scholarly journals A native prokaryotic voltage-dependent calcium channel with a novel selectivity filter sequence

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Takushi Shimomura ◽  
Yoshiki Yonekawa ◽  
Hitoshi Nagura ◽  
Michihiro Tateyama ◽  
Yoshinori Fujiyoshi ◽  
...  
Keyword(s):  
Action Potentials ◽  
Negative Charge ◽  
Selectivity Filter ◽  
Na Channels ◽  
Glycine Residue ◽  
Voltage Dependent Calcium Channel ◽  
Voltage Dependent ◽  
Living Organisms ◽  
Insight Into ◽  
Ca2 Channels

Voltage-dependent Ca2+ channels (Cavs) are indispensable for coupling action potentials with Ca2+ signaling in living organisms. The structure of Cavs is similar to that of voltage-dependent Na+ channels (Navs). It is known that prokaryotic Navs can obtain Ca2+ selectivity by negative charge mutations of the selectivity filter, but native prokaryotic Cavs had not yet been identified. We report the first identification of a native prokaryotic Cav, CavMr, whose selectivity filter contains a smaller number of negatively charged residues than that of artificial prokaryotic Cavs. A relative mutant whose selectivity filter was replaced with that of CavMr exhibits high Ca2+ selectivity. Mutational analyses revealed that the glycine residue of the CavMr selectivity filter is a determinant for Ca2+ selectivity. This glycine residue is well conserved among subdomains I and III of eukaryotic Cavs. These findings provide new insight into the Ca2+ selectivity mechanism that is conserved from prokaryotes to eukaryotes.

scholarly journals A native prokaryotic voltage-dependent calcium channel with a novel selectivity filter sequence

2019 ◽  
Author(s):  
Takushi Shimomura ◽  
Yoshiki Yonekawa ◽  
Hitoshi Nagura ◽  
Michihiro Tateyama ◽  
Yoshinori Fujiyoshi ◽  
...  
Keyword(s):  
Calcium Channel ◽  
Action Potentials ◽  
Negative Charge ◽  
Selectivity Filter ◽  
Glycine Residue ◽  
Voltage Dependent Calcium Channel ◽  
Charged Residues ◽  
Voltage Dependent ◽  
Living Organisms ◽  
Insight Into

AbstractVoltage-dependent Ca2+ channels (Cavs) are indispensable for coupling action potentials with Ca2+ signaling in living organisms. The structure of Cavs is similar to that of voltage-dependent Na+ channels (Navs). It is known that prokaryotic Navs can obtain Ca2+ selectivity by negative charge mutations of the selectivity filter, but native prokaryotic Cavs had not yet been identified.Here, we report the first identification of a native prokaryotic Cav, CavMr, and its relative, NavPp. Although CavMr contains a smaller number of negatively charged residues in the selectivity filter than artificial prokaryotic Cavs, CavMr exhibits high Ca2+ selectivity. In contrast, NavPp, which has similar filter sequence to artificial Cavs, mainly allows Na+ to permeate. Interestingly, a NavPp mutant whose selectivity filter was replaced with that of CavMr exhibits high Ca2+ selectivity. Mutational analyses revealed that the glycine residue of the CavMr selectivity filter is a determinant for Ca2+ selectivity. This glycine residue is well conserved among subdomains I and III of eukaryotic Cavs.These findings provide new insight into the Ca2+ selectivity mechanism conserved from prokaryotes to eukaryotes.

scholarly journals Optical measurement of conduction in single demyelinated axons.

1990 ◽  
Vol 95 (5) ◽  
pp. 867-889 ◽  
Author(s):  
P Shrager ◽  
C T Rubinstein
Keyword(s):  
Schwann Cells ◽  
Action Potentials ◽  
Spontaneous Recovery ◽  
Demyelinating Disease ◽  
Optical Measurement ◽  
Recovery Of Function ◽  
Na Channels ◽  
Cable Properties ◽  
Probability Of Success ◽  
Voltage Dependent

Demyelination was initiated in Xenopus sciatic nerves by an intraneural injection of lysolecithin over a 2-3-mm region. During the next week macrophages and Schwann cells removed all remaining damaged myelin by phagocytosis. Proliferating Schwann cells then began to remyelinate the axons, with the first few lamellae appearing 13 d after surgery. Action potentials were recorded optically through the use of a potential-sensitive dye. Signals could be detected both at normal nodes of Ranvier and within demyelinated segments. Before remyelination, conduction through the lesion occurred in only a small fraction of the fibers. However, in these particular cases we could demonstrate continuous (nonsaltatory) conduction at very low velocities over long (greater than one internode) lengths of demyelinated axons. We have previously found through loose patch clamp experiments that the internodal axolemma contains voltage-dependent Na+ channels at a density approximately 4% of that at the nodes. These channels alone, however, are insufficient for successful conduction past the transition point between myelinated and demyelinated regions. Small improvements in the passive cable properties of the axon, adequate for propagation at this site, can be realized through the close apposition of macrophages and Schwann cells. As the initial lamellae of myelin appear, the probability of success at the transition zone increases rapidly, though the conduction velocity through the demyelinated segment is not appreciably changed. A detailed computational model is used to test the relative roles of the internodal Na+ channels and the new extracellular layer. The results suggest a possible mechanism that may contribute to the spontaneous recovery of function often seen in demyelinating disease.

An active membrane model of the cerebellar Purkinje cell. I. Simulation of current clamps in slice

1994 ◽  
Vol 71 (1) ◽  
pp. 375-400 ◽  
Author(s):  
E. De Schutter ◽  
J. M. Bower
Keyword(s):  
Purkinje Cell ◽  
Ionic Channels ◽  
K Channel ◽  
Na Channels ◽  
Plateau Potentials ◽  
Spike Generation ◽  
K Channels ◽  
Voltage Dependent ◽  
P Type ◽  
Ca2 Channels

1. A detailed compartmental model of a cerebellar Purkinje cell with active dendritic membrane was constructed. The model was based on anatomic reconstructions of single Purkinje cells and included 10 different types of voltage-dependent channels described by Hodgkin-Huxley equations, derived from Purkinje cell-specific voltage-clamp data where available. These channels included a fast and persistent Na+ channel, three voltage-dependent K+ channels, T-type and P-type Ca2+ channels, and two types of Ca(2+)-activated K+ channels. 2. The ionic channels were distributed differentially over three zones of the model, with Na+ channels in the soma, fast K+ channels in the soma and main dendrite, and Ca2+ channels and Ca(2+)-activated K+ channels in the entire dendrite. Channel densities in the model were varied until it could reproduce Purkinje cell responses to current injections in the soma or dendrite, as observed in slice recordings. 3. As in real Purkinje cells, the model generated two types of spiking behavior. In response to small current injections the model fired exclusively fast somatic spikes. These somatic spikes were caused by Na+ channels and repolarized by the delayed rectifier. When higher-amplitude current injections were given, sodium spiking increased in frequency until the model generated large dendritic Ca2+ spikes. Analysis of membrane currents underlying this behavior showed that these Ca2+ spikes were caused by the P-type Ca2+ channel and repolarized by the BK-type Ca(2+)-activated K+ channel. As in pharmacological blocking experiments, removal of Na+ channels abolished the fast spikes and removal of Ca2+ channels removed Ca2+ spiking. 4. In addition to spiking behavior, the model also produced slow plateau potentials in both the dendrite and soma. These longer-duration potentials occurred in response to both short and prolonged current steps. Analysis of the model demonstrated that the plateau potentials in the soma were caused by the window current component of the fast Na+ current, which was much larger than the current through the persistent Na+ channels. Plateau potentials in the dendrite were carried by the same P-type Ca2+ channel that was also responsible for Ca2+ spike generation. The P channel could participate in both model functions because of the low-threshold K2-type Ca(2+)-activated K+ channel, which dynamically changed the threshold for dendritic spike generation through a negative feedback loop with the activation kinetics of the P-type Ca2+ channel. 5. These model responses were robust to changes in the densities of all of the ionic channels.(ABSTRACT TRUNCATED AT 400 WORDS)

Local Anesthetics Modulate Neuronal Calcium Signaling through Multiple Sites of Action

2003 ◽  
Vol 98 (5) ◽  
pp. 1139-1146 ◽  
Author(s):  
Fang Xu ◽  
Zayra Garavito-Aguilar ◽  
Esperanza Recio-Pinto ◽  
Jin Zhang ◽  
Thomas J. J. Blanck
Keyword(s):  
Local Anesthetics ◽  
K Channel ◽  
Na Channels ◽  
Channel Blockade ◽  
K Channels ◽  
Voltage Dependent ◽  
Sites Of Action ◽  
A Site ◽  
Ca2 Channels

Background Local anesthetics (LAs) are known to inhibit voltage-dependent Na+ channels, as well as K+ and Ca2+ channels, but with lower potency. Since cellular excitability and responsiveness are largely determined by intracellular Ca2+ availability, sites along the Ca2+ signaling pathways may be targets of LAs. This study was aimed to investigate the LA effects on depolarization and receptor-mediated intracellular Ca2+ changes and to examine the role of Na+ and K+ channels in such functional responses. Methods Effects of bupivacaine, ropivacaine, mepivacaine, and lidocaine (0.1-2.3 mm) on evoked [Ca2+](i) transients were investigated in neuronal SH-SY5Y cell suspensions using Fura-2 as the intracellular Ca2+ indicator. Potassium chloride (KCl, 100 mm) and carbachol (1 mm) were individually or sequentially applied to evoke increases in intracellular Ca2+. Coapplication of LA and Na+/K+ channel blockers was used to evaluate the role of Na+ and K+ channels in the LA effect on the evoked [Ca2+](i) transients. Results All four LAs concentration-dependently inhibited both KCl- and carbachol-evoked [Ca2+](i) transients with the potency order bupivacaine > ropivacaine > lidocaine >/= mepivacaine. The carbachol-evoked [Ca2+](i) transients were more sensitive to LAs without than with a KCl prestimulation, whereas the LA-effect on the KCl-evoked [Ca2+](i) transients was not uniformly affected by a carbachol prestimulation. Na+ channel blockade did not alter the evoked [Ca2+](i) transients with or without a LA. In the absence of LA, K+ channel blockade increased the KCl-, but decreased the carbachol-evoked [Ca2+](i) transients. A coapplication of LA and K+ channel blocker resulted in larger inhibition of both KCl- and carbachol-evoked [Ca2+](i) transients than by LA alone. Conclusions Different and overlapping sites of action of LAs are involved in inhibiting the KCl- and carbachol-evoked [Ca2+](i) transients, including voltage-dependent Ca2+ channels, a site associated with the caffeine-sensitive Ca2+ store and a possible site associated with the IP(3)-sensitive Ca2+ store, and a site in the muscarinic pathway. K+ channels, but not Na+ channels, seem to modulate the evoked [Ca2+](i) transients, as well as the LA-effects on such responses.

Inward currents underlying action potentials in rat adrenal chromaffin cells

1996 ◽  
Vol 76 (2) ◽  
pp. 1195-1211 ◽  
Author(s):  
B. Hollins ◽  
S. R. Ikeda
Keyword(s):  
Ion Channels ◽  
Voltage Clamp ◽  
Action Potentials ◽  
Chromaffin Cells ◽  
Na Channels ◽  
Adrenal Chromaffin Cells ◽  
Voltage Dependent ◽  
Slope Factor ◽  
Inward Currents ◽  
Adrenal Chromaffin

1. Current- and voltage-clamp studies were conducted on isolated rat adrenal chromaffin cells to identify the voltage-dependent ion channels mediating inward currents. 2. Mean resting membrane potential of the isolated cells was -62 +/- 3 (SE) mV. Evoked action potentials were both Na+ and Ca2+ based, and whole cell voltage-clamp studies in normal saline revealed an inward-rectifier-type current. 3. Na+ channels were studied in isolation and showed a half-inactivation of -60 +/- 2 mV with a slope factor of -6 mV and a half-activation of -26.8 +/- 2 mV with a slope factor of 6.5 +/- 0.7 mV. 4. Isolated Ca2+ currents, elicited in 10 mM external Ca2+, revealed a T-type current in a subset of cells. Ca2+ currents were sensitive to both N- and L-type channel antagonists, and blockade of the current by the L-type channel antagonist nimodipine and the N-type channel antagonist omega-conotoxin GVIA revealed a third Ca2+-current component that was unaffected by the P-type channel antagonist omega-agatoxin IVA. 5. Ca2+ currents were facilitated 5-20% by a depolarizing prepulse, and facilitation was completely blocked by nimodipine. The effects of the dihydropyridine L-type channel agonist, (+)202-791 and depolarizing prepulses on the currents were additive. 6. The results of this study show that the properties of voltage-dependent ion channels in rat chromaffin cells differ from those reported in their counterparts in bovine chromaffin cells. Na+ channels differ in activation and inactivation properties and Ca2+ channels differ in activation, sensitivity to antagonists, and the magnitude of voltage-dependent facilitation.

scholarly journals The effects of microenvironment on the redifferentiation of regenerating neurones: neurite architecture, acetylcholine receptors and Ca2+ channel distribution

1987 ◽  
Vol 132 (1) ◽  
pp. 111-131
Author(s):  
M. E. Spira ◽  
D. Zeldes ◽  
B. Hochner ◽  
A. Dormann
Keyword(s):  
Action Potentials ◽  
Acetylcholine Receptors ◽  
Membrane Properties ◽  
Environmental Cues ◽  
Freezing And Thawing ◽  
Intrinsic Factors ◽  
Extrinsic Cues ◽  
Voltage Dependent ◽  
Synaptic Receptors ◽  
Ca2 Channels

Severed adult neurones, which are capable of regrowth, encounter different microenvironments from those encountered during development. Moreover, adult neurones may respond in a different manner from developing neurones to the same environmental cues. Thus, the recovery of the integrative and transmission capabilities (which depend on the neuronal architecture, passive and active membrane properties, and synaptic receptor distribution) by a regenerating adult neurone may not be complete. In the present review, we examine several aspects of the outcome of the interaction between the microenvironment and regrowing neurones using the cockroach giant interneurones (GINs) as a model system. We demonstrate that whereas extrinsic cues govern the morphological redifferentiation and distribution of synaptic receptors, the distribution of voltage-dependent Ca2+ channels is to a large extent determined by intrinsic factors. The pathway of regrowth and the architecture of regenerating GINs were studied by examination of intracellularly stained fibres. The environments provided by the connectives and ganglia are different. The elongating sprouts in the connective appeared as smooth cylinders. Within the ganglionic domain, the main longitudinal sprouts emitted neurites which extended and branched into the neuropile. The local cues for branching of neurites were eliminated by freezing and thawing of the ganglia prior to the arrival of the growing tips. The failure to extend neurites under these conditions is attributed to the elimination of extrinsic signals for morphological redifferentiation of the fibres, since the same fibres emit neurites in anterior ganglia which have not been subjected to freezing and thawing. The distribution of acetylcholine receptors (AChRs) on the GINs was mapped by ionophoretic application of ACh. In both the intact and regenerating GINs receptors were located only on the neurites. Freezing and thawing of a ganglion eliminated the local signals for insertion and/or activation of AChRs on the neurites. Thus, both the morphological redifferentiation and the distribution of AChRs are affected by the microenvironment. Voltage-dependent Ca2+ channels were detected after intracellular injection of tetraethylammonium into the GIN and in the presence of tetrodotoxin (TTX) and Ba2+ in the extracellular space. The regrowing axon tips always revealed large barium action potentials independent of the CNS microenvironment. This observation is consistent with the hypothesis that Ca2+ plays an important role in the growth process. However, increased Ba2+ responsiveness was also observed in axonal segments proximal to the region of neuronal extension.(ABSTRACT TRUNCATED AT 400 WORDS)

A model for dendritic Ca2+ accumulation in hippocampal pyramidal neurons based on fluorescence imaging measurements

1994 ◽  
Vol 71 (3) ◽  
pp. 1065-1077 ◽  
Author(s):  
D. B. Jaffe ◽  
W. N. Ross ◽  
J. E. Lisman ◽  
N. Lasser-Ross ◽  
H. Miyakawa ◽  
...  
Keyword(s):  
Fluorescence Imaging ◽  
Action Potentials ◽  
Hippocampal Neurons ◽  
Pyramidal Neurons ◽  
Na Channels ◽  
Hippocampal Pyramidal Neurons ◽  
K Channels ◽  
Voltage Gated ◽  
Synaptic Potentials ◽  
Ca2 Channels

1. High-speed fluorescence imaging was used to measure intracellular Ca2+ concentration ([Ca2+]i) changes in hippocampal neurons injected with the Ca(2+)-sensitive indicator fura-2 during intrasomatic and synaptic stimulation. The results of these experiments were used to construct a biophysical model of [Ca2+]i dynamics in hippocampal neurons. 2. A compartmental model of a pyramidal neuron was constructed incorporating published passive membrane properties of these cells, three types of voltage-gated Ca2+ channels characterized from adult hippocampal neurons, voltage-gated Na+ and K+ currents, and mechanisms for Ca2+ buffering and extrusion. 3. In hippocampal pyramidal neurons imaging of Na+ entry during electrical activity suggests that Na+ channels, at least in sufficient density to sustain action potentials, are localized in the soma and the proximal part of the apical dendritic tree. The model, which incorporates this distribution, demonstrates that action potentials attenuate steeply in passive distal dendritic compartments or distal dendritic compartments containing Ca2+ and K+ channels. This attenuation was affected by intracellular resistivity but not membrane resistivity. 4. Consistent with fluorescence imaging experiments, a non-uniform distribution of Ca2+ accumulation was generated by Ca2+ entry through voltage-gated Ca2+ channels opened by decrementally propagating Na+ action potentials. Consequently, the largest increases in [C2+]i were produced in the proximal dendrites. Distal voltage-gated Ca2+ currents were activated by broad, almost isopotential action potentials produced by reducing the overall density of K+ channels. 5. Simulations of subthreshold synaptic stimulation produced dendritic Ca2+ entry by the activation of voltage-gated Ca2+ channels. In the model these Ca2+ signals were localized near the site of synaptic input because of the attenuation of synaptic potentials with distance from the site of origin and the steep voltage-dependence of Ca2+ channel activation. 6. These simulations support the hypotheses generated from experimental evidence regarding the differential distribution of voltage-gated Ca2+ and Na+ channels in hippocampal neurons and the resulting voltage-gated Ca2+ accumulation from action and synaptic potentials.

scholarly journals Author response: A native prokaryotic voltage-dependent calcium channel with a novel selectivity filter sequence

2019 ◽  
Author(s):  
Takushi Shimomura ◽  
Yoshiki Yonekawa ◽  
Hitoshi Nagura ◽  
Michihiro Tateyama ◽  
Yoshinori Fujiyoshi ◽  
...  
Keyword(s):  
Calcium Channel ◽  
Selectivity Filter ◽  
Author Response ◽  
Voltage Dependent Calcium Channel ◽  
Voltage Dependent
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