scholarly journals Voltage control of Ca2+ permeation through N-type calcium (CaV2.2) channels

2014 ◽  
Vol 144 (3) ◽  
pp. 207-220 ◽  
Author(s):  
Zafir Buraei ◽  
Haoya Liang ◽  
Keith S. Elmslie
Keyword(s):  
Binding Sites ◽  
Neurotransmitter Release ◽  
Action Potentials ◽  
Channel Blocker ◽  
Cellular Functions ◽  
Voltage Gated ◽  
High Affinity ◽  
Step Model ◽  
Independent Test ◽  
Voltage Dependent

Voltage-gated calcium (CaV) channels deliver Ca2+ to trigger cellular functions ranging from cardiac muscle contraction to neurotransmitter release. The mechanism by which these channels select for Ca2+ over other cations is thought to involve multiple Ca2+-binding sites within the pore. Although the Ca2+ affinity and cation preference of these sites have been extensively investigated, the effect of voltage on these sites has not received the same attention. We used a neuronal preparation enriched for N-type calcium (CaV2.2) channels to investigate the effect of voltage on Ca2+ flux. We found that the EC50 for Ca2+ permeation increases from 13 mM at 0 mV to 240 mM at 60 mV, indicating that, during permeation, Ca2+ ions sense the electric field. These data were nicely reproduced using a three-binding-site step model. Using roscovitine to slow CaV2.2 channel deactivation, we extended these measurements to voltages <0 mV. Permeation was minimally affected at these hyperpolarized voltages, as was predicted by the model. As an independent test of voltage effects on permeation, we examined the Ca2+-Ba2+ anomalous mole fraction (MF) effect, which was both concentration and voltage dependent. However, the Ca2+-Ba2+ anomalous MF data could not be reproduced unless we added a fourth site to our model. Thus, Ca2+ permeation through CaV2.2 channels may require at least four Ca2+-binding sites. Finally, our results suggest that the high affinity of Ca2+ for the channel helps to enhance Ca2+ influx at depolarized voltages relative to other ions (e.g., Ba2+ or Na+), whereas the absence of voltage effects at negative potentials prevents Ca2+ from becoming a channel blocker. Both effects are needed to maximize Ca2+ influx over the voltages spanned by action potentials.

scholarly journals The structural biology of voltage-gated calcium channel function and regulation

2006 ◽  
Vol 34 (5) ◽  
pp. 887-893 ◽  
Author(s):  
F. Van Petegem ◽  
D.L. Minor
Keyword(s):  
Neurotransmitter Release ◽  
Action Potentials ◽  
Molecular Switches ◽  
Voltage Gated ◽  
Calcium Dependent ◽  
Voltage Gated Calcium Channels ◽  
Dependent Inactivation ◽  
Voltage Dependent ◽  
Soluble Calcium ◽  
Molecular Framework

Voltage-gated calcium channels (CaVs) are large (∼0.5 MDa), multisubunit, macromolecular machines that control calcium entry into cells in response to membrane potential changes. These molecular switches play pivotal roles in cardiac action potentials, neurotransmitter release, muscle contraction, calcium-dependent gene transcription and synaptic transmission. CaVs possess self-regulatory mechanisms that permit them to change their behaviour in response to activity, including voltage-dependent inactivation, calcium-dependent inactivation and calcium-dependent facilitation. These processes arise from the concerted action of different channel domains with CaV β-subunits and the soluble calcium sensor calmodulin. Until recently, nothing was known about the CaV structure at high resolution. Recent crystallographic work has revealed the first glimpses at the CaV molecular framework and set a new direction towards a detailed mechanistic understanding of CaV function.

scholarly journals Voltage-dependent inward currents in smooth muscle cells of skeletal muscle arterioles

2017 ◽  
Author(s):  
Alexandra V. Ulyanova ◽  
Roman E. Shirokov
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Action Potentials ◽  
Channel Blocker ◽  
Muscle Cells ◽  
Maximal Velocity ◽  
Voltage Gated ◽  
Voltage Dependent ◽  
Inward Currents

AbstractVoltage-dependent inward currents responsible for the depolarizing phase of action potentials were characterized in smooth muscle cells of 4th order arterioles in mouse skeletal muscle. Currents through L-type Ca2+ channels were expected to be dominant; however, action potentials were not eliminated in nominally Ca2+-free bathing solution or by addition of L-type Ca2+ channel blocker nifedipine (10 μM). Instead, Na+ channel blocker tetrodotoxin (TTX, 1 μM) reduced the maximal velocity of the upstroke at low, but not at normal (2 mM), Ca2+ in the bath. The magnitude of TTX-sensitive currents recorded with 140 mM Na+ was about 20 pA/pF. TTX-sensitive currents decreased five-fold when Ca2+ increased from 2 to 10 mM. The currents reduced three-fold in the presence of 10 mM caffeine, but remained unaltered by 1 mM of isobutylmethylxanthine (IBMX). In addition to L-type Ca2+ currents (15 pA/pF in 20 mM Ca2+), we also found Ca2+ currents that are resistant to 10 μM nifedipine (5 pA/pF in 20 mM Ca2+). Based on their biophysical properties, these Ca2+ currents are likely to be through voltage-gated T-type Ca2+ channels. Our results suggest that Na+ and at least two types (T- and L-) of Ca2+ voltage-gated channels contribute to depolarization of smooth muscle cells in skeletal muscle arterioles. Voltage-gated Na+ channels appear to be under a tight control by Ca2+ signaling.

Two Types of Voltage-Gated K+ Currents in Dissociated Heart Ventricular Muscle Cells of the Snail Lymnaea stagnalis

1999 ◽  
Vol 82 (5) ◽  
pp. 2415-2427 ◽  
Author(s):  
M. S. Yeoman ◽  
P. R. Benjamin
Keyword(s):  
Voltage Clamp ◽  
Action Potentials ◽  
Muscle Cells ◽  
Delayed Rectifier ◽  
Voltage Sensitivity ◽  
Ventricular Muscle ◽  
Current Time ◽  
Voltage Gated ◽  
Time To Peak ◽  
Voltage Dependent

We have used a combination of current-clamp and voltage-clamp techniques to characterize the electrophysiological properties of enzymatically dissociated Lymnaea heart ventricle cells. Dissociated ventricular muscle cells had average resting membrane potentials of −55 ± 5 mV. When hyperpolarized to potentials between −70 and −63 mV, ventricle cells were capable of firing repetitive action potentials (8.5 ± 1.2 spikes/min) that failed to overshoot 0 mV. The action potentials were either simple spikes or more complex spike/plateau events. The latter were always accompanied by strong contractions of the muscle cell. The waveform of the action potentials were shown to be dependent on the presence of extracellular Ca2+ and K+ ions. With the use of the single-electrode voltage-clamp technique, two types of voltage-gated K+ currents were identified that could be separated by differences in their voltage sensitivity and time-dependent kinetics. The first current activated between −50 and −40 mV. It was relatively fast to activate (time-to-peak; 13.7 ± 0.7 ms at +40 mV) and inactivated by 53.3 ± 4.9% during a maintained 200-ms depolarization. It was fully available for activation below −80 mV and was completely inactivated by holding potentials more positive than −40 mV. It was completely blocked by 5 mM 4-aminopyridine (4-AP) and by concentrations of tetraethylammonium chloride (TEA) >10 mM. These properties characterize this current as a member of the A-type family of voltage-dependent K+ currents. The second voltage-gated K+ current activated at more depolarized potentials (−30 to −20 mV). It activated slower than the A-type current (time-to-peak; 74.1 ± 3.9 ms at +40 mV) and showed little inactivation (6.2 ± 2.1%) during a maintained 200-ms depolarization. The current was fully available for activation below −80 mV with a proportion of the current still available for activation at potentials as positive as 0 mV. The current was completely blocked by 1–3 mM TEA. These properties characterize this current as a member of the delayed rectifier family of voltage-dependent K+ currents. The slow activation rates and relatively depolarized activation thresholds of the two K+ currents are suggestive that their main role is to contribute to the repolarization phase of the action potential.

scholarly journals Structure of the saxiphilin:saxitoxin (STX) complex reveals a convergent molecular recognition strategy for paralytic toxins

2019 ◽  
Vol 5 (6) ◽  
pp. eaax2650 ◽  
Author(s):  
Tien-Jui Yen ◽  
Marco Lolicato ◽  
Rhiannon Thomas-Tran ◽  
J. Du Bois ◽  
Daniel L. Minor
Keyword(s):  
Molecular Recognition ◽  
Binding Site ◽  
Binding Sites ◽  
Paralytic Shellfish Poisoning ◽  
Shellfish Poisoning ◽  
Voltage Gated ◽  
Toxin Binding ◽  
High Affinity ◽  
Paralytic Shellfish ◽  
Protein Sensors

Dinoflagelates and cyanobacteria produce saxitoxin (STX), a lethal bis-guanidinium neurotoxin causing paralytic shellfish poisoning. A number of metazoans have soluble STX-binding proteins that may prevent STX intoxication. However, their STX molecular recognition mechanisms remain unknown. Here, we present structures of saxiphilin (Sxph), a bullfrog high-affinity STX-binding protein, alone and bound to STX. The structures reveal a novel high-affinity STX-binding site built from a “proto-pocket” on a transferrin scaffold that also bears thyroglobulin domain protease inhibitor repeats. Comparison of Sxph and voltage-gated sodium channel STX-binding sites reveals a convergent toxin recognition strategy comprising a largely rigid binding site where acidic side chains and a cation-π interaction engage STX. These studies reveal molecular rules for STX recognition, outline how a toxin-binding site can be built on a naïve scaffold, and open a path to developing protein sensors for environmental STX monitoring and new biologics for STX intoxication mitigation.

scholarly journals Functions of Presynaptic Voltage-gated Calcium Channels

Function ◽  
2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Annette C Dolphin
Keyword(s):  
Calcium Channels ◽  
Neurotransmitter Release ◽  
Action Potentials ◽  
Voltage Dependence ◽  
Kinetic Properties ◽  
Excitable Cells ◽  
Single Action ◽  
Voltage Gated ◽  
Voltage Gated Calcium Channels ◽  
Vesicular Release

Abstract Voltage-gated calcium channels are the principal conduits for depolarization-mediated Ca2+ entry into excitable cells. In this review, the biophysical properties of the relevant members of this family of channels, those that are present in presynaptic terminals, will be discussed in relation to their function in mediating neurotransmitter release. Voltage-gated calcium channels have properties that ensure they are specialized for particular roles, for example, differences in their activation voltage threshold, their various kinetic properties, and their voltage-dependence of inactivation. All these attributes play into the ability of the various voltage-gated calcium channels to participate in different patterns of presynaptic vesicular release. These include synaptic transmission resulting from single action potentials, and longer-term changes mediated by bursts or trains of action potentials, as well as release resulting from graded changes in membrane potential in specialized sensory synapses.

scholarly journals Intracellular Mg2+ Enhances the Function of Bk-Type Ca2+-Activated K+ Channels

2001 ◽  
Vol 118 (5) ◽  
pp. 589-606 ◽  
Author(s):  
Jingyi Shi ◽  
Jianmin Cui
Keyword(s):  
Binding Sites ◽  
Neurotransmitter Release ◽  
Single Channel ◽  
Bk Channels ◽  
Bk Channel ◽  
Dependent Manner ◽  
Physiological Conditions ◽  
Channel Function ◽  
Open Conformation ◽  
Voltage Dependent

BK channels modulate neurotransmitter release due to their activation by voltage and Ca2+. Intracellular Mg2+ also modulates BK channels in multiple ways with opposite effects on channel function. Previous single-channel studies have shown that Mg2+ blocks the pore of BK channels in a voltage-dependent manner. We have confirmed this result by studying macroscopic currents of the mslo1 channel. We find that Mg2+ activates mslo1 BK channels independently of Ca2+ and voltage by preferentially binding to their open conformation. The mslo3 channel, which lacks Ca2+ binding sites in the tail, is not activated by Mg2+. However, coexpression of the mslo1 core and mslo3 tail produces channels with Mg2+ sensitivity similar to mslo1 channels, indicating that Mg2+ sites differ from Ca2+ sites. We discovered that Mg2+ also binds to Ca2+ sites and competitively inhibits Ca2+-dependent activation. Quantitative computation of these effects reveals that the overall effect of Mg2+ under physiological conditions is to enhance BK channel function.

Modulation of Transmitter Release by Action Potential Duration at the Hippocampal CA3-CA1 Synapse

1999 ◽  
Vol 81 (1) ◽  
pp. 288-298 ◽  
Author(s):  
Jing Qian ◽  
Peter Saggau
Keyword(s):  
Action Potential ◽  
Synaptic Transmission ◽  
Action Potential Duration ◽  
Transmitter Release ◽  
Neurotransmitter Release ◽  
Channel Blocker ◽  
Transmission Curve ◽  
Similar Reduction ◽  
Voltage Dependent ◽  
Hippocampal Ca3

Qian, Jing and Peter Saggau. Modulation of transmitter release by action potential duration at the hippocampal CA3-CA1 synapse. J. Neurophysiol. 81: 288–298, 1999. Presynaptic Ca2+ influx through voltage-dependent Ca2+ channels triggers neurotransmitter release. Action potential duration plays a determinant role in the dynamics of presynaptic Ca2+ influx. In this study, the presynaptic Ca2+ influx was optically measured with a low-affinity Ca2+ indicator (Furaptra). The effect of action potential duration on Ca2+ influx and transmitter release was investigated. The K+ channel blocker 4-aminopyridine (4-AP) was applied to broaden the action potential and thereby increase presynaptic Ca2+ influx. This increase of Ca2+ influx appeared to be much less effective in enhancing transmitter release than raising the extracellular Ca2+ concentration. 4-AP did not change the Ca2+ dependence of transmitter release but instead shifted the synaptic transmission curve toward larger total Ca2+ influx. These results suggest that changing the duration of Ca2+ influx is not equivalent to changing its amplitude in locally building up an effective Ca2+ concentration near the Ca2+ sensor of the release machinery. Furthermore, in the presence of 4-AP, the N-type Ca2+ channel blocker ωCgTx GVIA was much less effective in blocking transmitter release. This phenomenon was not simply due to a saturation of the release machinery by the increased overall Ca2+ influx because a similar reduction of Ca2+ influx by application of the nonspecific Ca2+ channel blocker Cd2+ resulted in much more inhibition of transmitter release. Rather, the different potencies of ω-CgTx GVIA and Cd2+ in inhibiting transmitter release suggest that the Ca2+ sensor is possibly located at a distance from a cluster of Ca2+ channels such that it is sensitive to the location of Ca2+ channels within the cluster.

High-affinity binding sites for [3H]saxitoxin are associated with voltage-dependent sodium channels in portal vein smooth muscle

1990 ◽  
Vol 184 (2-3) ◽  
pp. 315-319 ◽  
Author(s):  
Jean Mironneau ◽  
Cécile Martin ◽  
Serge Arnaudeau ◽  
Khadija Jmari ◽  
Lala Rakotoarisoa ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Portal Vein ◽  
Sodium Channels ◽  
Binding Sites ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
Voltage Dependent
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