V 1.5 mutants. We used whole-cell patch clamp to assess the biophysical properties of Na V 1.5 wild-type (WT), ∆KPQ, E1784K, 1795insD and Q1909R mutants in human embryonic kidney 293 cells transiently transfected with the Na V 1.5 α subunit (WT or mutants), β1 subunit and enhanced green fluorescent protein. Voltage-dependence and kinetics were measured at cytosolic calcium levels of approximately 0, 500 and 2500�

Calmodulins from Schistosoma mansoni: Biochemical analysis and interaction with IQ-motifs from voltage-gated calcium channels.

PubMed

Thomas, Charlotte M; Timson, David J

2018-05-17

The trematode Schistosoma mansoni is a causative agent of schistosomiasis, the second most common parasitic disease of humans after malaria. Calcium homeostasis and calcium-mediated signalling pathways are of particular interest in this species. The drug of choice for treating schistosomiasis, praziquantel, disrupts the regulation of calcium uptake and there is interest in exploiting calcium-mediated processes for future drug discovery. Calmodulin is a calcium sensing protein, present in most eukaryotes. It is a critical regulator of processes as diverse as muscle contraction, cell division and, partly through interaction with voltage-gated calcium channels, intra-cellular calcium concentrations. S. mansoni expresses two highly similar calmodulins - SmCaM1 and SmCaM2. Both proteins interact with calcium, manganese, cadmium (II), iron (II) and lead ions in native gel electrophoresis. These ions also cause conformational changes in the proteins resulting in the exposure of a more hydrophobic surface (as demonstrated by anilinonaphthalene-8-sulfonate fluorescence assays). The proteins are primarily dimeric in the absence of calcium ions, but monomeric in the presence of this ion. Both SmCaM1 and SmCaM2 interact with a peptide corresponding to an IQ-motif derived from the α-subunit of the voltage-gated calcium channel SmCa v 1B (residues 1923-1945). Both proteins bound with slightly higher affinity in the presence of calcium ions. However, there was no difference between the affinities of the two proteins for the peptide. This interaction could be antagonised by chlorpromazine and trifluoperazine, but not praziquantel or thiamylal. Interestingly no interaction could be detected with the other three IQ-motifs identified in S. mansoni voltage-gated ion calcium channels. Copyright © 2018 Elsevier Ltd. All rights reserved.

Voltage-dependent calcium channel involvement in NMDA-induced activation of NOS.

PubMed

Alagarsamy, S; Johnson, K M

1995-11-13

We have previously shown that N-methyl-D-aspartate (NMDA) increases nitric oxide synthase (NOS) activity in rat frontal cortex; however, the actual mechanism of this activation has not been addressed. Tetrodotoxin (TTX; 0.05 microM) inhibited NMDA-activated NOS, suggesting that TTX-sensitive Na+ channels are interposed between the NMDA receptors and the NOS cellular compartment. The NMDA response was also blocked by voltage-dependent Ca2+ channel (VDCC) blockers including Cd2+, Co2+, funnel web spider toxin (FTX) and omega-Aga IVa, but not by nifedipine or omega-conotoxin. These data suggest that Ca2+ flux through P- and/or Q-type VDCC subsequent to NMDA-induced depolarization may be at least as important for NOS activation as Ca2+ entry through the NMDA receptor.

Identification of a Novel EF-Loop in the N-terminus of TRPM2 Channel Involved in Calcium Sensitivity

PubMed Central

Luo, Yuhuan; Yu, Xiafei; Ma, Cheng; Luo, Jianhong; Yang, Wei

2018-01-01

As an oxidative stress sensor, transient receptor potential melastatin 2 (TRPM2) channel is involved in many physiological and pathological processes including warmth sensing, ischemia injury, inflammatory diseases and diabetes. Intracellular calcium is critical for TRPM2 channel activation and the IQ-like motif in the N-terminus has been shown to be important by mediating calmodulin binding. Sequence analysis predicted two potential EF-loops in the N-terminus of TRPM2. Site-directed mutagenesis combining with functional assay showed that substitution with alanine of several residues, most of which are conserved in the typical EF-loop, including D267, D278, D288, and E298 dramatically reduced TRPM2 channel currents. By further changing the charges or side chain length of these conserved residues, our results indicate that the negative charge of D267 and the side chain length of D278 are critical for calcium-induced TRPM2 channel activation. G272I mutation also dramatically reduced the channel currents, suggesting that this site is critical for calcium-induced TRPM2 channel activation. Furthermore, D267A mutant dramatically reduced the currents induced by calcium alone compared with that by ADPR, indicating that D267 residue in D267–D278 motif is the most important site for calcium sensitivity of TRPM2. In addition, inside-out recordings showed that mutations at D267, G272, D278, and E298 had no effect on single-channel conductance. Taken together, our data indicate that D267–D278 motif in the N-terminus as a novel EF-loop is critical for calcium-induced TRPM2 channel activation.

Magnolol inhibits colonic motility through down-regulation of voltage-sensitive L-type Ca2+ channels of colonic smooth muscle cells in rats.

PubMed

Zhang, Man; Zang, Kai-Hong; Luo, Jia-Lie; Leung, Fung-Ping; Huang, Yu; Lin, Cheng-Yuan; Yang, Zhi-Jun; Lu, Ai-Ping; Tang, Xu-Dong; Xu, Hong-Xi; Sung, Joseph Jao-yiu; Bian, Zhao-Xiang

2013-11-15

This study aimed to investigate the effect of magnolol (5,5'-diallyl-2,2'-biphenyldiol) on contraction in distal colonic segments of rats and the underlying mechanisms. Colonic segments were mounted in organ baths for isometric force measurement. Whole-cell voltage-sensitive L-type Ca(2+) currents were recorded on isolated single colonic smooth muscle cells using patch-clamp technique. The spontaneous contractions and acetylcholine (ACh)- and Bay K 8644-induced contractions were inhibited by magnolol (3-100 μM). In the presence of Bay K8644 (100 nM), magnolol (10-100 μM) inhibited the contraction induced by 10 μM ACh. By contrast, tetrodotoxin (100 nM) and Nώ-nitro-L-arginine methyl ester (L-NAME 100 μM) did not change the inhibitory effect of magnolol (10 μM). In addition, magnolol (3-100 μM) inhibited the L-type Ca(2+) currents. The present results suggest that magnolol inhibits colonic smooth muscle contraction through downregulating L-type Ca(2+) channel activity. Copyright © 2013 Elsevier GmbH. All rights reserved.

Inhibition of Voltage-Gated Calcium Channels as Common Mode of Action for (Mixtures of) Distinct Classes of Insecticides

PubMed Central

Meijer, Marieke; Dingemans, Milou M.L.; van den Berg, Martin; Westerink, Remco H.S.

2014-01-01

Humans are exposed to distinct structural classes of insecticides with different neurotoxic modes of action. Because calcium homeostasis is essential for proper neuronal function and development, we investigated the effects of insecticides from different classes (pyrethroid: (α-)cypermethrin; organophosphate: chlorpyrifos; organochlorine: endosulfan; neonicotinoid: imidacloprid) and mixtures thereof on the intracellular calcium concentration ([Ca2+]i). Effects of acute (20 min) exposure to (mixtures of) insecticides on basal and depolarization-evoked [Ca2+]i were studied in vitro with Fura-2-loaded PC12 cells and high resolution single-cell fluorescence microscopy. The data demonstrate that cypermethrin, α-cypermethrin, endosulfan, and chlorpyrifos concentration-dependently decreased depolarization-evoked [Ca2+]i, with 50% (IC50) at 78nM, 239nM, 250nM, and 899nM, respectively. Additionally, acute exposure to chlorpyrifos or endosulfan (10μM) induced a modest increase in basal [Ca2+]i, amounting to 68 ± 8nM and 53 ± 8nM, respectively. Imidacloprid did not disturb basal or depolarization-evoked [Ca2+]i at 10μM. Following exposure to binary mixtures, effects on depolarization-evoked [Ca2+]i were within the expected effect additivity range, whereas the effect of the tertiary mixture was less than this expected additivity effect range. These results demonstrate that different types of insecticides inhibit depolarization-evoked [Ca2+]i in PC12 cells by inhibiting voltage-gated calcium channels (VGCCs) in vitro at concentrations comparable with human occupational exposure levels. Moreover, the effective concentrations in this study are below those for earlier described modes of action. Because inhibition of VGCCs appears to be a common and potentially additive mode of action of several classes of insecticides, this target should be considered in neurotoxicity risk assessment studies. PMID:24913802

Calcium-dependent inactivation of calcium channels in cochlear hair cells of the chicken

PubMed Central

Lee, Seunghwan; Briklin, Olga; Hiel, Hakim; Fuchs, Paul

2007-01-01

Voltage-gated calcium channels support both spontaneous and sound-evoked neurotransmitter release from ribbon synapses of cochlear hair cells. A variety of regulatory mechanisms must cooperate to ensure the appropriate level of activity in the restricted pool of synaptic calcium channels (∼100) available to each synaptic ribbon. One potential feedback mechanism, calcium-dependent inactivation (CDI) of voltage-gated, L-type calcium channels, can be modulated by calmodulin-like calcium-binding proteins. CDI of voltage-gated calcium current was studied in hair cells of the chicken's basilar papilla (analogous to the mammalian cochlea) after blocking the predominant potassium conductances. For inactivating currents produced by 2.5 s steps to the peak of the current–voltage relation (1 mm EGTA internal calcium buffer), single exponential fits yielded an average decay time constant of 1.92 ± 0.18 s (mean ±s.e.m., n = 12) at 20–22°C, while recovery occurred with a half-time of ∼10 s. Inactivation produced no change in reversal potential, arguing that the observed relaxation did not result from alternative processes such as calcium accumulation or activation of residual potassium currents. Substitution of external calcium with barium greatly reduced inactivation, while inhibition of endoplasmic calcium pumps with t-benzohydroquinone (BHQ) or thapsigargin made inactivation occur faster and to a greater extent. Raising external calcium 10-fold (from 2 to 20 mm) increased peak current 3-fold, but did not alter the extent or time course of CDI. However, increasing levels of internal calcium buffer consistently reduced the rate and extent of inactivation. With 1 mm EGTA buffering and in 2 mm external calcium, the available pool of calcium channels was half-inactivated near the resting membrane potential (−50 mV). CDI may be further regulated by calmodulin-like calcium-binding proteins (CaBPs). mRNAs for several CaBPs are expressed in chicken cochlear tissue, and

A voltage-division-type low-jitter self-triggered repetition-rate switch.

PubMed

Su, Jian-Cang; Zeng, Bo; Gao, Peng-Cheng; Li, Rui; Wu, Xiao-Long; Zhao, Liang

2016-10-01

A voltage-division-type (V/N) low-jitter self-triggered multi-stage switch is put forward. It comprises of a triggered corona gap, several quasi-uniform-field gaps, and an inversion inductor. When the corona gap is in the stage of self-breakdown, the multi-stage gaps are triggered and the switch is closed via an over-voltage. This type of V/N switch has the advantage of compact structure since the auxiliary components like the gas-blowing system and the triggered system are eliminated from the whole system. It also has advantages such as low breakdown jitter and high energy efficiency. The dependence of the self-triggered voltage on the over-voltage factor and the switch operating voltage is deduced. A switch of this type is designed and fabricated and experiments to research its characteristics are conducted. The results show that this switch can operate on a voltage of 1 MV at 50 Hz and can generate 1000 successive pulses with a jitter as low as 3% and an energy efficiency as high as 90%. This V/N switch can work under a high repetition rate with a long lifetime.

THE CRITICAL ROLE OF VOLTAGE-DEPENDENT CALCIUM CHANNEL IN AXONAL REPAIR FOLLOWING MECHANICAL TRAUMA

PubMed Central

Nehrt, Ashley; Rodgers, Richard; Shapiro, Scott; Borgens, Richard; Shi, Riyi

2009-01-01

Membrane disruption following mechanical injury likely plays a critical role in the pathology of spinal cord trauma. It is known that intracellular calcium is a key factor that is essential to membrane resealing. However, the differential role of calcium influx through the injury site and through voltage dependent calcium channels (VDCC) has not been examined in detail. Using a well established ex vivo guinea pig spinal cord white matter preparation, we have found that axonal membrane resealing was significantly inhibited following transection or compression in the presence of cadmiun, a non-specific calcium channel blocker, or nimodipine, a specific L-type calcium channel blocker. Membrane resealing was assessed by the changes of membrane potential and compound action potential (CAP), and exclusion of horseradish peroxidase 60 minutes following trauma. Furthermore, 1 μM BayK 8644, a VDCC agonist, significantly enhanced membrane resealing. Interestingly, this effect was completely abolished when the concentration of BayK 8644 was increased to 30 μM. These data suggest that VDCC play a critical role in membrane resealing. Further, there is likely an appropriate range of calcium influx through VDCC which ensures effective axonal membrane resealing. Since elevated intracellular calcium has also been linked to axonal deterioration, blockage of VDCC is proposed to be a clinical treatment for various injuries. The knowledge gained in this study will likely help us better understand the role of calcium in various CNS trauma, which is critical for designing new approaches or perhaps optimizing the effectiveness of existing methods in the treatment of CNS trauma. PMID:17448606

Endothelin induces two types of contractions of rat uterus: phasic contractions by way of voltage-dependent calcium channels and developing contractions through a second type of calcium channels

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

Kozuka, M.; Ito, T.; Hirose, S.

1989-02-28

Effects of endothelin on nonvascular smooth muscle have been examined using rat uterine horns and two modes of endothelin action have been revealed. Endothelin (0.3 nM) caused rhythmic contractions of isolated uterus in the presence of extracellular calcium. The rhythmic contractions were completely inhibited by calcium channel antagonists. These characteristics of endothelin-induced contractions were very similar to those induced by oxytocin. Binding assays using /sup 125/I-endothelin showed that endothelin and the calcium channel blockers did not compete for the binding sites. However, endothelin was unique in that it caused, in addition to rhythmic contractions, a slowly developing monophasic contraction thatmore » was insensitive to calcium channel blockers. This developing contraction became dominant at higher concentrations of endothelin and was also calcium dependent.« less

AKAP150-mediated TRPV1 sensitization is disrupted by calcium/calmodulin.

PubMed

Chaudhury, Sraboni; Bal, Manjot; Belugin, Sergei; Shapiro, Mark S; Jeske, Nathaniel A

2011-05-14

The transient receptor potential vanilloid type1 (TRPV1) is expressed in nociceptive sensory neurons and is sensitive to phosphorylation. A-Kinase Anchoring Protein 79/150 (AKAP150) mediates phosphorylation of TRPV1 by Protein Kinases A and C, modulating channel activity. However, few studies have focused on the regulatory mechanisms that control AKAP150 association with TRPV1. In the present study, we identify a role for calcium/calmodulin in controlling AKAP150 association with, and sensitization of, TRPV1. In trigeminal neurons, intracellular accumulation of calcium reduced AKAP150 association with TRPV1 in a manner sensitive to calmodulin antagonism. This was also observed in transfected Chinese hamster ovary (CHO) cells, providing a model for conducting molecular analysis of the association. In CHO cells, the deletion of the C-terminal calmodulin-binding site of TRPV1 resulted in greater association with AKAP150, and increased channel activity. Furthermore, the co-expression of wild-type calmodulin in CHOs significantly reduced TRPV1 association with AKAP150, as evidenced by total internal reflective fluorescence-fluorescence resonance energy transfer (TIRF-FRET) analysis and electrophysiology. Finally, dominant-negative calmodulin co-expression increased TRPV1 association with AKAP150 and increased basal and PKA-sensitized channel activity. the results from these studies indicate that calcium/calmodulin interferes with the association of AKAP150 with TRPV1, potentially extending resensitization of the channel.

Progress in the structural understanding of voltage-gated calcium channel (CaV) function and modulation

PubMed Central

Findeisen, Felix

2010-01-01

Voltage-gated calcium channels (CaVs) are large, transmembrane multiprotein complexes that couple membrane depolarization to cellular calcium entry. These channels are central to cardiac action potential propagation, neurotransmitter and hormone release, muscle contraction and calcium-dependent gene transcription. Over the past six years, the advent of high-resolution structural studies of CaV components from different isoforms and CaV modulators has begun to reveal the architecture that underlies the exceptionally rich feedback modulation that controls CaV action. These descriptions of CaV molecular anatomy have provided new, structure-based insights into the mechanisms by which particular channel elements affect voltage-dependent inactivation (VDI), calcium-dependent inactivation (CDI) and calcium-dependent facilitation (CDF). The initial successes have been achieved through structural studies of soluble channel domains and modulator proteins and have proven most powerful when paired with biochemical and functional studies that validate ideas inspired by the structures. Here, we review the progress in this growing area and highlight some key open challenges for future efforts. PMID:21139419

Construction and use of a zebrafish heart voltage and calcium optical mapping system, with integrated electrocardiogram and programmable electrical stimulation

PubMed Central

Lin, Eric; Craig, Calvin; Lamothe, Marcel; Sarunic, Marinko V.; Beg, Mirza Faisal

2015-01-01

Zebrafish are increasingly being used as a model of vertebrate cardiology due to mammalian-like cardiac properties in many respects. The size and fecundity of zebrafish make them suitable for large-scale genetic and pharmacological screening. In larger mammalian hearts, optical mapping is often used to investigate the interplay between voltage and calcium dynamics and to investigate their respective roles in arrhythmogenesis. This report outlines the construction of an optical mapping system for use with zebrafish hearts, using the voltage-sensitive dye RH 237 and the calcium indicator dye Rhod-2 using two industrial-level CCD cameras. With the use of economical cameras and a common 532-nm diode laser for excitation, the rate dependence of voltage and calcium dynamics within the atrial and ventricular compartments can be simultaneously determined. At 140 beats/min, the atrial action potential duration was 36 ms and the transient duration was 53 ms. With the use of a programmable electrical stimulator, a shallow rate dependence of 3 and 4 ms per 100 beats/min was observed, respectively. In the ventricle the action potential duration was 109 ms and the transient duration was 124 ms, with a steeper rate dependence of 12 and 16 ms per 100 beats/min. Synchronous electrocardiograms and optical mapping recordings were recorded, in which the P-wave aligns with the atrial voltage peak and R-wave aligns with the ventricular peak. A simple optical pathway and imaging chamber are detailed along with schematics for the in-house construction of the electrocardiogram amplifier and electrical stimulator. Laboratory procedures necessary for zebrafish heart isolation, cannulation, and loading are also presented. PMID:25740339

Calmodulin and calcium differentially regulate the neuronal Nav1.1 voltage-dependent sodium channel

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

Gaudioso, Christelle; Carlier, Edmond; Youssouf, Fahamoe

2011-07-29

Highlights: {yields} Both Ca{sup ++}-Calmodulin (CaM) and Ca{sup ++}-free CaM bind to the C-terminal region of Nav1.1. {yields} Ca{sup ++} and CaM have both opposite and convergent effects on I{sub Nav1.1}. {yields} Ca{sup ++}-CaM modulates I{sub Nav1.1} amplitude. {yields} CaM hyperpolarizes the voltage-dependence of activation, and increases the inactivation rate. {yields} Ca{sup ++} alone antagonizes CaM for both effects, and depolarizes the voltage-dependence of inactivation. -- Abstract: Mutations in the neuronal Nav1.1 voltage-gated sodium channel are responsible for mild to severe epileptic syndromes. The ubiquitous calcium sensor calmodulin (CaM) bound to rat brain Nav1.1 and to the human Nav1.1 channelmore » expressed by a stably transfected HEK-293 cell line. The C-terminal region of the channel, as a fusion protein or in the yeast two-hybrid system, interacted with CaM via a consensus C-terminal motif, the IQ domain. Patch clamp experiments on HEK1.1 cells showed that CaM overexpression increased peak current in a calcium-dependent way. CaM had no effect on the voltage-dependence of fast inactivation, and accelerated the inactivation kinetics. Elevating Ca{sup ++} depolarized the voltage-dependence of fast inactivation and slowed down the fast inactivation kinetics, and for high concentrations this effect competed with the acceleration induced by CaM alone. Similarly, the depolarizing action of calcium antagonized the hyperpolarizing shift of the voltage-dependence of activation due to CaM overexpression. Fluorescence spectroscopy measurements suggested that Ca{sup ++} could bind the Nav1.1 C-terminal region with micromolar affinity.« less

Attenuated response of L-type calcium current to nitric oxide in atrial fibrillation.

PubMed

Rozmaritsa, Nadiia; Christ, Torsten; Van Wagoner, David R; Haase, Hannelore; Stasch, Johannes-Peter; Matschke, Klaus; Ravens, Ursula

2014-03-01

Nitric oxide (NO) synthesized by cardiomyocytes plays an important role in the regulation of cardiac function. Here, we studied the impact of NO signalling on calcium influx in human right atrial myocytes and its relation to atrial fibrillation (AF). Right atrial appendages (RAAs) were obtained from patients in sinus rhythm (SR) and AF. The biotin-switch technique was used to evaluate endogenous S-nitrosylation of the α1C subunit of L-type calcium channels. Comparing SR to AF, S-nitrosylation of Ca(2+) channels was similar. Direct effects of the NO donor S-nitroso-N-acetyl-penicillamine (SNAP) on L-type calcium current (ICa,L) were studied in cardiomyocytes with standard voltage-clamp techniques. In SR, ICa,L increased with SNAP (100 µM) by 48%, n/N = 117/56, P < 0.001. The SNAP effect on ICa,L involved activation of soluble guanylate cyclase and protein kinase A. Specific inhibition of phosphodiesterase (PDE)3 with cilostamide (1 µM) enhanced ICa,L to a similar extent as SNAP. However, when cAMP was elevated by PDE3 inhibition or β-adrenoceptor stimulation, SNAP reduced ICa,L, pointing to cGMP-cAMP cross-regulation. In AF, the stimulatory effect of SNAP on ICa,L was attenuated, while its inhibitory effect on isoprenaline- or cilostamide-stimulated current was preserved. cGMP elevation with SNAP was comparable between the SR and AF group. Moreover, the expression of PDE3 and soluble guanylate cyclase was not reduced in AF. NO exerts dual effects on ICa,L in SR with an increase of basal and inhibition of cAMP-stimulated current, and in AF NO inhibits only stimulated ICa,L. We conclude that in AF, cGMP regulation of PDE2 is preserved, but regulation of PDE3 is lost.

[Effects of the monosaccharide derivative 8RN-DAGal on the putative P-type calcium channel expressed in Xenopus oocytes].

PubMed

Fournier, F; Charpentier, G; Lahyani, A; Bruner, J; Czternasty, G; Marlot, D; Ronco, G; Villa, P; Brule, G

1993-01-01

P-type calcium channels are expressed in Xenopus oocytes after injection of rat cerebellar mRNA. The FTX and omega-Aga-IVa toxins extracted from Agelenopsis aperta venom are known to inhibit the activity of this channel. The present results demonstrate that 8RN-DAGal is also a antagonist of P-type calcium channels. The inhibition of the current, obtained with Ba2+, as charge carrier, is voltage dependent.

Accuracy criteria voltage electrocardiography left ventricular hypertrophy to distinguish types of left ventricular hypertrophy geometry

NASA Astrophysics Data System (ADS)

Tumbur, O.; Safri, Z.; Hassan, R.

2018-03-01

Different types of left ventricular hypertrophy geometry are associated with different risk of cardiovascular disease. The purpose of this study was to determine the role of various ECG voltages of LVH to distinguish the type of LVH geometry. A cross-sectional study from June to November 2015, 100 patients in Adam Malik Hospital Medan. The result of LVH ECG criteria of Sokolow-Lyon was not met then obtained normal left ventricular geometry with 60% sensitivity, 72.22% specificity, and 71% accuracy. The eccentric type of LVH is obtained when the Cornel Voltage is not met; the sensitivity is 25%, specificity 71.88%, and 55% accuracy. Concentric geometric hypertrophy when the RV6/V5> 1 ratio is satisfied, the sensitivity is 55.56%, specificity 56.36%, and 56% accuracy. The RV6/V5>1 ratio was not met, the concentric geometry type of hypertrophy remodeling was determined with a sensitivity of 55.56%, specificity 49.45%, and 50% accuracy. Conclusions, various LVHECG criteria distinguish the type of LVH geometry. Sokolow-Lyon and Cornel Voltage sensitivity and specificity are better than the RV6/V5 ratio.

AKAP150-mediated TRPV1 sensitization is disrupted by calcium/calmodulin

PubMed Central

2011-01-01

Background The transient receptor potential vanilloid type1 (TRPV1) is expressed in nociceptive sensory neurons and is sensitive to phosphorylation. A-Kinase Anchoring Protein 79/150 (AKAP150) mediates phosphorylation of TRPV1 by Protein Kinases A and C, modulating channel activity. However, few studies have focused on the regulatory mechanisms that control AKAP150 association with TRPV1. In the present study, we identify a role for calcium/calmodulin in controlling AKAP150 association with, and sensitization of, TRPV1. Results In trigeminal neurons, intracellular accumulation of calcium reduced AKAP150 association with TRPV1 in a manner sensitive to calmodulin antagonism. This was also observed in transfected Chinese hamster ovary (CHO) cells, providing a model for conducting molecular analysis of the association. In CHO cells, the deletion of the C-terminal calmodulin-binding site of TRPV1 resulted in greater association with AKAP150, and increased channel activity. Furthermore, the co-expression of wild-type calmodulin in CHOs significantly reduced TRPV1 association with AKAP150, as evidenced by total internal reflective fluorescence-fluorescence resonance energy transfer (TIRF-FRET) analysis and electrophysiology. Finally, dominant-negative calmodulin co-expression increased TRPV1 association with AKAP150 and increased basal and PKA-sensitized channel activity. Conclusions the results from these studies indicate that calcium/calmodulin interferes with the association of AKAP150 with TRPV1, potentially extending resensitization of the channel. PMID:21569553

Highly efficient and low voltage silver nanowire-based OLEDs employing a n-type hole injection layer.

PubMed

Lee, Hyungjin; Lee, Donghwa; Ahn, Yumi; Lee, Eun-Woo; Park, Lee Soon; Lee, Youngu

2014-08-07

Highly flexible and efficient silver nanowire-based organic light-emitting diodes (OLEDs) have been successfully fabricated by employing a n-type hole injection layer (HIL). The silver nanowire-based OLEDs without light outcoupling structures exhibited excellent device characteristics such as extremely low turn-on voltage (3.6 V) and high current and power efficiencies (44.5 cd A(-1) and 35.8 lm W(-1)). In addition, flexible OLEDs with the silver nanowire transparent conducting electrode (TCE) and n-type HIL fabricated on plastic substrates showed remarkable mechanical flexibility as well as device performance.

Voltage inactivation of Ca2+ entry and secretion associated with N- and P/Q-type but not L-type Ca2+ channels of bovine chromaffin cells

PubMed Central

Villarroya, Mercedes; Olivares, Román; Ruíz, Ana; Cano-Abad, María F; de Pascual, Ricardo; Lomax, Richard B; López, Manuela G; Mayorgas, Inés; Gandía, Luis; García, Antonio G

1999-01-01

In this study we pose the question of why the bovine adrenal medullary chromaffin cell needs various subtypes (L, N, P, Q) of the neuronal high-voltage activated Ca2+ channels to control a given physiological function, i.e. the exocytotic release of catecholamines. One plausible hypothesis is that Ca2+ channel subtypes undergo different patterns of inactivation during cell depolarization. The net Ca2+ uptake (measured using 45Ca2+) into hyperpolarized cells (bathed in a nominally Ca2+-free solution containing 1·2 mM K+) after application of a Ca2+ pulse (5 s exposure to 100 mM K+ and 2 mM Ca2+), amounted to 0·65 ± 0·02 fmol cell−1; in depolarized cells (bathed in nominally Ca2+-free solution containing 100 mM K+) the net Ca2+ uptake was 0·16 ± 0·01 fmol cell−1. This was paralleled by a dramatic reduction of the increase in the cytosolic Ca2+ concentration, [Ca2+]i, caused by Ca2+ pulses applied to fura-2-loaded single cells, from 1181 ± 104 nM in hyperpolarized cells to 115 ± 9 nM in depolarized cells. A similar decrease was observed when studying catecholamine release. Secretion was decreased when K+ concentration was increased from 1·2 to 100 mM; the Ca2+ pulse caused, when comparing the extreme conditions, the secretion of 807 ± 35 nA of catecholamines in hyperpolarized cells and 220 ± 19 nA in depolarized cells. The inactivation by depolarization of Ca2+ entry and secretion occluded the blocking effects of combined ω-conotoxin GVIA (1 μM) and ω-agatoxin IVA (2 μM), thus suggesting that depolarization caused a selective inactivation of the N- and P/Q-type Ca2+ channels. This was strengthened by two additional findings: (i) nifedipine (3 μM), an L-type Ca2+ channel blocker, suppressed the fraction of Ca2+ entry (24 %) and secretion (27 %) left unblocked by depolarization; (ii) FPL64176 (3 μM), an L-type Ca2+ channel ‘activator’, dramatically enhanced the entry of Ca2+ and the secretory response in depolarized cells. In voltage

Sigma-1 receptor agonist increases axon outgrowth of hippocampal neurons via voltage-gated calcium ions channels.

PubMed

Li, Dong; Zhang, Shu-Zhuo; Yao, Yu-Hong; Xiang, Yun; Ma, Xiao-Yun; Wei, Xiao-Li; Yan, Hai-Tao; Liu, Xiao-Yan

2017-12-01

Sigma-1 receptors (Sig-1Rs) are unique endoplasmic reticulum proteins that have been implicated in both neurodegenerative and ischemic diseases, such as Alzheimer's disease and stroke. Accumulating evidence has suggested that Sig-1R plays a role in neuroprotection and axon outgrowth. The underlying mechanisms of Sig-1R-mediated neuroprotection have been well elucidated. However, the mechanisms underlying the effects of Sig-1R on axon outgrowth are not fully understood. To clarify this issue, we utilized immunofluorescence to compare the axon lengths of cultured naïve hippocampal neurons before and after the application of the Sig-1R agonist, SA4503. Then, electrophysiology and immunofluorescence were used to examine voltage-gated calcium ion channel (VGCCs) currents in the cell membranes and growth cones. We found that Sig-1R activation dramatically enhanced the axonal length of the naïve hippocampal neurons. Application of the Sig-1R antagonist NE100 and gene knockdown techniques both demonstrated the effects of Sig-1R. The growth-promoting effect of SA4503 was accompanied by the inhibition of voltage-gated Ca 2+ influx and was recapitulated by incubating the neurons with the L-type, N-type, and P/Q-type VGCC blockers, nimodipine, MVIIA and ω-agatoxin IVA, respectively. This effect was unrelated to glial cells. The application of SA4503 transformed the growth cone morphologies from complicated to simple, which favored axon outgrowth. Sig-1R activation can enhance axon outgrowth and may have a substantial influence on neurogenesis and neurodegenerative diseases. © 2017 John Wiley & Sons Ltd.

New design of a passive type RADFET reader for enhanced sensitivity

NASA Astrophysics Data System (ADS)

Lee, Dae-Hee

2016-07-01

We present a new design of a passive type RADFET reader with enhanced radiation sensitivity. Using a electostatic plate, we have applied a static electric field to the gate voltage, which impacts a positive biasing on the p-type MOSFET. The resultant effect shows that 1.8 times of radiation sensitivity increased when we measured the threshold voltage shift of the RADFET exposed to 30 krad irradiation. We discuss further about the characteristic changes of a RADFET according to the positive biasing on the gate voltage.

Intrathecal P/Q- and R-type calcium channel blockades on spinal substance P release and c-Fos expression

PubMed Central

Terashima, Tetsuji; Xu, Qinghao; Yamaguchi, Shigeki; Yaksh, Tony L.

2013-01-01

Intrathecal (IT) studies have shown that several voltage sensitive calcium channels (VSCCs), such as the L-, N- and T-type may play roles in nociception and that of these only the N-type regulates primary afferent substance P (SP) release. However, the actions of other VSCCs at the spinal level are not well known. We investigated the roles of spinal P/Q- and R-type VSCCs, by IT administration of R-type (SNX-482) and P/Q-type (ω-agatoxin IVA) VSCC blockers on intraplantar formalin-evoked flinching, SP release from primary afferents and c-Fos expression in spinal dorsal horn. Intraplantar injection of formalin (2.5%, 50 µL) produced an intense, characteristic biphasic paw flinching response. In rats with IT catheters, IT SNX-482 (0.5 µg) reduced formalin-evoked paw flinching in both phase 1 and 2 compared with vehicle. Intraplantar formalin caused robust neurokinin 1 receptor (NK1r) internalization (indicating SP release) and c-Fos expression in the ipsilateral dorsal horn, which were blocked by IT SNX-482. IT ω-agatoxin IVA (0.03, 0.125 and 0.5 µg) did not reduce formalin-evoked paw flinching or c-Fos expression at any doses, with higher doses resulting in motor dysfunction. Thus, we demonstrated that blockade of spinal R-type, but not P/Q type VSCCs attenuated formalin-induced pain behavior, NK1r internalization and c-Fos expression in the superficial dorsal horn. This study supports a role for Cav2.3 in presynaptic neurotransmitter release from peptidergic nociceptive afferents and pain behaviors. PMID:23810829

In Situ Optical Mapping of Voltage and Calcium in the Heart

PubMed Central

Ewart, Paul; Ashley, Euan A.; Loew, Leslie M.; Kohl, Peter; Bollensdorff, Christian; Woods, Christopher E.

2012-01-01

Electroanatomic mapping the interrelation of intracardiac electrical activation with anatomic locations has become an important tool for clinical assessment of complex arrhythmias. Optical mapping of cardiac electrophysiology combines high spatiotemporal resolution of anatomy and physiological function with fast and simultaneous data acquisition. If applied to the clinical setting, this could improve both diagnostic potential and therapeutic efficacy of clinical arrhythmia interventions. The aim of this study was to explore this utility in vivo using a rat model. To this aim, we present a single-camera imaging and multiple light-emitting-diode illumination system that reduces economic and technical implementation hurdles to cardiac optical mapping. Combined with a red-shifted calcium dye and a new near-infrared voltage-sensitive dye, both suitable for use in blood-perfused tissue, we demonstrate the feasibility of in vivo multi-parametric imaging of the mammalian heart. Our approach combines recording of electrophysiologically-relevant parameters with observation of structural substrates and is adaptable, in principle, to trans-catheter percutaneous approaches. PMID:22876327

Calcium sensitive ring-like oligomers formed by synaptotagmin

PubMed Central

Wang, Jing; Bello, Oscar; Auclair, Sarah M.; Wang, Jing; Coleman, Jeff; Pincet, Frederic; Krishnakumar, Shyam S.; Sindelar, Charles V.; Rothman, James E.

2014-01-01

The synaptic vesicle protein synaptotagmin-1 (SYT) is required to couple calcium influx to the membrane fusion machinery. However, the structural mechanism underlying this process is unclear. Here we report an unexpected circular arrangement (ring) of SYT’s cytosolic domain (C2AB) formed on lipid monolayers in the absence of free calcium ions as revealed by electron microscopy. Rings vary in diameter from 18–43 nm, corresponding to 11–26 molecules of SYT. Continuous stacking of the SYT rings occasionally converts both lipid monolayers and bilayers into protein-coated tubes. Helical reconstruction of the SYT tubes shows that one of the C2 domains (most likely C2B, based on its biochemical properties) interacts with the membrane and is involved in ring formation, and the other C2 domain points radially outward. SYT rings are disrupted rapidly by physiological concentrations of free calcium but not by magnesium. Assuming that calcium-free SYT rings are physiologically relevant, these results suggest a simple and novel mechanism by which SYT regulates neurotransmitter release: The ring acts as a spacer to prevent the completion of the soluble N-ethylmaleimide–sensitive factor activating protein receptor (SNARE) complex assembly, thereby clamping fusion in the absence of calcium. When the ring disassembles in the presence of calcium, fusion proceeds unimpeded. PMID:25201968

P/Q-type and T-type voltage-gated calcium channels are involved in the contraction of mammary and brain blood vessels from hypertensive patients.

PubMed

Thuesen, A D; Lyngsø, K S; Rasmussen, L; Stubbe, J; Skøtt, O; Poulsen, F R; Pedersen, C B; Rasmussen, L M; Hansen, P B L

2017-03-01

Calcium channel blockers are widely used in cardiovascular diseases. Besides L-type channels, T- and P/Q-type calcium channels are involved in the contraction of human renal blood vessels. It was hypothesized that T- and P/Q-type channels are involved in the contraction of human brain and mammary blood vessels. Internal mammary arteries from bypass surgery patients and cerebral arterioles from patients with brain tumours with and without hypertension were tested in a myograph and perfusion set-up. PCR and immunohistochemistry were performed on isolated blood vessels. The P/Q-type antagonist ω-agatoxin IVA (10 -8  mol L -1 ) and the T-type calcium blocker mibefradil (10 -7  mol L -1 ) inhibited KCl depolarization-induced contraction in mammary arteries from hypertensive patients with no effect on blood vessels from normotensive patients. ω-Agatoxin IVA decreased contraction in cerebral arterioles from hypertensive patients. L-type blocker nifedipine abolished the contraction in mammary arteries. PCR analysis showed expression of P/Q-type (Ca v 2.1), T-type (Ca v 3.1 and Ca v 3.2) and L-type (Ca v 1.2) calcium channels in mammary and cerebral arteries. Immunohistochemical labelling of mammary and cerebral arteries revealed the presence of Ca v 2.1 in endothelial and smooth muscle cells. Ca v 3.1 was also detected in mammary arteries. P/Q- and T-type Ca v are present in human internal mammary arteries and in cerebral penetrating arterioles. P/Q- and T-type calcium channels are involved in the contraction of mammary arteries from hypertensive patients but not from normotensive patients. Furthermore, in cerebral arterioles P/Q-type channels importance was restricted to hypertensive patients might lead to that T- and P/Q-type channels could be a new target in hypertensive patients. © 2016 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Long-term effects of L- and N-type calcium channel blocker on uric acid levels and left atrial volume in hypertensive patients.

PubMed

Masaki, Mitsuru; Mano, Toshiaki; Eguchi, Akiyo; Fujiwara, Shohei; Sugahara, Masataka; Hirotani, Shinichi; Tsujino, Takeshi; Komamura, Kazuo; Koshiba, Masahiro; Masuyama, Tohru

2016-11-01

Left ventricular (LV) diastolic dysfunction is associated with hypertension and hyperuricemia. However, it is not clear whether the L- and N-type calcium channel blocker will improve LV diastolic dysfunction through the reduction of uric acid. The aim of this study was to investigate the effects of anti-hypertensive therapy, the L- and N-type calcium channel blocker, cilnidipine or the L-type calcium channel blocker, amlodipine, on left atrial reverse remodeling and uric acid in hypertensive patients. We studied 62 patients with untreated hypertension, randomly assigned to cilnidipine or amlodipine for 48 weeks. LV diastolic function was assessed with the left atrial volume index (LAVI), mitral early diastolic wave (E), tissue Doppler early diastolic velocity (E') and the ratio (E/E'). Serum uric acid levels were measured before and after treatment. After treatment, systolic and diastolic blood pressures equally dropped in both groups. LAVI, E/E', heart rate and uric acid levels decreased at 48 weeks in the cilnidipine group but not in the amlodipine group. The % change from baseline to 48 weeks in LAVI, E wave, E/E' and uric acid levels were significantly lower in the cilnidipine group than in the amlodipine group. Larger %-drop in uric acid levels were associated with larger %-reduction of LAVI (p < 0.01). L- and N-type calcium channel blocker but not L-type calcium channel blocker may improve LV diastolic function in hypertensive patients, at least partially through the decrease in uric acid levels.

L-type calcium channels refine the neural population code of sound level.

PubMed

Grimsley, Calum Alex; Green, David Brian; Sivaramakrishnan, Shobhana

2016-12-01

The coding of sound level by ensembles of neurons improves the accuracy with which listeners identify how loud a sound is. In the auditory system, the rate at which neurons fire in response to changes in sound level is shaped by local networks. Voltage-gated conductances alter local output by regulating neuronal firing, but their role in modulating responses to sound level is unclear. We tested the effects of L-type calcium channels (Ca L : Ca V 1.1-1.4) on sound-level coding in the central nucleus of the inferior colliculus (ICC) in the auditory midbrain. We characterized the contribution of Ca L to the total calcium current in brain slices and then examined its effects on rate-level functions (RLFs) in vivo using single-unit recordings in awake mice. Ca L is a high-threshold current and comprises ∼50% of the total calcium current in ICC neurons. In vivo, Ca L activates at sound levels that evoke high firing rates. In RLFs that increase monotonically with sound level, Ca L boosts spike rates at high sound levels and increases the maximum firing rate achieved. In different populations of RLFs that change nonmonotonically with sound level, Ca L either suppresses or enhances firing at sound levels that evoke maximum firing. Ca L multiplies the gain of monotonic RLFs with dynamic range and divides the gain of nonmonotonic RLFs with the width of the RLF. These results suggest that a single broad class of calcium channels activates enhancing and suppressing local circuits to regulate the sensitivity of neuronal populations to sound level. Copyright © 2016 the American Physiological Society.

L-type calcium channels refine the neural population code of sound level

PubMed Central

Grimsley, Calum Alex; Green, David Brian

2016-01-01

The coding of sound level by ensembles of neurons improves the accuracy with which listeners identify how loud a sound is. In the auditory system, the rate at which neurons fire in response to changes in sound level is shaped by local networks. Voltage-gated conductances alter local output by regulating neuronal firing, but their role in modulating responses to sound level is unclear. We tested the effects of L-type calcium channels (CaL: CaV1.1–1.4) on sound-level coding in the central nucleus of the inferior colliculus (ICC) in the auditory midbrain. We characterized the contribution of CaL to the total calcium current in brain slices and then examined its effects on rate-level functions (RLFs) in vivo using single-unit recordings in awake mice. CaL is a high-threshold current and comprises ∼50% of the total calcium current in ICC neurons. In vivo, CaL activates at sound levels that evoke high firing rates. In RLFs that increase monotonically with sound level, CaL boosts spike rates at high sound levels and increases the maximum firing rate achieved. In different populations of RLFs that change nonmonotonically with sound level, CaL either suppresses or enhances firing at sound levels that evoke maximum firing. CaL multiplies the gain of monotonic RLFs with dynamic range and divides the gain of nonmonotonic RLFs with the width of the RLF. These results suggest that a single broad class of calcium channels activates enhancing and suppressing local circuits to regulate the sensitivity of neuronal populations to sound level. PMID:27605536

Calcium influx through L-type channels attenuates skeletal muscle contraction via inhibition of adenylyl cyclases.

PubMed

Menezes-Rodrigues, Francisco Sandro; Pires-Oliveira, Marcelo; Duarte, Thiago; Paredes-Gamero, Edgar Julian; Chiavegatti, Tiago; Godinho, Rosely Oliveira

2013-11-15

Skeletal muscle contraction is triggered by acetylcholine induced release of Ca(2+) from sarcoplasmic reticulum. Although this signaling pathway is independent of extracellular Ca(2+), L-type voltage-gated calcium channel (Cav) blockers have inotropic effects on frog skeletal muscles which occur by an unknown mechanism. Taking into account that skeletal muscle fiber expresses Ca(+2)-sensitive adenylyl cyclase (AC) isoforms and that cAMP is able to increase skeletal muscle contraction force, we investigated the role of Ca(2+) influx on mouse skeletal muscle contraction and the putative crosstalk between extracellular Ca(2+) and intracellular cAMP signaling pathways. The effects of Cav blockers (verapamil and nifedipine) and extracellular Ca(2+) chelator EGTA were evaluated on isometric contractility of mouse diaphragm muscle under direct electrical stimulus (supramaximal voltage, 2 ms, 0.1 Hz). Production of cAMP was evaluated by radiometric assay while Ca(2+) transients were assessed by confocal microscopy using L6 cells loaded with fluo-4/AM. Ca(2+) channel blockers verapamil and nifedipine had positive inotropic effect, which was mimicked by removal of extracellular Ca(+2) with EGTA or Ca(2+)-free Tyrode. While phosphodiesterase inhibitor IBMX potentiates verapamil positive inotropic effect, it was abolished by AC inhibitors SQ22536 and NYK80. Finally, the inotropic effect of verapamil was associated with increased intracellular cAMP content and mobilization of intracellular Ca(2+), indicating that positive inotropic effects of Ca(2+) blockers depend on cAMP formation. Together, our results show that extracellular Ca(2+) modulates skeletal muscle contraction, through inhibition of Ca(2+)-sensitive AC. The cross-talk between extracellular calcium and cAMP-dependent signaling pathways appears to regulate the extent of skeletal muscle contraction responses. © 2013 Published by Elsevier B.V.

PYRETHROID INDUCED ALTERATIONS IN TRANSCRIPTION OF CALCIUM RESPONSIVE AND IMMEDIATE EARLY GENES IN VIVO.

EPA Science Inventory

Multiple molecular targets for pyrethroid insecticides have been evaluated in in vitro preparations, including but not limited to voltage-sensitive sodium channels (VSSCs), voltage-sensitive calcium channels (VSCCs), GABAergic receptors, ATPases and mitochondrial respiratory chai...

CaV3.1 isoform of T-type calcium channels supports excitability of rat and mouse ventral tegmental area neurons.

PubMed

Tracy, Matthew E; Tesic, Vesna; Stamenic, Tamara Timic; Joksimovic, Srdjan M; Busquet, Nicolas; Jevtovic-Todorovic, Vesna; Todorovic, Slobodan M

2018-03-23

Recent data have implicated voltage-gated calcium channels in the regulation of the excitability of neurons within the mesolimbic reward system. While the attention of most research has centered on high voltage L-type calcium channel activity, the presence and role of the low voltage-gated T-type calcium channel (T-channels) has not been well explored. Hence, we investigated T-channel properties in the neurons of the ventral tegmental area (VTA) utilizing wild-type (WT) rats and mice, Ca V 3.1 knock-out (KO) mice, and TH-eGFP knock-in (KI) rats in acute horizontal brain slices of adolescent animals. In voltage-clamp experiments, we first assessed T-channel activity in WT rats with characteristic properties of voltage-dependent activation and inactivation, as well as characteristic crisscrossing patterns of macroscopic current kinetics. T-current kinetics were similar in WT mice and WT rats but T-currents were abolished in Ca V 3.1 KO mice. In ensuing current-clamp experiments, we observed the presence of hyperpolarization-induced rebound burst firing in a subset of neurons in WT rats, as well as dopaminergic and non-dopaminergic neurons in TH-eGFP KI rats. Following the application of a pan-selective T-channel blocker TTA-P2, rebound bursting was significantly inhibited in all tested cells. In a behavioral assessment, the acute locomotor increase induced by a MK-801 (Dizocilpine) injection in WT mice was abolished in Ca V 3.1 KO mice, suggesting a tangible role for 3.1 T-type channels in drug response. We conclude that pharmacological targeting of Ca V 3.1 isoform of T-channels may be a novel approach for the treatment of disorders of mesolimbic reward system. Copyright © 2018. Published by Elsevier Ltd.

A new mode of regulation of N-type inactivation in a Caenorhabditis elegans voltage-gated potassium channel.

PubMed

Cai, Shi-Qing; Sesti, Federico

2007-06-22

N-type inactivation in voltage-gated K+ (Kv) channels is a widespread means to modulate neuronal excitability and signaling. Here we have shown a novel mechanism of N-type inactivation in a Caenorhabditis elegans Kv channel. The N-terminal sequence of KVS-1 contains a domain of 22 amino acids that resembles the inactivation ball in A-type channels, which is preceded by a domain of eighteen amino acids. Wild type KVS-1 currents can be described as A-type; however, their kinetics are significantly (approximately 5-fold) slower. When the putative inactivation ball is deleted, the current becomes non-inactivating. Inactivation is restored in non-inactivating channels by diffusion of the missing inactivation domain in the cytoplasm. Deletion of the domain in front of the ball speeds inactivation kinetics approximately 5-fold. We conclude that KVS-1 is the first example of a novel type of Kv channel simultaneously possessing an N-inactivating ball preceded by an N inactivation regulatory domain (NIRD) that acts to slow down inactivation through steric mechanisms.

Synthetic peptides corresponding to human follicle-stimulating hormone (hFSH)-beta-(1-15) and hFSH-beta-(51-65) induce uptake of 45Ca++ by liposomes: evidence for calcium-conducting transmembrane channel formation

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

Grasso, P.; Santa-Coloma, T.A.; Reichert, L.E. Jr.

1991-06-01

We have previously described FSH receptor-mediated influx of 45Ca++ in cultured Sertoli cells from immature rats and receptor-enriched proteoliposomes via activation of voltage-sensitive and voltage-independent calcium channels. We have further shown that this effect of FSH does not require cholera toxin- or pertussis toxin-sensitive guanine nucleotide binding protein or activation of adenylate cyclase. In the present study, we have identified regions of human FSH-beta-subunit which appear to be involved in mediating calcium influx. We screened 11 overlapping peptide amides representing the entire primary structure of hFSH-beta-subunit for their effects on 45Ca++ flux in FSH receptor-enriched proteoliposomes. hFSH-beta-(1-15) and hFSH-beta-(51-65) inducedmore » uptake of 45Ca++ in a concentration-related manner. This effect of hFSH-beta-(1-15) and hFSH-beta-(51-65) was also observed in liposomes lacking incorporated FSH receptor. Reducing membrane fluidity by incubating liposomes (containing no receptor) with hFSH-beta-(1-15) or hFSH-beta-(51-65) at temperatures lower than the transition temperatures of their constituent phospholipids resulted in no significant (P greater than 0.05) difference in 45Ca++ uptake. The effectiveness of the calcium ionophore A23187, however, was abolished. Ruthenium red, a voltage-independent calcium channel antagonist, was able to completely block uptake of 45Ca++ induced by hFSH-beta-(1-15) and hFSH-beta-(51-65) whereas nifedipine, a calcium channel blocker specific for L-type voltage-sensitive calcium channels, was without effect. These results suggest that in addition to its effect on voltage-sensitive calcium channel activity, interaction of FSH with its receptor may induce formation of transmembrane aqueous channels which also facilitate influx of extracellular calcium.« less

Calcium triggers reversal of calmodulin on nested anti-parallel sites in the IQ motif of the neuronal voltage-dependent sodium channel NaV1.2.

PubMed

Hovey, Liam; Fowler, C Andrew; Mahling, Ryan; Lin, Zesen; Miller, Mark Stephen; Marx, Dagan C; Yoder, Jesse B; Kim, Elaine H; Tefft, Kristin M; Waite, Brett C; Feldkamp, Michael D; Yu, Liping; Shea, Madeline A

2017-05-01

Several members of the voltage-gated sodium channel family are regulated by calmodulin (CaM) and ionic calcium. The neuronal voltage-gated sodium channel Na V 1.2 contains binding sites for both apo (calcium-depleted) and calcium-saturated CaM. We have determined equilibrium dissociation constants for rat Na V 1.2 IQ motif [IQRAYRRYLLK] binding to apo CaM (~3nM) and (Ca 2+ ) 4 -CaM (~85nM), showing that apo CaM binding is favored by 30-fold. For both apo and (Ca 2+ ) 4 -CaM, NMR demonstrated that Na V 1.2 IQ motif peptide (Na V 1.2 IQp ) exclusively made contacts with C-domain residues of CaM (CaM C ). To understand how calcium triggers conformational change at the CaM-IQ interface, we determined a solution structure (2M5E.pdb) of (Ca 2+ ) 2 -CaM C bound to Na V 1.2 IQp . The polarity of (Ca 2+ ) 2 -CaM C relative to the IQ motif was opposite to that seen in apo CaM C -Na v 1.2 IQp (2KXW), revealing that CaM C recognizes nested, anti-parallel sites in Na v 1.2 IQp . Reversal of CaM may require transient release from the IQ motif during calcium binding, and facilitate a re-orientation of CaM N allowing interactions with non-IQ Na V 1.2 residues or auxiliary regulatory proteins interacting in the vicinity of the IQ motif. Copyright © 2017 Elsevier B.V. All rights reserved.

The alpha2-delta protein: an auxiliary subunit of voltage-dependent calcium channels as a recognized drug target.

PubMed

Thorpe, Andrew J; Offord, James

2010-07-01

Currently, there are two drugs on the market, gabapentin (Neurontin) and pregabalin (Lyrica), that are proposed to exert their therapeutic effect through binding to the alpha2-delta subunit of voltage-sensitive calcium channels. This activity was unexpected, as the alpha2-delta subunit had previously been considered not to be a pharmacological target. In this review, the role of the alpha2-delta subunits is discussed and the mechanism of action of the alpha2-delta ligands in vitro and in vivo is summarized. Finally, new insights into the mechanism of drugs that bind to this protein are discussed.

Calcium influx is required for endocytotic membrane retrieval

PubMed Central

Vogel, Steven S.; Smith, Robert M.; Baibakov, Boris; Ikebuchi, Yoshihide; Lambert, Nevin A.

1999-01-01

Cells use endocytotic membrane retrieval to compensate for excess surface membrane after exocytosis. Retrieval is thought to be calcium-dependent, but the source of this calcium is not known. We found that, in sea urchin eggs, endocytotic membrane retrieval required extracellular calcium. Inhibitors of P-type calcium channels—cadmium, ω-conotoxin MVIIC, ω-agatoxin IVA, and ω-agatoxin TK—blocked membrane retrieval; selective inhibitors of N-type and L-type channels did not. Treatment with calcium ionophores overcame agatoxin inhibition in a calcium-dependent manner. Cadmium blocked membrane retrieval when applied during the first 5 minutes after fertilization, the period when the membrane potential is depolarized. We conclude that calcium influx through ω-agatoxin-sensitive channels plays a key role in signaling for endocytotic membrane retrieval. PMID:10220411

Regulation of CaV2 calcium channels by G protein coupled receptors

PubMed Central

Zamponi, Gerald W.; Currie, Kevin P.M.

2012-01-01

Voltage gated calcium channels (Ca2+ channels) are key mediators of depolarization induced calcium influx into excitable cells, and thereby play pivotal roles in a wide array of physiological responses. This review focuses on the inhibition of CaV2 (N- and P/Q-type) Ca2+-channels by G protein coupled receptors (GPCRs), which exerts important autocrine/paracrine control over synaptic transmission and neuroendocrine secretion. Voltage-dependent inhibition is the most widespread mechanism, and involves direct binding of the G protein βγ dimer (Gβγ) to the α1 subunit of CaV2 channels. GPCRs can also recruit several other distinct mechanisms including phosphorylation, lipid signaling pathways, and channel trafficking that result in voltage-independent inhibition. Current knowledge of Gβγ-mediated inhibition is reviewed, including the molecular interactions involved, determinants of voltage-dependence, and crosstalk with other cell signaling pathways. A summary of recent developments in understanding the voltage-independent mechanisms prominent in sympathetic and sensory neurons is also included. PMID:23063655

Calcium sensitivity of residual force enhancement in rabbit skinned fibers.

PubMed

Joumaa, V; Herzog, W

2014-08-15

Isometric force after active stretch of muscles is higher than the purely isometric force at the corresponding length. This property is termed residual force enhancement. Active force in skeletal muscle depends on calcium attachment characteristics to the regulatory proteins. Passive force has been shown to influence calcium attachment characteristics, specifically the sarcomere length dependence of calcium sensitivity. Since one of the mechanisms proposed to explain residual force enhancement is the increase in passive force that results from engagement of titin upon activation and stretch, our aim was to test if calcium sensitivity of residual force enhancement was different from that of its corresponding purely isometric contraction and if such a difference was related to the molecular spring titin. Force-pCa curves were established in rabbit psoas skinned fibers for reference and residual force-enhanced states at a sarcomere length of 3.0 μm 1) in a titin-intact condition, 2) after treatment with trypsin to partially eliminate titin, and 3) after treatment with trypsin and osmotic compression with dextran T-500 to decrease the lattice spacing in the absence of titin. The force-pCa curves of residual force enhancement were shifted to the left compared with their corresponding controls in titin-intact fibers, indicating increased calcium sensitivity. No difference in calcium sensitivity was observed between reference and residual force-enhanced contractions in trypsin-treated and osmotically compressed trypsin-treated fibers. Furthermore, calcium sensitivity after osmotic compression was lower than that observed for residual force enhancement in titin-intact skinned fibers. These results suggest that titin-based passive force regulates the increase in calcium sensitivity of residual force enhancement by a mechanism other than reduction of the myofilament lattice spacing. Copyright © 2014 the American Physiological Society.

Action of insecticidal N-alkylamides at site 2 of the voltage-sensitive sodium channel

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

Ottea, J.A.; Payne, G.T.; Soderlund, D.M.

1990-08-01

Nine synthetic N-alkylamides were examined as inhibitors of the specific binding of ({sup 3}H)batrachotoxinin A 20{alpha}-benzoate (({sup 3}H)BTX-B) to sodium channels and as activators of sodium uptake in mouse brain synaptoneurosomes. In the presence of scorpion (Leiurus quinquestriatus) venom, the six insecticidal analogues were active as both inhibitors of ({sup 3}H)BTX-B binding and stimulators of sodium uptake. These findings are consistent with an action of these compounds at the alkaloid activator recognition site (site 2) of the voltage-sensitive sodium channel. The three noninsecticidal N-alkylamides also inhibited ({sup 3}H)BTX-B binding but were ineffective as activators of sodium uptake. Concentration-response studies revealedmore » that some of the insecticidal amides also enhanced sodium uptake through a second, high-affinity interaction that does not involve site 2, but this secondary effect does not appear to be correlated with insecticidal activity. The activities of N-alkylamides as sodium channel activators were influenced by the length of the alkenyl chain and the location of unsaturation within the molecule. These results further define the actions of N-alkylamides on sodium channels and illustrate the significance of the multiple binding domains of the sodium channel as target sites for insect control agents.« less

Extracellular protons enable activation of the calcium-dependent chloride channel TMEM16A.

PubMed

Cruz-Rangel, Silvia; De Jesús-Pérez, José J; Aréchiga-Figueroa, Iván A; Rodríguez-Menchaca, Aldo A; Pérez-Cornejo, Patricia; Hartzell, H Criss; Arreola, Jorge

2017-03-01

that protons regulate TMEM16A by tuning its open probability without modifying the single channel current. We found a robust reduction of the proton effect at high [Ca 2+ ] i . To identify protonation targets we mutated all extracellular glutamate and histidine residues and 4 of 11 aspartates. Most mutants were sensitive to protons. However, mutation that substituted glutamic acid (E) for glutamine (Q) at amino acid position 623 (E623Q) displayed a titration curve shifted to the left relative to wild type channels and the I Cl was nearly insensitive to proton concentrations between 10 -5.5 and 10 -9.0  m. Additionally, I Cl of the mutant containing an aspartic acid (D) to asparagine (N) substitution at position 405 (D405N) mutant was partially inhibited by a proton concentration of 10 -5.5  m, but 10 -9.0  m produced the same effect as in wild type. Based on our findings we propose that external protons titrate glutamic acid 623, which enables voltage activation of TMEM16A at non-saturating [Ca 2+ ] i . © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

Modulation of A-type potassium channels by a family of calcium sensors.

PubMed

An, W F; Bowlby, M R; Betty, M; Cao, J; Ling, H P; Mendoza, G; Hinson, J W; Mattsson, K I; Strassle, B W; Trimmer, J S; Rhodes, K J

2000-02-03

In the brain and heart, rapidly inactivating (A-type) voltage-gated potassium (Kv) currents operate at subthreshold membrane potentials to control the excitability of neurons and cardiac myocytes. Although pore-forming alpha-subunits of the Kv4, or Shal-related, channel family form A-type currents in heterologous cells, these differ significantly from native A-type currents. Here we describe three Kv channel-interacting proteins (KChIPs) that bind to the cytoplasmic amino termini of Kv4 alpha-subunits. We find that expression of KChIP and Kv4 together reconstitutes several features of native A-type currents by modulating the density, inactivation kinetics and rate of recovery from inactivation of Kv4 channels in heterologous cells. All three KChIPs co-localize and co-immunoprecipitate with brain Kv4 alpha-subunits, and are thus integral components of native Kv4 channel complexes. The KChIPs have four EF-hand-like domains and bind calcium ions. As the activity and density of neuronal A-type currents tightly control responses to excitatory synaptic inputs, these KChIPs may regulate A-type currents, and hence neuronal excitability, in response to changes in intracellular calcium.

Discrimination of clinically significant calcium salts using MARS spectral CT

NASA Astrophysics Data System (ADS)

Kirkbride, T. E.; Raja, A.; Mueller, K.; Bateman, C. J.; Becce, F.; Anderson, N.

2017-03-01

Calcium compounds within tissues are usually a sign of pathology, and calcium crystal type is often a pointer to the diagnosis. There are clinical advantages in being able to determine the quantity and type of calcifications non-invasively in cardiovascular, genitourinary and musculoskeletal disorders, and treatment differs depending on the crystal type and quantity. The problem arises when trying to distinguish between different calcium compounds within the same image due to their similar attenuation properties. There are spectroscopic differences between calcium salts at very low energies. As calcium oxalate and calcium hydroxyapatite can co-exist in breast and musculoskeletal pathologies of the breast, we wished to determine whether Spectral CT could distinguish between them in the same image at clinical X-ray energy ranges. Energy thresholds of 15, 22, 29 and 36keV and tube voltages of 50, 80 and 110kVp were chosen, and images were analysed to determine the percentage difference in the attenuation coefficients of calcium hydroxyapatite samples at concentrations of 54.3, 211.7, 808.5 and 1169.3mg/ml, and calcium oxalate at a concentration of 2000 mg/ml. The two lower concentrations of calcium hydroxyapatite were distinguishable from calcium oxalate at all energies and all tube voltages, whereas the ability to discriminate oxalate from hydroxyapatite at higher concentrations was dependent on the threshold energy but only mildly dependent on the tube voltage used. Spectral CT shows promise for distinguishing clinically important calcium salts.

Kinetic and pharmacological properties distinguishing three types of calcium currents in chick sensory neurones.

PubMed Central

Fox, A P; Nowycky, M C; Tsien, R W

1987-01-01

1. Calcium currents in cultured dorsal root ganglion (d.r.g.) cells were studied with the whole-cell patch-clamp technique. Using experimental conditions that suppressed Na+ and K+ currents, and 3-10 mM-external Ca2+ or Ba2+, we distinguished three distinct types of calcium currents (L, T and N) on the basis of voltage-dependent kinetics and pharmacology. 2. Component L activates at relatively positive test potentials (t.p. greater than -10 mV) and shows little inactivation during a 200 ms depolarization. It is completely reprimed at a holding potential (h.p.) of -60 mV, and can be isolated by using a more depolarized h.p. (-40 mV) to inactivate the other two types of calcium currents. 3. Component T can be seen in isolation with weak test pulses. It begins activating at potentials more positive than -70 mV and inactivates quickly and completely during a maintained depolarization (time constant, tau approximately 20-50 ms). The current amplitude and the rate of decay increase with stronger depolarizations until both reach a maximum at approximately -40 mV. Inactivation is complete at h.p. greater than -60 mV and is progressively removed between -60 and -95 mV. 4. Component N activates at relatively strong depolarizations (t.p. greater than -20 mV) and decays with time constants ranging from 50 to 110 ms. Inactivation is removed over a very broad range of holding potentials (h.p. between -40 and -110 mV). 5. With 10 mM-EGTA in the pipette solution, substitution of Ba2+ for Ca2+ as the charge carrier does not alter the rates of activation or relaxation of any component. However, T-type channels are approximately equally permeable to Ca2+ and Ba2+, while L-type and N-type channels are both much more permeable to Ba2+. 6. Component N cannot be explained by current-dependent inactivation of L current resulting from recruitment of extra L-type channels at negative holding potentials: raising the external Ba2+ concentration to 110 mM greatly increases the amplitude of L

Progress in the structural understanding of voltage-gated calcium channel (CaV) function and modulation.

PubMed

Minor, Daniel L; Findeisen, Felix

2010-01-01

Voltage-gated calcium channels (CaVs) are large, transmembrane multiprotein complexes that couple membrane depolarization to cellular calcium entry. These channels are central to cardiac action potential propagation, neurotransmitter and hormone release, muscle contraction, and calcium-dependent gene transcription. Over the past six years, the advent of high-resolution structural studies of CaV components from different isoforms and CaV modulators has begun to reveal the architecture that underlies the exceptionally rich feedback modulation that controls CaV action. These descriptions of CaV molecular anatomy have provided new, structure-based insights into the mechanisms by which particular channel elements affect voltage-dependent inactivation (VDI), calcium‑dependent inactivation (CDI), and calcium‑dependent facilitation (CDF). The initial successes have been achieved through structural studies of soluble channel domains and modulator proteins and have proven most powerful when paired with biochemical and functional studies that validate ideas inspired by the structures. Here, we review the progress in this growing area and highlight some key open challenges for future efforts.

Comparative performance of a genetically-encoded voltage indicator and a blue voltage sensitive dye for large scale cortical voltage imaging

PubMed Central

Mutoh, Hiroki; Mishina, Yukiko; Gallero-Salas, Yasir; Knöpfel, Thomas

2015-01-01

Traditional small molecule voltage sensitive dye indicators have been a powerful tool for monitoring large scale dynamics of neuronal activities but have several limitations including the lack of cell class specific targeting, invasiveness and difficulties in conducting longitudinal studies. Recent advances in the development of genetically-encoded voltage indicators have successfully overcome these limitations. Genetically-encoded voltage indicators (GEVIs) provide sufficient sensitivity to map cortical representations of sensory information and spontaneous network activities across cortical areas and different brain states. In this study, we directly compared the performance of a prototypic GEVI, VSFP2.3, with that of a widely used small molecule voltage sensitive dye (VSD), RH1691, in terms of their ability to resolve mesoscopic scale cortical population responses. We used three synchronized CCD cameras to simultaneously record the dual emission ratiometric fluorescence signal from VSFP2.3 and RH1691 fluorescence. The results show that VSFP2.3 offers more stable and less invasive recording conditions, while the signal-to-noise level and the response dynamics to sensory inputs are comparable to RH1691 recordings. PMID:25964738

Acute exposure to progesterone attenuates cardiac contraction by modifying myofilament calcium sensitivity in the female mouse heart

PubMed Central

Feridooni, Hirad A.; MacDonald, Jennifer K.; Ghimire, Anjali; Pyle, W. Glen

2017-01-01

Acute application of progesterone attenuates cardiac contraction, although the underlying mechanisms are unclear. We investigated whether progesterone modified contraction in isolated ventricular myocytes and identified the Ca2+ handling mechanisms involved in female C57BL/6 mice (6–9 mo; sodium pentobarbital anesthesia). Cells were field-stimulated (4 Hz; 37°C) and exposed to progesterone (0.001–10.0 μM) or vehicle (35 min). Ca2+ transients (fura-2) and cell shortening were recorded simultaneously. Maximal concentrations of progesterone inhibited peak contraction by 71.4% (IC50 = 160 ± 50 nM; n = 12) and slowed relaxation by 75.4%. By contrast, progesterone had no effect on amplitudes or time courses of underlying Ca2+ transients. Progesterone (1 µM) also abbreviated action potential duration. When the duration of depolarization was controlled by voltage-clamp, progesterone attenuated contraction and slowed relaxation but did not affect Ca2+ currents, Ca2+ transients, sarcoplasmic reticulum (SR) content, or fractional release of SR Ca2+. Actomyosin MgATPase activity was assayed in myofilaments from hearts perfused with progesterone (1 μM) or vehicle (35 min). While maximal responses to Ca2+ were not affected by progesterone, myofilament Ca2+ sensitivity was reduced (EC50 = 0.94 ± 0.01 µM for control, n = 7 vs. 1.13 ± 0.05 μM for progesterone, n = 6; P < 0.05) and progesterone increased phosphorylation of myosin binding protein C. The effects on contraction were inhibited by lonaprisan (progesterone receptor antagonist) and levosimendan (Ca2+ sensitizer). Unlike results in females, progesterone had no effect on contraction or myofilament Ca2+ sensitivity in age-matched male mice. These data indicate that progesterone reduces myofilament Ca2+ sensitivity in female hearts, which may exacerbate manifestations of cardiovascular disease late in pregnancy when progesterone levels are high. NEW & NOTEWORTHY We investigated myocardial effects of acute

The effects of crustacean cardioactive peptide on locust oviducts are calcium-dependent.

PubMed

Donini, Andrew; Lange, Angela B

2002-04-01

The role of calcium as a second messenger in the crustacean cardioactive peptide (CCAP)-induced contractions of the locust oviducts was investigated. Incubation of the oviducts in a calcium-free saline containing, a preferential calcium cation chelator, or an extracellular calcium channel blocker, abolished CCAP-induced contractions, indicating that the effects of CCAP on the oviducts are calcium-dependent. In contrast, sodium free saline did not affect CCAP-induced contractions. Co-application of CCAP to the oviducts with preferential L-type voltage-dependent calcium channel blockers reduced CCAP-induced contractions by 32-54%. Two preferential T-type voltage-dependent calcium channel blockers both inhibited CCAP-induced oviduct contractions although affecting different components of the contractions. Amiloride decreased the tonic component of CCAP-induced contractions by 40-55% and flunarizine dihydrochloride decreased the frequency of CCAP-induced phasic contractions by as much as 65%, without affecting tonus. Flunarizine dihydrochloride did not alter the proctolin-induced contractions of the oviducts. Results suggest that the actions of CCAP are partially mediated by voltage-dependent calcium channels similar to vertebrate L-type and T-type channels. High-potassium saline does not abolish CCAP-induced contractions indicating the presence of receptor-operated calcium channels that mediate the actions of CCAP on the oviducts. The involvement of calcium from intracellular stores in CCAP-induced contractions of the oviducts is likely since, an intracellular calcium antagonist decreased CCAP-induced contractions by 30-35%.

Metabotropic and ionotropic glutamate receptors regulate calcium channel currents in salamander retinal ganglion cells

PubMed Central

Shen, Wen; Slaughter, Malcolm M

1998-01-01

Glutamate suppressed high-voltage-activated barium currents (IBa,HVA) in tiger salamander retinal ganglion cells. Both ionotropic (iGluR) and metabotropic (mGluR) receptors contributed to this calcium channel inhibition. Trans-ACPD (1-aminocyclopentane-trans-1S,3R-dicarboxylic acid), a broad-spectrum metabotropic glutamate receptor agonist, suppressed a dihydropyridine-sensitive barium current. Kainate, an ionotropic glutamate receptor agonist, reduced an ω-conotoxin GVIA-sensitive current. The relative effectiveness of selective agonists indicated that the predominant metabotropic receptor was the L-2-amino-4-phosphonobutyrate (l-AP4)-sensitive, group III receptor. This receptor reversed the action of forskolin, but this was not responsible for calcium channel suppression. l-AP4 raised internal calcium concentration. Antagonists of phospholipase C, inositol trisphosphate (IP3) receptors and ryanodine receptors inhibited the action of metabotropic agonists, indicating that group III receptor transduction was linked to this pathway. The action of kainate was partially suppressed by BAPTA, by calmodulin antagonists and by blockers of calmodulin-dependent phosphatase. Suppression by kainate of the calcium channel current was more rapid when calcium was the charge carrier, instead of barium. The results indicate that calcium influx through kainate-sensitive glutamate receptors can activate calmodulin, which stimulates phosphatases that may directly suppress voltage-sensitive calcium channels. Thus, ionotropic and metabotropic glutamate receptors inhibit distinct calcium channels. They could act synergistically, since both increase internal calcium. These pathways provide negative feedback that can reduce calcium influx when ganglion cells are depolarized. PMID:9660896

Cav1.2 and Cav1.3 L‐type calcium channels independently control short‐ and long‐term sensitization to pain

PubMed Central

Radwani, Houda; Lopez‐Gonzalez, Maria José; Cattaert, Daniel; Roca‐Lapirot, Olivier; Dobremez, Eric; Bouali‐Benazzouz, Rabia; Eiríksdóttir, Emelía; Langel, Ülo; Favereaux, Alexandre; Errami, Mohammed; Landry, Marc

2016-01-01

Key points L‐type calcium channels in the CNS exist as two subunit forming channels, Cav1.2 and Cav1.3, which are involved in short‐ and long‐term plasticity.We demonstrate that Cav1.3 but not Cav1.2 is essential for wind‐up.These results identify Cav1.3 as a key conductance responsible for short‐term sensitization in physiological pain transmission.We confirm the role of Cav1.2 in a model of long‐term plasticity associated with neuropathic pain.Up‐regulation of Cav1.2 and down‐regultation of Cav1.3 in neuropathic pain underlies the switch from physiology to pathology.Finally, the results of the present study reveal that therapeutic targeting molecular pathways involved in wind‐up may be not relevant in the treatment of neuropathy. Abstract Short‐term central sensitization to pain temporarily increases the responsiveness of nociceptive pathways after peripheral injury. In dorsal horn neurons (DHNs), short‐term sensitization can be monitored through the study of wind‐up. Wind‐up, a progressive increase in DHNs response following repetitive peripheral stimulations, depends on the post‐synaptic L‐type calcium channels. In the dorsal horn of the spinal cord, two L‐type calcium channels are present, Cav1.2 and Cav1.3, each displaying specific kinetics and spatial distribution. In the present study, we used a mathematical model of DHNs in which we integrated the specific patterns of expression of each Cav subunits. This mathematical approach reveals that Cav1.3 is necessary for the onset of wind‐up, whereas Cav1.2 is not and that synaptically triggered wind‐up requires NMDA receptor activation. We then switched to a biological preparation in which we knocked down Cav subunits and confirmed the prominent role of Cav1.3 in both naive and spinal nerve ligation model of neuropathy (SNL). Interestingly, although a clear mechanical allodynia dependent on Cav1.2 expression was observed after SNL, the amplitude of wind‐up was decreased

Voltage-Sensitive Fluorescence of Indocyanine Green in the Heart

PubMed Central

Martišienė, Irma; Mačianskienė, Regina; Treinys, Rimantas; Navalinskas, Antanas; Almanaitytė, Mantė; Karčiauskas, Dainius; Kučinskas, Audrius; Grigalevičiūtė, Ramunė; Zigmantaitė, Vilma; Benetis, Rimantas; Jurevičius, Jonas

2016-01-01

So far, the optical mapping of cardiac electrical signals using voltage-sensitive fluorescent dyes has only been performed in experimental studies because these dyes are not yet approved for clinical use. It was recently reported that the well-known and widely used fluorescent dye indocyanine green (ICG), which has FDA approval, exhibits voltage sensitivity in various tissues, thus raising hopes that electrical activity could be optically mapped in the clinic. The aim of this study was to explore the possibility of using ICG to monitor cardiac electrical activity. Optical mapping experiments were performed on Langendorff rabbit hearts stained with ICG and perfused with electromechanical uncouplers. The residual contraction force and electrical action potentials were recorded simultaneously. Our research confirms that ICG is a voltage-sensitive dye with a dual-component (fast and slow) response to membrane potential changes. The fast component of the optical signal (OS) can have opposite polarities in different parts of the fluorescence spectrum. In contrast, the polarity of the slow component remains the same throughout the entire spectrum. Separating the OS into these components revealed two different voltage-sensitivity mechanisms for ICG. The fast component of the OS appears to be electrochromic in nature, whereas the slow component may arise from the redistribution of the dye molecules within or around the membrane. Both components quite accurately track the time of electrical signal propagation, but only the fast component is suitable for estimating the shape and duration of action potentials. Because ICG has voltage-sensitive properties in the entire heart, we suggest that it can be used to monitor cardiac electrical behavior in the clinic. PMID:26840736

Reversible calcium alloying enables a practical room-temperature rechargeable calcium-ion battery with a high discharge voltage

NASA Astrophysics Data System (ADS)

Wang, Meng; Jiang, Chunlei; Zhang, Songquan; Song, Xiaohe; Tang, Yongbing; Cheng, Hui-Ming

2018-06-01

Calcium-ion batteries (CIBs) are attractive candidates for energy storage because Ca2+ has low polarization and a reduction potential (-2.87 V versus standard hydrogen electrode, SHE) close to that of Li+ (-3.04 V versus SHE), promising a wide voltage window for a full battery. However, their development is limited by difficulties such as the lack of proper cathode/anode materials for reversible Ca2+ intercalation/de-intercalation, low working voltages (<2 V), low cycling stability, and especially poor room-temperature performance. Here, we report a CIB that can work stably at room temperature in a new cell configuration using graphite as the cathode and tin foils as the anode as well as the current collector. This CIB operates on a highly reversible electrochemical reaction that combines hexafluorophosphate intercalation/de-intercalation at the cathode and a Ca-involved alloying/de-alloying reaction at the anode. An optimized CIB exhibits a working voltage of up to 4.45 V with capacity retention of 95% after 350 cycles.

High-Sensitivity GaN Microchemical Sensors

NASA Technical Reports Server (NTRS)

Son, Kyung-ah; Yang, Baohua; Liao, Anna; Moon, Jeongsun; Prokopuk, Nicholas

2009-01-01

Systematic studies have been performed on the sensitivity of GaN HEMT (high electron mobility transistor) sensors using various gate electrode designs and operational parameters. The results here show that a higher sensitivity can be achieved with a larger W/L ratio (W = gate width, L = gate length) at a given D (D = source-drain distance), and multi-finger gate electrodes offer a higher sensitivity than a one-finger gate electrode. In terms of operating conditions, sensor sensitivity is strongly dependent on transconductance of the sensor. The highest sensitivity can be achieved at the gate voltage where the slope of the transconductance curve is the largest. This work provides critical information about how the gate electrode of a GaN HEMT, which has been identified as the most sensitive among GaN microsensors, needs to be designed, and what operation parameters should be used for high sensitivity detection.

Mini-dystrophin restores L-type calcium currents in skeletal muscle of transgenic mdx mice

PubMed Central

Friedrich, O; Both, M; Gillis, J M; Chamberlain, J S; Fink, RHA

2004-01-01

L-type calcium currents (iCa) were recorded using the two-microelectrode voltage-clamp technique in single short toe muscle fibres of three different mouse strains: (i) C57/SV129 wild-type mice (wt); (ii) mdx mice (an animal model for Duchenne muscular dystrophy; and (iii) transgenically engineered mini-dystrophin (MinD)-expressing mdx mice. The activation and inactivation properties of iCa were examined in 2- to 18-month-old animals. Ca2+ current densities at 0 mV in mdx fibres increased with age, but were always significantly smaller compared to age-matched wild-type fibres. Time-to-peak (TTP) of iCa was prolonged in mdx fibres compared to wt fibres. MinD fibres always showed similar TTP and current amplitudes compared to age-matched wt fibres. In all three genotypes, the voltage-dependent inactivation and deactivation of iCa were similar. Intracellular resting calcium concentration ([Ca2+]i) and the distribution of dihydropyridine binding sites were also not different in young animals of all three genotypes, whereas iCa was markedly reduced in mdx fibres. We conclude, that dystrophin influences L-type Ca2+ channels via a direct or indirect linkage which may be disrupted in mdx mice and may be crucial for proper excitation–contraction coupling initiating Ca2+ release from the sarcoplasmic reticulum. This linkage seems to be fully restored in the presence of mini-dystrophin. PMID:14594987

Voltage Dependence of a Neuromodulator-Activated Ionic Current.

PubMed

Gray, Michael; Golowasch, Jorge

2016-01-01

The neuromodulatory inward current (IMI) generated by crab Cancer borealis stomatogastric ganglion neurons is an inward current whose voltage dependence has been shown to be crucial in the activation of oscillatory activity of the pyloric network of this system. It has been previously shown that IMI loses its voltage dependence in conditions of low extracellular calcium, but that this effect appears to be regulated by intracellular calmodulin. Voltage dependence is only rarely regulated by intracellular signaling mechanisms. Here we address the hypothesis that the voltage dependence of IMI is mediated by intracellular signaling pathways activated by extracellular calcium. We demonstrate that calmodulin inhibitors and a ryanodine antagonist can reduce IMI voltage dependence in normal Ca(2+), but that, in conditions of low Ca(2+), calmodulin activators do not restore IMI voltage dependence. Further, we show evidence that CaMKII alters IMI voltage dependence. These results suggest that calmodulin is necessary but not sufficient for IMI voltage dependence. We therefore hypothesize that the Ca(2+)/calmodulin requirement for IMI voltage dependence is due to an active sensing of extracellular calcium by a GPCR family calcium-sensing receptor (CaSR) and that the reduction in IMI voltage dependence by a calmodulin inhibitor is due to CaSR endocytosis. Supporting this, preincubation with an endocytosis inhibitor prevented W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride)-induced loss of IMI voltage dependence, and a CaSR antagonist reduced IMI voltage dependence. Additionally, myosin light chain kinase, which is known to act downstream of the CaSR, seems to play a role in regulating IMI voltage dependence. Finally, a Gβγ-subunit inhibitor also affects IMI voltage dependence, in support of the hypothesis that this process is regulated by a G-protein-coupled CaSR.

Calcium influences sensitivity to growth inhibition induced by a cell surface sialoglycopeptide

NASA Technical Reports Server (NTRS)

Betz, N. A.; Fattaey, H. K.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

1994-01-01

While studies concerning mitogenic factors have been an important area of research for many years, much less is understood about the mechanisms of action of cell surface growth inhibitors. We have purified an 18 kDa cell surface sialoglycopeptide growth inhibitor (CeReS-18) which can reversibly inhibit the proliferation of diverse cell types. The studies discussed in this article show that three mouse keratinocyte cell lines exhibit sixty-fold greater sensitivity than other fibroblasts and epithelial-like cells to CeReS-18-induced growth inhibition. Growth inhibition induced by CeReS-18 treatment is a reversible process, and the three mouse keratinocyte cell lines exhibited either single or multiple cell cycle arrest points, although a predominantly G0/G1 cell cycle arrest point was exhibited in Swiss 3T3 fibroblasts. The sensitivity of the mouse keratinocyte cell lines to CeReS-18-induced growth inhibition was not affected by the degree of tumorigenic progression in the cell lines and was not due to differences in CeReS-18 binding affinity or number of cell surface receptors per cell. However, the sensitivity of both murine fibroblasts and keratinocytes could be altered by changing the extracellular calcium concentration, such that increased extracellular calcium concentrations resulted in decreased sensitivity to CeReS-18-induced proliferation inhibition. Thus the increased sensitivity of the murine keratinocyte cell lines to CeReS-18 could be ascribed to the low calcium concentration used in their propagation. Studies are currently under way investigating the role of calcium in CeReS-18-induced growth arrest. The CeReS-18 may serve as a very useful tool to study negative growth control and the signal transduction events associated with cell cycling.

G protein modulation of CaV2 voltage-gated calcium channels.

PubMed

Currie, Kevin P M

2010-01-01

Voltage-gated Ca(2+) channels translate the electrical inputs of excitable cells into biochemical outputs by controlling influx of the ubiquitous second messenger Ca(2+) . As such the channels play pivotal roles in many cellular functions including the triggering of neurotransmitter and hormone release by CaV2.1 (P/Q-type) and CaV2.2 (N-type) channels. It is well established that G protein coupled receptors (GPCRs) orchestrate precise regulation neurotransmitter and hormone release through inhibition of CaV2 channels. Although the GPCRs recruit a number of different pathways, perhaps the most prominent, and certainly most studied among these is the so-called voltage-dependent inhibition mediated by direct binding of Gβγ to the α1 subunit of CaV2 channels. This article will review the basics of Ca(2+) -channels and G protein signaling, and the functional impact of this now classical inhibitory mechanism on channel function. It will also provide an update on more recent developments in the field, both related to functional effects and crosstalk with other signaling pathways, and advances made toward understanding the molecular interactions that underlie binding of Gβγ to the channel and the voltage-dependence that is a signature characteristic of this mechanism.

Active Dendrites and Differential Distribution of Calcium Channels Enable Functional Compartmentalization of Golgi Cells.

PubMed

Rudolph, Stephanie; Hull, Court; Regehr, Wade G

2015-11-25

Interneurons are essential to controlling excitability, timing, and synaptic integration in neuronal networks. Golgi cells (GoCs) serve these roles at the input layer of the cerebellar cortex by releasing GABA to inhibit granule cells (grcs). GoCs are excited by mossy fibers (MFs) and grcs and provide feedforward and feedback inhibition to grcs. Here we investigate two important aspects of GoC physiology: the properties of GoC dendrites and the role of calcium signaling in regulating GoC spontaneous activity. Although GoC dendrites are extensive, previous studies concluded they are devoid of voltage-gated ion channels. Hence, the current view holds that somatic voltage signals decay passively within GoC dendrites, and grc synapses onto distal dendrites are not amplified and are therefore ineffective at firing GoCs because of strong passive attenuation. Using whole-cell recording and calcium imaging in rat slices, we find that dendritic voltage-gated sodium channels allow somatic action potentials to activate voltage-gated calcium channels (VGCCs) along the entire dendritic length, with R-type and T-type VGCCs preferentially located distally. We show that R- and T-type VGCCs located in the dendrites can boost distal synaptic inputs and promote burst firing. Active dendrites are thus critical to the regulation of GoC activity, and consequently, to the processing of input to the cerebellar cortex. In contrast, we find that N-type channels are preferentially located near the soma, and control the frequency and pattern of spontaneous firing through their close association with calcium-activated potassium (KCa) channels. Thus, VGCC types are differentially distributed and serve specialized functions within GoCs. Interneurons are essential to neural processing because they modulate excitability, timing, and synaptic integration within circuits. At the input layer of the cerebellar cortex, a single type of interneuron, the Golgi cell (GoC), carries these functions. The

Thrombospondin-4 divergently regulates voltage-gated Ca2+ channel subtypes in sensory neurons after nerve injury.

PubMed

Pan, Bin; Guo, Yuan; Wu, Hsiang-En; Park, John; Trinh, Van Nancy; Luo, Z David; Hogan, Quinn H

2016-09-01

Loss of high-voltage-activated (HVA) calcium current (ICa) and gain of low-voltage-activated (LVA) ICa after painful peripheral nerve injury cause elevated excitability in sensory neurons. Nerve injury is also accompanied by increased expression of the extracellular matrix glycoprotein thrombospondin-4 (TSP4), and interruption of TSP4 function can reverse or prevent behavioral hypersensitivity after injury. We therefore investigated TSP4 regulation of ICa in dorsal root ganglion (DRG) neurons. During depolarization adequate to activate HVA ICa, TSP4 decreases both N- and L-type ICa and the associated intracellular calcium transient. In contrast, TSP4 increases ICa and the intracellular calcium signal after low-voltage depolarization, which we confirmed is due to ICa through T-type channels. These effects are blocked by gabapentin, which ameliorates neuropathic pain by targeting the α2δ1 calcium subunit. Injury-induced changes of HVA and LVA ICa are attenuated in TSP4 knockout mice. In the neuropathic pain model of spinal nerve ligation, TSP4 application did not further regulate ICa of injured DRG neurons. Taken together, these findings suggest that elevated TSP4 after peripheral nerve injury may contribute to hypersensitivity of peripheral sensory systems by decreasing HVA and increasing LVA in DRG neurons by targeting the α2δ1 calcium subunit. Controlling TSP4 overexpression in peripheral sensory neurons may be a target for analgesic drug development for neuropathic pain.

P/Q-type calcium channels activate neighboring calcium-dependent potassium channels in mouse motor nerve terminals.

PubMed

Protti, D A; Uchitel, O D

1997-08-01

The identity of the voltage-dependent calcium channels (VDCC), which trigger the Ca2+-gated K+ currents (IK(Ca)) in mammalian motor nerve terminals, was investigated by means of perineurial recordings. The effects of Ca2+ chelators with different binding kinetics on the activation of IK(Ca) were also examined. The calcium channel blockers of the P/Q family, omega-agatoxin IVA (omega-Aga-IVA) and funnel-web spider toxin (FTX), have been shown to exert a strong blocking effect on IK(Ca). In contrast, nitrendipine and omega-conotoxin GVIA (omega-CgTx) did not affect the Ca2+-activated K+ currents. The intracellular action of the fast Ca2+ buffers BAPTA and DM-BAPTA prevented the activation of the IK(Ca), while the slow Ca2+ buffer EGTA was ineffective at blocking it. These data indicate that P/Q-type VDCC mediate the Ca2+ influx which activates IK(Ca). The spatial association between Ca2+ and Ca2+-gated K+ channels is discussed, on the basis of the differential effects of the fast and slow Ca2+ chelators.

Grafting voltage and pharmacological sensitivity in potassium channels.

PubMed

Lan, Xi; Fan, Chunyan; Ji, Wei; Tian, Fuyun; Xu, Tao; Gao, Zhaobing

2016-08-01

A classical voltage-gated ion channel consists of four voltage-sensing domains (VSDs). However, the roles of each VSD in the channels remain elusive. We developed a GVTDT (Graft VSD To Dimeric TASK3 channels that lack endogenous VSDs) strategy to produce voltage-gated channels with a reduced number of VSDs. TASK3 channels exhibit a high host tolerance to VSDs of various voltage-gated ion channels without interfering with the intrinsic properties of the TASK3 selectivity filter. The constructed channels, exemplified by the channels grafted with one or two VSDs from Kv7.1 channels, exhibit classical voltage sensitivity, including voltage-dependent opening and closing. Furthermore, the grafted Kv7.1 VSD transfers the potentiation activity of benzbromarone, an activator that acts on the VSDs of the donor channels, to the constructed channels. Our study indicates that one VSD is sufficient to voltage-dependently gate the pore and provides new insight into the roles of VSDs.

Localized accumulation of cytosolic calcium near the fused sperm is associated with the calcium- and voltage-dependent block of sperm entry in the sea urchin egg.

PubMed

Ivonnet, Pedro I; Mohri, Tatsuma; McCulloh, David H

2017-10-01

Interaction of the sperm and egg depolarizes the egg membrane, allowing the sperm to enter; however, if the egg membrane is not allowed to depolarize from its resting potential (e.g., by voltage-clamp), the sperm will not enter. Previous studies demonstrated that sperm entry into sea urchin eggs that are voltage-clamped at negative membrane potentials is regulated both by the egg's membrane potential and a voltage-dependent influx of calcium into the egg. In these cases, electrical or cytoplasmic continuity (sperm-egg membrane fusion) occurs at negative membrane potentials, but subsequent loss of cytoplasmic continuity results in failure of sperm entry (unfusion). The work presented herein examined where, in relation to the sperm, and when, in relation to the sperm-induced electrophysiological events, the egg's calcium influx occurs, and how these events relate to successful or failed sperm entry. When sperm entered the egg, elevation of intracellular calcium concentration ([Ca 2+ ] i ) began near the fused sperm on average 5.9 s after sperm-egg membrane fusion. Conversely, when sperm failed to enter the egg, [Ca 2+ ] i elevated near the site of sperm-egg fusion on average 0.7 s after sperm-egg membrane fusion, which is significantly earlier than in eggs for which sperm entered. Therefore, the accumulation of calcium near the site of sperm-egg fusion is spatially and temporally consistent with the mechanism that may be responsible for loss of cytoplasmic continuity and failure of sperm entry. © 2017 Wiley Periodicals, Inc.

Touch responsiveness in zebrafish requires voltage-gated calcium channel 2.1b

PubMed Central

Low, Sean E.; Woods, Ian G.; Lachance, Mathieu; Ryan, Joel; Saint-Amant, Louis

2012-01-01

The molecular and physiological basis of the touch-unresponsive zebrafish mutant fakir has remained elusive. Here we report that the fakir phenotype is caused by a missense mutation in the gene encoding voltage-gated calcium channel 2.1b (CACNA1Ab). Injection of RNA encoding wild-type CaV2.1 restores touch responsiveness in fakir mutants, whereas knockdown of CACNA1Ab via morpholino oligonucleotides recapitulates the fakir mutant phenotype. Fakir mutants display normal current-evoked synaptic communication at the neuromuscular junction but have attenuated touch-evoked activation of motor neurons. NMDA-evoked fictive swimming is not affected by the loss of CaV2.1b, suggesting that this channel is not required for motor pattern generation. These results, coupled with the expression of CACNA1Ab by sensory neurons, suggest that CaV2.1b channel activity is necessary for touch-evoked activation of the locomotor network in zebrafish. PMID:22490555

Ziconotide: neuronal calcium channel blocker for treating severe chronic pain.

PubMed

Miljanich, G P

2004-12-01

Ziconotide (PRIALT) is a neuroactive peptide in the final stages of clinical development as a novel non-opioid treatment for severe chronic pain. It is the synthetic equivalent of omega-MVIIA, a component of the venom of the marine snail, Conus magus. The mechanism of action underlying ziconotide's therapeutic profile derives from its potent and selective blockade of neuronal N-type voltage-sensitive calcium channels (N-VSCCs). Direct blockade of N-VSCCs inhibits the activity of a subset of neurons, including pain-sensing primary nociceptors. This mechanism of action distinguishes ziconotide from all other analgesics, including opioid analgesics. In fact, ziconotide is potently anti-nociceptive in animal models of pain in which morphine exhibits poor anti-nociceptive activity. Moreover, in contrast to opiates, tolerance to ziconotide is not observed. Clinical studies of ziconotide in more than 2,000 patients reveal important correlations to ziconotide's non-clinical pharmacology. For example, ziconotide provides significant pain relief to severe chronic pain sufferers who have failed to obtain relief from opiate therapy and no evidence of tolerance to ziconotide is seen in these patients. Contingent on regulatory approval, ziconotide will be the first in a new class of neurological drugs: the N-type calcium channel blockers, or NCCBs. Its novel mechanism of action as a non-opioid analgesic suggests ziconotide has the potential to play a valuable role in treatment regimens for severe chronic pain. If approved for clinical use, ziconotide will further validate the neuroactive venom peptides as a source of new and useful medicines.

Voltage Dependence of a Neuromodulator-Activated Ionic Current123

PubMed Central

2016-01-01

Abstract The neuromodulatory inward current (IMI) generated by crab Cancer borealis stomatogastric ganglion neurons is an inward current whose voltage dependence has been shown to be crucial in the activation of oscillatory activity of the pyloric network of this system. It has been previously shown that IMI loses its voltage dependence in conditions of low extracellular calcium, but that this effect appears to be regulated by intracellular calmodulin. Voltage dependence is only rarely regulated by intracellular signaling mechanisms. Here we address the hypothesis that the voltage dependence of IMI is mediated by intracellular signaling pathways activated by extracellular calcium. We demonstrate that calmodulin inhibitors and a ryanodine antagonist can reduce IMI voltage dependence in normal Ca2+, but that, in conditions of low Ca2+, calmodulin activators do not restore IMI voltage dependence. Further, we show evidence that CaMKII alters IMI voltage dependence. These results suggest that calmodulin is necessary but not sufficient for IMI voltage dependence. We therefore hypothesize that the Ca2+/calmodulin requirement for IMI voltage dependence is due to an active sensing of extracellular calcium by a GPCR family calcium-sensing receptor (CaSR) and that the reduction in IMI voltage dependence by a calmodulin inhibitor is due to CaSR endocytosis. Supporting this, preincubation with an endocytosis inhibitor prevented W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride)-induced loss of IMI voltage dependence, and a CaSR antagonist reduced IMI voltage dependence. Additionally, myosin light chain kinase, which is known to act downstream of the CaSR, seems to play a role in regulating IMI voltage dependence. Finally, a Gβγ-subunit inhibitor also affects IMI voltage dependence, in support of the hypothesis that this process is regulated by a G-protein-coupled CaSR. PMID:27257619

Complementary p- and n-type polymer doping for ambient stable graphene inverter.

PubMed

Yun, Je Moon; Park, Seokhan; Hwang, Young Hwan; Lee, Eui-Sup; Maiti, Uday; Moon, Hanul; Kim, Bo-Hyun; Bae, Byeong-Soo; Kim, Yong-Hyun; Kim, Sang Ouk

2014-01-28

Graphene offers great promise to complement the inherent limitations of silicon electronics. To date, considerable research efforts have been devoted to complementary p- and n-type doping of graphene as a fundamental requirement for graphene-based electronics. Unfortunately, previous efforts suffer from undesired defect formation, poor controllability of doping level, and subtle environmental sensitivity. Here we present that graphene can be complementary p- and n-doped by simple polymer coating with different dipolar characteristics. Significantly, spontaneous vertical ordering of dipolar pyridine side groups of poly(4-vinylpyridine) at graphene surface can stabilize n-type doping at room-temperature ambient condition. The dipole field also enhances and balances the charge mobility by screening the impurity charge effect from the bottom substrate. We successfully demonstrate ambient stable inverters by integrating p- and n-type graphene transistors, which demonstrated clear voltage inversion with a gain of 0.17 at a 3.3 V input voltage. This straightforward polymer doping offers diverse opportunities for graphene-based electronics, including logic circuits, particularly in mechanically flexible form.

Reversible loss of gravitropic sensitivity in maize roots after tip application of calcium chelators

NASA Technical Reports Server (NTRS)

Lee, J. S.; Mulkey, T. J.; Evans, M. L.

1983-01-01

The application of calcium chelating agents (EDTA or EGTA) to the tips of maize roots caused a loss of gravitropic sensitivity. When the chelator was replaced with calcium chloride, gravitropic sensitivity was restored. Asymmetric application of calcium chloride near the tip of a vertical root caused curvature toward the calcium source. When the calcium was applied to the upper surface of the tip of a root oriented horizontally, the root curved upward even though control roots exhibited strong downward curvature. Application of calcium chloride to the tips of decapped roots, which are known to be gravitropically insensitive, did not restore gravitropic sensitivity. However, asymmetric application of calcium chloride near the tips of decapped roots caused curvature toward the calcium source. Calcium may play a key role in linking gravity detection to gravitropic curvature in roots.

Apo states of calmodulin and CaBP1 control CaV1 voltage-gated calcium channel function through direct competition for the IQ domain.

PubMed

Findeisen, Felix; Rumpf, Christine H; Minor, Daniel L

2013-09-09

In neurons, binding of calmodulin (CaM) or calcium-binding protein 1 (CaBP1) to the CaV1 (L-type) voltage-gated calcium channel IQ domain endows the channel with diametrically opposed properties. CaM causes calcium-dependent inactivation and limits calcium entry, whereas CaBP1 blocks calcium-dependent inactivation (CDI) and allows sustained calcium influx. Here, we combine isothermal titration calorimetry with cell-based functional measurements and mathematical modeling to show that these calcium sensors behave in a competitive manner that is explained quantitatively by their apo-state binding affinities for the IQ domain. This competition can be completely blocked by covalent tethering of CaM to the channel. Further, we show that Ca(2+)/CaM has a sub-picomolar affinity for the IQ domain that is achieved without drastic alteration of calcium-binding properties. The observation that the apo forms of CaM and CaBP1 compete with each other demonstrates a simple mechanism for direct modulation of CaV1 function and suggests a means by which excitable cells may dynamically tune CaV activity. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

Analysis of the Effects of Calcium or Magnesium on Voltage-Clamp Currents in Perfused Squid Axons Bathed in Solutions of High Potassium

PubMed Central

Rojas, Eduardo; Taylor, Robert E.; Atwater, Illani; Bezanilla, Francisco

1969-01-01

Isolated axons from the squid, Dosidicus gigas, were internally perfused with potassium fluoride solutions. Membrane currents were measured following step changes of membrane potential in a voltage-clamp arrangement with external isosmotic solution changes in the order: potassium-free artificial seawater; potassium chloride; potassium chloride containing 10, 25, 40 or 50, mM calcium or magnesium; and potassium-free artificial seawater. The following results suggest that the currents measured under voltage clamp with potassium outside and inside can be separated into two components and that one of them, the predominant one, is carried through the potassium system. (a) Outward currents in isosmotic potassium were strongly and reversibly reduced by tetraethylammonium chloride. (b) Without calcium or magnesium a progressive increase in the nontime-dependent component of the currents (leakage) occurred. (c) The restoration of calcium or magnesium within 15–30 min decreases this leakage. (d) With 50 mM divalent ions the steady-state current-voltage curve was nonlinear with negative resistance as observed in intact axons in isosmotic potassium. (e) The time-dependent components of the membrane currents were not clearly affected by calcium or magnesium. These results show a strong dependence of the leakage currents on external calcium or magnesium concentration but provide no support for the involvement of calcium or magnesium in the kinetics of the potassium system. PMID:5823216

Analysis of the effects of calcium or magnesium on voltage-clamp currents in perfused squid axons bathed in solutions of high potassium.

PubMed

Rojas, E; Taylor, R E; Atwater, I; Bezanilla, F

1969-10-01

Isolated axons from the squid, Dosidicus gigas, were internally perfused with potassium fluoride solutions. Membrane currents were measured following step changes of membrane potential in a voltage-clamp arrangement with external isosmotic solution changes in the order: potassium-free artificial seawater; potassium chloride; potassium chloride containing 10, 25, 40 or 50, mM calcium or magnesium; and potassium-free artificial seawater. The following results suggest that the currents measured under voltage clamp with potassium outside and inside can be separated into two components and that one of them, the predominant one, is carried through the potassium system. (a) Outward currents in isosmotic potassium were strongly and reversibly reduced by tetraethylammonium chloride. (b) Without calcium or magnesium a progressive increase in the nontime-dependent component of the currents (leakage) occurred. (c) The restoration of calcium or magnesium within 15-30 min decreases this leakage. (d) With 50 mM divalent ions the steady-state current-voltage curve was nonlinear with negative resistance as observed in intact axons in isosmotic potassium. (e) The time-dependent components of the membrane currents were not clearly affected by calcium or magnesium. These results show a strong dependence of the leakage currents on external calcium or magnesium concentration but provide no support for the involvement of calcium or magnesium in the kinetics of the potassium system.

Diffusion properties of conventional and calcium-sensitive MRI contrast agents in the rat cerebral cortex.

PubMed

Hagberg, Gisela E; Mamedov, Ilgar; Power, Anthony; Beyerlein, Michael; Merkle, Hellmut; Kiselev, Valerij G; Dhingra, Kirti; Kubìček, Vojtĕch; Angelovski, Goran; Logothetis, Nikos K

2014-01-01

Calcium-sensitive MRI contrast agents can only yield quantitative results if the agent concentration in the tissue is known. The agent concentration could be determined by diffusion modeling, if relevant parameters were available. We have established an MRI-based method capable of determining diffusion properties of conventional and calcium-sensitive agents. Simulations and experiments demonstrate that the method is applicable both for conventional contrast agents with a fixed relaxivity value and for calcium-sensitive contrast agents. The full pharmacokinetic time-course of gadolinium concentration estimates was observed by MRI before, during and after intracerebral administration of the agent, and the effective diffusion coefficient D* was determined by voxel-wise fitting of the solution to the diffusion equation. The method yielded whole brain coverage with a high spatial and temporal sampling. The use of two types of MRI sequences for sampling of the diffusion time courses was investigated: Look-Locker-based quantitative T(1) mapping, and T(1) -weighted MRI. The observation times of the proposed MRI method is long (up to 20 h) and consequently the diffusion distances covered are also long (2-4 mm). Despite this difference, the D* values in vivo were in agreement with previous findings using optical measurement techniques, based on observation times of a few minutes. The effective diffusion coefficient determined for the calcium-sensitive contrast agents may be used to determine local tissue concentrations and to design infusion protocols that maintain the agent concentration at a steady state, thereby enabling quantitative sensing of the local calcium concentration. Copyright © 2014 John Wiley & Sons, Ltd.

Characterization of L-type calcium channel activity in atrioventricular nodal myocytes from rats with streptozotocin-induced Diabetes mellitus

PubMed Central

Yuill, Kathryn H; Al Kury, Lina T; Howarth, Frank Christopher

2015-01-01

Cardiovascular complications are common in patients with Diabetes mellitus (DM). In addition to changes in cardiac muscle inotropy, electrical abnormalities are also commonly observed in these patients. We have previously shown that spontaneous cellular electrical activity is altered in atrioventricular nodal (AVN) myocytes, isolated from the streptozotocin (STZ) rat model of type-1 DM. In this study, utilizing the same model, we have characterized the changes in L-type calcium channel activity in single AVN myocytes. Ionic currents were recorded from AVN myocytes isolated from the hearts of control rats and from those with STZ-induced diabetes. Patch-clamp recordings were used to assess the changes in cellular electrical activity in individual myocytes. Type-1 DM significantly altered the cellular characteristics of L-type calcium current. A reduction in peak ICaL density was observed, with no corresponding changes in the activation parameters of the current. L-type calcium channel current also exhibited faster time-dependent inactivation in AVN myocytes from diabetic rats. A negative shift in the voltage dependence of inactivation was also evident, and a slowing of restitution parameters. These findings demonstrate that experimentally induced type-1 DM significantly alters AVN L-type calcium channel cellular electrophysiology. These changes in ion channel activity may contribute to the abnormalities in cardiac electrical function that are associated with high mortality levels in patients with DM. PMID:26603460

Neurotransmitter Release Can Be Stabilized by a Mechanism That Prevents Voltage Changes Near the End of Action Potentials from Affecting Calcium Currents

PubMed Central

Clarke, Stephen G.; Scarnati, Matthew S.

2016-01-01

At chemical synapses, presynaptic action potentials (APs) activate voltage-gated calcium channels, allowing calcium to enter and trigger neurotransmitter release. The duration, peak amplitude, and shape of the AP falling phase alter calcium entry, which can affect neurotransmitter release significantly. In many neurons, APs do not immediately return to the resting potential, but instead exhibit a period of depolarization or hyperpolarization referred to as an afterpotential. We hypothesized that presynaptic afterpotentials should alter neurotransmitter release by affecting the electrical driving force for calcium entry and calcium channel gating. In support of this, presynaptic calcium entry is affected by afterpotentials after standard instant voltage jumps. Here, we used the mouse calyx of Held synapse, which allows simultaneous presynaptic and postsynaptic patch-clamp recording, to show that the postsynaptic response is affected significantly by presynaptic afterpotentials after voltage jumps. We therefore tested the effects of presynaptic afterpotentials using simultaneous presynaptic and postsynaptic recordings and AP waveforms or real APs. Surprisingly, presynaptic afterpotentials after AP stimuli did not alter calcium channel responses or neurotransmitter release appreciably. We show that the AP repolarization time course causes afterpotential-induced changes in calcium driving force and changes in calcium channel gating to effectively cancel each other out. This mechanism, in which electrical driving force is balanced by channel gating, prevents changes in calcium influx from occurring at the end of the AP and therefore acts to stabilize synaptic transmission. In addition, this mechanism can act to stabilize neurotransmitter release when the presynaptic resting potential changes. SIGNIFICANCE STATEMENT The shape of presynaptic action potentials (APs), particularly the falling phase, affects calcium entry and small changes in calcium influx can produce large

Differential effects of tetracaine on two kinetic components of calcium release in frog skeletal muscle fibres.

PubMed Central

Pizarro, G; Csernoch, L; Uribe, I; Ríos, E

1992-01-01

1. Intramembrane charge movements and changes in intracellular calcium concentration were recorded simultaneously in voltage clamped cut skeletal muscle fibres of the frog in the presence and absence of tetracaine. 2. Extracellular application of 20 microM tetracaine reduced the increase in myoplasmic [Ca2+]. The effect on the underlying calcium release flux from the sarcoplasmic reticulum was to suppress the peak of the release while sparing the steady level attained at the end of 100 ms clamp depolarizations. 3. While the peak of the release flux at corresponding voltages was reduced by 62% after the addition of tetracaine, the rate of inactivation was the same when the pulses elicited release fluxes of similar amplitude. 4. Higher concentrations of tetracaine, 0.2 mM, abolished the calcium signal in stretched fibres whereas in slack fibres this concentration left a non-inactivating calcium release flux. 5. Lowering the extracellular pH antagonized the effect of the drug both on charge movements and on calcium signals. The permanently charged analogue tetracaine methobromide lacked effects on excitation-contraction coupling. 6. These results imply that the two kinetic components of calcium release flux have very different tetracaine sensitivities. They are also consistent with an intracellular site of action of the drug at low concentration. Taken together they strongly suggest that the inactivating and non-inactivating components of calcium release correspond to different pathways: one that inactivates, is sensitive to tetracaine and is controlled by calcium, and another that does not inactivate, is much less sensitive to tetracaine and is directly controlled by voltage. PMID:1297844

Zebrafish CaV2.1 Calcium Channels Are Tailored for Fast Synchronous Neuromuscular Transmission

PubMed Central

Naranjo, David; Wen, Hua; Brehm, Paul

2015-01-01

The CaV2.2 (N-type) and CaV2.1 (P/Q-type) voltage-dependent calcium channels are prevalent throughout the nervous system where they mediate synaptic transmission, but the basis for the selective presence at individual synapses still remains an open question. The CaV2.1 channels have been proposed to respond more effectively to brief action potentials (APs), an idea supported by computational modeling. However, the side-by-side comparison of CaV2.1 and CaV2.2 kinetics in intact neurons failed to reveal differences. As an alternative means for direct functional comparison we expressed zebrafish CaV2.1 and CaV2.2 α-subunits, along with their accessory subunits, in HEK293 cells. HEK cells lack calcium currents, thereby circumventing the need for pharmacological inhibition of mixed calcium channel isoforms present in neurons. HEK cells also have a simplified morphology compared to neurons, which improves voltage control. Our measurements revealed faster kinetics and shallower voltage-dependence of activation and deactivation for CaV2.1. Additionally, recordings of calcium current in response to a command waveform based on the motorneuron AP show, directly, more effective activation of CaV2.1. Analysis of calcium currents associated with the AP waveform indicate an approximately fourfold greater open probability (PO) for CaV2.1. The efficient activation of CaV2.1 channels during APs may contribute to the highly reliable transmission at zebrafish neuromuscular junctions. PMID:25650925

Resonant-Type Smooth Impact Drive Mechanism Actuator Operating at Lower Input Voltages

NASA Astrophysics Data System (ADS)

Morita, Takeshi; Nishimura, Takuma; Yoshida, Ryuichi; Hosaka, Hiroshi

2013-07-01

We report on the design and fabrication of a resonant-type smooth impact drive mechanism (SIDM) actuator based on a multilayered piezoelectric ceramic transducer. Conventional SIDMs use off-resonant sawtooth-shaped displacement in developing stick-slip motion of a slider, but require large input voltages for high-speed operation. In contrast, in resonant-type SIDMs, a quasi-sawtooth-shaped displacement is obtained by combining two resonant vibrational modes. This driving principle enables low input voltage operations. In combining the modes, their frequency ratio must be 1:2. To design and optimize the stator transducer to generate sawtooth-shaped displacements, a transfer matrix method was adopted. With a preload of 270 mN, the no-load speed was 40 mm/s under a driving voltage of 1.6 V (peak to peak). This input voltage was one-sixth that of previous SIDMs for the same performance. Concurrently, heat generation was significantly reduced because dielectric losses were suppressed under the lower input voltage operation.

Apo-states of calmodulin and CaBP1 control CaV1 voltage-gated calcium channel function through direct competition for the IQ domain

PubMed Central

Findeisen, Felix; Rumpf, Christine; Minor, Daniel L.

2013-01-01

In neurons, binding of calmodulin (CaM) or calcium-binding protein 1 (CaBP1) to the CaV1 (L-type) voltage-gated calcium channel IQ domain endows the channel with diametrically opposed properties. CaM causes calcium-dependent inactivation (CDI) and limits calcium entry, whereas CaBP1 blocks CDI and allows sustained calcium influx. Here, we combine isothermal titration calorimetry (ITC) with cell-based functional measurements and mathematical modeling to show that these calcium sensors behave in a competitive manner that is explained quantitatively by their apo-state binding affinities for the IQ domain. This competition can be completely blocked by covalent tethering of CaM to the channel. Further, we show that Ca2+/CaM has a sub-picomolar affinity for the IQ domain that is achieved without drastic alteration of calcium binding properties. The observation that the apo-forms of CaM and CaBP1 compete with each other demonstrates a simple mechanism for direct modulation of CaV1 function and suggests a means by which excitable cells may dynamically tune CaV activity. PMID:23811053

Voltage-Load Sensitivity Matrix Based Demand Response for Voltage Control in High Solar Penetration Distribution Feeders

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

Zhu, Xiangqi; Wang, Jiyu; Mulcahy, David

This paper presents a voltage-load sensitivity matrix (VLSM) based voltage control method to deploy demand response resources for controlling voltage in high solar penetration distribution feeders. The IEEE 123-bus system in OpenDSS is used for testing the performance of the preliminary VLSM-based voltage control approach. A load disaggregation process is applied to disaggregate the total load profile at the feeder head to each load nodes along the feeder so that loads are modeled at residential house level. Measured solar generation profiles are used in the simulation to model the impact of solar power on distribution feeder voltage profiles. Different casemore » studies involving various PV penetration levels and installation locations have been performed. Simulation results show that the VLSM algorithm performance meets the voltage control requirements and is an effective voltage control strategy.« less

Two-pore channels: Regulation by NAADP and customized roles in triggering calcium signals

PubMed Central

Patel, Sandip; Marchant, Jonathan; Brailoiu, Eugen

2010-01-01

NAADP is a potent regulator of cytosolic calcium levels. Much evidence suggests that NAADP activates a novel channel located on an acidic (lysosomal-like) calcium store, the mobilisation of which results in further calcium release from the endoplasmic reticulum. Here, we discuss the recent identification of a family of poorly characterized ion channels (the two-pore channels) as endo-lysosomal NAADP receptors. The generation of calcium signals by these channels is likened to those evoked by depolarisation during excitation-contraction coupling in muscle. We discuss the idea that two pore-channels can mediate a trigger release of calcium which is then amplified by calcium-induced calcium release from the endoplasmic reticulum. This is similar to the activation of voltage-sensitive calcium channels and subsequent mobilisation of sarcoplasmic reticulum calcium stores in cardiac tissue. We suggest that two-pore channels may physically interact with ryanodine receptors to account for more direct release of calcium from the endoplasmic reticulum in analogy with the conformational coupling of voltage-sensitive calcium channels and ryanodine receptors in skeletal muscle. Interaction of two-pore channels with other calcium release channels likely occurs between stores “trans-chatter” and possibly within the same store “cis-chatter”. We also speculate that trafficking of two-pore channels through the endolysosomal system facilitates interactions with calcium entry channels. Strategic placing of two-pore channels thus provides a versatile means of generating spatiotemporally complex cellular calcium signals. PMID:20621760

Structure-function of proteins interacting with the α1 pore-forming subunit of high-voltage-activated calcium channels

PubMed Central

Neely, Alan; Hidalgo, Patricia

2014-01-01

Openings of high-voltage-activated (HVA) calcium channels lead to a transient increase in calcium concentration that in turn activate a plethora of cellular functions, including muscle contraction, secretion and gene transcription. To coordinate all these responses calcium channels form supramolecular assemblies containing effectors and regulatory proteins that couple calcium influx to the downstream signal cascades and to feedback elements. According to the original biochemical characterization of skeletal muscle Dihydropyridine receptors, HVA calcium channels are multi-subunit protein complexes consisting of a pore-forming subunit (α1) associated with four additional polypeptide chains β, α2, δ, and γ, often referred to as accessory subunits. Twenty-five years after the first purification of a high-voltage calcium channel, the concept of a flexible stoichiometry to expand the repertoire of mechanisms that regulate calcium channel influx has emerged. Several other proteins have been identified that associate directly with the α1-subunit, including calmodulin and multiple members of the small and large GTPase family. Some of these proteins only interact with a subset of α1-subunits and during specific stages of biogenesis. More strikingly, most of the α1-subunit interacting proteins, such as the β-subunit and small GTPases, regulate both gating and trafficking through a variety of mechanisms. Modulation of channel activity covers almost all biophysical properties of the channel. Likewise, regulation of the number of channels in the plasma membrane is performed by altering the release of the α1-subunit from the endoplasmic reticulum, by reducing its degradation or enhancing its recycling back to the cell surface. In this review, we discuss the structural basis, interplay and functional role of selected proteins that interact with the central pore-forming subunit of HVA calcium channels. PMID:24917826

Optimization design on breakdown voltage of AlGaN/GaN high-electron mobility transistor

NASA Astrophysics Data System (ADS)

Yang, Liu; Changchun, Chai; Chunlei, Shi; Qingyang, Fan; Yuqian, Liu

2016-12-01

Simulations are carried out to explore the possibility of achieving high breakdown voltage of GaN HEMT (high-electron mobility transistor). GaN cap layers with gradual increase in the doping concentration from 2 × 1016 to 5 × 1019 cm-3 of N-type and P-type cap are investigated, respectively. Simulation results show that HEMT with P-doped GaN cap layer shows more potential to achieve higher breakdown voltage than N-doped GaN cap layer under the same doping concentration. This is because the ionized net negative space charges in P-GaN cap layer could modulate the surface electric field which makes more contribution to RESURF effect. Furthermore, a novel GaN/AlGaN/GaN HEMT with P-doped GaN buried layer in GaN buffer between gate and drain electrode is proposed. It shows enhanced performance. The breakdown voltage of the proposed structure is 640 V which is increased by 12% in comparison to UID (un-intentionally doped) GaN/AlGaN/GaN HEMT. We calculated and analyzed the distribution of electrons' density. It is found that the depleted region is wider and electric field maximum value is induced at the left edge of buried layer. So the novel structure with P-doped GaN buried layer embedded in GaN buffer has the better improving characteristics of the power devices. Project supported by the National Basic Research Program of China (No. 2014CB339900) and the Open Fund of Key Laboratory of Complex Electromagnetic Environment Science and Technology, China Academy of Engineering Physics (No. 2015-0214.XY.K).

The saponin monomer of dwarf lilyturf tuber, DT-13, reduces L-type calcium currents during hypoxia in adult rat ventricular myocytes.

PubMed

Tao, Jin; Wang, Hongyi; Zhou, Hong; Li, Shengnan

2005-10-28

The saponin monomer 13 of dwarf lilyturf tuber (DT-13), one of the saponin monomers of dwarf lilyturf tuber, has been found to have potent cardioprotective effects. In order to investigate the effects of DT-13 on L-type calcium currents (I(Ca,L)), exploring the mechanisms of DT-13's cardioprotective effects in the condition of pathophysiology, we directly measured the I(Ca,L) during hypoxia in the adult rat cardiac myocytes exposed to DT-13 using standard whole-cell patch-clamp recording technique. Our previous results showed that DT-13 exerted decreasing effects on the I(Ca,L) of the single adult rat cardiac myocytes. In the condition of hypoxia, the current density was inhibited by about 29% after exposure of the cells to DT-13 (0.1 micromol L(-1)) for 10 min, from 6.96+/-1.05 pA/pF to 4.38+/-0.35 pA/pF (n=5, P<0.05). This I(Ca,L)-inhibiting action of DT-13 was concentration-dependent and showed no frequency-dependence. DT-13 up-shifted the current-voltage (I-V) curve. Steady-state activation of I(Ca,L) was not affected markedly, and the half activation potential (V(0.5)) in the presence of DT-13 (0.1 micromol L(-1)) was also not significantly different. DT-13 at 0.1 micromol L(-1) markedly accelerated the voltage-dependent steady-state inactivation of calcium current and shifted the steady-state inactivation curve of I(Ca,L) to the left. In combination with previous reports, these results suggest that there might be a close relationship between the cardioprotective effects of DT-13 and L-type calcium channels in the condition of hypoxia.

Neurotransmitter Release Can Be Stabilized by a Mechanism That Prevents Voltage Changes Near the End of Action Potentials from Affecting Calcium Currents.

PubMed

Clarke, Stephen G; Scarnati, Matthew S; Paradiso, Kenneth G

2016-11-09

At chemical synapses, presynaptic action potentials (APs) activate voltage-gated calcium channels, allowing calcium to enter and trigger neurotransmitter release. The duration, peak amplitude, and shape of the AP falling phase alter calcium entry, which can affect neurotransmitter release significantly. In many neurons, APs do not immediately return to the resting potential, but instead exhibit a period of depolarization or hyperpolarization referred to as an afterpotential. We hypothesized that presynaptic afterpotentials should alter neurotransmitter release by affecting the electrical driving force for calcium entry and calcium channel gating. In support of this, presynaptic calcium entry is affected by afterpotentials after standard instant voltage jumps. Here, we used the mouse calyx of Held synapse, which allows simultaneous presynaptic and postsynaptic patch-clamp recording, to show that the postsynaptic response is affected significantly by presynaptic afterpotentials after voltage jumps. We therefore tested the effects of presynaptic afterpotentials using simultaneous presynaptic and postsynaptic recordings and AP waveforms or real APs. Surprisingly, presynaptic afterpotentials after AP stimuli did not alter calcium channel responses or neurotransmitter release appreciably. We show that the AP repolarization time course causes afterpotential-induced changes in calcium driving force and changes in calcium channel gating to effectively cancel each other out. This mechanism, in which electrical driving force is balanced by channel gating, prevents changes in calcium influx from occurring at the end of the AP and therefore acts to stabilize synaptic transmission. In addition, this mechanism can act to stabilize neurotransmitter release when the presynaptic resting potential changes. The shape of presynaptic action potentials (APs), particularly the falling phase, affects calcium entry and small changes in calcium influx can produce large changes in

Inhibition of recombinant Ca(v)3.1 (alpha(1G)) T-type calcium channels by the antipsychotic drug clozapine.

PubMed

Choi, Kee-Hyun; Rhim, Hyewhon

2010-01-25

Low voltage-activated T-type calcium channels are involved in the regulation of the neuronal excitability, and could be subject to many antipsychotic drugs. The effects of clozapine, an atypical antipsychotic drug, on recombinant Ca(v)3.1 T-type calcium channels heterologously expressed in human embryonic kidney 293 cells were examined using whole-cell patch-clamp recordings. At a standard holding potential of -100 mV, clozapine inhibited Ca(v)3.1 currents with an IC(50) value of 23.7+/-1.3 microM in a use-dependent manner. However, 10 microM clozapine inhibited more than 50% of the Ca(v)3.1 currents in recordings at a more physiologically relevant holding potential of -75 mV. Clozapine caused a significant hyperpolarizing shift in the steady-state inactivation curve of the Ca(v)3.1 channels, which is presumably the main mechanism accounting for the inhibition of the Ca(v)3.1 currents. In addition, clozapine slowed Ca(v)3.1 deactivation and inactivation kinetics but not activation kinetics. Clozapine-induced changes in deactivation and inactivation rates of the Ca(v)3.1 channel gating would likely facilitate calcium influx via Ca(v)3.1 T-type calcium channels. Thus, clozapine may exert its therapeutic and/or side effects by altering cell's excitability and firing properties through actions on T-type calcium channels.

Listening to membrane potential: photoacoustic voltage-sensitive dye recording.

PubMed

Zhang, Haichong K; Yan, Ping; Kang, Jeeun; Abou, Diane S; Le, Hanh N D; Jha, Abhinav K; Thorek, Daniel L J; Kang, Jin U; Rahmim, Arman; Wong, Dean F; Boctor, Emad M; Loew, Leslie M

2017-04-01

Voltage-sensitive dyes (VSDs) are designed to monitor membrane potential by detecting fluorescence changes in response to neuronal or muscle electrical activity. However, fluorescence imaging is limited by depth of penetration and high scattering losses, which leads to low sensitivity in vivo systems for external detection. By contrast, photoacoustic (PA) imaging, an emerging modality, is capable of deep tissue, noninvasive imaging by combining near-infrared light excitation and ultrasound detection. Here, we show that voltage-dependent quenching of dye fluorescence leads to a reciprocal enhancement of PA intensity. We synthesized a near-infrared photoacoustic VSD (PA-VSD), whose PA intensity change is sensitive to membrane potential. In the polarized state, this cyanine-based probe enhances PA intensity while decreasing fluorescence output in a lipid vesicle membrane model. A theoretical model accounts for how the experimental PA intensity change depends on fluorescence and absorbance properties of the dye. These results not only demonstrate PA voltage sensing but also emphasize the interplay of both fluorescence and absorbance properties in the design of optimized PA probes. Together, our results demonstrate PA sensing as a potential new modality for recording and external imaging of electrophysiological and neurochemical events in the brain.

Listening to membrane potential: photoacoustic voltage-sensitive dye recording

NASA Astrophysics Data System (ADS)

Zhang, Haichong K.; Yan, Ping; Kang, Jeeun; Abou, Diane S.; Le, Hanh N. D.; Jha, Abhinav K.; Thorek, Daniel L. J.; Kang, Jin U.; Rahmim, Arman; Wong, Dean F.; Boctor, Emad M.; Loew, Leslie M.

2017-04-01

Voltage-sensitive dyes (VSDs) are designed to monitor membrane potential by detecting fluorescence changes in response to neuronal or muscle electrical activity. However, fluorescence imaging is limited by depth of penetration and high scattering losses, which leads to low sensitivity in vivo systems for external detection. By contrast, photoacoustic (PA) imaging, an emerging modality, is capable of deep tissue, noninvasive imaging by combining near-infrared light excitation and ultrasound detection. Here, we show that voltage-dependent quenching of dye fluorescence leads to a reciprocal enhancement of PA intensity. We synthesized a near-infrared photoacoustic VSD (PA-VSD), whose PA intensity change is sensitive to membrane potential. In the polarized state, this cyanine-based probe enhances PA intensity while decreasing fluorescence output in a lipid vesicle membrane model. A theoretical model accounts for how the experimental PA intensity change depends on fluorescence and absorbance properties of the dye. These results not only demonstrate PA voltage sensing but also emphasize the interplay of both fluorescence and absorbance properties in the design of optimized PA probes. Together, our results demonstrate PA sensing as a potential new modality for recording and external imaging of electrophysiological and neurochemical events in the brain.

Cell-type-specific modelling of intracellular calcium signalling: a urothelial cell model.

PubMed

Appleby, Peter A; Shabir, Saqib; Southgate, Jennifer; Walker, Dawn

2013-09-06

Calcium signalling plays a central role in regulating a wide variety of cell processes. A number of calcium signalling models exist in the literature that are capable of reproducing a variety of experimentally observed calcium transients. These models have been used to examine in more detail the mechanisms underlying calcium transients, but very rarely has a model been directly linked to a particular cell type and experimentally verified. It is important to show that this can be achieved within the general theoretical framework adopted by these models. Here, we develop a framework designed specifically for modelling cytosolic calcium transients in urothelial cells. Where possible, we draw upon existing calcium signalling models, integrating descriptions of components known to be important in this cell type from a number of studies in the literature. We then add descriptions of several additional pathways that play a specific role in urothelial cell signalling, including an explicit ionic influx term and an active pumping mechanism that drives the cytosolic calcium concentration to a target equilibrium. The resulting one-pool model of endoplasmic reticulum (ER)-dependent calcium signalling relates the cytosolic, extracellular and ER calcium concentrations and can generate a wide range of calcium transients, including spikes, bursts, oscillations and sustained elevations in the cytosolic calcium concentration. Using single-variate robustness and multivariate sensitivity analyses, we quantify how varying each of the parameters of the model leads to changes in key features of the calcium transient, such as initial peak amplitude and the frequency of bursting or spiking, and in the transitions between bursting- and plateau-dominated modes. We also show that, novel to our urothelial cell model, the ionic and purinergic P2Y pathways make distinct contributions to the calcium transient. We then validate the model using human bladder epithelial cells grown in monolayer cell

The low threshold voltage n-type silicon transistors based on a polymer/silica nanocomposite gate dielectric: The effect of annealing temperatures on their operation

NASA Astrophysics Data System (ADS)

Hashemi, Adeleh; Bahari, Ali; Ghasemi, Shahram

2017-09-01

In this work, povidone/silica nanocomposite dielectric layers were deposited on the n-type Si (100) substrates for application in n-type silicon field-effect transistors (FET). Thermogravimetric analysis (TGA) indicated that strong chemical interactions between polymer and silica nanoparticles were created. In order to examine the effect of annealing temperatures on chemical interactions and nanostructure properties, annealing process was done at 423-513 K. Atomic force microscopy (AFM) images show the very smooth surfaces with very low surface roughness (0.038-0.088 nm). The Si2p and C1s core level photoemission spectra were deconvoluted to the chemical environments of Si and C atoms respectively. The obtained results of deconvoluted X-ray photoelectron spectroscopy (XPS) spectra revealed a high percentage of silanol hydrogen bonds in the sample which was not annealed. These bonds were inversed to stronger covalence bonds (siloxan bonds) at annealing temperature of 423 K. By further addition of temperature, siloxan bonds were shifted to lower binding energy of about 1 eV and their intensity were abated at annealing temperature of 513 K. The electrical characteristics were extracted from current-Voltage (I-V) and capacitance-voltage (C-V) measurements in metal-insulator-semiconductor (MIS) structure. The all n-type Si transistors showed very low threshold voltages (-0.24 to 1 V). The formation of the strongest cross-linking at nanostructure of dielectric film annealed at 423 K caused resulted in an un-trapped path for the transport of charge carriers yielding the lowest threshold voltage (0.08 V) and the highest electron mobility (45.01 cm2/V s) for its FET. By increasing the annealing temperature (473 and 513 K) on the nanocomposite dielectric films, the values of the average surface roughness, the capacitance and the FET threshold voltage increased and the value of FET electron field-effect mobility decreased.

Development of type-I/type-II hybrid dye sensitizer with both pyridyl group and catechol unit as anchoring group for type-I/type-II dye-sensitized solar cell.

PubMed

Ooyama, Yousuke; Furue, Kensuke; Enoki, Toshiaki; Kanda, Masahiro; Adachi, Yohei; Ohshita, Joji

2016-11-09

A type-I/type-II hybrid dye sensitizer with a pyridyl group and a catechol unit as the anchoring group has been developed and its photovoltaic performance in dye-sensitized solar cells (DSSCs) is investigated. The sensitizer has the ability to adsorb on a TiO 2 electrode through both the coordination bond at Lewis acid sites and the bidentate binuclear bridging linkage at Brønsted acid sites on the TiO 2 surface, which makes it possible to inject an electron into the conduction band of the TiO 2 electrode by the intramolecular charge-transfer (ICT) excitation (type-I pathway) and by the photoexcitation of the dye-to-TiO 2 charge transfer (DTCT) band (type-II pathway). It was found that the type-I/type-II hybrid dye sensitizer adsorbed on TiO 2 film exhibits a broad photoabsorption band originating from ICT and DTCT characteristics. Here we reveal the photophysical and electrochemical properties of the type-I/type-II hybrid dye sensitizer bearing a pyridyl group and a catechol unit, along with its adsorption modes onto TiO 2 film, and its photovoltaic performance in type-I/type-II DSSC, based on optical (photoabsorption and fluorescence spectroscopy) and electrochemical measurements (cyclic voltammetry), density functional theory (DFT) calculation, FT-IR spectroscopy of the dyes adsorbed on TiO 2 film, photocurrent-voltage (I-V) curves, incident photon-to-current conversion efficiency (IPCE) spectra, and electrochemical impedance spectroscopy (EIS) for DSSC.

Direct Interaction of CaVβ with Actin Up-regulates L-type Calcium Currents in HL-1 Cardiomyocytes*

PubMed Central

Stölting, Gabriel; de Oliveira, Regina Campos; Guzman, Raul E.; Miranda-Laferte, Erick; Conrad, Rachel; Jordan, Nadine; Schmidt, Silke; Hendriks, Johnny; Gensch, Thomas; Hidalgo, Patricia

2015-01-01

Expression of the β-subunit (CaVβ) is required for normal function of cardiac L-type calcium channels, and its up-regulation is associated with heart failure. CaVβ binds to the α1 pore-forming subunit of L-type channels and augments calcium current density by facilitating channel opening and increasing the number of channels in the plasma membrane, by a poorly understood mechanism. Actin, a key component of the intracellular trafficking machinery, interacts with Src homology 3 domains in different proteins. Although CaVβ encompasses a highly conserved Src homology 3 domain, association with actin has not yet been explored. Here, using co-sedimentation assays and FRET experiments, we uncover a direct interaction between CaVβ and actin filaments. Consistently, single-molecule localization analysis reveals streaklike structures composed by CaVβ2 that distribute over several micrometers along actin filaments in HL-1 cardiomyocytes. Overexpression of CaVβ2-N3 in HL-1 cells induces an increase in L-type current without altering voltage-dependent activation, thus reflecting an increased number of channels in the plasma membrane. CaVβ mediated L-type up-regulation, and CaVβ-actin association is prevented by disruption of the actin cytoskeleton with cytochalasin D. Our study reveals for the first time an interacting partner of CaVβ that is directly involved in vesicular trafficking. We propose a model in which CaVβ promotes anterograde trafficking of the L-type channels by anchoring them to actin filaments in their itinerary to the plasma membrane. PMID:25533460

Effects of Combined Calcium and Vitamin D Supplementation on Insulin Secretion, Insulin Sensitivity and β-Cell Function in Multi-Ethnic Vitamin D-Deficient Adults at Risk for Type 2 Diabetes: A Pilot Randomized, Placebo-Controlled Trial

PubMed Central

Gagnon, Claudia; Daly, Robin M.; Carpentier, André; Lu, Zhong X.; Shore-Lorenti, Catherine; Sikaris, Ken; Jean, Sonia; Ebeling, Peter R.

2014-01-01

Objectives To examine whether combined vitamin D and calcium supplementation improves insulin sensitivity, insulin secretion, β-cell function, inflammation and metabolic markers. Design 6-month randomized, placebo-controlled trial. Participants Ninety-five adults with serum 25-hydroxyvitamin D [25(OH)D] ≤55 nmol/L at risk of type 2 diabetes (with prediabetes or an AUSDRISK score ≥15) were randomized. Analyses included participants who completed the baseline and final visits (treatment n = 35; placebo n = 45). Intervention Daily calcium carbonate (1,200 mg) and cholecalciferol [2,000–6,000 IU to target 25(OH)D >75 nmol/L] or matching placebos for 6 months. Measurements Insulin sensitivity (HOMA2%S, Matsuda index), insulin secretion (insulinogenic index, area under the curve (AUC) for C-peptide) and β-cell function (Matsuda index x AUC for C-peptide) derived from a 75 g 2-h OGTT; anthropometry; blood pressure; lipid profile; hs-CRP; TNF-α; IL-6; adiponectin; total and undercarboxylated osteocalcin. Results Participants were middle-aged adults (mean age 54 years; 69% Europid) at risk of type 2 diabetes (48% with prediabetes). Compliance was >80% for calcium and vitamin D. Mean serum 25(OH)D concentration increased from 48 to 95 nmol/L in the treatment group (91% achieved >75 nmol/L), but remained unchanged in controls. There were no significant changes in insulin sensitivity, insulin secretion and β-cell function, or in inflammatory and metabolic markers between or within the groups, before or after adjustment for potential confounders including waist circumference and season of recruitment. In a post hoc analysis restricted to participants with prediabetes, a significant beneficial effect of vitamin D and calcium supplementation on insulin sensitivity (HOMA%S and Matsuda) was observed. Conclusions Daily vitamin D and calcium supplementation for 6 months may not change OGTT-derived measures of insulin sensitivity, insulin secretion and �

N- and P-type Ca2+ channels are involved in acetylcholine release at a neuroneuronal synapse: only the N-type channel is the target of neuromodulators.

PubMed Central

Fossier, P; Baux, G; Tauc, L

1994-01-01

Cholinergic transmission in an identified neuro-neuronal synapse of the Aplysia buccal ganglion was depressed by application of a partially purified extract of the funnel-web-spider venom (FTx) or of its synthetic analog (sFTx). This specific blocker of voltage-dependent P-type Ca2+ channels did not interfere with the effect of the N-type Ca2+ channel blocker omega-conotoxin, which could further decrease synaptic transmission after a previous application of FTx. Similar results were obtained when the reversal order of application of these two Ca2+ channel blockers was used. Both P- and N-type Ca2+ currents trigger acetylcholine release in the presynaptic neuron. The neuromodulatory effects of FMRF-amide, histamine, and buccalin on transmitter release disappeared after the blockade of the N-type Ca2+ channels but remained still effective in the presence of FTx. These results indicate that only N-type Ca2+ channels appear to be sensitive to the neuromodulators we have identified. PMID:7910963

N- and P-type Ca2+ channels are involved in acetylcholine release at a neuroneuronal synapse: only the N-type channel is the target of neuromodulators.

PubMed

Fossier, P; Baux, G; Tauc, L

1994-05-24

Cholinergic transmission in an identified neuro-neuronal synapse of the Aplysia buccal ganglion was depressed by application of a partially purified extract of the funnel-web-spider venom (FTx) or of its synthetic analog (sFTx). This specific blocker of voltage-dependent P-type Ca2+ channels did not interfere with the effect of the N-type Ca2+ channel blocker omega-conotoxin, which could further decrease synaptic transmission after a previous application of FTx. Similar results were obtained when the reversal order of application of these two Ca2+ channel blockers was used. Both P- and N-type Ca2+ currents trigger acetylcholine release in the presynaptic neuron. The neuromodulatory effects of FMRF-amide, histamine, and buccalin on transmitter release disappeared after the blockade of the N-type Ca2+ channels but remained still effective in the presence of FTx. These results indicate that only N-type Ca2+ channels appear to be sensitive to the neuromodulators we have identified.

Benzodiazepine-induced hippocampal CA1 neuron alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic acid (AMPA) receptor plasticity linked to severity of withdrawal anxiety: differential role of voltage-gated calcium channels and N-methyl-D-aspartic acid receptors.

PubMed

Xiang, Kun; Tietz, Elizabeth I

2007-09-01

Withdrawal from 1-week oral administration of the benzodiazepine, flurazepam (FZP) is associated with increased alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic acid (AMPA) receptor (AMPAR) miniature excitatory postsynaptic currents (mEPSCs) but reduction of N-methyl-D-aspartic acid (NMDA) receptor (NMDAR)-evoked (e)EPSCs in hippocampal CA1 neurons. A positive correlation was observed between increased AMPAR-mediated mEPSC amplitude and anxiety-like behavior in 1-day FZP-withdrawn rats. These effects were disrupted by systemic AMPAR antagonist administration (GYKI-52466, 0.5 mg/kg, intraperitoneal) at withdrawal onset, strengthening the hypothesis that CA1 neuron AMPAR-mediated hyperexcitability is a central component of a functional anatomic circuit associated with the expression of withdrawal anxiety. Abolition of AMPAR current upregulation in 2-day FZP withdrawn rats by GYKI-52466 injection also reversed the reduction in NMDAR-mediated eEPSC amplitude in CA1 neurons from the same rats, suggesting that downregulation of NMDAR function may serve a protective, negative-feedback role to prevent AMPAR-mediated neuronal overexcitation. NMDAR antagonist administration (MK-801, 0.25 mg/kg intraperitoneally) had no effect on modifying increased glutamatergic strength or on withdrawal anxiety, whereas injection of an L-type voltage-gated calcium channel antagonist, nimodipine (10 mg/kg, intraperitoneally) averted AMPAR current enhancement and anxiety-like behavior, suggesting that these manifestations may be initiated by a voltage-gated calcium channel-dependent signal transduction pathway. An evidence-based model of likely cellular mechanisms in the hippocampus contributing to benzodiazepine withdrawal anxiety was proposed implicating regulation of multiple CA1 neuron ion channels.

Voltage-sensitive dye recording from networks of cultured neurons

NASA Astrophysics Data System (ADS)

Chien, Chi-Bin

This thesis describes the development and testing of a sensitive apparatus for recording electrical activity from microcultures of rat superior cervical ganglion (SCG) neurons by using voltage-sensitive fluorescent dyes.The apparatus comprises a feedback-regulated mercury arc light source, an inverted epifluorescence microscope, a novel fiber-optic camera with discrete photodiode detectors, and low-noise preamplifiers. Using an NA 0.75 objective and illuminating at 10 W/cm2 with the 546 nm mercury line, a typical SCG neuron stained with the styryl dye RH423 gives a detected photocurrent of 1 nA; the light source and optical detectors are quiet enough that the shot noise in this photocurrent--about.03% rms--dominates. The design, theory, and performance of this dye-recording apparatus are discussed in detail.Styryl dyes such as RH423 typically give signals of 1%/100 mV on these cells; the signals are linear in membrane potential, but do not appear to arise from a purely electrochromic mechanism. Given this voltage sensitivity and the noise level of the apparatus, it should be possible to detect both action potentials and subthreshold synaptic potentials from SCG cell bodies. In practice, dye recording can easily detect action potentials from every neuron in an SCG microculture, but small synaptic potentials are obscured by dye signals from the dense network of axons.In another microculture system that does not have such long and complex axons, this dye-recording apparatus should be able to detect synaptic potentials, making it possible to noninvasively map the synaptic connections in a microculture, and thus to study long-term synaptic plasticity.

Distribution of ABO Blood Groups and Coronary Artery Calcium.

PubMed

Wang, Yao; Zhou, Bing-Yang; Zhu, Cheng-Gang; Guo, Yuan-Lin; Wu, Na-Qiong; Qing, Ping; Gao, Ying; Liu, Geng; Dong, Qian; Li, Jian-Jun

2017-06-01

ABO blood groups have been confirmed to be associated with cardiovascular diseases such as coronary artery disease. However, whether ABO blood group is correlated with coronary artery calcium (CAC) is still unknown. 301 patients with coronary artery calcium score (CACS) assessed by computed tomography were consecutively enrolled and divided into two groups: with calcium group (CACS>0, n=104) and without calcium group (CACS=0, n=197). Distribution of ABO blood groups was evaluated between the two groups. The percentage of A blood type was significantly higher (p=0.008) and O blood type was significantly lower (p=0.037) in the calcium group. Univariate regression analysis showed that age, total cholesterol, low density lipoprotein cholesterol, high-sensitivity C-reactive protein, A blood type were positively correlated with CAC, and O blood type was inversely associated with CAC. Multivariate regression analysis showed that A blood type was independently associated with CAC (odds ratio: 2.217, 95% confidence interval: 1.260-3.900, p=0.006) even after further adjustment for variables that were clearly different between the two groups. Our data has suggested for the first time that A blood type was an independent risk marker for CAC. Copyright © 2016 Australian and New Zealand Society of Cardiac and Thoracic Surgeons (ANZSCTS) and the Cardiac Society of Australia and New Zealand (CSANZ). Published by Elsevier B.V. All rights reserved.

Changes in force and calcium sensitivity in the developing avian heart.

PubMed

Godt, R E; Fogaça, R T; Nosek, T M

1991-11-01

The aim of this study was to characterize the development of the contractile properties of intact and chemically skinned muscle from chicken heart and to compare these characteristics with those of developing mammalian heart reported by others. Small trabeculae were dissected from left ventricles of Arbor Acre chickens between embryonic day 7 and young adulthood (7 weeks post-hatching). At all ages, increasing extracellular calcium (0.45-3.6 mM) progressively increased twitch force of electrically stimulated trabeculae. Twitch force at 1.8 mM extracellular calcium, normalized to cross-sectional area, increased to a maximum at 1 day post-hatching, remained constant through 3 weeks post-hatching, but then decreased at 7 weeks post-hatching. The maximal calcium-activated force of trabeculae chemically skinned with Triton X-100 detergent increased to a maximum 2 days before the time of hatching and was not significantly changed up to 7 weeks post-hatching. Over the ages studied, average twitch force in 1.8 mM calcium was between 26 and 66% of maximal calcium-activated force after skinning, suggesting that the contractile apparatus is not fully activated during the twitch in normal Ringer. In skinned trabeculae, the calcium sensitivity of the contractile apparatus was higher in the embryo than in the young adult. These age-dependent changes in calcium sensitivity are correlated with isoform switching in troponin T. A decrease in pH from 7.0 to 6.5 decreased the calcium sensitivity of the contractile apparatus to a greater degree in skinned trabeculae from young adult hearts than in those from embryonic hearts. This change in susceptibility to acidosis is temporally associated with isoform switching in troponin I.(ABSTRACT TRUNCATED AT 250 WORDS)

[Nonuniform distribution and contribution of the P- and P/Q-type calcium channels to short-term inhibitory synaptic transmission in cultured hippocampal neurons].

PubMed

Mizerna, O P; Fedulova, S A; Veselovs'kyĭ, M S

2010-01-01

In the present study, we investigated the sensitivity of GABAergic short-term plasticity to the selective P- and P/Q-type calcium channels blocker omega-agatoxin-IVA. To block the P-type channels we used 30 nM of this toxin and 200 nM of the toxin was used to block the P/Q channel types. The evoked inhibitory postsynaptic currents (eIPSC) were studied using patch-clamp technique in whole-cell configuration in postsynaptic neuron and local extracellular stimulation of single presynaptic axon by rectangular pulse. The present data show that the contribution of P- and P/Q-types channels to GABAergic synaptic transmission in cultured hippocampal neurons are 30% and 45%, respectively. It was shown that the mediate contribution of the P- and P/Q-types channels to the amplitudes of eIPSC is different to every discovered neuron. It means that distribution of these channels is non-uniform. To study the short-term plasticity of inhibitory synaptic transmission, axons of presynaptic neurons were paired-pulse stimulated with the interpulse interval of 150 ms. Neurons demonstrated both the depression and facilitation. The application of 30 nM and 200 nM of the blocker decreased the depression and increased facilitation to 8% and 11%, respectively. In addition, we found that the mediate contribution of the P- and P/Q-types channels to realization of synaptic transmission after the second stimuli is 4% less compared to that after the first one. Therefore, blocking of both P- and P/Q-types calcium channels can change the efficiency of synaptic transmission. In this instance it facilitates realization of the transmission via decreased depression or increased facilitation. These results confirm that the P- and P/Q-types calcium channels are involved in regulation of the short-term inhibitory synaptic plasticity in cultured hippocampal neurons.

Synchronous circadian voltage rhythms with asynchronous calcium rhythms in the suprachiasmatic nucleus

PubMed Central

Enoki, Ryosuke; Oda, Yoshiaki; Mieda, Michihiro; Ono, Daisuke; Honma, Sato; Honma, Ken-ichi

2017-01-01

The suprachiasmatic nucleus (SCN), the master circadian clock, contains a network composed of multiple types of neurons which are thought to form a hierarchical and multioscillator system. The molecular clock machinery in SCN neurons drives membrane excitability and sends time cue signals to various brain regions and peripheral organs. However, how and at what time of the day these neurons transmit output signals remain largely unknown. Here, we successfully visualized circadian voltage rhythms optically for many days using a genetically encoded voltage sensor, ArcLightD. Unexpectedly, the voltage rhythms are synchronized across the entire SCN network of cultured slices, whereas simultaneously recorded Ca2+ rhythms are topologically specific to the dorsal and ventral regions. We further found that the temporal order of these two rhythms is cell-type specific: The Ca2+ rhythms phase-lead the voltage rhythms in AVP neurons but Ca2+ and voltage rhythms are nearly in phase in VIP neurons. We confirmed that circadian firing rhythms are also synchronous and are coupled with the voltage rhythms. These results indicate that SCN networks with asynchronous Ca2+ rhythms produce coherent voltage rhythms. PMID:28270612

Statistical identification of stimulus-activated network nodes in multi-neuron voltage-sensitive dye optical recordings.

PubMed

Fathiazar, Elham; Anemuller, Jorn; Kretzberg, Jutta

2016-08-01

Voltage-Sensitive Dye (VSD) imaging is an optical imaging method that allows measuring the graded voltage changes of multiple neurons simultaneously. In neuroscience, this method is used to reveal networks of neurons involved in certain tasks. However, the recorded relative dye fluorescence changes are usually low and signals are superimposed by noise and artifacts. Therefore, establishing a reliable method to identify which cells are activated by specific stimulus conditions is the first step to identify functional networks. In this paper, we present a statistical method to identify stimulus-activated network nodes as cells, whose activities during sensory network stimulation differ significantly from the un-stimulated control condition. This method is demonstrated based on voltage-sensitive dye recordings from up to 100 neurons in a ganglion of the medicinal leech responding to tactile skin stimulation. Without relying on any prior physiological knowledge, the network nodes identified by our statistical analysis were found to match well with published cell types involved in tactile stimulus processing and to be consistent across stimulus conditions and preparations.

High temperature sensitivity is intrinsic to voltage-gated potassium channels

PubMed Central

Yang, Fan; Zheng, Jie

2014-01-01

Temperature-sensitive transient receptor potential (TRP) ion channels are members of the large tetrameric cation channels superfamily but are considered to be uniquely sensitive to heat, which has been presumed to be due to the existence of an unidentified temperature-sensing domain. Here we report that the homologous voltage-gated potassium (Kv) channels also exhibit high temperature sensitivity comparable to that of TRPV1, which is detectable under specific conditions when the voltage sensor is functionally decoupled from the activation gate through either intrinsic mechanisms or mutations. Interestingly, mutations could tune Shaker channel to be either heat-activated or heat-deactivated. Therefore, high temperature sensitivity is intrinsic to both TRP and Kv channels. Our findings suggest important physiological roles of heat-induced variation in Kv channel activities. Mechanistically our findings indicate that temperature-sensing TRP channels may not contain a specialized heat-sensor domain; instead, non-obligatory allosteric gating permits the intrinsic heat sensitivity to drive channel activation, allowing temperature-sensitive TRP channels to function as polymodal nociceptors. DOI: http://dx.doi.org/10.7554/eLife.03255.001 PMID:25030910

Sulfhydryl oxidation modifies the calcium dependence of ryanodine-sensitive calcium channels of excitable cells.

PubMed Central

Marengo, J J; Hidalgo, C; Bull, R

1998-01-01

The calcium dependence of ryanodine-sensitive single calcium channels was studied after fusing with planar lipid bilayers sarcoendoplasmic reticulum vesicles isolated from excitable tissues. Native channels from mammalian or amphibian skeletal muscle displayed three different calcium dependencies, cardiac (C), mammalian skeletal (MS), and low fractional open times (low Po), as reported for channels from brain cortex. Native channels from cardiac muscle presented only the MS and C dependencies. Channels with the MS or low Po behaviors showed bell-shaped calcium dependencies, but the latter had fractional open times of <0.1 at all [Ca2+]. Channels with C calcium dependence were activated by [Ca2+] < 10 microM and were not inhibited by increasing cis [Ca2+] up to 0.5 mM. After oxidation with 2,2'-dithiodipyridine or thimerosal, channels with low Po or MS dependencies increased their activity. These channels modified their calcium dependencies sequentially, from low Po to MS and C, or from MS to C. Reduction with glutathione of channels with C dependence (native or oxidized) decreased their fractional open times in 0.5 mM cis [Ca2+], from near unity to 0.1-0.3. These results show that all native channels displayed at least two calcium dependencies regardless of their origin, and that these changed after treatment with redox reagents. PMID:9512024

Photocontrol of Voltage-Gated Ion Channel Activity by Azobenzene Trimethylammonium Bromide in Neonatal Rat Cardiomyocytes

PubMed Central

Frolova, Sheyda R.; Gaiko, Olga; Tsvelaya, Valeriya A.; Pimenov, Oleg Y.; Agladze, Konstantin I.

2016-01-01

The ability of azobenzene trimethylammonium bromide (azoTAB) to sensitize cardiac tissue excitability to light was recently reported. The dark, thermally relaxed trans- isomer of azoTAB suppressed spontaneous activity and excitation propagation speed, whereas the cis- isomer had no detectable effect on the electrical properties of cardiomyocyte monolayers. As the membrane potential of cardiac cells is mainly controlled by activity of voltage-gated ion channels, this study examined whether the sensitization effect of azoTAB was exerted primarily via the modulation of voltage-gated ion channel activity. The effects of trans- and cis- isomers of azoTAB on voltage-dependent sodium (INav), calcium (ICav), and potassium (IKv) currents in isolated neonatal rat cardiomyocytes were investigated using the whole-cell patch-clamp technique. The experiments showed that azoTAB modulated ion currents, causing suppression of sodium (Na+) and calcium (Ca2+) currents and potentiation of net potassium (K+) currents. This finding confirms that azoTAB-effect on cardiac tissue excitability do indeed result from modulation of voltage-gated ion channels responsible for action potential. PMID:27015602

A Rationally Designed, General Strategy for Membrane Orientation of Photoinduced Electron Transfer-Based Voltage-Sensitive Dyes.

PubMed

Kulkarni, Rishikesh U; Yin, Hang; Pourmandi, Narges; James, Feroz; Adil, Maroof M; Schaffer, David V; Wang, Yi; Miller, Evan W

2017-02-17

Voltage imaging with fluorescent dyes offers promise for interrogating the complex roles of membrane potential in coordinating the activity of neurons in the brain. Yet, low sensitivity often limits the broad applicability of optical voltage indicators. In this paper, we use molecular dynamics (MD) simulations to guide the design of new, ultrasensitive fluorescent voltage indicators that use photoinduced electron transfer (PeT) as a voltage-sensing switch. MD simulations predict an approximately 16% increase in voltage sensitivity resulting purely from improved alignment of dye with the membrane. We confirm this theoretical finding by synthesizing 9 new voltage-sensitive (VoltageFluor, or VF) dyes and establishing that all of them display the expected improvement of approximately 19%. This synergistic outworking of theory and experiment enabled computational and theoretical estimation of VF dye orientation in lipid bilayers and has yielded the most sensitive PeT-based VF dye to date. We use this new voltage indicator to monitor voltage spikes in neurons from rat hippocampus and human pluripotent-stem-cell-derived dopaminergic neurons.

Calcium responses to synaptically activated bursts of action potentials and their synapse-independent replay in cultured networks of hippocampal neurons.

PubMed

Bengtson, C Peter; Kaiser, Martin; Obermayer, Joshua; Bading, Hilmar

2013-07-01

Both synaptic N-methyl-d-aspartate (NMDA) receptors and voltage-operated calcium channels (VOCCs) have been shown to be critical for nuclear calcium signals associated with transcriptional responses to bursts of synaptic input. However the direct contribution to nuclear calcium signals from calcium influx through NMDA receptors and VOCCs has been obscured by their concurrent roles in action potential generation and synaptic transmission. Here we compare calcium responses to synaptically induced bursts of action potentials with identical bursts devoid of any synaptic contribution generated using the pre-recorded burst as the voltage clamp command input to replay the burst in the presence of blockers of action potentials or ionotropic glutamate receptors. Synapse independent replays of bursts produced nuclear calcium responses with amplitudes around 70% of their original synaptically generated signals and were abolished by the L-type VOCC blocker, verapamil. These results identify a major direct source of nuclear calcium from local L-type VOCCs whose activation is boosted by NMDA receptor dependent depolarization. The residual component of synaptically induced nuclear calcium signals which was both VOCC independent and NMDA receptor dependent showed delayed kinetics consistent with a more distal source such as synaptic NMDA receptors or internal stores. The dual requirement of NMDA receptors and L-type VOCCs for synaptic activity-induced nuclear calcium dependent transcriptional responses most likely reflects a direct somatic calcium influx from VOCCs whose activation is amplified by synaptic NMDA receptor-mediated depolarization and whose calcium signal is boosted by a delayed input from distal calcium sources mostly likely entry through NMDA receptors and release from internal stores. This article is part of a Special Issue entitled: 12th European Symposium on Calcium. Copyright © 2013 Elsevier B.V. All rights reserved.

Direct interaction of CaVβ with actin up-regulates L-type calcium currents in HL-1 cardiomyocytes.

PubMed

Stölting, Gabriel; de Oliveira, Regina Campos; Guzman, Raul E; Miranda-Laferte, Erick; Conrad, Rachel; Jordan, Nadine; Schmidt, Silke; Hendriks, Johnny; Gensch, Thomas; Hidalgo, Patricia

2015-02-20

Expression of the β-subunit (CaVβ) is required for normal function of cardiac L-type calcium channels, and its up-regulation is associated with heart failure. CaVβ binds to the α1 pore-forming subunit of L-type channels and augments calcium current density by facilitating channel opening and increasing the number of channels in the plasma membrane, by a poorly understood mechanism. Actin, a key component of the intracellular trafficking machinery, interacts with Src homology 3 domains in different proteins. Although CaVβ encompasses a highly conserved Src homology 3 domain, association with actin has not yet been explored. Here, using co-sedimentation assays and FRET experiments, we uncover a direct interaction between CaVβ and actin filaments. Consistently, single-molecule localization analysis reveals streaklike structures composed by CaVβ2 that distribute over several micrometers along actin filaments in HL-1 cardiomyocytes. Overexpression of CaVβ2-N3 in HL-1 cells induces an increase in L-type current without altering voltage-dependent activation, thus reflecting an increased number of channels in the plasma membrane. CaVβ mediated L-type up-regulation, and CaVβ-actin association is prevented by disruption of the actin cytoskeleton with cytochalasin D. Our study reveals for the first time an interacting partner of CaVβ that is directly involved in vesicular trafficking. We propose a model in which CaVβ promotes anterograde trafficking of the L-type channels by anchoring them to actin filaments in their itinerary to the plasma membrane. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Calcium current in isolated neonatal rat ventricular myocytes.

PubMed Central

Cohen, N M; Lederer, W J

1987-01-01

1. Calcium currents (ICa) from neonatal rat ventricular heart muscle cells grown in primary culture were examined using the 'whole-cell' voltage-clamp technique (Hamill, Marty, Neher, Sakmann & Sigworth, 1981). Examination of ICa was limited to one calcium channel type, 'L' type (Nilius, Hess, Lansman & Tsien, 1985), by appropriate voltage protocols. 2. We measured transient and steady-state components of ICa, and could generally describe ICa in terms of the steady-state activation (d infinity) and inactivation (f infinity) parameters. 3. We observed that the reduction of ICa by the calcium channel antagonist D600 can be explained by both a shift of d infinity to more positive potentials as well as a slight reduction of ICa conductance. D600 did not significantly alter either the rate of inactivation of ICa or the voltage dependence of f infinity. 4. The calcium channel modulator BAY K8644 shifted both d infinity and f infinity to more negative potentials. Additionally, BAY K8644 increased the rate of inactivation at potentials between +5 and +55 mV. Furthermore, BAY K8644 also increased ICa conductance, a change consistent with a promotion of 'mode 2' calcium channel activity (Hess, Lansman & Tsien, 1984). 5. We conclude that, as predicted by d infinity and f infinity, there is a significant steady-state component of ICa ('window current') at plateau potentials in neonatal rat heart cells. Modulation of the steady-state and transient components of ICa by various agents can be attributed both to specific alterations in d infinity and f infinity and to more complicated alterations in the mode of calcium channel activity. PMID:2451004

Association between Genetic Polymorphisms in Cav2.3 (R-type) Ca2+ Channels and Fentanyl Sensitivity in Patients Undergoing Painful Cosmetic Surgery

PubMed Central

Fukuda, Ken-ichi; Kasai, Shinya; Hasegawa, Junko; Hayashida, Masakazu; Minami, Masabumi; Ikeda, Kazutaka

2013-01-01

Individual differences in the sensitivity to fentanyl, a widely used opioid analgesic, lead to different proper doses of fentanyl, which can hamper effective pain treatment. Voltage-activated Ca2+ channels (VACCs) play a crucial role in the nervous system by controlling membrane excitability and calcium signaling. Cav2.3 (R-type) VACCs have been especially thought to play critical roles in pain pathways and the analgesic effects of opioids. However, unknown is whether single-nucleotide polymorphisms (SNPs) of the human CACNA1E (calcium channel, voltage-dependent, R type, alpha 1E subunit) gene that encodes Cav2.3 VACCs influence the analgesic effects of opioids. Thus, the present study examined associations between fentanyl sensitivity and SNPs in the human CACNA1E gene in 355 Japanese patients who underwent painful orofacial cosmetic surgery, including bone dissection. We first conducted linkage disequilibrium (LD) analyses of 223 SNPs in a region that contains the CACNA1E gene using genomic samples from 100 patients, and a total of 13 LD blocks with 42 Tag SNPs were observed within and around the CACNA1E gene region. In the preliminary study using the same 100 genomic samples, only the rs3845446 A/G SNP was significantly associated with perioperative fentanyl use among these 42 Tag SNPs. In a confirmatory study using the other 255 genomic samples, this SNP was also significantly associated with perioperative fentanyl use. Thus, we further analyzed associations between genotypes of this SNP and all of the clinical data using a total of 355 samples. The rs3845446 A/G SNP was associated with intraoperative fentanyl use, 24 h postoperative fentanyl requirements, and perioperative fentanyl use. Subjects who carried the minor G allele required significantly less fentanyl for pain control compared with subjects who did not carry this allele. Although further validation is needed, the present findings show the possibility of the involvement of CACNA1E gene polymorphisms

Extended-gate-type IGZO electric-double-layer TFT immunosensor with high sensitivity and low operation voltage

NASA Astrophysics Data System (ADS)

Liang, Lingyan; Zhang, Shengnan; Wu, Weihua; Zhu, Liqiang; Xiao, Hui; Liu, Yanghui; Zhang, Hongliang; Javaid, Kashif; Cao, Hongtao

2016-10-01

An immunosensor is proposed based on the indium-gallium-zinc-oxide (IGZO) electric-double-layer thin-film transistor (EDL TFT) with a separating extended gate. The IGZO EDL TFT has a field-effect mobility of 24.5 cm2 V-1 s-1 and an operation voltage less than 1.5 V. The sensors exhibit the linear current response to label-free target immune molecule in the concentrations ranging from 1.6 to 368 × 10-15 g/ml with a detection limit of 1.6 × 10-15 g/ml (0.01 fM) under an ultralow operation voltage of 0.5 V. The IGZO TFT component demonstrates a consecutive assay stability and recyclability due to the unique structure with the separating extended gate. With the excellent electrical properties and the potential for plug-in-card-type multifunctional sensing, extended-gate-type IGZO EDL TFTs can be promising candidates for the development of a label-free biosensor for public health applications.

Recording membrane potential changes through photoacoustic voltage sensitive dye

NASA Astrophysics Data System (ADS)

Zhang, Haichong K.; Kang, Jeeun; Yan, Ping; Abou, Diane S.; Le, Hanh N. D.; Thorek, Daniel L. J.; Kang, Jin U.; Gjedde, Albert; Rahmim, Arman; Wong, Dean F.; Loew, Leslie M.; Boctor, Emad M.

2017-03-01

Monitoring of the membrane potential is possible using voltage sensitive dyes (VSD), where fluorescence intensity changes in response to neuronal electrical activity. However, fluorescence imaging is limited by depth of penetration and high scattering losses, which leads to low sensitivity in vivo systems for external detection. In contrast, photoacoustic (PA) imaging, an emerging modality, is capable of deep tissue, noninvasive imaging by combining near infrared light excitation and ultrasound detection. In this work, we develop the theoretical concept whereby the voltage-dependent quenching of dye fluorescence leads to a reciprocal enhancement of PA intensity. Based on this concept, we synthesized a novel near infrared photoacoustic VSD (PA-VSD) whose PA intensity change is sensitive to membrane potential. In the polarized state, this cyanine-based probe enhances PA intensity while decreasing fluorescence output in a lipid vesicle membrane model. With a 3-9 μM VSD concentration, we measured a PA signal increase in the range of 5.3 % to 18.1 %, and observed a corresponding signal reduction in fluorescence emission of 30.0 % to 48.7 %. A theoretical model successfully accounts for how the experimental PA intensity change depends on fluorescence and absorbance properties of the dye. These results not only demonstrate the voltage sensing capability of the dye, but also indicate the necessity of considering both fluorescence and absorbance spectral sensitivities in order to optimize the characteristics of improved photoacoustic probes. Together, our results demonstrate photoacoustic sensing as a potential new modality for sub-second recording and external imaging of electrophysiological and neurochemical events in the brain.

Threshold voltage tuning in AlGaN/GaN HFETs with p-type Cu2O gate synthesized by magnetron reactive sputtering

NASA Astrophysics Data System (ADS)

Wang, Lei; Li, Liuan; Xie, Tian; Wang, Xinzhi; Liu, Xinke; Ao, Jin-Ping

2018-04-01

In present study, copper oxide films were prepared at different sputtering powers (10-100 W) using magnetron reactive sputtering. The crystalline structure, surface morphologies, composition, and optical band gap of the as-grown films are dependent on sputtering power. As the sputtering power decreasing from 100 to 10 W, the composition of films changed from CuO to quasi Cu2O domination. Moreover, when the sputtering power is 10 W, a relative high hole carrier density and high-surface-quality quasi Cu2O thin film can be achieved. AlGaN/GaN HFETs were fabricated with the optimized p-type quasi Cu2O film as gate electrode, the threshold voltage of the device shows a 0.55 V positive shift, meanwhile, a lower gate leakage current, a higher ON/OFF drain current ratio of ∼108, a higher electron mobility (1465 cm2/Vs), and a lower subthreshold slope of 74 mV/dec are also achieved, compared with the typical Ni/Au-gated HFETs. Therefore, Cu2O have a great potential to develop high performance p-type gate AlGaN/GaN HFETs.

Reciprocal Regulation of Reactive Oxygen Species and Phospho-CREB Regulates Voltage Gated Calcium Channel Expression during Mycobacterium tuberculosis Infection

PubMed Central

Selvakumar, Arti; Antony, Cecil; Singhal, Jhalak; Tiwari, Brijendra K.; Singh, Yogendra; Natarajan, Krishnamurthy

2014-01-01

Our previous work has demonstrated the roles played by L-type Voltage Gated Calcium Channels (VGCC) in regulating Mycobacterium tuberculosis (M. tb) survival and pathogenesis. Here we decipher mechanisms and pathways engaged by the pathogen to regulate VGCC expression in macrophages. We show that M. tb and its antigen Rv3416 use phospho-CREB (pCREB), Reactive Oxygen Species (ROS), Protein Kinase C (PKC) and Mitogen Activated Protein Kinase (MAPK) to modulate VGCC expression in macrophages. siRNA mediated knockdown of MyD88, IRAK1, IRAK2 or TRAF6 significantly inhibited antigen mediated VGCC expression. Inhibiting Protein Kinase C (PKC) or MEK-ERK1/2 further increased VGCC expression. Interestingly, inhibiting intracellular calcium release upregulated antigen mediated VGCC expression, while inhibiting extracellular calcium influx had no significant effect. siRNA mediated knockdown of transcription factors c-Jun, SOX5 and CREB significantly inhibited Rv3416 mediated VGCC expression. A dynamic reciprocal cross-regulation between ROS and pCREB was observed that in turn governed VGCC expression with ROS playing a limiting role in the process. Further dissection of the mechanisms such as the interplay between ROS and pCREB would improve our understanding of the regulation of VGCC expression during M. tb infection. PMID:24797940

Mechanisms of pyrethroid insecticide-induced stimulation of calcium influx in neocortical neurons

EPA Science Inventory

Pyrethroid insecticides bind to voltage-gated sodium channels (VGSCs) and modify their gating kinetics, thereby disrupting neuronal function. Pyrethroids have also been reported to alter the function of other channel types, including activation of voltage-gated Ca2+ calcium chann...

Modulation of low-voltage-activated T-type Ca²⁺ channels.

PubMed

Zhang, Yuan; Jiang, Xinghong; Snutch, Terrance P; Tao, Jin

2013-07-01

Low-voltage-activated T-type Ca²⁺ channels contribute to a wide variety of physiological functions, most predominantly in the nervous, cardiovascular and endocrine systems. Studies have documented the roles of T-type channels in sleep, neuropathic pain, absence epilepsy, cell proliferation and cardiovascular function. Importantly, novel aspects of the modulation of T-type channels have been identified over the last few years, providing new insights into their physiological and pathophysiological roles. Although there is substantial literature regarding modulation of native T-type channels, the underlying molecular mechanisms have only recently begun to be addressed. This review focuses on recent evidence that the Ca(v)3 subunits of T-type channels, Ca(v)3.1, Ca(v)3.2 and Ca(v)3.3, are differentially modulated by a multitude of endogenous ligands including anandamide, monocyte chemoattractant protein-1, endostatin, and redox and oxidizing agents. The review also provides an overview of recent knowledge gained concerning downstream pathways involving G-protein-coupled receptors. This article is part of a Special Issue entitled: Calcium channels. Copyright © 2012 Elsevier B.V. All rights reserved.

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)

Recent advances of high voltage AlGaN/GaN power HFETs

NASA Astrophysics Data System (ADS)

Uemoto, Yasuhiro; Ueda, Tetsuzo; Tanaka, Tsuyoshi; Ueda, Daisuke

2009-02-01

We review our recent advances of GaN-based high voltage power transistors. These are promising owing to low on-state resistance and high breakdown voltage taking advantages of superior material properties. However, there still remain a couple of technical issues to be solved for the GaN devices to replace the existing Si-based power devices. The most critical issue is to achieve normally-off operation which is strongly desired for the safety operation, however, it has been very difficult because of the built-in polarization electric field. Our new device called GIT (Gate Injection Transistor) utilizing conductivity modulation successfully achieves the normally-off operation keeping low on-state resistance. The fabricated GIT on a Si substrate exhibits threshold voltage of +1.0V. The obtained on-state resistance and off-state breakdown voltage were 2.6mΩ•cm2 and 800V, respectively. Remaining technical issue is to further increase the breakdown voltage. So far, the reported highest off-state breakdown voltage of AlGaN/GaN HFETs has been 1900V. Overcoming these issues by a novel device structure, we have demonstrated the world highest breakdown voltages of 10400V using thick poly-crystalline AlN as a passivation film and Via-holes through sapphire which enable very efficient layout of the lateral HFET array avoiding any undesired breakdown of passivation films. Since conventional wet or dry etching cannot be used for chemically stable sapphire, high power pulsed laser is used to form the via-holes. The presented GaN power devices demonstrate that GaN is advantageous for high voltage power switching applications replacing currently used Si-based power MOSFETs and IGBTs.

A molecule-based genetic association approach implicates a range of voltage-gated calcium channels associated with schizophrenia.

PubMed

Li, Wen; Fan, Chun Chieh; Mäki-Marttunen, Tuomo; Thompson, Wesley K; Schork, Andrew J; Bettella, Francesco; Djurovic, Srdjan; Dale, Anders M; Andreassen, Ole A; Wang, Yunpeng

2018-06-01

Traditional genome-wide association studies (GWAS) have successfully detected genetic variants associated with schizophrenia. However, only a small fraction of heritability can be explained. Gene-set/pathway-based methods can overcome limitations arising from single nucleotide polymorphism (SNP)-based analysis, but most of them place constraints on size which may exclude highly specific and functional sets, like macromolecules. Voltage-gated calcium (Ca v ) channels, belonging to macromolecules, are composed of several subunits whose encoding genes are located far away or even on different chromosomes. We combined information about such molecules with GWAS data to investigate how functional channels associated with schizophrenia. We defined a biologically meaningful SNP-set based on channel structure and performed an association study by using a validated method: SNP-set (sequence) kernel association test. We identified eight subtypes of Ca v channels significantly associated with schizophrenia from a subsample of published data (N = 56,605), including the L-type channels (Ca v 1.1, Ca v 1.2, Ca v 1.3), P-/Q-type Ca v 2.1, N-type Ca v 2.2, R-type Ca v 2.3, T-type Ca v 3.1, and Ca v 3.3. Only genes from Ca v 1.2 and Ca v 3.3 have been implicated by the largest GWAS (N = 82,315). Each subtype of Ca v channels showed relatively high chip heritability, proportional to the size of its constituent gene regions. The results suggest that abnormalities of Ca v channels may play an important role in the pathophysiology of schizophrenia and these channels may represent appropriate drug targets for therapeutics. Analyzing subunit-encoding genes of a macromolecule in aggregate is a complementary way to identify more genetic variants of polygenic diseases. This study offers the potential of power for discovery the biological mechanisms of schizophrenia. © 2018 Wiley Periodicals, Inc.

Activation of G-protein-coupled receptor 30 increases T-type calcium currents in trigeminal ganglion neurons via the cholera toxin-sensitive protein kinase A pathway.

PubMed

Yue, Jingxia; Zhang, Yi; Li, Xuemin; Gong, Shan; Tao, Jin; Jiang, Xinghong

2014-11-01

G protein-coupled receptor 30 (GPR30) is a seven transmembrane domain G protein coupled receptor. In our study, GPR30 expression was found in trigeminal ganglia (TG) in mice, detected by RT-PCR and western blotting. We examined the effects of GPR30 activation on T-type calcium channels using GPR30-specific compound 1 (G-1), a GPR30-selective agonist, in TG neurons and demonstrated that G-1 induced an increase in T-type calcium channel currents (T-currents) in TGs. Intracellular infusion of GDP-β-S and pre-treatment of the neurons with cholera toxin (CTX) blocked the effects of G-1, suggesting that the G(s)-protein was involved. Intracellular application of the protein kinase A (PKA) inhibitor PKI 6-22 or pretreatment of the neurons with H89 abolished G-1 -induced enhancement of T-currents in TG neurons. However, incubation with PKC inhibitor elicited no such effects. In conclusion, our study shows that activation of GPR30 by G-1 increases T-currents via the CTX-sensitive and PKA-dependent pathway.

p -n Junction Rectifying Characteristics of Purely n -Type GaN-Based Structures

NASA Astrophysics Data System (ADS)

Zuo, P.; Jiang, Y.; Ma, Z. G.; Wang, L.; Zhao, B.; Li, Y. F.; Yue, G.; Wu, H. Y.; Yan, H. J.; Jia, H. Q.; Wang, W. X.; Zhou, J. M.; Sun, Q.; Liu, W. M.; Ji, An-Chun; Chen, H.

2017-08-01

The GaN-based p -n junction rectifications are important in the development of high-power electronics. Here, we demonstrate that p -n junction rectifying characteristics can be realized with pure n -type structures by inserting an (In,Ga)N quantum well into the GaN /(Al ,Ga )N /GaN double heterostructures. Unlike the usual barriers, the insertion of an (In,Ga)N quantum well, which has an opposite polarization field to that of the (Al,Ga)N barrier, tailors significantly the energy bands of the system. The lifted energy level of the GaN spacer and the formation of the (In ,Ga )N /GaN interface barrier can improve the reverse threshold voltage and reduce the forward threshold voltage simultaneously, forming the p -n junction rectifying characteristics.

Human autoantibodies specific for the α1A calcium channel subunit reduce both P-type and Q-type calcium currents in cerebellar neurons

PubMed Central

Pinto, Ashwin; Gillard, Samantha; Moss, Fraser; Whyte, Kathryn; Brust, Paul; Williams, Mark; Stauderman, Ken; Harpold, Michael; Lang, Bethan; Newsom-Davis, John; Bleakman, David; Lodge, David; Boot, John

1998-01-01

The pharmacological properties of voltage-dependent calcium channel (VDCC) subtypes appear mainly to be determined by the α1 pore-forming subunit but, whether P-and Q-type VDCCs are encoded by the same α1 gene presently is unresolved. To investigate this, we used IgG antibodies to presynaptic VDCCs at motor nerve terminals that underlie muscle weakness in the autoimmune Lambert–Eaton myasthenic syndrome (LEMS). We first studied their action on changes in intracellular free Ca2+ concentration [Ca2+]i in human embryonic kidney (HEK293) cell lines expressing different combinations of human recombinant VDCC subunits. Incubation for 18 h with LEMS IgG (2 mg/ml) caused a significant dose-dependent reduction in the K+-stimulated [Ca2+]i increase in the α1A cell line but not in the α1B, α1C, α1D, and α1E cell lines, establishing the α1A subunit as the target for these autoantibodies. Exploiting this specificity, we incubated cultured rat cerebellar neurones with LEMS IgG and observed a reduction in P-type current in Purkinje cells and both P- and Q-type currents in granule cells. These data are consistent with the hypothesis that the α1A gene encodes for the pore-forming subunit of both P-type and Q-type VDCCs. PMID:9653186

Fast calcium transients translate the distribution and conduction of neural activity in different regions of a single sensory neuron.

PubMed

Purali, Nuhan

2017-09-01

In the present study, cytosolic calcium concentration changes were recorded in response to various forms of excitations, using the fluorescent calcium indicator dye OG-BAPTA1 together with the current or voltage clamp methods in stretch receptor neurons of crayfish. A single action potential evoked a rise in the resting calcium level in the axon and axonal hillock, whereas an impulse train or a large saturating current injection would be required to evoke an equivalent response in the dendrite region. Under voltage clamp conditions, amplitude differences between axon and dendrite responses vanished completely. The fast activation time and the modulation of the response by extracellular calcium concentration changes indicated that the evoked calcium transients might be mediated by calcium entry into the cytosol through a voltage-gated calcium channel. The decay of the responses was slow and sensitive to extracellular sodium and calcium concentrations as well as exposure to 1-10 mM NiCl 2 and 10-500 µM lanthanum. Thus, a sodium calcium exchanger and a calcium ATPase might be responsible for calcium extrusion from the cytosol. Present results indicate that the calcium indicator OG-BAPTA1 might be an efficient but indirect way of monitoring regional membrane potential differences in a single neuron.

Differential inhibition of N and P/Q Ca2+ currents by 5-HT1A and 5-HT1D receptors in spinal neurons of Xenopus larvae

PubMed Central

Sun, Qian-Quan; Dale, Nicholas

1998-01-01

In whole-cell patch clamp recordings made from non-sensory neurons acutely isolated from the spinal cord of Xenopus (stage 40–42) larvae, two forms of inhibition of the high voltage-activated (HVA) Ca2+ currents were produced by 5-HT. One was voltage dependent and associated with both slowing of the activation kinetics and shifting of the voltage dependence of the HVA currents. This inhibition was relieved by strong depolarizing prepulses. A second form of inhibition was neither associated with slowing of the activation kinetics nor relieved by depolarizing prepulses and was thus voltage independent. In all neurons examined, 5-HT (1 μM) reversibly reduced 34 ± 1.6 % (n = 102) of the HVA Ca2+ currents. In about 40 % of neurons, the inhibition was totally voltage independent. In another 5 %, the inhibition was totally voltage dependent. In the remaining neurons, inhibition was only partially (by around 40 %) relieved by a large depolarizing prepulse, suggesting that in these, the inhibition consisted of both voltage-dependent and -independent components. By using selective channel blockers, we found that 5-HT acted on both N- and P/Q-type channels. However, whereas the inhibition of P/Q-type currents was only voltage independent, the inhibition of N-type currents had both voltage-dependent and -independent components. The effects of 5-HT on HVA Ca2+ currents were mediated by 5-HT1A and 5-HT1D receptors. The 5-HT1A receptors not only preferentially caused voltage-independent inhibition, but did so by acting mainly on the ω-agatoxin-IVA-sensitive Ca2+ channels. In contrast, the 5-HT1D receptor produced both voltage-dependent and -independent inhibition and was preferentially coupled to ω-conotoxin-GVIA sensitive channels. This complexity of modulation may allow fine tuning of transmitter release and calcium signalling in the spinal circuitry of Xenopus larvae. PMID:9625870

μ-Opioid receptor activation inhibits N- and P-type Ca2+ channel currents in magnocellular neurones of the rat supraoptic nucleus

PubMed Central

Soldo, Brandi L; Moises, Hylan C

1998-01-01

The whole-cell voltage-clamp technique was used to examine opioid regulation of Ba2+ currents (IBa) through voltage-sensitive Ca2+ channels in isolated magnocellular supraoptic neurones (MNCs). The effects of local application of μ-, δ- or κ-opioid receptor selective agonists were examined on specific components of high voltage-activated (HVA) IBa, pharmacologically isolated by use of Ca2+ channel-subtype selective antagonists. The μ-opioid receptor selective agonist, DAMGO, suppressed HVA IBa (in 64/71 neurones) in a naloxone-reversible and concentration-dependent manner (EC50 = 170 nm, Emax = 19.5 %). The DAMGO-induced inhibition was rapid in onset, associated with kinetic slowing and voltage dependent, being reversed by strong depolarizing prepulses. Low-voltage activated (LVA) IBa was not modulated by DAMGO. Administration of κ- (U69 593) or δ-selective (DPDPE) opioid receptor agonists did not affect IBa. However, immunostaining of permeabilized MNCs with an antibody specific for κ1-opioid receptors revealed the presence of this opioid receptor subtype in a large number of isolated somata. μ-Opioid-induced inhibition in IBa was largely abolished after blockade of N-type and P-type channel currents by ω-conotoxin GVIA (1 μm) and ω-agatoxin IVA (100 nm), respectively. Quantitation of antagonist effects on DAMGO-induced reductions in IBa revealed that N- and P-type channels contributed roughly equally to the μ-opioid sensitive portion of total IBa. These results indicate that μ-opioid receptors are negatively coupled to N- and P-type Ca2+ channels in the somatodendritic regions of MNCs, possibly via a membrane-delimited G-protein-dependent pathway. They also support a scheme in which opioids may act in part to modulate cellular activity and regulate neurosecretory function by their direct action on the neuroendocrine neurones of the hypothalamic supraoptic neucleus. PMID:9824718

Postsynaptic N-type or P/Q-type calcium channels mediate long-term potentiation by group I metabotropic glutamate receptors in the trigeminal oralis.

PubMed

Weon, Haein; Kim, Tae Wan; Youn, Dong-Ho

2017-11-01

Both N-type and P/Q-type voltage-gated Ca 2+ channels (VGCCs) are involved in the induction of long-term potentiation (LTP), the long-lasting increase of synaptic strength, in the central nervous system. To provide further information on the roles of N-type and P/Q-type VGCCs in the induction of LTP at excitatory synapses of trigeminal primary afferents in the spinal trigeminal subnucleus oralis (Vo), we investigated whether they contribute to the induction of LTP by activation of group I metabotropic glutamate receptors (mGluRs). (S)-3,5-Dihydroxyphenylglycine (DHPG; 10μM for 5min), the group I mGluR agonist, was used to induce LTP of excitatory postsynaptic currents that were evoked in the Vo neurons by stimulating the trigeminal track. Weak blockade of the N-type or P/Q-type VGCCs by ω-conotoxin GVIA or ω-agatoxin IVA, respectively, which inhibited only 20-40% of Ca 2+ currents recorded in isolated trigeminal ganglion neurons but had no effect on the basal excitatory synaptic transmission, completely blocked the induction of LTP. In contrast, stronger blockade of the channels, which inhibited >50% of Ca 2+ currents and about 30% of basal synaptic transmission, resulted in the development of long-term depression (LTD), the long-lasting decrease of synaptic strength. Interestingly, the postsynaptic mechanism of DHPG-induced LTP, which was determined by paired-pulse ratio, disappeared when LTP was blocked, or LTD occurred, while a presynaptic mechanism still remained. Our data suggest that postsynaptic N-type and P/Q-type VGCCs mediate the DHPG-induced LTP at the trigeminal afferent synapses in the Vo. Copyright © 2017 Elsevier Inc. All rights reserved.

The Role of a Coronary Artery Calcium Scan in Type 1 Diabetes

PubMed Central

Eaton, R. Philip; Schade, David S.

2016-01-01

Abstract The coronary artery calcium (CAC) scan has recently emerged as a reproducible noninvasive test to detect asymptomatic atherosclerotic coronary artery disease. It has several advantages over the traditional cardiac stress testing modalities, including lower cost, greater sensitivity for nonobstructing coronary artery lesions, and excellent prognostic value when combined with the Framingham risk parameters. Its chief disadvantage is that it does not identify obstructing coronary artery lesions or noncalcified coronary artery plaque. A CAC scan utilizes a chest computed tomogram and computer software to calculate the amount of calcium in the four main coronary vessels. Calcium is deposited in coronary plaques so that the greater the calcium score, the greater the plaque burden. This, in turn, is the basis for predicting a 10–15-year risk of a cardiovascular event. Individuals with a zero calcium score have a very low 10-year risk of a cardiovascular event. Obtaining a calcium score in a diabetic patient permits rational decisions for prescribing statin therapy. In patients with a zero score, the initiation of statin therapy is not recommended because the 5-year incidence of atherosclerotic cardiovascular disease is so low. In patients with diabetes, it is recommended to repeat the calcium scan in 4–5 years to permit timely therapy in the event that the score becomes positive. Since statins mildly increase coronary calcium as part of the stabilization of plaque, a reduction in the calcium score should not be anticipated. However, progression of the calcium score by more than 15%/year (calculated from a repeat CAC scan) provides additional prognostic information of an indication of progression of atherosclerosis. In summary, the coronary calcium score is a major clinical advance for noninvasively detecting coronary artery disease and managing antiatherosclerotic therapy in type 1 diabetes. PMID:27585206

ZnT-1 enhances the activity and surface expression of T-type calcium channels through activation of Ras-ERK signaling.

PubMed

Mor, Merav; Beharier, Ofer; Levy, Shiri; Kahn, Joy; Dror, Shani; Blumenthal, Daniel; Gheber, Levi A; Peretz, Asher; Katz, Amos; Moran, Arie; Etzion, Yoram

2012-07-15

Zinc transporter-1 (ZnT-1) is a putative zinc transporter that confers cellular resistance from zinc toxicity. In addition, ZnT-1 has important regulatory functions, including inhibition of L-type calcium channels and activation of Raf-1 kinase. Here we studied the effects of ZnT-1 on the expression and function of T-type calcium channels. In Xenopus oocytes expressing voltage-gated calcium channel (CaV) 3.1 or CaV3.2, ZnT-1 enhanced the low-threshold calcium currents (I(caT)) to 182 ± 15 and 167.95 ± 9.27% of control, respectively (P < 0.005 for both channels). As expected, ZnT-1 also enhanced ERK phosphorylation. Coexpression of ZnT-1 and nonactive Raf-1 blocked the ZnT-1-mediated ERK phosphorylation and abolished the ZnT-1-induced augmentation of I(caT). In mammalian cells (Chinese hamster ovary), coexpression of CaV3.1 and ZnT-1 increased the I(caT) to 166.37 ± 6.37% compared with cells expressing CaV3.1 alone (P < 0.01). Interestingly, surface expression measurements using biotinylation or total internal reflection fluorescence microscopy indicated marked ZnT-1-induced enhancement of CaV3.1 surface expression. The MEK inhibitor PD-98059 abolished the ZnT-1-induced augmentation of surface expression of CaV3.1. In cultured murine cardiomyocytes (HL-1 cells), transient exposure to zinc, leading to enhanced ZnT-1 expression, also enhanced the surface expression of endogenous CaV3.1 channels. Consistently, in these cells, endothelin-1, a potent activator of Ras-ERK signaling, enhanced the surface expression of CaV3.1 channels in a PD-98059-sensitive manner. Our findings indicate that ZnT-1 enhances the activity of CaV3.1 and CaV3.2 through activation of Ras-ERK signaling. The augmentation of CaV3.1 currents by Ras-ERK activation is associated with enhanced trafficking of the channel to the plasma membrane.

Low-threshold voltage ultraviolet light-emitting diodes based on (Al,Ga)N metal-insulator-semiconductor structures

NASA Astrophysics Data System (ADS)

Liang, Yu-Han; Towe, Elias

2017-12-01

Al-rich III-nitride-based deep-ultraviolet (UV) (275-320 nm) light-emitting diodes are plagued with a low emission efficiency and high turn-on voltages. We report Al-rich (Al,Ga)N metal-insulator-semiconductor UV light-emitting Schottky diodes with low turn-on voltages of <3 V, which are about half those of typical (Al,Ga)N p-i-n diodes. Our devices use a thin AlN film as the insulator and an n-type Al0.58Ga0.42N film as the semiconductor. To improve the efficiency, we inserted a GaN quantum-well structure between the AlN insulator and the n-type Al x Ga1- x N semiconductor. The benefits of the quantum-well structure include the potential to tune the emission wavelength and the capability to confine carriers for more efficient radiative recombination.

Masters or slaves? Vesicle release machinery and the regulation of presynaptic calcium channels.

PubMed

Jarvis, Scott E; Zamponi, Gerald W

2005-05-01

Calcium entry through presynaptic voltage-gated calcium channels is essential for neurotransmitter release. The two major types of presynaptic calcium channels contain a synaptic protein interaction site that physically interacts with synaptic vesicle release proteins. This is thought to tighten the coupling between the sources of calcium entry and the neurotransmitter release machinery. Conversely, the binding of synaptic proteins to presynaptic calcium channels regulates calcium channel activity. Hence, presynaptic calcium channels act not only as the masters of the synaptic release process, but also as key targets for feedback inhibition.

Ferulic acid relaxed rat aortic, small mesenteric and coronary arteries by blocking voltage-gated calcium channel and calcium desensitization via dephosphorylation of ERK1/2 and MYPT1.

PubMed

Zhou, Zhong-Yan; Xu, Jia-Qi; Zhao, Wai-Rong; Chen, Xin-Lin; Jin, Yu; Tang, Nuo; Tang, Jing-Yi

2017-11-15

Ferulic acid, a natural ingredient presents in several Chinese Materia Medica such as Radix Angelicae Sinensis, has been identified as an important multifunctional and physiologically active small molecule. However, its pharmacological activity in different blood vessel types and underlying mechanisms are unclear. The present study was to investigate the vascular reactivity and the possible action mechanism of FA on aorta, small mesenteric arteries and coronary arteries isolated from Wistar rats. We found FA dose-dependently relieved the contraction of aorta, small mesenteric arteries and coronary arteries induced by different contractors, U46619, phenylephrine (Phe) and KCl. The relaxant effect of FA was not affected by L-NAME (eNOS inhibitor), ODQ (soluble guanylate cyclase inhibitor), and mechanical removal of endothelium in thoracic aortas. The contraction caused by 60mM KCl (60K) was concentration-dependently hindered by FA pretreatment in all three types of arteries. In Ca 2+ -free 60K solution, FA weakened Ca 2+ -related contraction in a concentration dependent manner. And FA relaxed both fluoride and phorbol ester which were PKC, ERK and Rho-kinase activators induced contraction in aortic rings with or without Ca 2+ in krebs solution. Western blotting experiments in A7r5 cells revealed that FA inhibited calcium sensitization via dephosphorylation of ERK1/2 and MYPT1. Furthermore, the relaxation effect of FA was attenuated by verapamil (calcium channel blocker), ERK inhibitor, and fasudil (ROCK inhibitor). These results provide evidence that FA exhibits endothelium-independent vascular relaxant effect in different types of arteries. The molecular mechanism of vasorelaxation activity of FA probably involved calcium channel inhibition and calcium desensitization. Copyright © 2017. Published by Elsevier B.V.

Dynamic modulation of spike timing-dependent calcium influx during corticostriatal upstates

PubMed Central

Evans, R. C.; Maniar, Y. M.

2013-01-01

The striatum of the basal ganglia demonstrates distinctive upstate and downstate membrane potential oscillations during slow-wave sleep and under anesthetic. The upstates generate calcium transients in the dendrites, and the amplitude of these calcium transients depends strongly on the timing of the action potential (AP) within the upstate. Calcium is essential for synaptic plasticity in the striatum, and these large calcium transients during the upstates may control which synapses undergo plastic changes. To investigate the mechanisms that underlie the relationship between calcium and AP timing, we have developed a realistic biophysical model of a medium spiny neuron (MSN). We have implemented sophisticated calcium dynamics including calcium diffusion, buffering, and pump extrusion, which accurately replicate published data. Using this model, we found that either the slow inactivation of dendritic sodium channels (NaSI) or the calcium inactivation of voltage-gated calcium channels (CDI) can cause high calcium corresponding to early APs and lower calcium corresponding to later APs. We found that only CDI can account for the experimental observation that sensitivity to AP timing is dependent on NMDA receptors. Additional simulations demonstrated a mechanism by which MSNs can dynamically modulate their sensitivity to AP timing and show that sensitivity to specifically timed pre- and postsynaptic pairings (as in spike timing-dependent plasticity protocols) is altered by the timing of the pairing within the upstate. These findings have implications for synaptic plasticity in vivo during sleep when the upstate-downstate pattern is prominent in the striatum. PMID:23843436

Voltage-gated calcium channel autoimmune cerebellar degeneration

PubMed Central

McKasson, Marilyn; Clawson, Susan A.; Hill, Kenneth E.; Wood, Blair; Carlson, Noel; Bromberg, Mark; Greenlee, John E.

2016-01-01

Objectives: To describe response to treatment in a patient with autoantibodies against voltage-gated calcium channels (VGCCs) who presented with autoimmune cerebellar degeneration and subsequently developed Lambert-Eaton myasthenic syndrome (LEMS), and to study the effect of the patient's autoantibodies on Purkinje cells in rat cerebellar slice cultures. Methods: Case report and study of rat cerebellar slice cultures incubated with patient VGCC autoantibodies. Results: A 53-year-old man developed progressive incoordination with ataxic speech. Laboratory evaluation revealed VGCC autoantibodies without other antineuronal autoantibodies. Whole-body PET scans 6 and 12 months after presentation detected no malignancy. The patient improved significantly with IV immunoglobulin G (IgG), prednisone, and mycophenolate mofetil, but worsened after IV IgG was halted secondary to aseptic meningitis. He subsequently developed weakness with electrodiagnostic evidence of LEMS. The patient's IgG bound to Purkinje cells in rat cerebellar slice cultures, followed by neuronal death. Reactivity of the patient's autoantibodies with VGCCs was confirmed by blocking studies with defined VGCC antibodies. Conclusions: Autoimmune cerebellar degeneration associated with VGCC autoantibodies may precede onset of LEMS and may improve with immunosuppressive treatment. Binding of anti-VGCC antibodies to Purkinje cells in cerebellar slice cultures may be followed by cell death. Patients with anti-VGCC autoantibodies may be at risk of irreversible neurologic injury over time, and treatment should be initiated early. PMID:27088118

Parathyroid hormone enhances fluid shear-induced [Ca2+]i signaling in osteoblastic cells through activation of mechanosensitive and voltage-sensitive Ca2+ channels

NASA Technical Reports Server (NTRS)

Ryder, K. D.; Duncan, R. L.

2001-01-01

Osteoblasts respond to both fluid shear and parathyroid hormone (PTH) with a rapid increase in intracellular calcium concentration ([Ca2+]i). Because both stimuli modulate the kinetics of the mechanosensitive cation channel (MSCC), we postulated PTH would enhance the [Ca2+]i response to fluid shear by increasing the sensitivity of MSCCs. After a 3-minute preflow at 1 dyne/cm2, MC3T3-E1 cells were subjected to various levels of shear and changes in [Ca2+]i were assessed using Fura-2. Pretreatment with 50 nM bovine PTH(1-34) [bPTH(1-34)] significantly enhanced the shear magnitude-dependent increase in [Ca2+]i. Gadolinium (Gd3+), an MSCC blocker, significantly inhibited the mean peak [Ca2+]i response to shear and shear + bPTH(1-34). Nifedipine (Nif), an L-type voltage-sensitive Ca2+ channel (VSCC) blocker, also significantly reduced the [Ca2+]i response to shear + bPTH(1-34), but not to shear alone, suggesting VSCC activation plays an interactive role in the action of these stimuli together. Activation of either the protein kinase C (PKC) or protein kinase A (PKA) pathways with specific agonists indicated that PKC activation did not alter the Ca2+ response to shear, whereas PKA activation significantly increased the [Ca2+]i response to lower magnitudes of shear. bPTH(1-34), which activates both pathways, induced the greatest [Ca2+]i response at each level of shear, suggesting an interaction of these pathways in this response. These data indicate that PTH significantly enhances the [Ca2+]i response to shear primarily via PKA modulation of the MSCC and VSCC.

Block of high-threshold calcium channels by the synthetic polyamines sFTX-3.3 and FTX-3.3.

PubMed

Norris, T M; Moya, E; Blagbrough, I S; Adams, M E

1996-10-01

A polyamine component of Agelenopsis aperta spider venom designated FTX is reported to be a selective antagonist of P-type calcium channels in the mammalian brain. Consequently, this component has frequently been used as a pharmacological tool to determine the presence, distribution, and function of P-type channels in physiological systems. We describe antagonism of calcium channels by the synthesized polyamine FTX-3.3, which has the proposed structure of natural FTX. We also examined a corresponding polyamine amide, sFTX-3.3. These polyamines are critically evaluated for antagonism of three high-threshold calcium channel subtypes in rat neurons through the use of the whole-cell patch-clamp technique. FTX-3.3 (IC50 = approximately 0.13 mM) is approximately twice as potent as sFTX-3.3 (IC50 = approximately 0.24 mM) against P-type channels and approximately 3-fold more potent against N-type channels (FTX-3.3, IC50 = approximately 0.24 mM; sFTX-3.3, IC50 = approximately 0.70 mM). Both polyamines also block L-type calcium channels with similar potencies. sFTX-3.3 (1 mM) and FTX-3.3 (0.5 mM) typically block 50% and 65% of Bay K8644-enhanced L-type current, respectively. Antagonism of each calcium channel subtype is voltage dependent, with less inhibition of Ba2+ currents at more-positive potentials. These data show that both sFTX-3.3 and FTX-3.3 antagonize P-, N-, and L-type calcium channels in mammalian Purkinje and superior cervical ganglia neurons with similar IC50 values.

Discovery and Development of Calcium Channel Blockers

PubMed Central

Godfraind, Théophile

2017-01-01

In the mid 1960s, experimental work on molecules under screening as coronary dilators allowed the discovery of the mechanism of calcium entry blockade by drugs later named calcium channel blockers. This paper summarizes scientific research on these small molecules interacting directly with L-type voltage-operated calcium channels. It also reports on experimental approaches translated into understanding of their therapeutic actions. The importance of calcium in muscle contraction was discovered by Sidney Ringer who reported this fact in 1883. Interest in the intracellular role of calcium arose 60 years later out of Kamada (Japan) and Heibrunn (USA) experiments in the early 1940s. Studies on pharmacology of calcium function were initiated in the mid 1960s and their therapeutic applications globally occurred in the the 1980s. The first part of this report deals with basic pharmacology in the cardiovascular system particularly in isolated arteries. In the section entitled from calcium antagonists to calcium channel blockers, it is recalled that drugs of a series of diphenylpiperazines screened in vivo on coronary bed precontracted by angiotensin were initially named calcium antagonists on the basis of their effect in depolarized arteries contracted by calcium. Studies on arteries contracted by catecholamines showed that the vasorelaxation resulted from blockade of calcium entry. Radiochemical and electrophysiological studies performed with dihydropyridines allowed their cellular targets to be identified with L-type voltage-operated calcium channels. The modulated receptor theory helped the understanding of their variation in affinity dependent on arterial cell membrane potential and promoted the terminology calcium channel blocker (CCB) of which the various chemical families are introduced in the paper. In the section entitled tissue selectivity of CCBs, it is shown that characteristics of the drug, properties of the tissue, and of the stimuli are important factors of

Cd(2+) sensitivity and permeability of a low voltage-activated Ca(2+) channel with CatSper-like selectivity filter.

PubMed

Garza-López, Edgar; Chávez, Julio César; Santana-Calvo, Carmen; López-González, Ignacio; Nishigaki, Takuya

2016-07-01

CatSper is a sperm-specific Ca(2+) channel that plays an essential role in the male fertility. However, its biophysical properties have been poorly characterized mainly due to its deficient heterologous expression. As other voltage-gated Ca(2+) channels (CaVs), CatSper possesses a conserved Ca(2+)-selective filter motif ([T/S]x[D/E]xW) in the pore region. Interestingly, CatSper conserves four aspartic acids (DDDD) as the negatively charged residues in this motif while high voltage-activated CaVs have four glutamic acids (EEEE) and low voltage-activated CaVs possess two glutamic acids and two aspartic acids (EEDD). Previous studies based on site-directed mutagenesis of L- and T-type channels showed that the number of D seems to have a negative correlation with their cadmium (Cd(2+)) sensitivity. These results suggest that CatSper (DDDD) would have low sensitivity to Cd(2+). To explore Cd(2+)-sensitivity and -permeability of CatSper, we performed two types of experiments: 1) Electrophysiological analysis of heterologously expressed human CaV3.1 channel and three pore mutants (DEDD, EDDD and DDDD), 2) Cd(2+) imaging of human spermatozoa with FluoZin-1. Electrophysiological studies showed a significant increase in Cd(2+) and manganese (Mn(2+)) currents through the CaV3.1 mutants as well as a reduction in the inhibitory effect of Cd(2+) on the Ca(2+) current. In fluorescence imaging with human sperm, we observed an increase in Cd(2+) influx potentiated by progesterone, a potent activator of CatSper. These results support our hypothesis, namely that Cd(2+)-sensitivity and -permeability are related to the absolute number of D in the Ca(2+)-selective filter independently to the type of the Cav channels. Copyright © 2016 Elsevier Ltd. All rights reserved.

NIFLUMIC ACID BLOCKS NATIVE AND RECOMBINANT T-TYPE CHANNELS

PubMed Central

Balderas, E; Arteaga-Tlecuitl, R; Rivera, M; Gomora, JC; Darszon, A

2012-01-01

Voltage-dependent calcium channels are widely distributed in animal cells, including spermatozoa. Calcium is fundamental in many sperm functions such as: motility, capacitation and the acrosome reaction, all essential for fertilization. Pharmacological evidence has suggested T-type calcium channels participate in the acrosome reaction. Niflumic acid (NA), a non-steroidal anti-inflammatory drug commonly used as chloride channel blocker, blocks T-currents in mouse spermatogenic cells and Cl− channels in testicular sperm. Here we examine the mechanism of NA blockade and explore if it can be used to separate the contribution of different CaV3 members previously detected in these cells. Electrophysiological patch-clamp recordings were performed in isolated mouse spermatogenic cells and in HEK cells heterologously expressing CaV3 channels. NA blocks mouse spermatogenic cell T-type currents with an IC50 of 73.5 µM, without major voltage-dependent effects. The NA blockade is more potent in the open and in the inactivated state than in the closed state of the T-type channels. Interestingly, we found that heterologously expressed CaV3.1 and CaV3.3 channels were more sensitive to NA than CaV3.2 channels, and this drug substantially slowed the recovery from inactivation of the three isoforms. Molecular docking modeling of drug-channel binding predicts that NA binds preferentially to the extracellular face of CaV3.1 channels. The biophysical characteristics of mouse spermatogenic cell T-type currents more closely resemble those from heterologously expressed CaV3.1 channels, including their sensitivity to NA. As CaV3.1 null mice maintain their spermatogenic cell T-currents, it is likely that a novel CaV3.2 isoform is responsible for them. PMID:21898399

Characterization and bioactivity of novel calcium antagonists - N-methoxy-benzyl haloperidol quaternary ammonium salt

PubMed Central

Chen, Yi-Cun; Zhu, Wei; Zhong, Shu-Ping; Zheng, Fu-Chun; Gao, Fen-Fei; Zhang, Yan-Mei; Xu, Han; Zheng, Yan-Shan; Shi, Gang-Gang

2015-01-01

BACKGROUND AND PURPOSE Calcium antagonists play an important role in clinical practice. However, most of them have serious side effects. We have synthesized a series of novel calcium antagonists, quaternary ammonium salt derivatives of haloperidol with N-p-methoxybenzyl (X1), N-m-methoxybenzyl (X2) and N-o-methoxybenzyl (X3) groups. The objective of this study was to investigate the bioactivity of these novel calcium antagonists, especially the vasodilation activity and cardiac side-effects. The possible working mechanisms of these haloperidol derivatives were also explored. EXPERIMENTAL APPROACH Novel calcium antagonists were synthesized by amination. Compounds were screened for their activity of vasodilation on isolated thoracic aortic ring of rats. Their cardiac side effects were explored. The patch-clamp, confocal laser microscopy and the computer-fitting molecular docking experiments were employed to investigate the possible working mechanisms of these calcium antagonists. RESULTS The novel calcium antagonists, X1, X2 and X3 showed stronger vasodilation effect and less cardiac side effect than that of classical calcium antagonists. They blocked L-type calcium channels with an potent effect order of X1 > X2 > X3. Consistently, X1, X2 and X3 interacted with different regions of Ca2+-CaM-CaV1.2 with an affinity order of X1 > X2 > X3. CONCLUSIONS The new halopedidol derivatives X1, X2 and X3 are novel calcium antagonists with stronger vasodilation effect and less cardiac side effect. They could have wide clinical application. PMID:26544729

46 CFR 111.79-13 - Different voltages and power types.

Code of Federal Regulations, 2012 CFR

2012-10-01

... 46 Shipping 4 2012-10-01 2012-10-01 false Different voltages and power types. 111.79-13 Section 111.79-13 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) ELECTRICAL ENGINEERING ELECTRIC SYSTEMS-GENERAL REQUIREMENTS Receptacles § 111.79-13 Different voltages and power types. If...

46 CFR 111.79-13 - Different voltages and power types.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 46 Shipping 4 2011-10-01 2011-10-01 false Different voltages and power types. 111.79-13 Section 111.79-13 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) ELECTRICAL ENGINEERING ELECTRIC SYSTEMS-GENERAL REQUIREMENTS Receptacles § 111.79-13 Different voltages and power types. If...

46 CFR 111.79-13 - Different voltages and power types.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 46 Shipping 4 2010-10-01 2010-10-01 false Different voltages and power types. 111.79-13 Section 111.79-13 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) ELECTRICAL ENGINEERING ELECTRIC SYSTEMS-GENERAL REQUIREMENTS Receptacles § 111.79-13 Different voltages and power types. If...

46 CFR 111.79-13 - Different voltages and power types.

Code of Federal Regulations, 2014 CFR

2014-10-01

... 46 Shipping 4 2014-10-01 2014-10-01 false Different voltages and power types. 111.79-13 Section 111.79-13 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) ELECTRICAL ENGINEERING ELECTRIC SYSTEMS-GENERAL REQUIREMENTS Receptacles § 111.79-13 Different voltages and power types. If...

46 CFR 111.79-13 - Different voltages and power types.

Code of Federal Regulations, 2013 CFR

2013-10-01

... 46 Shipping 4 2013-10-01 2013-10-01 false Different voltages and power types. 111.79-13 Section 111.79-13 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) ELECTRICAL ENGINEERING ELECTRIC SYSTEMS-GENERAL REQUIREMENTS Receptacles § 111.79-13 Different voltages and power types. If...

Calcium-controlled conformational choreography in the N-terminal half of adseverin

NASA Astrophysics Data System (ADS)

Chumnarnsilpa, Sakesit; Robinson, Robert C.; Grimes, Jonathan M.; Leyrat, Cedric

2015-09-01

Adseverin is a member of the calcium-regulated gelsolin superfamily of actin-binding proteins. Here we report the crystal structure of the calcium-free N-terminal half of adseverin (iA1-A3) and the Ca2+-bound structure of A3, which reveal structural similarities and differences with gelsolin. Solution small-angle X-ray scattering combined with ensemble optimization revealed a dynamic Ca2+-dependent equilibrium between inactive, intermediate and active conformations. Increasing calcium concentrations progressively shift this equilibrium from a main population of inactive conformation to the active form. Molecular dynamics simulations of iA1-A3 provided insights into Ca2+-induced destabilization, implicating a critical role for the A2 type II calcium-binding site and the A2A3 linker in the activation process. Finally, mutations that disrupt the A1/A3 interface increase Ca2+-independent F-actin severing by A1-A3, albeit at a lower efficiency than observed for gelsolin domains G1-G3. Together, these data address the calcium dependency of A1-A3 activity in relation to the calcium-independent activity of G1-G3.

Inhibition of parathyroid hormone release by maitotoxin, a calcium channel activator

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

Fitzpatrick, L.A.; Yasumoto, T.; Aurbach, G.D.

1989-01-01

Maitotoxin, a toxin derived from a marine dinoflagellate, is a potent activator of voltage-sensitive calcium channels. To further test the hypothesis that inhibition of PTH secretion by calcium is mediated via a calcium channel we studied the effect of maitotoxin on dispersed bovine parathyroid cells. Maitotoxin inhibited PTH release in a dose-dependent fashion, and inhibition was maximal at 1 ng/ml. Chelation of extracellular calcium by EGTA blocked the inhibition of PTH by maitotoxin. Maitotoxin enhanced the effects of the dihydropyridine calcium channel agonist (+)202-791 and increased the rate of radiocalcium uptake in parathyroid cells. Pertussis toxin, which ADP-ribosylates and inactivatesmore » a guanine nucleotide regulatory protein that interacts with calcium channels in the parathyroid cell, did not affect the inhibition of PTH secretion by maitotoxin. Maitotoxin, by its action on calcium channels allows entry of extracellular calcium and inhibits PTH release. Our results suggest that calcium channels are involved in the release of PTH. Inhibition of PTH release by maitotoxin is not sensitive to pertussis toxin, suggesting that maitotoxin may act distal to the site interacting with a guanine nucleotide regulatory protein, or maitotoxin could interact with other ions or second messengers to inhibit PTH release.« less

Activity of Palythoa caribaeorum Venom on Voltage-Gated Ion Channels in Mammalian Superior Cervical Ganglion Neurons.

PubMed

Lazcano-Pérez, Fernando; Castro, Héctor; Arenas, Isabel; García, David E; González-Muñoz, Ricardo; Arreguín-Espinosa, Roberto

2016-05-05

The Zoanthids are an order of cnidarians whose venoms and toxins have been poorly studied. Palythoa caribaeorum is a zoanthid commonly found around the Mexican coastline. In this study, we tested the activity of P. caribaeorum venom on voltage-gated sodium channel (NaV1.7), voltage-gated calcium channel (CaV2.2), the A-type transient outward (IA) and delayed rectifier (IDR) currents of KV channels of the superior cervical ganglion (SCG) neurons of the rat. These results showed that the venom reversibly delays the inactivation process of voltage-gated sodium channels and inhibits voltage-gated calcium and potassium channels in this mammalian model. The compounds responsible for these effects seem to be low molecular weight peptides. Together, these results provide evidence for the potential use of zoanthids as a novel source of cnidarian toxins active on voltage-gated ion channels.

Al0 0.3Ga 0.7N PN diode with breakdown voltage >1600 V

DOE PAGES

Allerman, A. A.; Armstrong, A. M.; Fischer, A. J.; ...

2016-07-21

Demonstration of Al0 0.3Ga 0.7N PN diodes grown with breakdown voltages in excess of 1600 V is reported. The total epilayer thickness is 9.1 μm and was grown by metal-organic vapour-phase epitaxy on 1.3-mm-thick sapphire in order to achieve crack-free structures. A junction termination edge structure was employed to control the lateral electric fields. A current density of 3.5 kA/cm 2 was achieved under DC forward bias and a reverse leakage current <3 nA was measured for voltages <1200 V. The differential on-resistance of 16 mΩ cm 2 is limited by the lateral conductivity of the n-type contact layer requiredmore » by the front-surface contact geometry of the device. An effective critical electric field of 5.9 MV/cm was determined from the epilayer properties and the reverse current–voltage characteristics. To our knowledge, this is the first aluminium gallium nitride (AlGaN)-based PN diode exhibiting a breakdown voltage in excess of 1 kV. Finally, we note that a Baliga figure of merit (V br 2/R spec,on) of 150 MW/cm 2 found is the highest reported for an AlGaN PN diode and illustrates the potential of larger-bandgap AlGaN alloys for high-voltage devices.« less

Variable N-type negative resistance in an injection-gated double-injection diode

NASA Technical Reports Server (NTRS)

Kapoor, A. K.; Henderson, H. T.

1981-01-01

Double-injection (DI) switching devices consist of p+ and n+ contacts (for hole and electron injection, respectively), separated by a near intrinsic semiconductor region containing deep traps. Under proper conditions, these devices exhibit S-type differential negative resistance (DNR) similar to silicon-controlled rectifiers. With the added influence of a p+ gate appropriately placed between the anode (p+) and cathode (n+), the current-voltage characteristic of the device has been manipulated for the first time to exhibit a variable N-type DNR. The anode current and the anode-to-cathode voltage levels at which this N-type DNR is observed can be varied by changing the gate-to-cathode bias. In essence, the classical S-type DI diode can be electronically transformed into an N-type diode. A first-order phenomenological model is proposed for the N-type DNR.

Mechanisms of Pyrethroid Insecticide-Induced Stimulation of Calcium Influx in Neocortical Neurons

PubMed Central

Cao, Zhengyu; Shafer, Timothy J.

2011-01-01

Pyrethroid insecticides bind to voltage-gated sodium channels (VGSCs) and modify their gating kinetics, thereby disrupting neuronal function. Pyrethroids have also been reported to alter the function of other channel types, including activation of voltage-gated calcium channels. Therefore, the present study compared the ability of 11 structurally diverse pyrethroids to evoke Ca2+ influx in primary cultures of mouse neocortical neurons. Nine pyrethroids (tefluthrin, deltamethrin, λ-cyhalothrin, β-cyfluthrin, esfenvalerate, S-bioallethrin, fenpropathrin, cypermethrin, and bifenthrin) produced concentration-dependent elevations in intracellular calcium concentration ([Ca2+]i) in neocortical neurons. Permethrin and resmethrin were without effect on [Ca2+]i. These pyrethroids displayed a range of efficacies on Ca2+ influx; however, the EC50 values for active pyrethroids all were within one order of magnitude. Tetrodotoxin blocked increases in [Ca2+]i caused by all nine active pyrethroids, indicating that the effects depended on VGSC activation. The pathways for deltamethrin- and tefluthrin-induced Ca2+ influx include N-methyl-d-aspartic acid receptors, L-type Ca2+ channels, and reverse mode of operation of the Na+/Ca2+ exchanger inasmuch as antagonists of these sites blocked deltamethrin-induced Ca2+ influx. These data demonstrate that pyrethroids stimulate Ca2+ entry into neurons subsequent to their actions on VGSCs. PMID:20881019

Flexible CMOS-Like Circuits Based on Printed P-Type and N-Type Carbon Nanotube Thin-Film Transistors.

PubMed

Zhang, Xiang; Zhao, Jianwen; Dou, Junyan; Tange, Masayoshi; Xu, Weiwei; Mo, Lixin; Xie, Jianjun; Xu, Wenya; Ma, Changqi; Okazaki, Toshiya; Cui, Zheng

2016-09-01

P-type and n-type top-gate carbon nanotube thin-film transistors (TFTs) can be selectively and simultaneously fabricated on the same polyethylene terephthalate (PET) substrate by tuning the types of polymer-sorted semiconducting single-walled carbon nanotube (sc-SWCNT) inks, along with low temperature growth of HfO 2 thin films as shared dielectric layers. Both the p-type and n-type TFTs show good electrical properties with on/off ratio of ≈10 5 , mobility of ≈15 cm 2 V -1 s -1 , and small hysteresis. Complementary metal oxide semiconductor (CMOS)-like logic gates and circuits based on as-prepared p-type and n-type TFTs have been achieved. Flexible CMOS-like inverters exhibit large noise margin of 84% at low voltage (1/2 V dd = 1.5 V) and maximum voltage gain of 30 at V dd of 1.5 V and low power consumption of 0.1 μW. Both of the noise margin and voltage gain are one of the best values reported for flexible CMOS-like inverters at V dd less than 2 V. The printed CMOS-like inverters work well at 10 kHz with 2% voltage loss and delay time of ≈15 μs. A 3-stage ring oscillator has also been demonstrated on PET substrates and the oscillation frequency of 3.3 kHz at V dd of 1 V is achieved. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Contractile properties and sarcoplasmic reticulum calcium content in type I and type II skeletal muscle fibres in active aged humans

PubMed Central

Lamboley, C R; Wyckelsma, V L; Dutka, T L; McKenna, M J; Murphy, R M; Lamb, G D

2015-01-01

This study examined the contractile properties and sarcoplasmic reticulum (SR) Ca2+ content in mechanically skinned vastus lateralis muscle fibres of Old (70 ± 4 years) and Young (22 ± 3 years) humans to investigate whether changes in muscle fibre properties contribute to muscle weakness in old age. In type II fibres of Old subjects, specific force was reduced by ∼17% and Ca2+ sensitivity was also reduced (pCa50 decreased ∼0.05 pCa units) relative to that in Young. S-Glutathionylation of fast troponin I (TnIf) markedly increased Ca2+ sensitivity in type II fibres, but the increase was significantly smaller in Old versus Young (+0.136 and +0.164 pCa unit increases, respectively). Endogenous and maximal SR Ca2+ content were significantly smaller in both type I and type II fibres in Old subjects. In fibres of Young, the SR could be nearly fully depleted of Ca2+ by a combined caffeine and low Mg2+ stimulus, whereas in fibres of Old the amount of non-releasable Ca2+ was significantly increased (by > 12% of endogenous Ca2+ content). Western blotting showed an increased proportion of type I fibres in Old subjects, and increased amounts of calsequestrin-2 and calsequestrin-like protein. The findings suggest that muscle weakness in old age is probably attributable in part to (i) an increased proportion of type I fibres, (ii) a reduction in both maximum specific force and Ca2+ sensitivity in type II fibres, and also a decreased ability of S-glutathionylation of TnIf to counter the fatiguing effects of metabolites on Ca2+ sensitivity, and (iii) a reduction in the amount of releasable SR Ca2+ in both fibre types. Key points Muscle weakness in old age is due in large part to an overall loss of skeletal muscle tissue, but it remains uncertain how much also stems from alterations in the properties of the individual muscle fibres. This study examined the contractile properties and amount of stored intracellular calcium in single muscle fibres of Old (70�

LRP1 influences trafficking of N-type calcium channels via interaction with the auxiliary α2δ-1 subunit

PubMed Central

Kadurin, Ivan; Rothwell, Simon W.; Lana, Beatrice; Nieto-Rostro, Manuela; Dolphin, Annette C.

2017-01-01

Voltage-gated Ca2+ (CaV) channels consist of a pore-forming α1 subunit, which determines the main functional and pharmacological attributes of the channel. The CaV1 and CaV2 channels are associated with auxiliary β- and α2δ-subunits. The molecular mechanisms involved in α2δ subunit trafficking, and the effect of α2δ subunits on trafficking calcium channel complexes remain poorly understood. Here we show that α2δ-1 is a ligand for the Low Density Lipoprotein (LDL) Receptor-related Protein-1 (LRP1), a multifunctional receptor which mediates trafficking of cargoes. This interaction with LRP1 is direct, and is modulated by the LRP chaperone, Receptor-Associated Protein (RAP). LRP1 regulates α2δ binding to gabapentin, and influences calcium channel trafficking and function. Whereas LRP1 alone reduces α2δ-1 trafficking to the cell-surface, the LRP1/RAP combination enhances mature glycosylation, proteolytic processing and cell-surface expression of α2δ-1, and also increase plasma-membrane expression and function of CaV2.2 when co-expressed with α2δ-1. Furthermore RAP alone produced a small increase in cell-surface expression of CaV2.2, α2δ-1 and the associated calcium currents. It is likely to be interacting with an endogenous member of the LDL receptor family to have these effects. Our findings now provide a key insight and new tools to investigate the trafficking of calcium channel α2δ subunits. PMID:28256585

[Aging-related ionic remodeling of L-type voltage dependent calcium channel in left atria of canine].

PubMed

Zhou, Xian-hui; Zhang, Jian; Gan, Tian-yi; Xu, Guo-jun; Tang, Bao-peng

2012-04-01

To investigate aging-related ionic remodeling of L-type voltage dependent calcium channel (LVDCC) in left atria of canine. Seven adult (2.0 - 2.5 years) and 10 aged (> 8 years) dogs were used. The current of LVDCC was recorded by patch clamp technique in the whole cell mode. The action potential duration (APD(90)), amplitude of action potential plateau (APA), I(Ca-L) peak current density of LVDCC were recorded. The mRNA and protein expressions of α1c subunit (Ca(V1.2)), sarcoplasmic reticulum Ca(2+)-ATPase (SECRA(2)), Calpain-I, ryanodine receptor (RYR(2)) were detected by quantitative RT-PCR and Western blot, respectively. I(Ca-L) peak current density [(-8.11 ± 0.54) pA/pF vs. (-14.04 ± 0.82) pA/pF, P < 0.05] was significantly reduced and action potential duration to 90% repolarization (APD(90)) significantly prolonged [(340.5 ± 10.1) ms vs. (320.0 ± 7.9) ms, P < 0.05] in aged group than in adult group. The mRNA gene expression level of Ca(V1.2) was significantly lower (0.90 ± 0.35 vs. 2.38 ± 0.40, P < 0.05) while mRNA expression of RYR(2) was significantly higher (4.39 ± 4.68 vs. 1.49 ± 1.69, P < 0.05) in the aged dogs than in the adult dogs. mRNA expression of SECRA(2) and Calpain-I was similar between the two groups. Similarly, the protein expression level of Ca(V1.2) was significantly lower (0.13 ± 0.10 vs. 0.29 ± 0.12, P < 0.05) while the protein expression level of RYR(2) was significantly higher (0.18 ± 0.21 vs. 0.08 ± 0.36, P < 0.05) in the aged dogs than in the adult dogs. Again, protein expression of SECRA(2), PLN(1) and Calpain-I was similar between the two groups. These data suggest that aging could induce mRNA and protein expression changes of Ca(V1.2) and RYR(2) of LVDCC which might serve as the molecular basis of I(Ca-L) remodeling in aged dogs and might be linked to the increased likelihood of developing atrial fibrillation (AF) in aged dogs.

Inhibition of Cav3.2 T-type Calcium Channels by Its Intracellular I-II Loop*

PubMed Central

Monteil, Arnaud; Chausson, Patrick; Boutourlinsky, Katia; Mezghrani, Alexandre; Huc-Brandt, Sylvaine; Blesneac, Iulia; Bidaud, Isabelle; Lemmers, Céline; Leresche, Nathalie; Lambert, Régis C.; Lory, Philippe

2015-01-01

Voltage-dependent calcium channels (Cav) of the T-type family (Cav3.1, Cav3.2, and Cav3.3) are activated by low threshold membrane depolarization and contribute greatly to neuronal network excitability. Enhanced T-type channel activity, especially Cav3.2, contributes to disease states, including absence epilepsy. Interestingly, the intracellular loop connecting domains I and II (I-II loop) of Cav3.2 channels is implicated in the control of both surface expression and channel gating, indicating that this I-II loop plays an important regulatory role in T-type current. Here we describe that co-expression of this I-II loop or its proximal region (Δ1-Cav3.2; Ser423–Pro542) together with recombinant full-length Cav3.2 channel inhibited T-type current without affecting channel expression and membrane incorporation. Similar T-type current inhibition was obtained in NG 108-15 neuroblastoma cells that constitutively express Cav3.2 channels. Of interest, Δ1-Cav3.2 inhibited both Cav3.2 and Cav3.1 but not Cav3.3 currents. Efficacy of Δ1-Cav3.2 to inhibit native T-type channels was assessed in thalamic neurons using viral transduction. We describe that T-type current was significantly inhibited in the ventrobasal neurons that express Cav3.1, whereas in nucleus reticularis thalami neurons that express Cav3.2 and Cav3.3 channels, only the fast inactivating T-type current (Cav3.2 component) was significantly inhibited. Altogether, these data describe a new strategy to differentially inhibit Cav3 isoforms of the T-type calcium channels. PMID:25931121

Spatial distribution of calcium-gated chloride channels in olfactory cilia.

PubMed

French, Donald A; Badamdorj, Dorjsuren; Kleene, Steven J

2010-12-30

In vertebrate olfactory receptor neurons, sensory cilia transduce odor stimuli into changes in neuronal membrane potential. The voltage changes are primarily caused by the sequential openings of two types of channel: a cyclic-nucleotide-gated (CNG) cationic channel and a calcium-gated chloride channel. In frog, the cilia are 25 to 200 µm in length, so the spatial distributions of the channels may be an important determinant of odor sensitivity. To determine the spatial distribution of the chloride channels, we recorded from single cilia as calcium was allowed to diffuse down the length of the cilium and activate the channels. A computational model of this experiment allowed an estimate of the spatial distribution of the chloride channels. On average, the channels were concentrated in a narrow band centered at a distance of 29% of the ciliary length, measured from the base of the cilium. This matches the location of the CNG channels determined previously. This non-uniform distribution of transduction proteins is consistent with similar findings in other cilia. On average, the two types of olfactory transduction channel are concentrated in the same region of the cilium. This may contribute to the efficient detection of weak stimuli.

Temperature Dependence Of Current-Voltage Characteristics Of Au/p-GaAsN Schottky Barrier Diodes, With Small N Content

NASA Astrophysics Data System (ADS)

Rangel-Kuoppa, Victor-Tapio; Reentilä, Outi; Sopanen, Markku; Lipsanen, Harri

2011-12-01

The temperature dependent current-voltage (IVT) measurements on Au Schottky barrier diodes made on intrinsically p-type GaAs1-xNx were carried out. Three samples with small N content (x = 0.5%, 0.7% and 1%) were studied. The temperature range was 10-320 K. All contacts were found to be of Schottky type. The ideality factor and the apparent barrier height calculated by using thermionic emission (TE) theory show a strong temperature dependence. The current voltage (IV) curves are fitted based on the TE theory, yielding a zero-bias carrier height (ΦB0) and a ideality factor (n) that decrease and increase with decreasing temperature, respectively. The linear fitting of ΦB0 vs n and its subsequent evaluation for n = 1 give a zero-bias ΦB0 in the order of 0.35-0.4 eV. From the reverse-bias IV study, it is found that the experimental carrier density (NA) values increase with increasing temperature and are in agreement with the intrinsic carrier concentration for GaAs.

P-type surface effects for thickness variation of 2um and 4um of n-type layer in GaN LED

NASA Astrophysics Data System (ADS)

Halim, N. S. A. Abdul; Wahid, M. H. A.; Hambali, N. A. M. Ahmad; Rashid, S.; Ramli, M. M.; Shahimin, M. M.

2017-09-01

The internal quantum efficiency of III-Nitrides group, GaN light-emitting diode (LED) has been considerably limited due to the insufficient hole injection and this is caused by the lack of performance p-type doping and low hole mobility. The low hole mobility makes the hole less energetic, thus reduced the performance operation of GaN LED itself. The internal quantum efficiency of GaN-based LED with surface roughness (texture) can be changed by texture size, density, and thickness of GaN film or by the combined effects of surface shape and thickness of GaN film. Besides, due to lack of p-type GaN, attempts to look forward the potential of GaN LED relied on the thickness of n-type layer and surface shape of p-type GaN layer. This work investigates the characteristics of GaN LED with undoped n-GaN layer of different thickness and the surface shape of p-type layer. The LEDs performance is significantly altered by modifying the thickness and shape. Enhancement of n-GaN layer has led to the annihilation of electrical conductivity of the chip. Different surface geometry governs the emission rate extensively. Internal quantum efficiency is also predominantly affected by the geometry of n-GaN layer which subjected to the current spreading. It is recorded that the IQE droop can be minimized by varying the thickness of the active layer without amplifying the forward voltage. Optimum forward voltage (I-V), total emission rate relationship with the injected current and internal quantum efficiency (IQE) for 2,4 µm on four different surfaces of p-type layer are also reported in this paper.

Single Nisoldipine-Sensitive Calcium Channels in Smooth Muscle Cells Isolated from Rabbit Mesenteric Artery

NASA Astrophysics Data System (ADS)

Worley, Jennings F.; Deitmer, Joachim W.; Nelson, Mark T.

1986-08-01

Single smooth muscle cells were enzymatically isolated from the rabbit mesenteric artery. At physiological levels of external Ca, these cells were relaxed and contracted on exposure to norepinephrine, caffeine, or high levels of potassium. The patch-clamp technique was used to measure unitary currents through single channels in the isolated cells. Single channels were selective for divalent cations and exhibited two conductance levels, 8 pS and 15 pS. Both types of channels were voltage-dependent, and channel activity occurred at potentials positive to -40 mV. The activity of both channel types was almost completely inhibited by 50 nM nisoldipine. These channels appear to be the pathways for voltage-dependent Ca influx in vascular smooth muscle and may be the targets of the clinically used dihydropyridines.

Restoration of Motor Defects Caused by Loss of Drosophila TDP-43 by Expression of the Voltage-Gated Calcium Channel, Cacophony, in Central Neurons.

PubMed

Lembke, Kayly M; Scudder, Charles; Morton, David B

2017-09-27

Defects in the RNA-binding protein, TDP-43, are known to cause a variety of neurodegenerative diseases, including amyotrophic lateral sclerosis and frontotemporal lobar dementia. A variety of experimental systems have shown that neurons are sensitive to TDP-43 expression levels, yet the specific functional defects resulting from TDP-43 dysregulation have not been well described. Using the Drosophila TDP-43 ortholog TBPH, we previously showed that TBPH-null animals display locomotion defects as third instar larvae. Furthermore, loss of TBPH caused a reduction in cacophony , a Type II voltage-gated calcium channel, expression and that genetically restoring cacophony in motor neurons in TBPH mutant animals was sufficient to rescue the locomotion defects. In the present study, we examined the relative contributions of neuromuscular junction physiology and the motor program to the locomotion defects and identified subsets of neurons that require cacophony expression to rescue the defects. At the neuromuscular junction, we showed mEPP amplitudes and frequency require TBPH. Cacophony expression in motor neurons rescued mEPP frequency but not mEPP amplitude. We also showed that TBPH mutants displayed reduced motor neuron bursting and coordination during crawling and restoring cacophony selectively in two pairs of cells located in the brain, the AVM001b/2b neurons, also rescued the locomotion and motor defects, but not the defects in neuromuscular junction physiology. These results suggest that the behavioral defects associated with loss of TBPH throughout the nervous system can be associated with defects in a small number of genes in a limited number of central neurons, rather than peripheral defects. SIGNIFICANCE STATEMENT TDP-43 dysfunction is a common feature in neurodegenerative diseases, including amyotrophic lateral sclerosis, frontotemporal lobar dementia, and Alzheimer's disease. Loss- and gain-of-function models have shown that neurons are sensitive to TDP-43

High-speed, random-access fluorescence microscopy: I. High-resolution optical recording with voltage-sensitive dyes and ion indicators.

PubMed

Bullen, A; Patel, S S; Saggau, P

1997-07-01

The design and implementation of a high-speed, random-access, laser-scanning fluorescence microscope configured to record fast physiological signals from small neuronal structures with high spatiotemporal resolution is presented. The laser-scanning capability of this nonimaging microscope is provided by two orthogonal acousto-optic deflectors under computer control. Each scanning point can be randomly accessed and has a positioning time of 3-5 microseconds. Sampling time is also computer-controlled and can be varied to maximize the signal-to-noise ratio. Acquisition rates up to 200k samples/s at 16-bit digitizing resolution are possible. The spatial resolution of this instrument is determined by the minimal spot size at the level of the preparation (i.e., 2-7 microns). Scanning points are selected interactively from a reference image collected with differential interference contrast optics and a video camera. Frame rates up to 5 kHz are easily attainable. Intrinsic variations in laser light intensity and scanning spot brightness are overcome by an on-line signal-processing scheme. Representative records obtained with this instrument by using voltage-sensitive dyes and calcium indicators demonstrate the ability to make fast, high-fidelity measurements of membrane potential and intracellular calcium at high spatial resolution (2 microns) without any temporal averaging.

High-speed, random-access fluorescence microscopy: I. High-resolution optical recording with voltage-sensitive dyes and ion indicators.

PubMed Central

Bullen, A; Patel, S S; Saggau, P

1997-01-01

The design and implementation of a high-speed, random-access, laser-scanning fluorescence microscope configured to record fast physiological signals from small neuronal structures with high spatiotemporal resolution is presented. The laser-scanning capability of this nonimaging microscope is provided by two orthogonal acousto-optic deflectors under computer control. Each scanning point can be randomly accessed and has a positioning time of 3-5 microseconds. Sampling time is also computer-controlled and can be varied to maximize the signal-to-noise ratio. Acquisition rates up to 200k samples/s at 16-bit digitizing resolution are possible. The spatial resolution of this instrument is determined by the minimal spot size at the level of the preparation (i.e., 2-7 microns). Scanning points are selected interactively from a reference image collected with differential interference contrast optics and a video camera. Frame rates up to 5 kHz are easily attainable. Intrinsic variations in laser light intensity and scanning spot brightness are overcome by an on-line signal-processing scheme. Representative records obtained with this instrument by using voltage-sensitive dyes and calcium indicators demonstrate the ability to make fast, high-fidelity measurements of membrane potential and intracellular calcium at high spatial resolution (2 microns) without any temporal averaging. Images FIGURE 6 PMID:9199810

Activity of Palythoa caribaeorum Venom on Voltage-Gated Ion Channels in Mammalian Superior Cervical Ganglion Neurons

PubMed Central

Lazcano-Pérez, Fernando; Castro, Héctor; Arenas, Isabel; García, David E.; González-Muñoz, Ricardo; Arreguín-Espinosa, Roberto

2016-01-01

The Zoanthids are an order of cnidarians whose venoms and toxins have been poorly studied. Palythoa caribaeorum is a zoanthid commonly found around the Mexican coastline. In this study, we tested the activity of P. caribaeorum venom on voltage-gated sodium channel (NaV1.7), voltage-gated calcium channel (CaV2.2), the A-type transient outward (IA) and delayed rectifier (IDR) currents of KV channels of the superior cervical ganglion (SCG) neurons of the rat. These results showed that the venom reversibly delays the inactivation process of voltage-gated sodium channels and inhibits voltage-gated calcium and potassium channels in this mammalian model. The compounds responsible for these effects seem to be low molecular weight peptides. Together, these results provide evidence for the potential use of zoanthids as a novel source of cnidarian toxins active on voltage-gated ion channels. PMID:27164140

5.0 kV breakdown-voltage vertical GaN p-n junction diodes

NASA Astrophysics Data System (ADS)

Ohta, Hiroshi; Hayashi, Kentaro; Horikiri, Fumimasa; Yoshino, Michitaka; Nakamura, Tohru; Mishima, Tomoyoshi

2018-04-01

A high breakdown voltage of 5.0 kV has been achieved for the first time in vertical GaN p-n junction diodes by using our newly developed guard-ring structures. A resistance device was inserted between the main diode portion and the guard-ring portion in a ring-shaped p-n diode to generate a voltage drop over the resistance device by leakage current flowing through the guard-ring portion under negatively biased conditions before breakdown. The voltage at the outer mesa edge of the guard-ring portion, where the electric field intensity is highest and the destructive breakdown usually occurs, is decreased by the voltage drop, so the electric field concentration in the portion is reduced. By adopting this structure, the breakdown voltage (V B) is raised by about 200 V. Combined with a low measured on-resistance (R on) of 1.25 mΩ cm2, Baliga’s figure of merit (V\\text{B}2/R\\text{on}) was as high as 20 GW/cm2.

The Role of Auxiliary Subunits for the Functional Diversity of Voltage-Gated Calcium Channels

PubMed Central

Campiglio, Marta; Flucher, Bernhard E

2015-01-01

Voltage-gated calcium channels (VGCCs) represent the sole mechanism to convert membrane depolarization into cellular functions like secretion, contraction, or gene regulation. VGCCs consist of a pore-forming α1 subunit and several auxiliary channel subunits. These subunits come in multiple isoforms and splice-variants giving rise to a stunning molecular diversity of possible subunit combinations. It is generally believed that specific auxiliary subunits differentially regulate the channels and thereby contribute to the great functional diversity of VGCCs. If auxiliary subunits can associate and dissociate from pre-existing channel complexes, this would allow dynamic regulation of channel properties. However, most auxiliary subunits modulate current properties very similarly, and proof that any cellular calcium channel function is indeed modulated by the physiological exchange of auxiliary subunits is still lacking. In this review we summarize available information supporting a differential modulation of calcium channel functions by exchange of auxiliary subunits, as well as experimental evidence in support of alternative functions of the auxiliary subunits. At the heart of the discussion is the concept that, in their native environment, VGCCs function in the context of macromolecular signaling complexes and that the auxiliary subunits help to orchestrate the diverse protein–protein interactions found in these calcium channel signalosomes. Thus, in addition to a putative differential modulation of current properties, differential subcellular targeting properties and differential protein–protein interactions of the auxiliary subunits may explain the need for their vast molecular diversity. J. Cell. Physiol. 999: 00–00, 2015. © 2015 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc. J. Cell. Physiol. 230: 2019–2031, 2015. © 2015 Wiley Periodicals, Inc. PMID:25820299

C-terminal modulatory domain controls coupling of voltage-sensing to pore opening in Cav1.3 L-type Ca(2+) channels.

PubMed

Lieb, Andreas; Ortner, Nadine; Striessnig, Jörg

2014-04-01

Activity of voltage-gated Cav1.3 L-type Ca(2+) channels is required for proper hearing as well as sinoatrial node and brain function. This critically depends on their negative activation voltage range, which is further fine-tuned by alternative splicing. Shorter variants miss a C-terminal regulatory domain (CTM), which allows them to activate at even more negative potentials than C-terminally long-splice variants. It is at present unclear whether this is due to an increased voltage sensitivity of the Cav1.3 voltage-sensing domain, or an enhanced coupling of voltage-sensor conformational changes to the subsequent opening of the activation gate. We studied the voltage-dependence of voltage-sensor charge movement (QON-V) and of current activation (ICa-V) of the long (Cav1.3L) and a short Cav1.3 splice variant (Cav1.342A) expressed in tsA-201 cells using whole cell patch-clamp. Charge movement (QON) of Cav1.3L displayed a much steeper voltage-dependence and a more negative half-maximal activation voltage than Cav1.2 and Cav3.1. However, a significantly higher fraction of the total charge had to move for activation of Cav1.3 half-maximal conductance (Cav1.3: 68%; Cav1.2: 52%; Cav3.1: 22%). This indicated a weaker coupling of Cav1.3 voltage-sensor charge movement to pore opening. However, the coupling efficiency was strengthened in the absence of the CTM in Cav1.342A, thereby shifting ICa-V by 7.2 mV to potentials that were more negative without changing QON-V. We independently show that the presence of intracellular organic cations (such as n-methyl-D-glucamine) induces a pronounced negative shift of QON-V and a more negative activation of ICa-V of all three channels. These findings illustrate that the voltage sensors of Cav1.3 channels respond more sensitively to depolarization than those of Cav1.2 or Cav3.1. Weak coupling of voltage sensing to pore opening is enhanced in the absence of the CTM, allowing short Cav1.342A splice variants to activate at lower voltages

Activation of brain nitric oxide synthase in depolarized human temporal cortex slices: differential role of voltage-sensitive calcium channels.

PubMed

Fontana, G; Fedele, E; Cossu, M; Munari, C; Raiteri, M

1997-11-01

1. Nitric oxide (NO) synthase activity was studied in slices of human temporal cortex samples obtained in neurosurgery by measuring the conversion of L-[3H]-arginine to L-[3H]-citrulline. 2. Elevation of extracellular K+ to 20, 35 or 60 mM concentration-dependently augmented L-[3H]-citrulline production. The response to 35 mM KCl was abolished by N(G)-nitro-L-arginine (100 microM) demonstrating NO synthase specific conversion of L-arginine to L-citrulline. Increasing extracellular MgCl2 concentration up to 10 mM also prevented the K+ (35 mM)-induced NO synthase activation, suggesting the absolute requirement of external calcium ions for enzyme activity. 3. However, the effect of high K+ (35 mM) on citrulline synthesis was insensitive to the antagonists of ionotropic and metabotropic glutamate receptors dizocilpine (MK-801), 6-nitro-7-sulphamoylbenzo(f)-quinoxaline-2-3-dione (NBQX) or L-2-amino-3-phosphonopropionic acid (L-AP3) as well as to the nicotinic receptor antagonist, mecamylamine. 4. The 35 mM K+ response was insensitive to omega-conotoxin GVIA (1 microM) and nifedipine (100 microM), but could be prevented in part by omega-agatoxin IVA (0.1 and 1 microM). The inhibition caused by 0.1 microM omega-agatoxin IVA (approximately 30%) was enhanced by adding omega-conotoxin GVIA (1 microM) or nifedipine (100 microM). Further inhibition (up to above 70%) could be observed when the three Ca2+ channel blockers were added together. Similarly, synthetic FTX 3.3 arginine polyamine (sFTX) prevented (50% at 100 microM) the K+-evoked NO synthase activation. This effect of sFTX was further enhanced (up to 70%) by adding 1 microM omega-conotoxin GVIA plus 100 microM nifedipine. No further inhibition could be observed upon addition of MK-801 or/and NBQX. 5. It was concluded that elevation of extracellular [K+] causes NO synthase activation by external Ca2+ entering cells mainly through channels of the P/Q-type. Other Ca2+ channels (L- and N-type) appear to contribute when P

Trigeminal ganglion neuron subtype-specific alterations of CaV2.1 calcium current and excitability in a Cacna1a mouse model of migraine

PubMed Central

Fioretti, B; Catacuzzeno, L; Sforna, L; Gerke-Duncan, M B; van den Maagdenberg, A M J M; Franciolini, F; Connor, M; Pietrobon, D

2011-01-01

Abstract Familial hemiplegic migraine type-1 (FHM1), a monogenic subtype of migraine with aura, is caused by gain-of-function mutations in CaV2.1 (P/Q-type) calcium channels. The consequences of FHM1 mutations on the trigeminovascular pathway that generates migraine headache remain largely unexplored. Here we studied the calcium currents and excitability properties of two subpopulations of small-diameter trigeminal ganglion (TG) neurons from adult wild-type (WT) and R192Q FHM1 knockin (KI) mice: capsaicin-sensitive neurons without T-type calcium currents (CS) and capsaicin-insensitive neurons characterized by the expression of T-type calcium currents (CI-T). Small TG neurons retrogradely labelled from the dura are mostly CS neurons, while CI-T neurons were not present in the labelled population. CS and CI-T neurons express CaV2.1 channels with different activation properties, and the CaV2.1 channels are differently affected by the FHM1 mutation in the two TG neuron subtypes. In CI-T neurons from FHM1 KI mice there was a larger P/Q-type current density following mild depolarizations, a larger action potential (AP)-evoked calcium current and a longer AP duration when compared to CI-T neurons from WT mice. In striking contrast, the P/Q-type current density, voltage dependence and kinetics were not altered by the FHM1 mutation in CS neurons. The excitability properties of mutant CS neurons were also unaltered. Congruently, the FHM1 mutation did not alter depolarization-evoked CGRP release from the dura mater, while CGRP release from the trigeminal ganglion was larger in KI compared to WT mice. Our findings suggest that the facilitation of peripheral mechanisms of CGRP action, such as dural vasodilatation and nociceptor sensitization at the meninges, does not contribute to the generation of headache in FHM1. PMID:22005682

A Neuron-Based Screening Platform for Optimizing Genetically-Encoded Calcium Indicators

PubMed Central

Schreiter, Eric R.; Hasseman, Jeremy P.; Tsegaye, Getahun; Fosque, Benjamin F.; Behnam, Reza; Shields, Brenda C.; Ramirez, Melissa; Kimmel, Bruce E.; Kerr, Rex A.; Jayaraman, Vivek; Looger, Loren L.; Svoboda, Karel; Kim, Douglas S.

2013-01-01

Fluorescent protein-based sensors for detecting neuronal activity have been developed largely based on non-neuronal screening systems. However, the dynamics of neuronal state variables (e.g., voltage, calcium, etc.) are typically very rapid compared to those of non-excitable cells. We developed an electrical stimulation and fluorescence imaging platform based on dissociated rat primary neuronal cultures. We describe its use in testing genetically-encoded calcium indicators (GECIs). Efficient neuronal GECI expression was achieved using lentiviruses containing a neuronal-selective gene promoter. Action potentials (APs) and thus neuronal calcium levels were quantitatively controlled by electrical field stimulation, and fluorescence images were recorded. Images were segmented to extract fluorescence signals corresponding to individual GECI-expressing neurons, which improved sensitivity over full-field measurements. We demonstrate the superiority of screening GECIs in neurons compared with solution measurements. Neuronal screening was useful for efficient identification of variants with both improved response kinetics and high signal amplitudes. This platform can be used to screen many types of sensors with cellular resolution under realistic conditions where neuronal state variables are in relevant ranges with respect to timing and amplitude. PMID:24155972

The L-type voltage-gated calcium channel CaV1.2 mediates fear extinction and modulates synaptic tone in the lateral amygdala.

PubMed

Temme, Stephanie J; Murphy, Geoffrey G

2017-11-01

L-type voltage-gated calcium channels (LVGCCs) have been implicated in both the formation and the reduction of fear through Pavlovian fear conditioning and extinction. Despite the implication of LVGCCs in fear learning and extinction, studies of the individual LVGCC subtypes, Ca V 1.2 and Ca V 1.3, using transgenic mice have failed to find a role of either subtype in fear extinction. This discontinuity between the pharmacological studies of LVGCCs and the studies investigating individual subtype contributions could be due to the limited neuronal deletion pattern of the Ca V 1.2 conditional knockout mice previously studied to excitatory neurons in the forebrain. To investigate the effects of deletion of Ca V 1.2 in all neuronal populations, we generated Ca V 1.2 conditional knockout mice using the synapsin1 promoter to drive Cre recombinase expression. Pan-neuronal deletion of Ca V 1.2 did not alter basal anxiety or fear learning. However, pan-neuronal deletion of Ca V 1.2 resulted in a significant deficit in extinction of contextual fear, implicating LVGCCs, specifically Ca V 1.2, in extinction learning. Further exploration on the effects of deletion of Ca V 1.2 on inhibitory and excitatory input onto the principle neurons of the lateral amygdala revealed a significant shift in inhibitory/excitatory balance. Together these data illustrate an important role of Ca V 1.2 in fear extinction and the synaptic regulation of activity within the amygdala. © 2017 Temme and Murphy; Published by Cold Spring Harbor Laboratory Press.

A temperature monitor circuit with small voltage sensitivity using a topology-reconfigurable ring oscillator

NASA Astrophysics Data System (ADS)

Kishimoto, Tadashi; Ishihara, Tohru; Onodera, Hidetoshi

2018-04-01

In this paper, we propose a temperature monitor circuit that exhibits a small supply voltage sensitivity adopting a circuit topology of a reconfigurable ring oscillator. The circuit topology of the monitor is crafted such that the oscillation frequency is determined by the amount of subthreshold leakage current, which has an exponential dependence on temperature. Another important characteristic of the monitor is its small supply voltage sensitivity. The measured oscillation frequency of a test chip fabricated in a 65 nm CMOS process varies only 2.6% under a wide range of supply voltages from 0.4 to 1.0 V at room temperature. The temperature estimation error ranges from -0.3 to 0.4 °C over a temperature range of 10 to 100 °C.

Spinocerebellum Ataxia Type 6: Molecular Mechanisms and Calcium Channel Genetics.

PubMed

Du, Xiaofei; Gomez, Christopher Manuel

2018-01-01

Spinocerebellar ataxia (SCA) type 6 is an autosomal dominant disease affecting cerebellar degeneration. Clinically, it is characterized by pure cerebellar dysfunction, slowly progressive unsteadiness of gait and stance, slurred speech, and abnormal eye movements with late onset. Pathological findings of SCA6 include a diffuse loss of Purkinje cells, predominantly in the cerebellar vermis. Genetically, SCA6 is caused by expansion of a trinucleotide CAG repeat in the last exon of longest isoform CACNA1A gene on chromosome 19p13.1-p13.2. Normal alleles have 4-18 repeats, while alleles causing disease contain 19-33 repeats. Due to presence of a novel internal ribosomal entry site (IRES) with the mRNA, CACNA1A encodes two structurally unrelated proteins with distinct functions within an overlapping open reading frame (ORF) of the same mRNA: (1) α1A subunit of P/Q-type voltage gated calcium channel; (2) α1ACT, a newly recognized transcription factor, with polyglutamine repeat at C-terminal end. Understanding the function of α1ACT in physiological and pathological conditions may elucidate the pathogenesis of SCA6. More importantly, the IRES, as the translational control element of α1ACT, provides a potential therapeutic target for the treatment of SCA6.

Ultra-compact Marx-type high-voltage generator

DOEpatents

Goerz, David A.; Wilson, Michael J.

2000-01-01

An ultra-compact Marx-type high-voltage generator includes individual high-performance components that are closely coupled and integrated into an extremely compact assembly. In one embodiment, a repetitively-switched, ultra-compact Marx generator includes low-profile, annular-shaped, high-voltage, ceramic capacitors with contoured edges and coplanar extended electrodes used for primary energy storage; low-profile, low-inductance, high-voltage, pressurized gas switches with compact gas envelopes suitably designed to be integrated with the annular capacitors; feed-forward, high-voltage, ceramic capacitors attached across successive switch-capacitor-switch stages to couple the necessary energy forward to sufficiently overvoltage the spark gap of the next in-line switch; optimally shaped electrodes and insulator surfaces to reduce electric field stresses in the weakest regions where dissimilar materials meet, and to spread the fields more evenly throughout the dielectric materials, allowing them to operate closer to their intrinsic breakdown levels; and uses manufacturing and assembly methods to integrate the capacitors and switches into stages that can be arranged into a low-profile Marx generator.

Contribution of Sialic Acid to the Voltage Dependence of Sodium Channel Gating

PubMed Central

Bennett, Eric; Urcan, Mary S.; Tinkle, Sally S.; Koszowski, Adam G.; Levinson, Simon R.

1997-01-01

A potential role for sialic acid in the voltage-dependent gating of rat skeletal muscle sodium channels (rSkM1) was investigated using Chinese hamster ovary (CHO) cells stably transfected with rSkM1. Changes in the voltage dependence of channel gating were observed after enzymatic (neuraminidase) removal of sialic acid from cells expressing rSkM1 and through the expression of rSkM1 in a sialylation-deficient cell line (lec2). The steady-state half-activation voltages (Va) of channels under each condition of reduced sialylation were ∼10 mV more depolarized than control channels. The voltage dependence of the time constants of channel activation and inactivation were also shifted in the same direction and by a similar magnitude. In addition, recombinant deletion of likely glycosylation sites from the rSkM1 sequence resulted in mutant channels that gated at voltages up to 10 mV more positive than wild-type channels. Thus three independent means of reducing channel sialylation show very similar effects on the voltage dependence of channel gating. Finally, steady-state activation voltages for channels subjected to reduced sialylation conditions were much less sensitive to the effects of external calcium than those measured under control conditions, indicating that sialic acid directly contributes to the negative surface potential. These results are consistent with an electrostatic mechanism by which external, negatively charged sialic acid residues on rSkM1 alter the electric field sensed by channel gating elements. PMID:9089440

Contractile properties and sarcoplasmic reticulum calcium content in type I and type II skeletal muscle fibres in active aged humans.

PubMed

Lamboley, C R; Wyckelsma, V L; Dutka, T L; McKenna, M J; Murphy, R M; Lamb, G D

2015-06-01

Muscle weakness in old age is due in large part to an overall loss of skeletal muscle tissue, but it remains uncertain how much also stems from alterations in the properties of the individual muscle fibres. This study examined the contractile properties and amount of stored intracellular calcium in single muscle fibres of Old (70 ± 4 years) and Young (22 ± 3 years) adults. The maximum level of force production (per unit cross-sectional area) in fast twitch fibres in Old subjects was lower than in Young subjects, and the fibres were also less sensitive to activation by calcium. The amount of calcium stored inside muscle fibres and available to trigger contraction was also lower in both fast- and slow-twitch muscle fibres in the Old subjects. These findings indicate that muscle weakness in old age stems in part from an impaired capacity for force production in the individual muscle fibres. This study examined the contractile properties and sarcoplasmic reticulum (SR) Ca(2+) content in mechanically skinned vastus lateralis muscle fibres of Old (70 ± 4 years) and Young (22 ± 3 years) humans to investigate whether changes in muscle fibre properties contribute to muscle weakness in old age. In type II fibres of Old subjects, specific force was reduced by ∼17% and Ca(2+) sensitivity was also reduced (pCa50 decreased ∼0.05 pCa units) relative to that in Young. S-Glutathionylation of fast troponin I (TnIf ) markedly increased Ca(2+) sensitivity in type II fibres, but the increase was significantly smaller in Old versus Young (+0.136 and +0.164 pCa unit increases, respectively). Endogenous and maximal SR Ca(2+) content were significantly smaller in both type I and type II fibres in Old subjects. In fibres of Young, the SR could be nearly fully depleted of Ca(2+) by a combined caffeine and low Mg(2+) stimulus, whereas in fibres of Old the amount of non-releasable Ca(2+) was significantly increased (by > 12% of endogenous Ca(2+) content). Western

Role of AlGaN/GaN interface traps on negative threshold voltage shift in AlGaN/GaN HEMT

NASA Astrophysics Data System (ADS)

Malik, Amit; Sharma, Chandan; Laishram, Robert; Bag, Rajesh Kumar; Rawal, Dipendra Singh; Vinayak, Seema; Sharma, Rajesh Kumar

2018-04-01

This article reports negative shift in the threshold-voltage in AlGaN/GaN high electron mobility transistor (HEMT) with application of reverse gate bias stress. The device is biased in strong pinch-off and low drain to source voltage condition for a fixed time duration (reverse gate bias stress), followed by measurement of transfer characteristics. Negative threshold voltage shift after application of reverse gate bias stress indicates the presence of more carriers in channel as compared to the unstressed condition. We propose the presence of AlGaN/GaN interface states to be the reason of negative threshold voltage shift, and developed a process to electrically characterize AlGaN/GaN interface states. We verified the results with Technology Computer Aided Design (TCAD) ATLAS simulation and got a good match with experimental measurements.

Effects of gold diffusion on n-type doping of GaAs nanowires.

PubMed

Tambe, Michael J; Ren, Shenqiang; Gradecak, Silvija

2010-11-10

The deposition of n-GaAs shells is explored as a method of n-type doping in GaAs nanowires grown by the Au-mediated metal-organic chemical vapor deposition. Core-shell GaAs/n-GaAs nanowires exhibit an unintended rectifying behavior that is attributed to the Au diffusion during the shell deposition based on studies using energy dispersive X-ray spectroscopy, current-voltage, capacitance-voltage, and Kelvin probe force measurements. Removing the gold prior to n-type shell deposition results in the realization of n-type GaAs nanowires without rectification. We directly correlate the presence of gold impurities to nanowire electrical properties and provide an insight into the role of seed particles on the properties of nanowires and nanowire heterostructures.

Visual Stimuli Evoked Action Potentials Trigger Rapidly Propagating Dendritic Calcium Transients in the Frog Optic Tectum Layer 6 Neurons.

PubMed

Svirskis, Gytis; Baranauskas, Gytis; Svirskiene, Natasa; Tkatch, Tatiana

2015-01-01

The superior colliculus in mammals or the optic tectum in amphibians is a major visual information processing center responsible for generation of orientating responses such as saccades in monkeys or prey catching avoidance behavior in frogs. The conserved structure function of the superior colliculus the optic tectum across distant species such as frogs, birds monkeys permits to draw rather general conclusions after studying a single species. We chose the frog optic tectum because we are able to perform whole-cell voltage-clamp recordings fluorescence imaging of tectal neurons while they respond to a visual stimulus. In the optic tectum of amphibians most visual information is processed by pear-shaped neurons possessing long dendritic branches, which receive the majority of synapses originating from the retinal ganglion cells. Since the first step of the retinal input integration is performed on these dendrites, it is important to know whether this integration is enhanced by active dendritic properties. We demonstrate that rapid calcium transients coinciding with the visual stimulus evoked action potentials in the somatic recordings can be readily detected up to the fine branches of these dendrites. These transients were blocked by calcium channel blockers nifedipine CdCl2 indicating that calcium entered dendrites via voltage-activated L-type calcium channels. The high speed of calcium transient propagation, >300 μm in <10 ms, is consistent with the notion that action potentials, actively propagating along dendrites, open voltage-gated L-type calcium channels causing rapid calcium concentration transients in the dendrites. We conclude that such activation by somatic action potentials of the dendritic voltage gated calcium channels in the close vicinity to the synapses formed by axons of the retinal ganglion cells may facilitate visual information processing in the principal neurons of the frog optic tectum.

Intact calcium signaling in adrenergic-deficient embryonic mouse hearts.

PubMed

Peoples, Jessica N; Taylor, David G; Katchman, Alexander N; Ebert, Steven N

2018-01-22

Mouse embryos that lack the ability to produce the adrenergic hormones, norepinephrine (NE) and epinephrine (EPI), due to disruption of the dopamine beta-hydroxylase (Dbh -/- ) gene inevitably perish from heart failure during mid-gestation. Since adrenergic stimulation is well-known to enhance calcium signaling in developing as well as adult myocardium, and impairments in calcium signaling are typically associated with heart failure, we hypothesized that adrenergic-deficient embryonic hearts would display deficiencies in cardiac calcium signaling relative to adrenergic-competent controls at a developmental stage immediately preceding the onset of heart failure, which first appears beginning or shortly after mouse embryonic day 10.5 (E10.5). To test this hypothesis, we used ratiometric fluorescent calcium imaging techniques to measure cytosolic calcium transients, [Ca 2+ ] i in isolated E10.5 mouse hearts. Our results show that spontaneous [Ca 2+ ] i oscillations were intact and robustly responded to a variety of stimuli including extracellular calcium (5 mM), caffeine (5 mM), and NE (100 nM) in a manner that was indistinguishable from controls. Further, we show similar patterns of distribution (via immunofluorescent histochemical staining) and activity (via patch-clamp recording techniques) for the major voltage-gated plasma membrane calcium channel responsible for the L-type calcium current, I Ca,L , in adrenergic-deficient and control embryonic cardiac cells. These results demonstrate that despite the absence of vital adrenergic hormones that consistently leads to embryonic lethality in vivo, intracellular and extracellular calcium signaling remain essentially intact and functional in embryonic mouse hearts through E10.5. These findings suggest that adrenergic stimulation is not required for the development of intracellular calcium oscillations or extracellular calcium signaling through I Ca,L and that aberrant calcium signaling does not likely contribute

Synaptic Calcium Regulation in Hair Cells of the Chicken Basilar Papilla

PubMed Central

Im, Gi Jung; Moskowitz, Howard S.; Lehar, Mohammed; Hiel, Hakim

2014-01-01

Cholinergic inhibition of hair cells occurs by activation of calcium-dependent potassium channels. A near-membrane postsynaptic cistern has been proposed to serve as a store from which calcium is released to supplement influx through the ionotropic ACh receptor. However, the time and voltage dependence of acetylcholine (ACh)-evoked potassium currents reveal a more complex relationship between calcium entry and release from stores. The present work uses voltage steps to regulate calcium influx during the application of ACh to hair cells in the chicken basilar papilla. When calcium influx was terminated at positive membrane potential, the ACh-evoked potassium current decayed exponentially over ∼100 ms. However, at negative membrane potentials, this current exhibited a secondary rise in amplitude that could be eliminated by dihydropyridine block of the voltage-gated calcium channels of the hair cell. Calcium entering through voltage-gated channels may transit through the postsynaptic cistern, since ryanodine and sarcoendoplasmic reticulum calcium-ATPase blockers altered the time course and magnitude of this secondary, voltage-dependent contribution to ACh-evoked potassium current. Serial section electron microscopy showed that efferent and afferent synaptic structures are juxtaposed, supporting the possibility that voltage-gated influx at afferent ribbon synapses influences calcium homeostasis during long-lasting cholinergic inhibition. In contrast, spontaneous postsynaptic currents (“minis”) resulting from stochastic efferent release of ACh were made briefer by ryanodine, supporting the hypothesis that the synaptic cistern serves primarily as a calcium barrier and sink during low-level synaptic activity. Hypolemmal cisterns such as that at the efferent synapse of the hair cell can play a dynamic role in segregating near-membrane calcium for short-term and long-term signaling. PMID:25505321

Low operation voltage of GaN-based LEDs with Al-doped ZnO upper contact directly on p-type GaN without insert layer

NASA Astrophysics Data System (ADS)

Chen, P. H.; Chen, Yu An; Chang, L. C.; Lai, W. C.; Kuo, Cheng Huang

2015-07-01

Al-doped ZnO (AZO) film was evaporated on double-side polished sapphire, p-GaN layers, n+-InGaN-GaN short-period superlattice (SPS) structures, and GaN-based light-emitting diodes (LEDs) by e-beam. The AZO film on the p-GaN layer after thermal annealing exhibited an extremely high transparency (98% at 450 nm) and a small specific contact resistance of 2.19 × 10-2 Ω cm2, which was almost the same as that of as-deposited AZO on n+-SPS structure. With 20 mA injection current, the forward voltages were 3.30 and 3.27 V, whereas the output powers were 4.32 and 4.07 mW for the LED with AZO on insert n+-SPS upper contact and the LED with AZO on p-GaN upper contact (without insert layer), respectively. The small specific contact resistance and low operation voltage of LED with AZO on p-GaN upper contact was achieved by rapid thermal annealing (RTA) process.

The allosteric site regulates the voltage sensitivity of muscarinic receptors.

PubMed

Hoppe, Anika; Marti-Solano, Maria; Drabek, Matthäus; Bünemann, Moritz; Kolb, Peter; Rinne, Andreas

2018-01-01

Muscarinic receptors (M-Rs) for acetylcholine (ACh) belong to the class A of G protein-coupled receptors. M-Rs are activated by orthosteric agonists that bind to a specific site buried in the M-R transmembrane helix bundle. In the active conformation, receptor function can be modulated either by allosteric modulators, which bind to the extracellular receptor surface or by the membrane potential via an unknown mechanism. Here, we compared the modulation of M 1 -Rs and M 3 -Rs induced by changes in voltage to their allosteric modulation by chemical compounds. We quantified changes in receptor signaling in single HEK 293 cells with a FRET biosensor for the G q protein cycle. In the presence of ACh, M 1 -R signaling was potentiated by voltage, similarly to positive allosteric modulation by benzyl quinolone carboxylic acid. Conversely, signaling of M 3 -R was attenuated by voltage or the negative allosteric modulator gallamine. Because the orthosteric site is highly conserved among M-Rs, but allosteric sites vary, we constructed "allosteric site" M 3 /M 1 -R chimeras and analyzed their voltage dependencies. Exchanging the entire allosteric sites eliminated the voltage sensitivity of ACh responses for both receptors, but did not affect their modulation by allosteric compounds. Furthermore, a point mutation in M 3 -Rs caused functional uncoupling of the allosteric and orthosteric sites and abolished voltage dependence. Molecular dynamics simulations of the receptor variants indicated a subtype-specific crosstalk between both sites, involving the conserved tyrosine lid structure of the orthosteric site. This molecular crosstalk leads to receptor subtype-specific voltage effects. Copyright © 2017 Elsevier Inc. All rights reserved.

Increase the threshold voltage of high voltage GaN transistors by low temperature atomic hydrogen treatment

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

Erofeev, E. V., E-mail: erofeev@micran.ru; Fedin, I. V.; Kutkov, I. V.

High-electron-mobility transistors (HEMTs) based on AlGaN/GaN epitaxial heterostructures are a promising element base for the fabrication of high voltage electronic devices of the next generation. This is caused by both the high mobility of charge carriers in the transistor channel and the high electric strength of the material, which makes it possible to attain high breakdown voltages. For use in high-power switches, normally off-mode GaN transistors operating under enhancement conditions are required. To fabricate normally off GaN transistors, one most frequently uses a subgate region based on magnesium-doped p-GaN. However, optimization of the p-GaN epitaxial-layer thickness and the doping levelmore » makes it possible to attain a threshold voltage of GaN transistors close to V{sub th} = +2 V. In this study, it is shown that the use of low temperature treatment in an atomic hydrogen flow for the p-GaN-based subgate region before the deposition of gate-metallization layers makes it possible to increase the transistor threshold voltage to V{sub th} = +3.5 V. The effects under observation can be caused by the formation of a dipole layer on the p-GaN surface induced by the effect of atomic hydrogen. The heat treatment of hydrogen-treated GaN transistors in a nitrogen environment at a temperature of T = 250°C for 12 h reveals no degradation of the transistor’s electrical parameters, which can be caused by the formation of a thermally stable dipole layer at the metal/p-GaN interface as a result of hydrogenation.« less

Limitations of threshold voltage engineering of AlGaN/GaN heterostructures by dielectric interface charge density and manipulation by oxygen plasma surface treatments

NASA Astrophysics Data System (ADS)

Lükens, G.; Yacoub, H.; Kalisch, H.; Vescan, A.

2016-05-01

The interface charge density between the gate dielectric and an AlGaN/GaN heterostructure has a significant impact on the absolute value and stability of the threshold voltage Vth of metal-insulator-semiconductor (MIS) heterostructure field effect transistor. It is shown that a dry-etching step (as typically necessary for normally off devices engineered by gate-recessing) before the Al2O3 gate dielectric deposition introduces a high positive interface charge density. Its origin is most likely donor-type trap states shifting Vth to large negative values, which is detrimental for normally off devices. We investigate the influence of oxygen plasma annealing techniques of the dry-etched AlGaN/GaN surface by capacitance-voltage measurements and demonstrate that the positive interface charge density can be effectively compensated. Furthermore, only a low Vth hysteresis is observable making this approach suitable for threshold voltage engineering. Analysis of the electrostatics in the investigated MIS structures reveals that the maximum Vth shift to positive voltages achievable is fundamentally limited by the onset of accumulation of holes at the dielectric/barrier interface. In the case of the Al2O3/Al0.26Ga0.74N/GaN material system, this maximum threshold voltage shift is limited to 2.3 V.

Additive effects of cilnidipine, an L-/N-type calcium channel blocker, and an angiotensin II receptor blocker on reducing cardiorenal damage in Otsuka Long-Evans Tokushima Fatty rats with type 2 diabetes mellitus.

PubMed

Mori, Yutaka; Aritomi, Shizuka; Niinuma, Kazumi; Nakamura, Tarou; Matsuura, Kenichi; Yokoyama, Junichi; Utsunomiya, Kazunori

2014-01-01

Cilnidipine (Cil), which is an L-/N-type calcium channel blocker (CCB), has been known to provide renal protection by decreasing the activity of the sympathetic nervous system (SNS) and the renin-angiotensin system. In this study, we compared the effects of the combination of Cil and amlodipine (Aml), which is an L-type CCB, with an angiotensin (Ang) II receptor blocker on diabetic cardiorenal damage in spontaneously type 2 diabetic rats. Seventeen-week-old Otsuka Long-Evans Tokushima Fatty rats were randomly assigned to receive Cil, Aml, valsartan (Val), Cil + Val, Aml + Val, or a vehicle (eight rats per group) for 22 weeks. Antihypertensive potencies were nearly equal among the CCB monotherapy groups and the combination therapy groups. The lowering of blood pressure by either treatment did not significantly affect the glycemic variables. However, exacerbations of renal and heart failure were significantly suppressed in rats administered Cil or Val, and additional suppression was observed in those administered Cil + Val. Although Val increased the renin-Ang system, Aml + Val treatment resulted in additional increases in these parameters, while Cil + Val did not show such effects. Furthermore, Cil increased the ratio of Ang-(1-7) to Ang-I, despite the fact that Val and Aml + Val decreased the Ang-(1-7) levels. These actions of Cil + Val might be due to their synergistic inhibitory effect on the activity of the SNS, and on aldosterone secretion through N-type calcium channel antagonism and Ang II receptor type 1 antagonism. Thus, Cil may inhibit the progression of cardiorenal disease in type 2 diabetes patients by acting as an N-type CCB and inhibiting the aldosterone secretion and SNS activation when these drugs were administered in combination with an Ang II receptor blocker.

The tobacco-specific carcinogen-operated calcium channel promotes lung tumorigenesis via IGF2 exocytosis in lung epithelial cells.

PubMed

Boo, Hye-Jin; Min, Hye-Young; Jang, Hyun-Ji; Yun, Hye Jeong; Smith, John Kendal; Jin, Quanri; Lee, Hyo-Jong; Liu, Diane; Kweon, Hee-Seok; Behrens, Carmen; Lee, J Jack; Wistuba, Ignacio I; Lee, Euni; Hong, Waun Ki; Lee, Ho-Young

2016-09-26

Nicotinic acetylcholine receptors (nAChRs) binding to the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces Ca 2+ signalling, a mechanism that is implicated in various human cancers. In this study, we investigated the role of NNK-mediated Ca 2+ signalling in lung cancer formation. We show significant overexpression of insulin-like growth factors (IGFs) in association with IGF-1R activation in human preneoplastic lung lesions in smokers. NNK induces voltage-dependent calcium channel (VDCC)-intervened calcium influx in airway epithelial cells, resulting in a rapid IGF2 secretion via the regulated pathway and thus IGF-1R activation. Silencing nAChR, α1 subunit of L-type VDCC, or various vesicular trafficking curators, including synaptotagmins and Rabs, or blockade of nAChR/VDCC-mediated Ca 2+ influx significantly suppresses NNK-induced IGF2 exocytosis, transformation and tumorigenesis of lung epithelial cells. Publicly available database reveals inverse correlation between use of calcium channel blockers and lung cancer diagnosis. Our data indicate that NNK disrupts the regulated pathway of IGF2 exocytosis and promotes lung tumorigenesis.

Current statins show calcium channel blocking activity through voltage gated channels.

PubMed

Ali, Niaz; Begum, Robina; Faisal, Muhammad Saleh; Khan, Aslam; Nabi, Muhammad; Shehzadi, Gulfam; Ullah, Shakir; Ali, Waqar

2016-09-21

significant right shift in IC50 for CCRCs (P ≤ 0.05). In case of verapamil, IC50 for control curves is -2.45 ± 0.06 [log (Ca (++)) M], while IC50 in presence of verapamil (0.1 μM) is -1.69 ± 0.05 [log (Ca (++)) M]. Statins produced right shift in the IC50 of CCRCs. The effects of statins are like that of effects of verapamil, a standard calcium channel blocker. Our findings suggest that current statins have calcium antagonistic effects that act on voltage gated calcium channels that may provide a rationale for cause muscle weakness and gastrointestinal disorders.

Resistive switching behaviors of Au/pentacene/Si-nanowire arrays/heavily doped n-type Si devices for memory applications

NASA Astrophysics Data System (ADS)

Tsao, Hou-Yen; Lin, Yow-Jon

2014-02-01

The fabrication of memory devices based on the Au/pentacene/heavily doped n-type Si (n+-Si), Au/pentacene/Si nanowires (SiNWs)/n+-Si, and Au/pentacene/H2O2-treated SiNWs/n+-Si structures and their resistive switching characteristics were reported. A pentacene memory structure using SiNW arrays as charge storage nodes was demonstrated. The Au/pentacene/SiNWs/n+-Si devices show hysteresis behavior. H2O2 treatment may lead to the hysteresis degradation. However, no hysteresis-type current-voltage characteristics were observed for Au/pentacene/n+-Si devices, indicating that the resistive switching characteristic is sensitive to SiNWs and the charge trapping effect originates from SiNWs. The concept of nanowires within the organic layer opens a promising direction for organic memory devices.

Bradykinin induced a positive chronotropic effect via stimulation of T- and L-type calcium currents in heart cells.

PubMed

El-Bizri, Nesrine; Bkaily, Ghassan; Wang, Shimin; Jacques, Danielle; Regoli, Domenico; D'Orléans-Juste, Pedro; Sukarieh, Rami

2003-03-01

Using Fluo-3 calcium dye confocal microscopy and spontaneously contracting embryonic chick heart cells, bradykinin (10(-10) M) was found to induce positive chronotropic effects by increasing the frequency of the transient increase of cytosolic and nuclear free Ca2+. Pretreatment of the cells with either B1 or B2 receptor antagonists (R126 and R817, respectively) completely prevented bradykinin (BK) induced positive chronotropic effects on spontaneously contracting single heart cells. Using the whole-cell voltage clamp technique and ionic substitution to separate the different ionic current species, our results showed that BK (10(-6) M) had no effect on fast Na+ inward current and delayed outward potassium current. However, both L- and T-type Ca2+ currents were found to be increased by BK in a dose-dependent manner (10(-10)-10(-7) M). The effects of BK on T- and L-type Ca2+ currents were partially blocked by the B1 receptor antagonist [Leu8]des-Arg9-BK (R592) (10(-7) M) and completely reversed by the B2 receptor antagonist D-Arg[Hyp3,D-Phe7,Leu8]BK (R-588) (10(-7) M) or pretreatment with pertussis toxin (PTX). These results demonstrate that BK induced a positive chronotropic effect via stimulation of T- and L-type Ca2+ currents in heart cells mainly via stimulation of B2 receptor coupled to PTX-sensitive G-proteins. The increase of both types of Ca2+ current by BK in heart cells may explain the positive inotropic and chronotropic effects of this hormone.

Na+/Ca2+ exchange in cardiac myocytes. Effect of ouabain on voltage dependence.

PubMed

Lee, H C; Clusin, W T

1987-02-01

Sarcolemmal sodium/calcium exchange activity was examined in individual chick embryonic myocardial cell aggregates that were loaded with quin 2. The baseline [Ca2+]i was 68 +/- 4 nM (n = 29). Abrupt superfusion with sodium-free lithium solution produced a fourfold increase in steady-state [Ca2+]i to 290 +/- 19 nM, which was reversible upon sodium restitution. Other methods of increasing [Ca2+]i such as KCl-depolarization or caffeine produced a dose-dependent increase in quin 2 fluorescence, accompanied by sustained contracture. The [Ca2+]i increase in zero sodium was linear, and its half-time (t1/2) of 15.1 +/- 0.1 s was similar to that of the sodium-free contracture (t1/2 = 14.4 +/- 0.5 s) under the same conditions. The sodium-dependent [Ca2+]i increase was not significantly greater when potassium served as the sodium substitute instead of lithium. This suggests that sodium/calcium exchange has little voltage dependence in this situation. However, in aggregates pretreated with ouabain (2.5 microM), the [Ca2+]i increase was almost threefold greater with potassium than with lithium (P less than 0.007). Ouabain therefore potentiated the effect of membrane potential on calcium influx. We propose that elevation of [Na2+]i is a prerequisite for voltage dependence of the sodium/calcium exchange under the conditions studied. Sodium loading will then drastically increase calcium influx during the action potential while inducing an outward membrane current that could accelerate repolarization.

Genetically-encoded fluorescent voltage sensors using the voltage-sensing domain of Nematostella and Danio phosphatases exhibit fast kinetics

PubMed Central

Baker, Bradley J.; Jin, Lei; Han, Zhou; Cohen, Lawrence B.; Popovic, Marko; Platisa, Jelena; Pieribone, Vincent

2012-01-01

A substantial increase in the speed of the optical response of genetically-encoded Fluorescent Protein voltage sensors (FP voltage sensors) was achieved by using the voltage-sensing phosphatase genes of Nematostella vectensis and Danio rerio. A potential N. vectensis voltage-sensing phosphatase was identified in silico. The voltage-sensing domain (S1–S4) of the N. vectensis homolog was used to create an FP voltage sensor called Nema. By replacing the phosphatase with a cerulean/citrine FRET pair, a new FP voltage sensor was synthesized with fast off kinetics (Tauoff <5 msec). However, the signal was small (ΔF/F= 0.6%/200 mV). FP voltage sensors using the D. rerio voltage-sensing phosphatase homolog, designated Zahra and Zahra 2, exhibited fast on and off kinetics within 2 msec of the time constants observed with the organic voltage-sensitive dye, di4-ANEPPS. Mutagenesis of the S4 region of the Danio FP voltage sensor shifted the voltage dependence to more negative potentials but did not noticeably affect the kinetics of the optical signal. PMID:22634212

Calmodulin regulates Cav3 T-type channels at their gating brake

PubMed Central

Taiakina, Valentina; Monteil, Arnaud; Piazza, Michael; Guan, Wendy; Stephens, Robert F.; Dieckmann, Thorsten; Guillemette, Joseph Guy; Spafford, J. David

2017-01-01

Calcium (Cav1 and Cav2) and sodium channels possess homologous CaM-binding motifs, known as IQ motifs in their C termini, which associate with calmodulin (CaM), a universal calcium sensor. Cav3 T-type channels, which serve as pacemakers of the mammalian brain and heart, lack a C-terminal IQ motif. We illustrate that T-type channels associate with CaM using co-immunoprecipitation experiments and single particle cryo-electron microscopy. We demonstrate that protostome invertebrate (LCav3) and human Cav3.1, Cav3.2, and Cav3.3 T-type channels specifically associate with CaM at helix 2 of the gating brake in the I–II linker of the channels. Isothermal titration calorimetry results revealed that the gating brake and CaM bind each other with high-nanomolar affinity. We show that the gating brake assumes a helical conformation upon binding CaM, with associated conformational changes to both CaM lobes as indicated by amide chemical shifts of the amino acids of CaM in 1H-15N HSQC NMR spectra. Intact Ca2+-binding sites on CaM and an intact gating brake sequence (first 39 amino acids of the I–II linker) were required in Cav3.2 channels to prevent the runaway gating phenotype, a hyperpolarizing shift in voltage sensitivities and faster gating kinetics. We conclude that the presence of high-nanomolar affinity binding sites for CaM at its universal gating brake and its unique form of regulation via the tuning of the voltage range of activity could influence the participation of Cav3 T-type channels in heart and brain rhythms. Our findings may have implications for arrhythmia disorders arising from mutations in the gating brake or CaM. PMID:28972185

Simplification and analysis of models of calcium dynamics based on IP3-sensitive calcium channel kinetics.

PubMed

Tang, Y; Stephenson, J L; Othmer, H G

1996-01-01

We study the models for calcium (Ca) dynamics developed in earlier studies, in each of which the key component is the kinetics of intracellular inositol-1,4,5-trisphosphate-sensitive Ca channels. After rapidly equilibrating steps are eliminated, the channel kinetics in these models are represented by a single differential equation that is linear in the state of the channel. In the reduced kinetic model, the graph of the steady-state fraction of conducting channels as a function of log10(Ca) is a bell-shaped curve. Dynamically, a step increase in inositol-1,4,5-trisphosphate induces an incremental increase in the fraction of conducting channels, whereas a step increase in Ca can either potentiate or inhibit channel activation, depending on the Ca level before and after the increase. The relationships among these models are discussed, and experimental tests to distinguish between them are given. Under certain conditions the models for intracellular calcium dynamics are reduced to the singular perturbed form epsilon dx/d tau = f(x, y, p), dy/d tau = g(x, y, p). Phase-plane analysis is applied to a generic form of these simplified models to show how different types of Ca response, such as excitability, oscillations, and a sustained elevation of Ca, can arise. The generic model can also be used to study frequency encoding of hormonal stimuli, to determine the conditions for stable traveling Ca waves, and to understand the effect of channel properties on the wave speed.

Skin Barrier and Calcium.

PubMed

Lee, Sang Eun; Lee, Seung Hun

2018-06-01

Epidermal barrier formation and the maintenance of barrier homeostasis are essential to protect us from the external environments and organisms. Moreover, impaired keratinocytes differentiation and dysfunctional skin barrier can be the primary causes or aggravating factors for many inflammatory skin diseases including atopic dermatitis and psoriasis. Therefore, understanding the regulation mechanisms of keratinocytes differentiation and skin barrier homeostasis is important to understand many skin diseases and establish an effective treatment strategy. Calcium ions (Ca 2+ ) and their concentration gradient in the epidermis are essential in regulating many skin functions, including keratinocyte differentiation, skin barrier formation, and permeability barrier homeostasis. Recent studies have suggested that the intracellular Ca 2+ stores such as the endoplasmic reticulum (ER) are the major components that form the epidermal calcium gradient and the ER calcium homeostasis is crucial for regulating keratinocytes differentiation, intercellular junction formation, antimicrobial barrier, and permeability barrier homeostasis. Thus, both Ca 2+ release from intracellular stores, such as the ER and Ca 2+ influx mechanisms are important in skin barrier. In addition, growing evidences identified the functional existence and the role of many types of calcium channels which mediate calcium flux in keratinocytes. In this review, the origin of epidermal calcium gradient and their role in the formation and regulation of skin barrier are focused. We also focus on the role of ER calcium homeostasis in skin barrier. Furthermore, the distribution and role of epidermal calcium channels, including transient receptor potential channels, store-operated calcium entry channel Orai1, and voltage-gated calcium channels in skin barrier are discussed.

Cyclic AMP-dependent regulation of P-type calcium channels expressed in Xenopus oocytes.

PubMed

Fournier, F; Bourinet, E; Nargeot, J; Charnet, P

1993-05-01

Xenopus oocytes injected with rat cerebellum mRNA, express voltage-dependent calcium channels (VDCC). These were identified as P-type Ca2+ channels by their insensitivity to dihydropyridines and omega-conotoxin and by their blockade by Agelenopsis aperta venom (containing the funnel-web spider toxins: FTX and omega-Aga-IV-A). Coinjection of cerebellar mRNA and antisense oligonucleotide complementary to the dihydropyridine-resistant brain Ca2+ channel, named BI [Mori Y. et al. (1991) Nature 350:398-402] or rbA [Starr T. V. B. et al. (1991) Proc Natl Acad Sci USA 88:5621-5625], strongly reduced the expressed Ba2+ current suggesting that these clones encode a P-type VDCC. The macroscopic Ca2+ channel activity was increased by direct intraoocyte injection of cAMP. This increase in current amplitude was concomitant with a slowing of current inactivation, and was attributed to activation of protein kinase A, since it could be antagonized by a peptidic inhibitor of this enzyme. Positive regulation of P-type VDCC could be of importance in Purkinje neurons and motor nerve terminals where this channel is predominant.

Synaptic calcium regulation in hair cells of the chicken basilar papilla.

PubMed

Im, Gi Jung; Moskowitz, Howard S; Lehar, Mohammed; Hiel, Hakim; Fuchs, Paul Albert

2014-12-10

Cholinergic inhibition of hair cells occurs by activation of calcium-dependent potassium channels. A near-membrane postsynaptic cistern has been proposed to serve as a store from which calcium is released to supplement influx through the ionotropic ACh receptor. However, the time and voltage dependence of acetylcholine (ACh)-evoked potassium currents reveal a more complex relationship between calcium entry and release from stores. The present work uses voltage steps to regulate calcium influx during the application of ACh to hair cells in the chicken basilar papilla. When calcium influx was terminated at positive membrane potential, the ACh-evoked potassium current decayed exponentially over ∼100 ms. However, at negative membrane potentials, this current exhibited a secondary rise in amplitude that could be eliminated by dihydropyridine block of the voltage-gated calcium channels of the hair cell. Calcium entering through voltage-gated channels may transit through the postsynaptic cistern, since ryanodine and sarcoendoplasmic reticulum calcium-ATPase blockers altered the time course and magnitude of this secondary, voltage-dependent contribution to ACh-evoked potassium current. Serial section electron microscopy showed that efferent and afferent synaptic structures are juxtaposed, supporting the possibility that voltage-gated influx at afferent ribbon synapses influences calcium homeostasis during long-lasting cholinergic inhibition. In contrast, spontaneous postsynaptic currents ("minis") resulting from stochastic efferent release of ACh were made briefer by ryanodine, supporting the hypothesis that the synaptic cistern serves primarily as a calcium barrier and sink during low-level synaptic activity. Hypolemmal cisterns such as that at the efferent synapse of the hair cell can play a dynamic role in segregating near-membrane calcium for short-term and long-term signaling. Copyright © 2014 the authors 0270-6474/14/3416688-10$15.00/0.

Retinoic acid induction of calcium channel expression in human NT2N neurons.

PubMed

Gao, Z Y; Xu, G; Stwora-Wojczyk, M M; Matschinsky, F M; Lee, V M; Wolf, B A

1998-06-18

Ca2+ channel expression and regulation of intracellular Ca2+ homeostasis were studied during retinoic acid (RA)-induced differentiation of the human teratocarcinoma cell line Ntera 2/C1.D1 (NT2- cells) into NT2N neurons, a unique model of human neurons in culture. The cytosolic Ca2+ level of undifferentiated NT2- cells was low (75 +/- 5 nM) and stable under basal conditions, and it was only marginally decreased (by 9%) upon removal of extracellular Ca2+. After 10 microM RA treatment, NT2- cells were irreversibly differentiated into a phenotype of neuron-like NT2N cells. Cytosolic Ca2+ level of NT2N neurons was higher (106 +/- 14 nM) than that of NT2- cells and spontaneously fluctuated (0.208 +/- 0.038 transients/min) under basal conditions. Although K+ increased 86Rb fluxes in both NT2- cells and NT2N neurons, it only increased cytosolic Ca2+ level in NT2N neurons. The K+-induced increase in cytosolic Ca2+ in NT2N neurons was antagonized by 0.1-10 microM nifedipine or verapamil, 5 microM omega-CgTx GVIA, but not by 1 microM omega-agatoxin IVA, 1 microM omega-agatoxin TK, 1 microM FTX-3.3, or 100 microM Ni+ implicating L- and N-type voltage-dependent Ca2+ channels. In L- and N-type channels, but not in P- and Q-types, mRNAs were expressed in NT2N neurons as well as NT2- cells. Quantitative analysis of L- and N-type Ca2+ protein levels showed major differences between NT2- cells and NT2N neurons. In NT2- cells, N-type Ca2+ channels were undetectable while L-type channels levels were fivefold lower compared to NT2N neurons. Our findings show that L- and N-type channels are expressed during differentiation of NT2- cells into neurons, and that these voltage-dependent Ca2+ channels have a major role in regulating intracellular Ca2+ homeostasis and neuronal excitability. Copyright 1998 Academic Press.

Calcium dynamics regulating the timing of decision-making in C. elegans.

PubMed

Tanimoto, Yuki; Yamazoe-Umemoto, Akiko; Fujita, Kosuke; Kawazoe, Yuya; Miyanishi, Yosuke; Yamazaki, Shuhei J; Fei, Xianfeng; Busch, Karl Emanuel; Gengyo-Ando, Keiko; Nakai, Junichi; Iino, Yuichi; Iwasaki, Yuishi; Hashimoto, Koichi; Kimura, Koutarou D

2017-05-23

Brains regulate behavioral responses with distinct timings. Here we investigate the cellular and molecular mechanisms underlying the timing of decision-making during olfactory navigation in Caenorhabditis elegans . We find that, based on subtle changes in odor concentrations, the animals appear to choose the appropriate migratory direction from multiple trials as a form of behavioral decision-making. Through optophysiological, mathematical and genetic analyses of neural activity under virtual odor gradients, we further find that odor concentration information is temporally integrated for a decision by a gradual increase in intracellular calcium concentration ([Ca 2+ ] i ), which occurs via L-type voltage-gated calcium channels in a pair of olfactory neurons. In contrast, for a reflex-like behavioral response, [Ca 2+ ] i rapidly increases via multiple types of calcium channels in a pair of nociceptive neurons. Thus, the timing of neuronal responses is determined by cell type-dependent involvement of calcium channels, which may serve as a cellular basis for decision-making.

Release of mitochondrial glutathione and calcium by a cyclosporin A-sensitive mechanism occurs without large amplitude swelling.

PubMed

Savage, M K; Reed, D J

1994-11-15

Treatment of isolated mitochondria with calcium and inorganic phosphate induces inner membrane permeability that is thought to be mediated through a non-selective, calcium-dependent pore. The inner membrane permeability results in the rapid efflux of small matrix solutes such as glutathione and calcium, loss of coupled functions, and large amplitude swelling. We have identified conditions of permeability transition without large amplitude swelling, a parameter often used to assess inner membrane permeability. The addition of either oligomycin, antimycin, or sulfide to incubation buffer containing calcium and inorganic phosphate abolished large-amplitude swelling of mitochondria but did not prevent inner membrane permeability as demonstrated by the release of mitochondrial glutathione and calcium. The release of both glutathione and calcium was inhibited by the addition of cyclosporin A, a potent inhibitor of permeability transition. Transmission electron microscopy analysis, combined with the glutathione and calcium release data, indicate that permeability transition can be observed in the absence of large-amplitude swelling. Permeability transition occurring both with and without large-amplitude swelling was accompanied by a collapse of the membrane potential. We conclude that cyclosporin A-sensitive permeability transition can occur without obvious morphological changes such as large-amplitude swelling. Monitoring the cyclosporin A-sensitive release of concentrated endogenous matrix solutes, such as GSH, may be a sensitive and useful indicator of permeability transition.

CRMP-2 peptide mediated decrease of high and low voltage-activated calcium channels, attenuation of nociceptor excitability, and anti-nociception in a model of AIDS therapy-induced painful peripheral neuropathy

PubMed Central

2012-01-01

Background The ubiquity of protein-protein interactions in biological signaling offers ample opportunities for therapeutic intervention. We previously identified a peptide, designated CBD3, that suppressed inflammatory and neuropathic behavioral hypersensitivity in rodents by inhibiting the ability of collapsin response mediator protein 2 (CRMP-2) to bind to N-type voltage-activated calcium channels (CaV2.2) [Brittain et al. Nature Medicine 17:822–829 (2011)]. Results and discussion Here, we utilized SPOTScan analysis to identify an optimized variation of the CBD3 peptide (CBD3A6K) that bound with greater affinity to Ca2+ channels. Molecular dynamics simulations demonstrated that the CBD3A6K peptide was more stable and less prone to the unfolding observed with the parent CBD3 peptide. This mutant peptide, conjugated to the cell penetrating motif of the HIV transduction domain protein TAT, exhibited greater anti-nociception in a rodent model of AIDS therapy-induced peripheral neuropathy when compared to the parent TAT-CBD3 peptide. Remarkably, intraperitoneal administration of TAT-CBD3A6K produced none of the minor side effects (i.e. tail kinking, body contortion) observed with the parent peptide. Interestingly, excitability of dissociated small diameter sensory neurons isolated from rats was also reduced by TAT-CBD3A6K peptide suggesting that suppression of excitability may be due to inhibition of T- and R-type Ca2+ channels. TAT-CBD3A6K had no effect on depolarization-evoked calcitonin gene related peptide (CGRP) release compared to vehicle control. Conclusions Collectively, these results establish TAT-CBD3A6K as a peptide therapeutic with greater efficacy in an AIDS therapy-induced model of peripheral neuropathy than its parent peptide, TAT-CBD3. Structural modifications of the CBD3 scaffold peptide may result in peptides with selectivity against a particular subset of voltage-gated calcium channels resulting in a multipharmacology of action on the target. PMID

Mechanisms of calcium sequestration by isolated Malpighian tubules of the house cricket Acheta domesticus.

PubMed

Browne, Austin; O'Donnell, Michael J

2018-01-01

Hemolymph calcium homeostasis in insects is achieved by the Malpighian tubules, primarily by sequestering excess Ca 2+ within internal calcium stores (Ca-rich granules) most often located within type I (principal) tubule cells. Using both the scanning ion-selective electrode technique and the Ramsay secretion assay this study provides the first measurements of basolateral and transepithelial Ca 2+ fluxes across the Malpighian tubules of an Orthopteran insect, the house cricket Acheta domesticus. Ca 2+ transport was specific to midtubule segments, where 97% of the Ca 2+ entering the tubule is sequestered within intracellular calcium stores and the remaining 3% is secreted into the lumen. Antagonists of voltage-gated (L-type) calcium channels decreased Ca 2+ influx ≥fivefold in adenosine 3',5'-cyclic monophosphate (cAMP)-stimulated tubules, suggesting basolateral Ca 2+ influx is facilitated by voltage-gated Ca 2+ channels. Increasing fluid secretion through manipulation of intracellular levels of cAMP or Ca 2+ had opposite effects on tubule Ca 2+ transport. The adenylyl cyclase-cAMP-PKA pathway promotes Ca 2+ sequestration whereas both 5-hydroxytryptamine and thapsigargin inhibited sequestration. Our results suggest that the midtubules of Acheta domesticus are dynamic calcium stores, which maintain hemolymph calcium concentration by manipulating rates of Ca 2+ sequestration through stimulatory (cAMP) and inhibitory (Ca 2+ ) regulatory pathways. © 2017 Wiley Periodicals, Inc.

Physiology and Evolution of Voltage-Gated Calcium Channels in Early Diverging Animal Phyla: Cnidaria, Placozoa, Porifera and Ctenophora

PubMed Central

Senatore, Adriano; Raiss, Hamad; Le, Phuong

2016-01-01

Voltage-gated calcium (Cav) channels serve dual roles in the cell, where they can both depolarize the membrane potential for electrical excitability, and activate transient cytoplasmic Ca2+ signals. In animals, Cav channels play crucial roles including driving muscle contraction (excitation-contraction coupling), gene expression (excitation-transcription coupling), pre-synaptic and neuroendocrine exocytosis (excitation-secretion coupling), regulation of flagellar/ciliary beating, and regulation of cellular excitability, either directly or through modulation of other Ca2+-sensitive ion channels. In recent years, genome sequencing has provided significant insights into the molecular evolution of Cav channels. Furthermore, expanded gene datasets have permitted improved inference of the species phylogeny at the base of Metazoa, providing clearer insights into the evolution of complex animal traits which involve Cav channels, including the nervous system. For the various types of metazoan Cav channels, key properties that determine their cellular contribution include: Ion selectivity, pore gating, and, importantly, cytoplasmic protein-protein interactions that direct sub-cellular localization and functional complexing. It is unclear when these defining features, many of which are essential for nervous system function, evolved. In this review, we highlight some experimental observations that implicate Cav channels in the physiology and behavior of the most early-diverging animals from the phyla Cnidaria, Placozoa, Porifera, and Ctenophora. Given our limited understanding of the molecular biology of Cav channels in these basal animal lineages, we infer insights from better-studied vertebrate and invertebrate animals. We also highlight some apparently conserved cellular functions of Cav channels, which might have emerged very early on during metazoan evolution, or perhaps predated it. PMID:27867359

Initial leakage current paths in the vertical-type GaN-on-GaN Schottky barrier diodes

NASA Astrophysics Data System (ADS)

Sang, Liwen; Ren, Bing; Sumiya, Masatomo; Liao, Meiyong; Koide, Yasuo; Tanaka, Atsushi; Cho, Yujin; Harada, Yoshitomo; Nabatame, Toshihide; Sekiguchi, Takashi; Usami, Shigeyoshi; Honda, Yoshio; Amano, Hiroshi

2017-09-01

Electrical characteristics of leakage current paths in vertical-type n-GaN Schottky barrier diodes (SBDs) on free-standing GaN substrates are investigated by using photon emission microscopy (PEM). The PEM mapping shows that the initial failure of the SBD devices at low voltages is due to the leakage current paths from polygonal pits in the GaN epilayers. It is observed that these polygonal pits originate from carbon impurity accumulation to the dislocations with a screw-type component by microstructure analysis. For the SBD without polygonal pits, no initial failure is observed and the first leakage appeals at the edge of electrodes as a result of electric field concentration. The mechanism of leakage at pits is explained in terms of trap assisted tunneling through fitting current-voltage characteristics.

TMEM16A is associated with voltage-gated calcium channels in mouse retina and its function is disrupted upon mutation of the auxiliary α2δ4 subunit

PubMed Central

Caputo, Antonella; Piano, Ilaria; Demontis, Gian Carlo; Bacchi, Niccolò; Casarosa, Simona; Santina, Luca Della; Gargini, Claudia

2015-01-01

Photoreceptors rely upon highly specialized synapses to efficiently transmit signals to multiple postsynaptic targets. Calcium influx in the presynaptic terminal is mediated by voltage-gated calcium channels (VGCC). This event triggers neurotransmitter release, but also gates calcium-activated chloride channels (TMEM), which in turn regulate VGCC activity. In order to investigate the relationship between VGCC and TMEM channels, we analyzed the retina of wild type (WT) and Cacna2d4 mutant mice, in which the VGCC auxiliary α2δ4 subunit carries a nonsense mutation, disrupting the normal channel function. Synaptic terminals of mutant photoreceptors are disarranged and synaptic proteins as well as TMEM16A channels lose their characteristic localization. In parallel, calcium-activated chloride currents are impaired in rods, despite unaltered TMEM16A protein levels. Co-immunoprecipitation revealed the interaction between VGCC and TMEM16A channels in the retina. Heterologous expression of these channels in tsA-201 cells showed that TMEM16A associates with the CaV1.4 subunit, and the association persists upon expression of the mutant α2δ4 subunit. Collectively, our experiments show association between TMEM16A and the α1 subunit of VGCC. Close proximity of these channels allows optimal function of the photoreceptor synaptic terminal under physiological conditions, but also makes TMEM16A channels susceptible to changes occurring to calcium channels. PMID:26557056

Impact of calcium-sensitive dyes on the beating properties and pharmacological responses of human iPS-derived cardiomyocytes using the calcium transient assay.

PubMed

Kopljar, Ivan; Hermans, An N; Teisman, Ard; Gallacher, David J; Lu, Hua Rong

Calcium-based screening of hiPS-CMs is a useful preclinical safety evaluation platform with the ability to generate robust signals that facilitates high-throughput screening and data analysis. However, due to the potential inherent toxicities, it is important to understand potential effects of different calcium-sensitive dyes on the hiPS-CMs model. We compared three calcium-sensitive fluorescence dyes (Cal520, ACTOne and Calcium 5) for their impact on the variability, the beating properties and the pharmacological responses of hiPS-CMs using the Hamamatsu FDSS/μCell imaging platform. Direct effects of three dyes on the electrophysiological properties of hiPS-CMs were evaluated with the multi-electrode array (MEA) Axion Maestro platform. We propose a specific experimental protocol for each dye which gives the most optimal assay conditions to minimize variability and possible adverse effects. We showed that Cal520 had the smallest effect on hiPS-CMs together with the longest-lasting stable amplitude signal (up to 4 h). Although all dyes had a (minor) acute effect on hiPS-CMs, in the form of reduced beat rate and prolonged field potential duration, the selection of the dye did not influence the pharmacological response of four cardioactive drugs (dofetilide, moxifloxacin, nimodipine and isoprenaline). In conclusion, we have documented that different calcium sensitive dyes have only minor direct (acute) effects on hiPS-CMs with Cal520 showing the least effects and the longest lasting signal amplitude. Importantly, drug-induced pharmacological responses in hiPS-CMs were comparable between the three dyes. These findings should help further improve the robustness of the hiPS-CMs-based calcium transient assay as a predictive, preclinical cardiac safety evaluation tool. Copyright © 2018 Elsevier Inc. All rights reserved.

Dye molecular structure device open-circuit voltage correlation in Ru(II) sensitizers with heteroleptic tridentate chelates for dye-sensitized solar cells.

PubMed

Wu, Kuan-Lin; Li, Cheng-Hsuan; Chi, Yun; Clifford, John N; Cabau, Lydia; Palomares, Emilio; Cheng, Yi-Ming; Pan, Hsiao-An; Chou, Pi-Tai

2012-05-02

Dicarboxyterpyridine chelates with π-conjugated pendant groups attached at the 5- or 6-position of the terminal pyridyl unit were synthesized. Together with 2,6-bis(5-pyrazolyl)pyridine, these were used successfully to prepare a series of novel heteroleptic, bis-tridentate Ru(II) sensitizers, denoted as TF-11-14. These dyes show excellent performance in dye-sensitized solar cells (DSCs) under AM1.5G simulated sunlight at a light intensity of 100 mW cm(-2) in comparison with a reference device containing [Ru(Htctpy)(NCS)(3)][TBA](3) (N749), where H(3)tctpy and TBA are 4,4',4"-tricarboxy-2,2':6',2"-terpyridine and tetra-n-butylammonium cation, respectively. In particular, the sensitizer TF-12 gave a short-circuit photocurrent of 19.0 mA cm(-2), an open-circuit voltage (V(OC)) of 0.71 V, and a fill factor of 0.68, affording an overall conversion efficiency of 9.21%. The increased conjugation conferred to the TF dyes by the addition of the π-conjugated pendant groups increases both their light-harvesting and photovoltaic energy conversion capability in comparison with N749. Detailed recombination processes in these devices were probed by various spectroscopic and dynamics measurements, and a clear correlation between the device V(OC) and the cell electron lifetime was established. In agreement with several other recent studies, the results demonstrate that high efficiencies can also be achieved with Ru(II) sensitizers that do not contain thiocyanate ancillaries. This bis-tridentate, dual-carboxy anchor configuration thus serves as a prototype for future omnibearing design of highly efficient Ru(II) sensitizers suited for use in DSCs. © 2012 American Chemical Society

A spot laser modulated resistance switching effect observed on n-type Mn-doped ZnO/SiO2/Si structure.

PubMed

Lu, Jing; Tu, Xinglong; Yin, Guilin; Wang, Hui; He, Dannong

2017-11-09

In this work, a spot laser modulated resistance switching (RS) effect is firstly observed on n-type Mn-doped ZnO/SiO 2 /Si structure by growing n-type Mn-doped ZnO film on Si wafer covered with a 1.2 nm native SiO 2 , which has a resistivity in the range of 50-80 Ω∙cm. The I-V curve obtained in dark condition evidences the structure a rectifying junction, which is further confirmed by placing external bias. Compared to the resistance state modulated by electric field only in dark (without illumination), the switching voltage driving the resistance state of the structure from one state to the other, shows clear shift under a spot laser illumination. Remarkably, the switching voltage shift shows a dual dependence on the illumination position and power of the spot laser. We ascribe this dual dependence to the electric filed produced by the redistribution of photo-generated carriers, which enhance the internal barrier of the hetero-junction. A complete theoretical analysis based on junction current and diffusion equation is presented. The dependence of the switching voltage on spot laser illumination makes the n-type Mn-doped ZnO/SiO 2 /Si structure sensitive to light, which thus allows for the integration of an extra functionality in the ZnO-based photoelectric device.

Calcium Homeostatasis and Mitochondrial Dysfunction in Dopaminergic Neurons of the Substantia Nigra

DTIC Science & Technology

2010-03-01

discovery that calcium entry through L-type channels during normal pacemaking elevates the sensitivity of SNc dopaminergic neurons to toxins; • the...discovery that L-type calcium channels participate in but are not necessary for pacemaking; • the discovery that serum concentration of the...FDA approved doses; • the discovery that calcium entry through L-type channels during pacemaking elevates mitochondrial oxidant stress and leads

Process dependency on threshold voltage of GaN MOSFET on AlGaN/GaN heterostructure

NASA Astrophysics Data System (ADS)

Wang, Qingpeng; Jiang, Ying; Miyashita, Takahiro; Motoyama, Shin-ichi; Li, Liuan; Wang, Dejun; Ohno, Yasuo; Ao, Jin-Ping

2014-09-01

GaN metal-oxide-semiconductor field-effect transistors (MOSFETs) with recessed gate on AlGaN/GaN heterostructure are reported in which the drain and source ohmic contacts were fabricated on the AlGaN/GaN heterostructure and the electron channel was formed on the GaN buffer layer by removing the AlGaN barrier layer. Negative threshold voltages were commonly observed in all devices. To investigate the reasons of the negative threshold voltages, different oxide thickness, etching gas and bias power of inductively-coupled plasma (ICP) system were utilized in the fabrication process of the GaN MOSFETs. It is found that positive charges of around 1 × 1012 q/cm2 exist near the interface at the just threshold condition in both silane- and tetraethylorthosilicate (TEOS)-based devices. It is also found that the threshold voltages do not obviously change with the different etching gas (SiCl4, BCl3 and two-step etching of SiCl4/Cl2) at the same ICP bias power level (20-25 W) and will become deeper when higher bias power is used in the dry recess process which may be related to the much serious ion bombardment damage. Furthermore, X-ray photoelectron spectroscopy (XPS) experiments were done to investigate the surface conditions. It is found that N 1s peaks become lower with higher bias power of the dry etching process. Also, silicon contamination was found and could be removed by HNO3/HF solution. It indicates that the nitrogen vacancies are mainly responsible for the negative threshold voltages rather than the silicon contamination. It demonstrates that optimization of the ICP recess conditions and improvement of the surface condition are still necessary to realize enhancement-mode GaN MOSFETs on AlGaN/GaN heterostructure.

PYK2: A Calcium-sensitive Protein Tyrosine Kinase Activated in Response to Fertilization of the Zebrafish Oocyte

PubMed Central

Sharma, Dipika; Kinsey, William H.

2012-01-01

Fertilization begins with binding and fusion of a sperm with the oocyte, a process that triggers a high amplitude calcium transient which propagates through the oocyte and stimulates a series of preprogrammed signal transduction events critical for zygote development. Identification of the pathways downstream of this calcium transient remains an important step in understanding the basis of zygote quality. The present study demonstrates that the calcium-calmodulin sensitive protein tyrosine kinase PYK2 is a target of the fertilization-induced calcium transient in the zebrafish oocyte and that it plays an important role in actin-mediated events critical for sperm incorporation. At fertilization, PYK2 was activated initially at the site of sperm-oocyte interaction and was closely associated with actin filaments forming the fertilization cone. Later PYK2 activation was evident throughout the entire oocyte cortex, however activation was most intense over the animal hemisphere. Fertilization-induced PYK2 activation could be blocked by suppressing calcium transients in the ooplasm via injection of BAPTA as a calcium chelator. PYK2 activation could be artificially induced in unfertilized oocytes by injection of IP3 at concentrations sufficient to induce calcium release. Functionally, suppression of PYK2 activity by chemical inhibition or by injection of a dominant-negative construct encoding the N-terminal ERM domain of PKY2 inhibited formation of an organized fertilization cone and reduced the frequency of successful sperm incorporation. Together, the above findings support a model in which PYK2 responds to the fertilization-induced calcium transient by promoting reorganization of the cortical actin cytoskeleton to form the fertilization cone. PMID:23084926

Genetically encoded fluorescent voltage sensors using the voltage-sensing domain of Nematostella and Danio phosphatases exhibit fast kinetics.

PubMed

Baker, Bradley J; Jin, Lei; Han, Zhou; Cohen, Lawrence B; Popovic, Marko; Platisa, Jelena; Pieribone, Vincent

2012-07-15

A substantial increase in the speed of the optical response of genetically encoded fluorescent protein voltage sensors (FP voltage sensors) was achieved by using the voltage-sensing phosphatase genes of Nematostella vectensis and Danio rerio. A potential N. vectensis voltage-sensing phosphatase was identified in silico. The voltage-sensing domain (S1-S4) of the N. vectensis homolog was used to create an FP voltage sensor called Nema. By replacing the phosphatase with a cerulean/citrine FRET pair, a new FP voltage sensor was synthesized with fast off kinetics (Tau(off)<5ms). However, the signal was small (ΔF/F=0.4%/200mV). FP voltage sensors using the D. rerio voltage-sensing phosphatase homolog, designated Zahra and Zahra 2, exhibited fast on and off kinetics within 2ms of the time constants observed with the organic voltage-sensitive dye, di4-ANEPPS. Mutagenesis of the S4 region of the Danio FP voltage sensor shifted the voltage dependence to more negative potentials but did not noticeably affect the kinetics of the optical signal. Copyright © 2012 Elsevier B.V. All rights reserved.

Action of aluminum, novel TPC1-type channel inhibitor, against salicylate-induced and cold-shock-induced calcium influx in tobacco BY-2 cells.

PubMed

Lin, Cun; Yu, Yawei; Kadono, Takashi; Iwata, Michiaki; Umemura, Kenji; Furuichi, Takuya; Kuse, Masaki; Isobe, Minoru; Yamamoto, Yoko; Matsumoto, Hideaki; Yoshizuka, Kazuharu; Kawano, Tomonori

2005-07-08

Previously, effect of Al ions on calcium signaling was assessed in tobacco cells expressing a Ca2+-monitoring luminescent protein, aequorin and a newly isolated putative plant Ca2+ channel protein from Arabidopsis thaliana, AtTPC1 (two-pore channel 1). TPC1 channels were shown to be the only channel known to be sensitive to Al and they are responsive to reactive oxygen species and cryptogein, a fungal elicitor protein. Thus, involvement of TPC1 channels in calcium signaling leading to development of plant defense mechanism has been suggested. Then, the use of Al as a specific inhibitor of TPC1-type plant calcium channels has been proposed. Here, using transgenic tobacco BY-2 cells expressing aequorin, we report on the evidence in support of the involvement of Al-sensitive signaling pathway requiring TPC1-type channel-dependent Ca2+ influx in response to salicylic acid, a key plant defense-inducing agent, but not to an elicitor prepared from the cell wall of rice blast disease fungus Magnaporthe grisea. In addition, involvement of Al-sensitive Ca2+ channels in response to cold shock was also tested. The data suggested that the elicitor used here induces the Ca2+ influx via Al-insensitive path, while salicylic acid and cold-shock-stimulate the influx of Ca2+ via Al-sensitive mechanism.

Role of voltage-gated L-type Ca2+ channel isoforms for brain function.

PubMed

Striessnig, J; Koschak, A; Sinnegger-Brauns, M J; Hetzenauer, A; Nguyen, N K; Busquet, P; Pelster, G; Singewald, N

2006-11-01

Voltage-gated LTCCs (L-type Ca2+ channels) are established drug targets for the treatment of cardiovascular diseases. LTCCs are also expressed outside the cardiovascular system. In the brain, LTCCs control synaptic plasticity in neurons, and DHP (dihydropyridine) LTCC blockers such as nifedipine modulate brain function (such as fear memory extinction and depression-like behaviour). Voltage-sensitive Ca2+ channels Cav1 .2 and Cav1.3 are the predominant brain LTCCs. As DHPs and other classes of organic LTCC blockers inhibit both isoforms, their pharmacological distinction is impossible and their individual contributions to defined brain functions remain largely unknown. Here, we summarize our recent experiments with two genetically modified mouse strains, which we generated to explore the individual biophysical features of Cav1.2 and Cav1.3 LTCCs and to determine their relative contributions to various physiological peripheral and neuronal functions. The results described here also allow predictions about the pharmacotherapeutic potential of isoform-selective LTCC modulators.

Low voltage operation of GaN vertical nanowire MOSFET

NASA Astrophysics Data System (ADS)

Son, Dong-Hyeok; Jo, Young-Woo; Seo, Jae Hwa; Won, Chul-Ho; Im, Ki-Sik; Lee, Yong Soo; Jang, Hwan Soo; Kim, Dae-Hyun; Kang, In Man; Lee, Jung-Hee

2018-07-01

GaN gate-all-around (GAA) vertical nanowire MOSFET (VNWMOSFET) with channel length of 300 nm and diameter of 120 nm, the narrowest GaN-based vertical nanowire transistor ever achieved from the top-down approach, was fabricated by utilizing anisotropic side-wall wet etching in TMAH solution and photoresist etch-back process. The VNWMOSFET exhibited output characteristics with very low saturation drain voltage of less than 0.5 V, which is hardly observed from the wide bandgap-based devices. Simulation results indicated that the narrow diameter of the VNWMOSFET with relatively short channel length is responsible for the low voltage operation. The VNWMOSFET also demonstrated normally-off mode with threshold voltage (VTH) of 0.7 V, extremely low leakage current of ∼10-14 A, low drain-induced barrier lowering (DIBL) of 125 mV/V, and subthreshold swing (SS) of 66-122 mV/decade. The GaN GAA VNWMOSFET with narrow channel diameter investigated in this work would be promising for new low voltage logic application. He has been a Professor with the School of Electrical Engineering and Computer Science, Kyungpook National University, Daegu, Korea, since 1993

Calcium-Mediated Apoptosis and Apoptotic Sensitization in Prostate Cancer

DTIC Science & Technology

2004-06-01

calcium- sensitive protease calpain, stimulating two distinct pathways that regulate phosphotyrosine-initiated cell signaling ( PTP1B ) or directly...trigger apoptosis (caspase 7). The role of caspase 7 and PTP1B in PC cell death and survival signaling was investigated using dominant negatives, siRNA...of a calpain-proteolyzed variant of PTP1B (tPTP1B) had minimal impact on growth-factor or cytokine-mediated tyrosine phosphorylation or cell

Modulation of voltage-gated channel currents by harmaline and harmane.

PubMed

Splettstoesser, Frank; Bonnet, Udo; Wiemann, Martin; Bingmann, Dieter; Büsselberg, Dietrich

2005-01-01

Harmala alkaloids are endogenous substances, which are involved in neurodegenerative disorders such as M. Parkinson, but some of them also have neuroprotective effects in the nervous system. While several sites of action at the cellular level (e.g. benzodiazepine receptors, 5-HT and GABA(A) receptors) have been identified, there is no report on how harmala alkaloids interact with voltage-gated membrane channels. The aim of this study was to investigate the effects of harmaline and harmane on voltage-activated calcium- (I(Ca(V))), sodium- (I(Na(V))) and potassium (I(K(V)))-channel currents, using the whole-cell patch-clamp method with cultured dorsal root ganglion neurones of 3-week-old rats. Currents were elicited by voltage steps from the holding potential to different command potentials. Harmaline and harmane reduced I(Ca(V)), I(Na(V)) and I(K(V)) concentration-dependent (10-500 microM) over the voltage range tested. I(Ca(V)) was reduced with an IC(50) of 100.6 microM for harmaline and by a significantly lower concentration of 75.8 microM (P<0.001, t-test) for harmane. The Hill coefficient was close to 1. Threshold concentration was around 10 microM for both substances. The steady state of inhibition of I(Ca(V)) by harmaline or harmane was reached within several minutes. The action was not use-dependent and at least partly reversible. It was mainly due to a reduction in the sustained calcium channel current (I(Ca(L+N))), while the transient voltage-gated calcium channel current (I(Ca(T))) was only partially affected. We conclude that harmaline and harmane are modulators of I(Ca(V)) in vitro. This might be related to their neuroprotective effects.

Modulation of voltage-gated channel currents by harmaline and harmane

PubMed Central

Splettstoesser, Frank; Bonnet, Udo; Wiemann, Martin; Bingmann, Dieter; Büsselberg, Dietrich

2004-01-01

Harmala alkaloids are endogenous substances, which are involved in neurodegenerative disorders such as M. Parkinson, but some of them also have neuroprotective effects in the nervous system. While several sites of action at the cellular level (e.g. benzodiazepine receptors, 5-HT and GABAA receptors) have been identified, there is no report on how harmala alkaloids interact with voltage-gated membrane channels. The aim of this study was to investigate the effects of harmaline and harmane on voltage-activated calcium- (ICa(V)), sodium- (INa(V)) and potassium (IK(V))-channel currents, using the whole-cell patch-clamp method with cultured dorsal root ganglion neurones of 3-week-old rats. Currents were elicited by voltage steps from the holding potential to different command potentials. Harmaline and harmane reduced ICa(V), INa(V) and IK(V) concentration-dependent (10–500 μM) over the voltage range tested. ICa(V) was reduced with an IC50 of 100.6 μM for harmaline and by a significantly lower concentration of 75.8 μM (P<0.001, t-test) for harmane. The Hill coefficient was close to 1. Threshold concentration was around 10 μM for both substances. The steady state of inhibition of ICa(V) by harmaline or harmane was reached within several minutes. The action was not use dependent and at least partly reversible. It was mainly due to a reduction in the sustained calcium channel current (ICa(L+N)), while the transient voltage-gated calcium channel current (ICa(T)) was only partially affected. We conclude that harmaline and harmane are modulators of ICa(V) in vitro. This might be related to their neuroprotective effects. PMID:15644868

Evaluation of modulating field of photoreflectance of surface-intrinsic-n+ type doped GaAs by using photoinduced voltage

NASA Astrophysics Data System (ADS)

Lee, W. Y.; Chien, J. Y.; Wang, D. P.; Huang, K. F.; Huang, T. C.

2002-04-01

Photoreflectance (PR) of surface-intrinsic-n+ type doped GaAs has been measured for various power densities of pump laser. The spectra exhibited many Franz-Keldysh oscillations, whereby the strength of electric field F in the undoped layer can be determined. The thus obtained Fs are subject to photovoltaic effect and are less than built-in field Fbi. In the previous work we have obtained the relation F≈Fbi-δF/2 when δF≪Fbi by using electroreflectance to simulate PR, where δF is the modulating field of the pump beam. In this work a method was devised to evaluate δF by using photoinduced voltages Vs and, hence, the relation can be verified by PR itself. The δFs obtained by Vs are also consistent with those of using imaginary part of fast Fourier transform of PR spectra.

Myosin light chain kinase controls voltage-dependent calcium channels in vascular smooth muscle.

PubMed

Martinsen, A; Schakman, O; Yerna, X; Dessy, C; Morel, N

2014-07-01

The Ca(2+)-dependent kinase myosin light chain kinase (MLCK) is the activator of smooth muscle contraction. In addition, it has been reported to be involved in Ca(2+) channel regulation in cultured cells, and we previously showed that the MLCK inhibitor ML-7 decreases arginine vasopressin (AVP)-induced Ca(2+) influx in rat aorta. This study was designed to investigate whether MLCK is involved in Ca(2+) regulation in resistance artery smooth muscle cell, which plays a major role in the control of blood pressure. As ML compounds were shown to have off-target effects, MLCK was downregulated by transfection with a small interfering RNA targeting MLCK (MLCK-siRNA) in rat small resistance mesenteric artery (RMA) and in the rat embryonic aortic cell line A7r5. Noradrenaline-induced contraction and Ca(2+) signal were significantly depressed in MLCK-siRNA compared to scramble-siRNA-transfected RMA. Contraction and Ca(2+) signal induced by high KCl and voltage-activated Ca(2+) current were also significantly decreased in MLCK-siRNA-transfected RMA, suggesting that MLCK depletion modifies voltage-operated Ca(2+) channels. KCl- and AVP-induced Ca(2+) signals and voltage-activated Ca(2+) current were decreased in MLCK-depleted A7r5 cells. Eventually, real-time quantitative PCR analysis indicated that in A7r5, MLCK controlled mRNA expression of CaV1.2 (L-type) and CaV3.1 (T-type) voltage-dependent Ca(2+) channels. Our results suggest that MLCK controls the transcription of voltage-dependent Ca(2+) channels in vascular smooth muscle cells.

Analysis of the Temperature Dependence of the Capacitance-Voltage and Conductance-Voltage Characteristics of Au/TiO2(rutile)/ n-Si Structures

NASA Astrophysics Data System (ADS)

KInacI, BarIş; Özçelik, Süleyman

2013-06-01

The capacitance-voltage-temperature ( C- V- T) and the conductance/angular frequency-voltage-temperature ( G/ω- V- T) characteristics of Au/TiO2(rutile)/ n-Si Schottky barrier diodes (SBDs) were investigated over the temperature range from 200 K to 380 K by considering the series resistance effect. Titanium dioxide (TiO2) was deposited on n-type silicon (Si) substrate using a direct-current (DC) magnetron sputtering system at 200°C. To improve the crystal quality, the deposited film was annealed at 900°C to promote a phase transition from the amorphous to rutile phase. The C -2 versus V plots gave a straight line in the reverse-bias region. The main electrical parameters, such as the doping concentration ( N D), Fermi energy level ( E F), depletion layer width ( W D), barrier height ( ф CV), and series resistance ( R S), of Au/TiO2(rutile)/ n-Si SBDs were calculated from the C- V- T and the G/ω- V- T characteristics. The obtained results show that ф CV, R S, and W D values decrease, while E F and N D values increase, with increasing temperature.

Mapping Calcium Rich Ejecta in Two Type Ia Supernovae

NASA Astrophysics Data System (ADS)

Fesen, Robert

2016-10-01

Type Ia supernovae (SNe Ia) are thermonuclear explosions of white dwarfs (WDs) in close binary systems with either a non-degenerate or WD companion. SN Ia explosion computations are quite challenging, involving a complex interplay of turbulent hydrodynamics, nuclear burning, conduction, radiative transfer in iron-group rich material and possibly magnetic fields leading to significant uncertainties. Several key questions about expansion asymmetries and the overall characteristics of SNe Ia could be resolved if one could obtain direct observations of the internal kinematics and elemental distributions of young SN Ia remnants.We propose to use WFC3/UVIS to obtain images of the normal Type Ia supernova remnant 0519-69.0 and the overluminous Type Ia supernova remnant 0509-67.5 in the LMC. The Ca II on-band F390M filter and off-band F336W and FQ422M filters will be used to determine the spatial extent and density distributions of the Ca-rich ejecta via resonance line absorption. Differences in the observed on and off band Ca II fluxes for LMC stars located behind these young 400 - 600 yr old remnants will yield calcium column density estimates for multiple lines-of-sight within these remnants. These results will be compared to the calcium distribution seen in SN 1885, a subluminous SN Ia in M31, already imaged by HST.The resulting calcium density distribution maps for both a normal and overluminous SN Ia events will provide powerful insights regarding the structure and kinematics of calcium-rich ejecta in three different type Ia subclass events, and unique empirical data with which to test current SN Ia explosion models.

Current-voltage characteristics of n-AlMgZnO/p-GaN junction diodes

NASA Astrophysics Data System (ADS)

Hsueh, Kuang-Po; Cheng, Po-Wei; Cheng, Yi-Chang; Sheu, Jinn-Kong; Yeh, Yu-Hsiang; Chiu, Hsien-Chin; Wang, Hsiang-Chun

2013-03-01

This study investigates the temperature dependence of the current-voltage (I-V) characteristics of Al-doped MgxZn1-xO/p-GaN junction diodes. Specifically, this study reports the deposition of n-type Al-doped MgxZn1-xO (AMZO) films on p-GaN using a radio-frequency (RF) magnetron sputtering system followed by annealing at 700, 800, 900, and 1000 °C in a nitrogen ambient for 60 seconds, respectively. The AMZO/GaN films were thereafter analyzed using Hall measurement and the x-ray diffraction (XRD) patterns. The XRD results show that the diffraction angles of the annealed AMZO films remain the same as that of GaN without shifting. The n-AMZO/p-GaN diode with 900 °C annealing had the lowest leakage current in forward and reverse bias. However, the leakage current of the diodes did not change significantly with an increase in annealing temperatures. These findings show that the n-AMZO/p-GaN junction diode is feasible for GaN-based heterojunction bipolar transistors (HBTs) and UV light-emitting diodes (LEDs).

Hysteresis effect implicates calcium cycling as a mechanism of repolarization alternans.

PubMed

Walker, Mariah L; Wan, Xiaoping; Kirsch, Glenn E; Rosenbaum, David S

2003-11-25

T-wave alternans is due to alternation of membrane repolarization at the cellular level and is a risk factor for sudden cardiac death. Recently, a hysteresis effect has been reported in patients whereby T-wave alternans, once induced by rapid heart rate, persists even when heart rate is subsequently slowed. We hypothesized that alternans hysteresis is an intrinsic property of cardiac myocytes, directly related to an underlying mechanism for repolarization alternans that involves intracellular calcium cycling. Stepwise pacing was used to induce alternans in Langendorff-perfused guinea pig hearts from which optical action potentials were recorded simultaneously at 256 ventricular sites with voltage-sensitive dyes and in whole-cell patch-clamped cardiac myocytes treated with or without BAPTA-AM (1,2-bis[2-aminophenoxy]ethane-N,N,N',N'-tetraacetic acid tetrakis [acetoxymethyl ester]). Alternans hysteresis was observed in every isolated heart: threshold heart rate for alternans was 280+/-12 bpm, but during subsequent deceleration of pacing, alternans persisted to significantly slower heart rates (238+/-5 bpm, P<0.05). Optical mapping showed that this effect also applied to the threshold for spatially discordant alternans (313+/-2.2 bpm during acceleration versus 250+/-6.6 bpm during deceleration, P<0.05). Alternans hysteresis was also observed in isolated cardiac myocytes. Moreover, calcium chelation by BAPTA-AM raised the threshold for alternans and inhibited hysteresis in a dose-dependent manner with no effect on baseline action potential duration. Alternans hysteresis is an intrinsic property of cardiac myocytes that can lead to persistence of arrhythmogenic discordant alternans even after heart rate is slowed. These results also support an important underlying role of calcium cycling in the mechanism of alternans.

Regulation of Cardiac ATP-sensitive Potassium Channel Surface Expression by Calcium/Calmodulin-dependent Protein Kinase II*

PubMed Central

Sierra, Ana; Zhu, Zhiyong; Sapay, Nicolas; Sharotri, Vikas; Kline, Crystal F.; Luczak, Elizabeth D.; Subbotina, Ekaterina; Sivaprasadarao, Asipu; Snyder, Peter M.; Mohler, Peter J.; Anderson, Mark E.; Vivaudou, Michel; Zingman, Leonid V.; Hodgson-Zingman, Denice M.

2013-01-01

Cardiac ATP-sensitive potassium (KATP) channels are key sensors and effectors of the metabolic status of cardiomyocytes. Alteration in their expression impacts their effectiveness in maintaining cellular energy homeostasis and resistance to injury. We sought to determine how activation of calcium/calmodulin-dependent protein kinase II (CaMKII), a central regulator of calcium signaling, translates into reduced membrane expression and current capacity of cardiac KATP channels. We used real-time monitoring of KATP channel current density, immunohistochemistry, and biotinylation studies in isolated hearts and cardiomyocytes from wild-type and transgenic mice as well as HEK cells expressing wild-type and mutant KATP channel subunits to track the dynamics of KATP channel surface expression. Results showed that activation of CaMKII triggered dynamin-dependent internalization of KATP channels. This process required phosphorylation of threonine at 180 and 224 and an intact 330YSKF333 endocytosis motif of the KATP channel Kir6.2 pore-forming subunit. A molecular model of the μ2 subunit of the endocytosis adaptor protein, AP2, complexed with Kir6.2 predicted that μ2 docks by interaction with 330YSKF333 and Thr-180 on one and Thr-224 on the adjacent Kir6.2 subunit. Phosphorylation of Thr-180 and Thr-224 would favor interactions with the corresponding arginine- and lysine-rich loops on μ2. We concluded that calcium-dependent activation of CaMKII results in phosphorylation of Kir6.2, which promotes endocytosis of cardiac KATP channel subunits. This mechanism couples the surface expression of cardiac KATP channels with calcium signaling and reveals new targets to improve cardiac energy efficiency and stress resistance. PMID:23223335

A new type of accelerator power supply based on voltage-type space vector PWM rectification technology

NASA Astrophysics Data System (ADS)

Wu, Fengjun; Gao, Daqing; Shi, Chunfeng; Huang, Yuzhen; Cui, Yuan; Yan, Hongbin; Zhang, Huajian; Wang, Bin; Li, Xiaohui

2016-08-01

To solve the problems such as low input power factor, a large number of AC current harmonics and instable DC bus voltage due to the diode or thyristor rectifier used in an accelerator power supply, particularly in the Heavy Ion Research Facility in Lanzhou-Cooler Storage Ring (HIRFL-CSR), we designed and built up a new type of accelerator power supply prototype base on voltage-type space vector PWM (SVPWM) rectification technology. All the control strategies are developed in TMS320C28346, which is a digital signal processor from TI. The experimental results indicate that an accelerator power supply with a SVPWM rectifier can solve the problems above well, and the output performance such as stability, tracking error and ripple current meet the requirements of the design. The achievement of prototype confirms that applying voltage-type SVPWM rectification technology in an accelerator power supply is feasible; and it provides a good reference for design and build of this new type of power supply.

Neurotensin effects on N-type calcium currents among rat pallidal neurons: an electrophysiological and immunohistochemical study.

PubMed

Martorana, Alessandro; Martella, Giuseppina; D'Angelo, Vincenza; Fusco, Francesca Romana; Spadoni, Francesca; Bernardi, Giorgio; Stefani, Alessandro

2006-10-01

The tridecapeptide neurotensin (NT) is involved in the modulation of dopamine (DA)-mediated functions in the nigrostriatal and mesocorticolimbic pathways. Its relevance in mammalian globus pallidus (GP) is questioned. A recent electrophysiological study on GP slices described NT-mediated robust membrane depolarization, depending upon the suppression of potassium conductance and/or the activation of cation current. Here, we have studied whether NT also affected high-voltage-activated calcium (Ca(2+)) currents, by means of whole-cell recordings on isolated GP neurons. In our hands, the full peptide and the segment NT8-13 reversibly inhibited N-like Ca(2+) current in about 60% of the recorded dissociated neurons, irrespective of their capacitance. The NT-mediated modulation showed no desensitization and was antagonized by the NT1 antagonists SR48692 and SR142948. These results imply an abundant expression of NTS(1) on GP cell somata. Then, we performed a light and immunofluorescence-confocal microscopy study of NTS(1) localization among GP neurons. We found that NTS(1) is localized in about 56% of GP neurons in both subpopulations of neurons, namely parvalbumin positive and negative. We conclude that NT, likely released from the striatal terminals in GP, acts through the postsynaptic NTS(1) preferentially localized in the lateral aspects of the GP. These data suggest a new implication (neither merely presynaptic nor simply "excitatory") for NT in the modulation of GP firing pattern. In addition, NT might have a role in affecting the interplay among the endogenous release of GABA/glutamate and DA. This hypothesis might have implications on both sensori-motor and associative functions of the GP and should be tested in DA-denervated disease models.

Calcium-induced calcium release in rod photoreceptor terminals boosts synaptic transmission during maintained depolarization

PubMed Central

Cadetti, Lucia; Bryson, Eric J.; Ciccone, Cory A.; Rabl, Katalin; Thoreson, Wallace B.

2008-01-01

We examined the contribution of calcium-induced calcium release (CICR) to synaptic transmission from rod photoreceptor terminals. Whole-cell recording and confocal calcium imaging experiments were conducted on rods with intact synaptic terminals in a retinal slice preparation from salamander. Low concentrations of ryanodine stimulated calcium increases in rod terminals, consistent with the presence of ryanodine receptors. Application of strong depolarizing steps (−70 to −10 mV) exceeding 200 ms or longer in duration evoked a wave of calcium that spread across the synaptic terminals of voltage-clamped rods. This secondary calcium increase was blocked by high concentrations of ryanodine, indicating it was due to CICR. Ryanodine (50 μM) had no significant effect on rod calcium current (Ica) although it slightly diminished rod light-evoked voltage responses. Bath application of 50 μM ryanodine strongly inhibited light-evoked currents in horizontal cells. Whether applied extracellularly or delivered into the rod cell through the patch pipette, ryanodine (50 μM) also inhibited excitatory post-synaptic currents (EPSCs) evoked in horizontal cells by depolarizing steps applied to rods. Ryanodine caused a preferential reduction in the later portions of EPSCs evoked by depolarizing steps of 200 ms or longer. These results indicate that CICR enhances calcium increases in rod terminals evoked by sustained depolarization, which in turn acts to boost synaptic exocytosis from rods. PMID:16819987

Calcium Homeostasis and Cone Signaling Are Regulated by Interactions between Calcium Stores and Plasma Membrane Ion Channels

PubMed Central

Bartoletti, Theodore M.; Huang, Wei; Akopian, Abram; Thoreson, Wallace B.; Krizaj, David

2009-01-01

Calcium is a messenger ion that controls all aspects of cone photoreceptor function, including synaptic release. The dynamic range of the cone output extends beyond the activation threshold for voltage-operated calcium entry, suggesting another calcium influx mechanism operates in cones hyperpolarized by light. We have used optical imaging and whole-cell voltage clamp to measure the contribution of store-operated Ca2+ entry (SOCE) to Ca2+ homeostasis and its role in regulation of neurotransmission at cone synapses. Mn2+ quenching of Fura-2 revealed sustained divalent cation entry in hyperpolarized cones. Ca2+ influx into cone inner segments was potentiated by hyperpolarization, facilitated by depletion of intracellular Ca2+ stores, unaffected by pharmacological manipulation of voltage-operated or cyclic nucleotide-gated Ca2+ channels and suppressed by lanthanides, 2-APB, MRS 1845 and SKF 96365. However, cation influx through store-operated channels crossed the threshold for activation of voltage-operated Ca2+ entry in a subset of cones, indicating that the operating range of inner segment signals is set by interactions between store- and voltage-operated Ca2+ channels. Exposure to MRS 1845 resulted in ∼40% reduction of light-evoked postsynaptic currents in photopic horizontal cells without affecting the light responses or voltage-operated Ca2+ currents in simultaneously recorded cones. The spatial pattern of store-operated calcium entry in cones matched immunolocalization of the store-operated sensor STIM1. These findings show that store-operated channels regulate spatial and temporal properties of Ca2+ homeostasis in vertebrate cones and demonstrate their role in generation of sustained excitatory signals across the first retinal synapse. PMID:19696927

The Calmodulin-Binding, Short Linear Motif, NSCaTE Is Conserved in L-Type Channel Ancestors of Vertebrate Cav1.2 and Cav1.3 Channels

PubMed Central

Taiakina, Valentina; Boone, Adrienne N.; Fux, Julia; Senatore, Adriano; Weber-Adrian, Danielle

2013-01-01

NSCaTE is a short linear motif of (xWxxx(I or L)xxxx), composed of residues with a high helix-forming propensity within a mostly disordered N-terminus that is conserved in L-type calcium channels from protostome invertebrates to humans. NSCaTE is an optional, lower affinity and calcium-sensitive binding site for calmodulin (CaM) which competes for CaM binding with a more ancient, C-terminal IQ domain on L-type channels. CaM bound to N- and C- terminal tails serve as dual detectors to changing intracellular Ca2+ concentrations, promoting calcium-dependent inactivation of L-type calcium channels. NSCaTE is absent in some arthropod species, and is also lacking in vertebrate L-type isoforms, Cav1.1 and Cav1.4 channels. The pervasiveness of a methionine just downstream from NSCaTE suggests that L-type channels could generate alternative N-termini lacking NSCaTE through the choice of translational start sites. Long N-terminus with an NSCaTE motif in L-type calcium channel homolog LCav1 from pond snail Lymnaea stagnalis has a faster calcium-dependent inactivation than a shortened N-termini lacking NSCaTE. NSCaTE effects are present in low concentrations of internal buffer (0.5 mM EGTA), but disappears in high buffer conditions (10 mM EGTA). Snail and mammalian NSCaTE have an alpha-helical propensity upon binding Ca2+-CaM and can saturate both CaM N-terminal and C-terminal domains in the absence of a competing IQ motif. NSCaTE evolved in ancestors of the first animals with internal organs for promoting a more rapid, calcium-sensitive inactivation of L-type channels. PMID:23626724

Induction of divalent cation permeability by heterologous expression of a voltage sensor domain.

PubMed

Arima, Hiroki; Tsutsui, Hidekazu; Sakamoto, Ayako; Yoshida, Manabu; Okamura, Yasushi

2018-01-06

The voltage sensor domain (VSD) is a protein domain that confers sensitivity to membrane potential in voltage-gated ion channels as well as the voltage-sensing phosphatase. Although VSDs have long been considered to function as regulatory units acting on adjacent effectors, recent studies have revealed the existence of direct ion permeation paths in some mutated VSDs and in the voltage-gated proton channel. In this study, we show that calcium currents are evoked upon membrane hyperpolarization in cells expressing a VSD derived from an ascidian voltage-gated ion channel superfamily. Unlike the previously reported omega-pore in the Shaker K + channel and rNav1.4, mutations are not required. From electrophysiological experiments in heterologous expression systems, we found that the conductance is directly mediated by the VSD itself and is carried by both monovalent and divalent cations. This is the first report of divalent cation permeation through a VSD-like structure. Copyright © 2018 Elsevier B.V. All rights reserved.

Current–voltage characteristics of high-voltage 4H-SiC p{sup +}–n{sub 0}–n{sup +} diodes in the avalanche breakdown mode

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

Ivanov, P. A., E-mail: Pavel.Ivanov@mail.ioffe.ru; Potapov, A. S.; Samsonova, T. P.

p{sup +}–n{sub 0}–n{sup +} 4H-SiC diodes with homogeneous avalanche breakdown at 1860 V are fabricated. The pulse current–voltage characteristics are measured in the avalanche-breakdown mode up to a current density of 4000 A/cm{sup 2}. It is shown that the avalanche-breakdown voltage increases with increasing temperature. The following diode parameters are determined: the avalanche resistance (8.6 × 10{sup –2} Ω cm{sup 2}), the electron drift velocity in the n{sub 0} base at electric fields higher than 10{sup 6} V/cm (7.8 × 10{sup 6} cm/s), and the relative temperature coefficient of the breakdown voltage (2.1 × 10{sup –4} K{sup –1}).

The tobacco-specific carcinogen-operated calcium channel promotes lung tumorigenesis via IGF2 exocytosis in lung epithelial cells

PubMed Central

Boo, Hye-Jin; Min, Hye-Young; Jang, Hyun-Ji; Yun, Hye Jeong; Smith, John Kendal; Jin, Quanri; Lee, Hyo-Jong; Liu, Diane; Kweon, Hee-Seok; Behrens, Carmen; Lee, J. Jack; Wistuba, Ignacio I.; Lee, Euni; Hong, Waun Ki; Lee, Ho-Young

2016-01-01

Nicotinic acetylcholine receptors (nAChRs) binding to the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces Ca2+ signalling, a mechanism that is implicated in various human cancers. In this study, we investigated the role of NNK-mediated Ca2+ signalling in lung cancer formation. We show significant overexpression of insulin-like growth factors (IGFs) in association with IGF-1R activation in human preneoplastic lung lesions in smokers. NNK induces voltage-dependent calcium channel (VDCC)-intervened calcium influx in airway epithelial cells, resulting in a rapid IGF2 secretion via the regulated pathway and thus IGF-1R activation. Silencing nAChR, α1 subunit of L-type VDCC, or various vesicular trafficking curators, including synaptotagmins and Rabs, or blockade of nAChR/VDCC-mediated Ca2+ influx significantly suppresses NNK-induced IGF2 exocytosis, transformation and tumorigenesis of lung epithelial cells. Publicly available database reveals inverse correlation between use of calcium channel blockers and lung cancer diagnosis. Our data indicate that NNK disrupts the regulated pathway of IGF2 exocytosis and promotes lung tumorigenesis. PMID:27666821

A calcium-channel homologue required for adaptation to dopamine and serotonin in Caenorhabditis elegans.

PubMed

Schafer, W R; Kenyon, C J

1995-05-04

Processing and storage of information by the nervous system requires the ability to modulate the response of excitable cells to neurotransmitter. A simple process of this type, known as adaptation or desensitization, occurs when prolonged stimulation triggers processes that attenuate the response to neurotransmitter. Here we report that the Caenorhabditis elegans gene unc-2 is required for adaptation to two neurotransmitters, dopamine and serotonin. A loss-of-function mutation in unc-2 resulted in failure to adapt either to paralysis by dopamine or to stimulation of egg laying by serotonin. In addition, unc-2 mutants displayed behaviours similar to those induced by serotonin treatment. We found that unc-2 encodes a homologue of a voltage-sensitive calcium-channel alpha-1 subunit. Expression of unc-2 occurs in two types of neurons implicated in the control of egg laying, a behaviour regulated by serotonin. Unc-2 appears to be required in modulatory neurons to downregulate the response of the egg-laying muscles to serotonin. We propose that adaptation to serotonin occurs through activation of an Unc-2-dependent calcium influx, which modulates the postsynaptic response to serotonin, perhaps by inhibiting the release of a potentiating neuropeptide.

Tuning the threshold voltage of carbon nanotube transistors by n-type molecular doping for robust and flexible complementary circuits

PubMed Central

Wang, Huiliang; Wei, Peng; Li, Yaoxuan; Han, Jeff; Lee, Hye Ryoung; Naab, Benjamin D.; Liu, Nan; Wang, Chenggong; Adijanto, Eric; Tee, Benjamin C.-K.; Morishita, Satoshi; Li, Qiaochu; Gao, Yongli; Cui, Yi; Bao, Zhenan

2014-01-01

Tuning the threshold voltage of a transistor is crucial for realizing robust digital circuits. For silicon transistors, the threshold voltage can be accurately controlled by doping. However, it remains challenging to tune the threshold voltage of single-wall nanotube (SWNT) thin-film transistors. Here, we report a facile method to controllably n-dope SWNTs using 1H-benzoimidazole derivatives processed via either solution coating or vacuum deposition. The threshold voltages of our polythiophene-sorted SWNT thin-film transistors can be tuned accurately and continuously over a wide range. Photoelectron spectroscopy measurements confirmed that the SWNT Fermi level shifted to the conduction band edge with increasing doping concentration. Using this doping approach, we proceeded to fabricate SWNT complementary inverters by inkjet printing of the dopants. We observed an unprecedented noise margin of 28 V at VDD = 80 V (70% of 1/2VDD) and a gain of 85. Additionally, robust SWNT complementary metal−oxide−semiconductor inverter (noise margin 72% of 1/2VDD) and logic gates with rail-to-rail output voltage swing and subnanowatt power consumption were fabricated onto a highly flexible substrate. PMID:24639537

Calcium dynamics regulating the timing of decision-making in C. elegans

PubMed Central

Tanimoto, Yuki; Yamazoe-Umemoto, Akiko; Fujita, Kosuke; Kawazoe, Yuya; Miyanishi, Yosuke; Yamazaki, Shuhei J; Fei, Xianfeng; Busch, Karl Emanuel; Gengyo-Ando, Keiko; Nakai, Junichi; Iino, Yuichi; Iwasaki, Yuishi; Hashimoto, Koichi; Kimura, Koutarou D

2017-01-01

Brains regulate behavioral responses with distinct timings. Here we investigate the cellular and molecular mechanisms underlying the timing of decision-making during olfactory navigation in Caenorhabditis elegans. We find that, based on subtle changes in odor concentrations, the animals appear to choose the appropriate migratory direction from multiple trials as a form of behavioral decision-making. Through optophysiological, mathematical and genetic analyses of neural activity under virtual odor gradients, we further find that odor concentration information is temporally integrated for a decision by a gradual increase in intracellular calcium concentration ([Ca2+]i), which occurs via L-type voltage-gated calcium channels in a pair of olfactory neurons. In contrast, for a reflex-like behavioral response, [Ca2+]i rapidly increases via multiple types of calcium channels in a pair of nociceptive neurons. Thus, the timing of neuronal responses is determined by cell type-dependent involvement of calcium channels, which may serve as a cellular basis for decision-making. DOI: http://dx.doi.org/10.7554/eLife.21629.001 PMID:28532547

ATP Releasing Connexin 30 Hemichannels Mediate Flow-Induced Calcium Signaling in the Collecting Duct

PubMed Central

Svenningsen, Per; Burford, James L.; Peti-Peterdi, János

2013-01-01

ATP in the renal tubular fluid is an important regulator of salt and water reabsorption via purinergic calcium signaling that involves the P2Y2 receptor, ENaC, and AQP2. Recently, we have shown that connexin (Cx) 30 hemichannels are localized to the non-junctional apical membrane of cells in the distal nephron-collecting duct (CD) and release ATP into the tubular fluid upon mechanical stimuli, leading to reduced salt and water reabsorption. Cx30−/− mice show salt-dependent elevations in BP and impaired pressure-natriuresis. Thus, we hypothesized that increased tubular flow rate leads to Cx30-dependent purinergic intracellular calcium ([Ca2+]i) signaling in the CD. Cortical CDs (CCDs) from wild type and Cx30−/− mice were freshly dissected and microperfused in vitro. Using confocal fluorescence imaging and the calcium-sensitive fluorophore pair Fluo-4 and Fura Red, we found that increasing tubular flow rate from 2 to 20 nl/min caused a significant 2.1-fold elevation in [Ca2+]i in wild type CCDs. This response was blunted in Cx30−/− CCDs ([Ca2+]i increased only 1.2-fold, p < 0.0001 vs. WT, n = 6 each). To further test our hypothesis we performed CD [Ca2+]i imaging in intact mouse kidneys in vivo using multiphoton microscopy and micropuncture delivery of the calcium-sensitive fluorophore Rhod-2. We found intrinsic, spontaneous [Ca2+]i oscillations in free-flowing CDs of wild type but not Cx30−/− mice. The [Ca2+]i oscillations were sensitive also to P2-receptor inhibition by suramin. Taken together, these data confirm that mechanosensitive Cx30 hemichannels mediate tubular ATP release and purinergic calcium signaling in the CD which mechanism plays an important role in the regulation of CD salt and water reabsorption. PMID:24137132

Calcium-activated potassium channels in insect pacemaker neurons as unexpected target site for the novel fumigant dimethyl disulfide.

PubMed

Gautier, Hélène; Auger, Jacques; Legros, Christian; Lapied, Bruno

2008-01-01

Dimethyl disulfide (DMDS), a plant-derived insecticide, is a promising fumigant as a substitute for methyl bromide. To further understand the mode of action of DMDS, we examined its effect on cockroach octopaminergic neurosecretory cells, called dorsal unpaired median (DUM) neurons, using whole-cell patch-clamp technique, calcium imaging and antisense oligonucleotide strategy. At low concentration (1 microM), DMDS modified spontaneous regular spike discharge into clear bursting activity associated with a decrease of the amplitude of the afterhyperpolarization. This effect led us to suspect alterations of calcium-activated potassium currents (IKCa) and [Ca(2+)](i) changes. We showed that DMDS reduced amplitudes of both peak transient and sustained components of the total potassium current. IKCa was confirmed as a target of DMDS by using iberiotoxin, cadmium chloride, and pSlo antisense oligonucleotide. In addition, we showed that DMDS induced [Ca(2+)](i) rise in Fura-2-loaded DUM neurons. Using calcium-free solution, and (R,S)-(3,4-dihydro-6,7-dimethoxy-isoquinoline-1-yl)-2-phenyl-N,N-di-[2-(2,3,4-trimethoxy-phenyl)ethyl]-acetamide (LOE 908) [an inhibitor of transient receptor potential (TRP)gamma], we demonstrated that TRPgamma initiated calcium influx. By contrast, omega-conotoxin GVIA (an inhibitor of N-type high-voltage-activated calcium channels), did not affect the DMDS-induced [Ca(2+)](i) rise. Finally, the participation of the calcium-induced calcium release mechanism was investigated using thapsigargin, caffeine, and ryanodine. Our study revealed that DMDS-induced elevation in [Ca(2+)](i) modulated IKCa in an unexpected bell-shaped manner via intracellular calcium. In conclusion, DMDS affects multiple targets, which could be an effective way to improve pest control efficacy of fumigation.

Photo-sensitization of ZnS nanoparticles with renowned ruthenium dyes N3, N719 and Z907 for application in solid state dye sensitized solar cells: A comparative study.

PubMed

Nosheen, Erum; Shah, Syed Mujtaba; Hussain, Hazrat; Murtaza, Ghulam

2016-09-01

This article presents a comprehensive relative report on the grafting of ZnS with renowned ruthenium ((Ru) dyes i.e. N3, N719 and Z907) and gives insight into their charge transfer interaction and sensitization mechanism for boosting solar cell efficiency. Influence of dye concentration on cell performance is also reported here. ZnS nanoparticles synthesized by a simple coprecipitation method with an average particle size of 15±2nm were characterized by X-ray diffraction (XRD), field emission scanning electron microscopy (FESEM), Elemental dispersive X-ray analysis (EDAX), tunneling electron microscopy (TEM) and UV-Visible (UV-Vis) spectroscopy. UV-Vis, photoluminescence (PL) and Fourier transform infra-red (FT-IR) spectroscopy confirms the successful grafting of these dyes over ZnS nanoparticles surface. Low-energy metal-to-ligand charge-transfer transition (MLCT) bands of dyes are mainly affected on grafting over the nanoparticle surface. Moreover their current voltage (I-V) results confirm the efficiency enhancement in ZnS solid state dye sensitized solar cells (SSDSSCs) owing to effective sensitization of this material with Ru dyes and helps in finding the optimum dye concentration for nanoparticles sensitization. Highest rise in overall solar cell efficiency i.e. 64% of the reference device has been observed for 0.3mM N719-ZnS sample owing to increased open circuit voltage (Voc) and fill factor (FF). Experimental and proposed results were found in good agreement with each other. Copyright © 2016 Elsevier B.V. All rights reserved.

The probability of quantal secretion near a single calcium channel of an active zone.

PubMed Central

Bennett, M R; Farnell, L; Gibson, W G

2000-01-01

A Monte Carlo analysis has been made of calcium dynamics and quantal secretion at microdomains in which the calcium reaches very high concentrations over distances of <50 nm from a channel and for which calcium dynamics are dominated by diffusion. The kinetics of calcium ions in microdomains due to either the spontaneous or evoked opening of a calcium channel, both of which are stochastic events, are described in the presence of endogenous fixed and mobile buffers. Fluctuations in the number of calcium ions within 50 nm of a channel are considerable, with the standard deviation about half the mean. Within 10 nm of a channel these numbers of ions can give rise to calcium concentrations of the order of 100 microM. The temporal changes in free calcium and calcium bound to different affinity indicators in the volume of an entire varicosity or bouton following the opening of a single channel are also determined. A Monte Carlo analysis is also presented of how the dynamics of calcium ions at active zones, after the arrival of an action potential and the stochastic opening of a calcium channel, determine the probability of exocytosis from docked vesicles near the channel. The synaptic vesicles in active zones are found docked in a complex with their calcium-sensor associated proteins and a voltage-sensitive calcium channel, forming a secretory unit. The probability of quantal secretion from an isolated secretory unit has been determined for different distances of an open calcium channel from the calcium sensor within an individual unit: a threefold decrease in the probability of secretion of a quantum occurs with a doubling of the distance from 25 to 50 nm. The Monte Carlo analysis also shows that the probability of secretion of a quantum is most sensitive to the size of the single-channel current compared with its sensitivity to either the binding rates of the sites on the calcium-sensor protein or to the number of these sites that must bind a calcium ion to trigger

Phosphorylation of purified mitochondrial Voltage-Dependent Anion Channel by c-Jun N-terminal Kinase-3 modifies channel voltage-dependence.

PubMed

Gupta, Rajeev; Ghosh, Subhendu

2017-06-01

Voltage-Dependent Anion Channel (VDAC) phosphorylated by c-Jun N-terminal Kinase-3 (JNK3) was incorporated into the bilayer lipid membrane. Single-channel electrophysiological properties of the native and the phosphorylated VDAC were compared. The open probability versus voltage curve of the native VDAC displayed symmetry around the voltage axis, whereas that of the phosphorylated VDAC showed asymmetry. This result indicates that phosphorylation by JNK3 modifies voltage-dependence of VDAC.

Modulating the Voltage-sensitivity of a Genetically Encoded Voltage Indicator

PubMed Central

Jung, Arong; Rajakumar, Dhanarajan; Yoon, Bong-June

2017-01-01

Saturation mutagenesis was performed on a single position in the voltage-sensing domain (VSD) of a genetically encoded voltage indicator (GEVI). The VSD consists of four transmembrane helixes designated S1-S4. The V220 position located near the plasma membrane/extracellular interface had previously been shown to affect the voltage range of the optical signal. Introduction of polar amino acids at this position reduced the voltage-dependent optical signal of the GEVI. Negatively charged amino acids slightly reduced the optical signal by 33 percent while positively charge amino acids at this position reduced the optical signal by 80%. Surprisingly, the range of V220D was similar to that of V220K with shifted optical responses towards negative potentials. In contrast, the V220E mutant mirrored the responses of the V220R mutation suggesting that the length of the side chain plays in role in determining the voltage range of the GEVI. Charged mutations at the 219 position all behaved similarly slightly shifting the optical response to more negative potentials. Charged mutations to the 221 position behaved erratically suggesting interactions with the plasma membrane and/or other amino acids in the VSD. Introduction of bulky amino acids at the V220 position increased the range of the optical response to include hyperpolarizing signals. Combining The V220W mutant with the R217Q mutation resulted in a probe that reduced the depolarizing signal and enhanced the hyperpolarizing signal which may lead to GEVIs that only report neuronal inhibition. PMID:29093633

Modulating the Voltage-sensitivity of a Genetically Encoded Voltage Indicator.

PubMed

Jung, Arong; Rajakumar, Dhanarajan; Yoon, Bong-June; Baker, Bradley J

2017-10-01

Saturation mutagenesis was performed on a single position in the voltage-sensing domain (VSD) of a genetically encoded voltage indicator (GEVI). The VSD consists of four transmembrane helixes designated S1-S4. The V220 position located near the plasma membrane/extracellular interface had previously been shown to affect the voltage range of the optical signal. Introduction of polar amino acids at this position reduced the voltage-dependent optical signal of the GEVI. Negatively charged amino acids slightly reduced the optical signal by 33 percent while positively charge amino acids at this position reduced the optical signal by 80%. Surprisingly, the range of V220D was similar to that of V220K with shifted optical responses towards negative potentials. In contrast, the V220E mutant mirrored the responses of the V220R mutation suggesting that the length of the side chain plays in role in determining the voltage range of the GEVI. Charged mutations at the 219 position all behaved similarly slightly shifting the optical response to more negative potentials. Charged mutations to the 221 position behaved erratically suggesting interactions with the plasma membrane and/or other amino acids in the VSD. Introduction of bulky amino acids at the V220 position increased the range of the optical response to include hyperpolarizing signals. Combining The V220W mutant with the R217Q mutation resulted in a probe that reduced the depolarizing signal and enhanced the hyperpolarizing signal which may lead to GEVIs that only report neuronal inhibition.

Sequential N-Terminal Pro-B-Type Natriuretic Peptide and High-Sensitivity Cardiac Troponin Measurements During Albumin Replacement in Patients With Severe Sepsis or Septic Shock.

PubMed

Masson, Serge; Caironi, Pietro; Fanizza, Caterina; Carrer, Sara; Caricato, Anselmo; Fassini, Paola; Vago, Tarcisio; Romero, Marilena; Tognoni, Gianni; Gattinoni, Luciano; Latini, Roberto

2016-04-01

Myocardial dysfunction is a frequent complication in patients with severe sepsis and can worsen the prognosis. We investigated whether circulating biomarkers related to myocardial function and injury predicted outcome and were associated with albumin replacement. A multicenter, randomized clinical trial about albumin replacement in severe sepsis or septic shock (the Albumin Italian Outcome Sepsis trial). Forty ICUs in Italy. Nine hundred and ninety-five patients with severe sepsis or septic shock. Randomization to albumin and crystalloid solutions or crystalloid solutions alone. Plasma concentrations of N- terminal pro-B-type natriuretic peptide and high-sensitivity cardiac troponin T were measured 1, 2, and 7 days after enrollment. We tested the relationship of single marker measurements or changes over time with clinical events, organ dysfunctions, albumin replacement, and ICU or 90-day mortality in the overall population and after stratification by shock. N-terminal pro-B-type natriuretic peptide levels were abnormal in 97.4% of the patients and high-sensitivity cardiac troponin T in 84.5%, with higher concentrations in those with shock. After extensive adjustments, N-terminal pro-B-type natriuretic peptide concentrations predicted ICU or 90-day mortality, better than high-sensitivity cardiac troponin T. Early changes in N-terminal pro-B-type natriuretic peptide or high-sensitivity cardiac troponin T concentrations were independently associated with subsequent mortality in patients with shock. Patients given albumin had significantly higher N-terminal pro-B-type natriuretic peptide levels; in addition, early rise in N-terminal pro-B-type natriuretic peptide was associated with a better outcome in this subgroup. Circulating N-terminal pro-B-type natriuretic peptide and high-sensitivity cardiac troponin T are frequently elevated in severe sepsis or septic shock and have relevant prognostic value, which may be important in monitoring the clinical efficacy of

Surface sensitization mechanism on negative electron affinity p-GaN nanowires

NASA Astrophysics Data System (ADS)

Diao, Yu; Liu, Lei; Xia, Sihao; Feng, Shu; Lu, Feifei

2018-03-01

The surface sensitization is the key to prepare negative electron affinity photocathode. The thesis emphasizes on the study of surface sensitization mechanism of p-type doping GaN nanowires utilizing first principles based on density function theory. The adsorption energy, work function, dipole moment, geometry structure, electronic structure and optical properties of Mg-doped GaN nanowires surfaces with various coverages of Cs atoms are investigated. The GaN nanowire with Mg doped in core position is taken as the sensitization base. At the initial stage of sensitization, the best adsorption site for Cs atom on GaN nanowire surface is BN, the bridge site of two adjacent N atoms. Surface sensitization generates a p-type internal surface with an n-type surface state, introducing a band bending region which can help reduce surface barrier and work function. With increasing Cs coverage, work functions decrease monotonously and the "Cs-kill" phenomenon disappears. For Cs coverage of 0.75 ML and 1 ML, the corresponding sensitization systems reach negative electron affinity state. Through surface sensitization, the absorption curves are red shifted and the absorption coefficient is cut down. All theoretical calculations can guide the design of negative electron affinity Mg doped GaN nanowires photocathode.

Electromagnetic fields act via activation of voltage-gated calcium channels to produce beneficial or adverse effects

PubMed Central

Pall, Martin L

2013-01-01

The direct targets of extremely low and microwave frequency range electromagnetic fields (EMFs) in producing non-thermal effects have not been clearly established. However, studies in the literature, reviewed here, provide substantial support for such direct targets. Twenty-three studies have shown that voltage-gated calcium channels (VGCCs) produce these and other EMF effects, such that the L-type or other VGCC blockers block or greatly lower diverse EMF effects. Furthermore, the voltage-gated properties of these channels may provide biophysically plausible mechanisms for EMF biological effects. Downstream responses of such EMF exposures may be mediated through Ca2+/calmodulin stimulation of nitric oxide synthesis. Potentially, physiological/therapeutic responses may be largely as a result of nitric oxide-cGMP-protein kinase G pathway stimulation. A well-studied example of such an apparent therapeutic response, EMF stimulation of bone growth, appears to work along this pathway. However, pathophysiological responses to EMFs may be as a result of nitric oxide-peroxynitrite-oxidative stress pathway of action. A single such well-documented example, EMF induction of DNA single-strand breaks in cells, as measured by alkaline comet assays, is reviewed here. Such single-strand breaks are known to be produced through the action of this pathway. Data on the mechanism of EMF induction of such breaks are limited; what data are available support this proposed mechanism. Other Ca2+-mediated regulatory changes, independent of nitric oxide, may also have roles. This article reviews, then, a substantially supported set of targets, VGCCs, whose stimulation produces non-thermal EMF responses by humans/higher animals with downstream effects involving Ca2+/calmodulin-dependent nitric oxide increases, which may explain therapeutic and pathophysiological effects. PMID:23802593

Design of high breakdown voltage vertical GaN p-n diodes with high-K/low-K compound dielectric structure for power electronics applications

NASA Astrophysics Data System (ADS)

Du, Jiangfeng; Li, Zhenchao; Liu, Dong; Bai, Zhiyuan; Liu, Yang; Yu, Qi

2017-11-01

In this work, a vertical GaN p-n diode with a high-K/low-K compound dielectric structure (GaN CD-VGD) is proposed and designed to achieve a record high breakdown voltage (BV) with a low specific on-resistance (Ron,sp). By introducing compound dielectric structure, the electric field near the p-n junction interface is suppressed due to the effects of high-K passivation layer, and a new electric field peak is induced into the n-type drift region, because of a discontinuity of electrical field at the interface of high-K and low-K layer. Therefore the distribution of electric field in GaN p-n diode becomes more uniform and an enhancement of breakdown voltage can be achieved. Numerical simulations demonstrate that GaN CD-VGD with a BV of 10650 V and a Ron,sp of 14.3 mΩ cm2, resulting in a record high figure-of-merit of 8 GW/cm2.

Spin injection in n-type resonant tunneling diodes.

PubMed

Orsi Gordo, Vanessa; Herval, Leonilson Ks; Galeti, Helder Va; Gobato, Yara Galvão; Brasil, Maria Jsp; Marques, Gilmar E; Henini, Mohamed; Airey, Robert J

2012-10-25

We have studied the polarized resolved photoluminescence of n-type GaAs/AlAs/GaAlAs resonant tunneling diodes under magnetic field parallel to the tunnel current. Under resonant tunneling conditions, we have observed two emission lines attributed to neutral (X) and negatively charged excitons (X-). We have observed a voltage-controlled circular polarization degree from the quantum well emission for both lines, with values up to -88% at 15 T at low voltages which are ascribed to an efficient spin injection from the 2D gases formed at the accumulation layers.

Activation of brain nitric oxide synthase in depolarized human temporal cortex slices: differential role of voltage-sensitive calcium channels

PubMed Central

Fontana, Giovanni; Fedele, Ernesto; Cossu, Massimo; Munari, Claudio; Raiteri, Maurizio

1997-01-01

Nitric oxide (NO) synthase activity was studied in slices of human temporal cortex samples obtained in neurosurgery by measuring the conversion of L-[3H]-arginine to L-[3H]-citrulline. Elevation of extracellular K+ to 20, 35 or 60 mM concentration-dependently augmented L-[3H]-citrulline production. The response to 35 mM KCl was abolished by NG-nitro-L-arginine (100 μM) demonstrating NO synthase specific conversion of L-arginine to L-citrulline. Increasing extracellular MgCl2 concentration up to 10 mM also prevented the K+ (35 mM)-induced NO synthase activation, suggesting the absolute requirement of external calcium ions for enzyme activity. However, the effect of high K+ (35 mM) on citrulline synthesis was insensitive to the antagonists of ionotropic and metabotropic glutamate receptors dizocilpine (MK-801), 6-nitro-7-sulphamoylbenzo(f)quinoxaline-2-3-dione (NBQX) or L-2-amino-3-phosphonopropionic acid (L-AP3) as well as to the nicotinic receptor antagonist, mecamylamine. The 35 mM K+ response was insensitive to ω-conotoxin GVIA (1 μM) and nifedipine (100 μM), but could be prevented in part by ω-agatoxin IVA (0.1 and 1 μM). The inhibition caused by 0.1 μM ω-agatoxin IVA (∼30%) was enhanced by adding ω-conotoxin GVIA (1 μM) or nifedipine (100 μM). Further inhibition (up to above 70%) could be observed when the three Ca2+ channel blockers were added together. Similarly, synthetic FTX 3.3 arginine polyamine (sFTX) prevented (50% at 100 μM) the K+-evoked NO synthase activation. This effect of sFTX was further enhanced (up to 70%) by adding 1 μM ω-conotoxin GVIA plus 100 μM nifedipine. No further inhibition could be observed upon addition of MK-801 or/and NBQX. It was concluded that elevation of extracellular [K+] causes NO synthase activation by external Ca+ entering cells mainly through channels of the P/Q-type. Other Ca2+ channels (L- and N-type) appear to contribute when P/Q-channels are blocked. PMID:9384511

Mechanisms Responsible for ω-Pore Currents in Cav Calcium Channel Voltage-Sensing Domains.

PubMed

Monteleone, Stefania; Lieb, Andreas; Pinggera, Alexandra; Negro, Giulia; Fuchs, Julian E; Hofer, Florian; Striessnig, Jörg; Tuluc, Petronel; Liedl, Klaus R

2017-10-03

Mutations of positively charged amino acids in the S4 transmembrane segment of a voltage-gated ion channel form ion-conducting pathways through the voltage-sensing domain, named ω-current. Here, we used structure modeling and MD simulations to predict pathogenic ω-currents in Ca V 1.1 and Ca V 1.3 Ca 2+ channels bearing several S4 charge mutations. Our modeling predicts that mutations of Ca V 1.1-R1 (R528H/G, R897S) or Ca V 1.1-R2 (R900S, R1239H) linked to hypokalemic periodic paralysis type 1 and of Ca V 1.3-R3 (R990H) identified in aldosterone-producing adenomas conducts ω-currents in resting state, but not during voltage-sensing domain activation. The mechanism responsible for the ω-current and its amplitude depend on the number of charges in S4, the position of the mutated S4 charge and countercharges, and the nature of the replacing amino acid. Functional characterization validates the modeling prediction showing that Ca V 1.3-R990H channels conduct ω-currents at hyperpolarizing potentials, but not upon membrane depolarization compared with wild-type channels. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Modulation of nano-selenium on tetrodotoxin-sensitive voltage-gated sodium currents in rat dorsal root ganglion neurons.

PubMed

Yuan, Huijun; Lan, Tonghan; Lin, Jiarui

2005-01-01

Nano-Selenium, a novel Nano technology production, was demonstrated to be useful in medical and scientific researches. Here, we investigated the effects of Nano-Selenium on tetrodotoxin-sensitive (TTX-S) voltage-dependent Na⁺channels in isolated rat dorsal root ganglion neurons, using whole-cell patch-clamp method. Nano-Selenium irreversibly decreased TTX-S Na⁺current (INa) in a concentration-dependent manner and shifted the maximum of the current/voltage relationship from -67mV to -52mV, without modifying the threshold potential of the current. Nano-Selenium shifted the steady-state activation and inactivation curves to the left. In the contrast of Na2SeO3, the inhibition effect of 1nM Nano-Se was much stronger. The cell treated with 1nM Na2SeO3firstly, still respond to futher addition of 1nM Nano-Selenium. These results prove Nano-Selenium to be a novel antiagonist, acted within the channel pore, not on or near the exterior surface of the channel protein where it would experience the membrane electric field, which possesses a distinct binding site from Na2SeO3.

Impact of calcium on N1 influenza neuraminidase dynamics and binding free energy.

PubMed

Lawrenz, Morgan; Wereszczynski, Jeff; Amaro, Rommie; Walker, Ross; Roitberg, Adrian; McCammon, J Andrew

2010-08-15

The highly pathogenic influenza strains H5N1 and H1N1 are currently treated with inhibitors of the viral surface protein neuraminidase (N1). Crystal structures of N1 indicate a conserved, high affinity calcium binding site located near the active site. The specific role of this calcium in the enzyme mechanism is unknown, though it has been shown to be important for enzymatic activity and thermostability. We report molecular dynamics (MD) simulations of calcium-bound and calcium-free N1 complexes with the inhibitor oseltamivir (marketed as the drug Tamiflu), independently using both the AMBER FF99SB and GROMOS96 force fields, to give structural insight into calcium stabilization of key framework residues. Y347, which demonstrates similar sampling patterns in the simulations of both force fields, is implicated as an important N1 residue that can "clamp" the ligand into a favorable binding pose. Free energy perturbation and thermodynamic integration calculations, using two different force fields, support the importance of Y347 and indicate a +3 to +5 kcal/mol change in the binding free energy of oseltamivir in the absence of calcium. With the important role of structure-based drug design for neuraminidase inhibitors and the growing literature on emerging strains and subtypes, inclusion of this calcium for active site stability is particularly crucial for computational efforts such as homology modeling, virtual screening, and free energy methods. 2010 Wiley-Liss, Inc.

Effect of applied voltage on surface properties of anodised titanium in mixture of β-glycerophosphate (β-GP) and calcium acetate (CA)

NASA Astrophysics Data System (ADS)

Chuan, Lee Te; Rathi, Muhammad Fareez Mohamad; Abidin, Muhamad Yusuf Zainal; Abdullah, Hasan Zuhudi; Idris, Maizlinda Izwana

2015-07-01

Anodic oxidation is a surface modification method which combines electric field driven metal and oxygen ion diffusion for formation of oxide layer on the anode surface. This method has been widely used to modify the surface morphology of biomaterial especially titanium. This study aimed to investigate the effect of applied voltage on titanium. Specifically, the titanium foil was anodised in mixture of β-glycerophosphate disodium salt pentahydrate (β-GP) and calcium acetate monohydrate (CA) with different applied voltage (50-350 V), electrolyte concentration (0.04 M β-GP + 0.4 M CA), anodising time (10minutes) and current density (50 and 70 mA.cm-2) at room temperature. Surface oxide properties of anodised titanium were characterised by digital single-lens reflex camera (DSLR camera), field emission scanning electron microscope (FESEM) and atomic force microscopy (AFM). At lower applied voltage (≤150 V), surface of titanium foils were relatively smooth. With increasing applied voltage (≥250 V), the oxide layer became more porous and donut-shaped pores were formed on the surface of titanium foils. The AFM results indicated that the surface roughness of anodised titanium increases with increasing of applied voltage. The porous and rough surface is able to promote the osseointegration and reduce the suffering time of patient.

Beyond voltage-gated ion channels: Voltage-operated membrane proteins and cellular processes.

PubMed

Zhang, Jianping; Chen, Xingjuan; Xue, Yucong; Gamper, Nikita; Zhang, Xuan

2018-04-18

Voltage-gated ion channels were believed to be the only voltage-sensitive proteins in excitable (and some non-excitable) cells for a long time. Emerging evidence indicates that the voltage-operated model is shared by some other transmembrane proteins expressed in both excitable and non-excitable cells. In this review, we summarize current knowledge about voltage-operated proteins, which are not classic voltage-gated ion channels as well as the voltage-dependent processes in cells for which single voltage-sensitive proteins have yet to be identified. Particularly, we will focus on the following. (1) Voltage-sensitive phosphoinositide phosphatases (VSP) with four transmembrane segments homologous to the voltage sensor domain (VSD) of voltage-gated ion channels; VSPs are the first family of proteins, other than the voltage-gated ion channels, for which there is sufficient evidence for the existence of the VSD domain; (2) Voltage-gated proton channels comprising of a single voltage-sensing domain and lacking an identified pore domain; (3) G protein coupled receptors (GPCRs) that mediate the depolarization-evoked potentiation of Ca 2+ mobilization; (4) Plasma membrane (PM) depolarization-induced but Ca 2+ -independent exocytosis in neurons. (5) Voltage-dependent metabolism of phosphatidylinositol 4,5-bisphosphate (PtdIns[4,5]P 2 , PIP 2 ) in the PM. These recent discoveries expand our understanding of voltage-operated processes within cellular membranes. © 2018 Wiley Periodicals, Inc.

TiN coated aluminum electrodes for DC high voltage electron guns

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

Mamun, Md Abdullah A.; Elmustafa, Abdelmageed A.; Taus, Rhys

Preparing electrodes made of metals like stainless steel, for use inside DC high voltage electron guns, is a labor-intensive and time-consuming process. In this paper, the authors report the exceptional high voltage performance of aluminum electrodes coated with hard titanium nitride (TiN). The aluminum electrodes were comparatively easy to manufacture and required only hours of mechanical polishing using silicon carbide paper, prior to coating with TiN by a commercial vendor. The high voltage performance of three TiN-coated aluminum electrodes, before and after gas conditioning with helium, was compared to that of bare aluminum electrodes, and electrodes manufactured from titanium alloymore » (Ti-6AI-4V). Following gas conditioning, each TiN-coated aluminum electrode reached -225 kV bias voltage while generating less than 100 pA of field emission (<10 pA) using a 40 mm cathode/anode gap, corresponding to field strength of 13.7 MV/m. Smaller gaps were studied to evaluate electrode performance at higher field strength with the best performing TiN-coated aluminum electrode reaching ~22.5 MV/m with field emission less than 100 pA. These results were comparable to those obtained from our best-performing electrodes manufactured from stainless steel, titanium alloy and niobium, as reported in references cited below. The TiN coating provided a very smooth surface and with mechanical properties of the coating (hardness and modulus) superior to those of stainless steel, titanium-alloy, and niobium electrodes. These features likely contributed to the improved high voltage performance of the TiN-coated aluminum electrodes.« less