Calcium Channels In Nerve Terminals Research Articles

The Number and Distinct Clustering Patterns of Voltage-Gated Calcium Channels in Nerve Terminals.

Upon the arrival of action potentials at nerve terminals, neurotransmitters are released from synaptic vesicles (SVs) by exocytosis. CaV2.1, 2.2, and 2.3 are the major subunits of the voltage-gated calcium channel (VGCC) responsible for increasing intraterminal calcium levels and triggering SV exocytosis in the central nervous system (CNS) synapses. The two-dimensional analysis of CaV2 distributions using sodium dodecyl sulfate (SDS)-digested freeze-fracture replica labeling (SDS-FRL) has revealed their numbers, densities, and nanoscale clustering patterns in individual presynaptic active zones. The variation in these properties affects the coupling of VGCCs with calcium sensors on SVs, synaptic efficacy, and temporal precision of transmission. In this study, we summarize how the morphological parameters of CaV2 distribution obtained using SDS-FRL differ depending on the different types of synapses and could correspond to functional properties in synaptic transmission.

Effects of IgG anti-GM1 monoclonal antibodies on neuromuscular transmission and calcium channel binding in rat neuromuscular junctions.

Guillain-Barré syndrome is a type of acute inflammatory neuropathy that causes ataxia and is associated with the IgG anti-GM1 antibody. However, the pathogenic role of the IgG anti-GM1 antibody and calcium channels in neuromuscular junctions (NMJs) remains unclear. Thus, the aim of the present study was to investigate the effects of the IgG anti-GM1 monoclonal antibody (mAb) on spontaneous muscle action potentials (SMAPs), and the effects of calcium channel blockers, in a rat spinal cord-muscle co-culture system. In addition, the binding of IgG anti-GM1 mAb to calcium channels was investigated in the rat hemidiaphragm. The frequency of SMAPs in the innervated muscle cells was acutely inhibited by the IgG anti-GM1 mAb; however, this effect was blocked by the N-type calcium channel blocker, ω-conotoxin GVIA (30 nM). Furthermore, the P/Q-type calcium channel blocker, ω-agatoxin IVA (10 nM), was found to partially block the IgG anti-GM1 mAb-induced inhibitory effect in the spinal cord-muscle co-culture system. Immunohistochemical analysis of the rat hemidiaphragm indicated that IgG anti-GM1 mAb binding overlapped with anti-Cav2.2 (α1B) antibody binding in the nerve terminal. In addition, IgG anti-GM1 mAb binding partially overlapped with anti-Cav2.1 (α1A) antibody binding. Thus, the results demonstrated that the IgG anti-GM1 mAb binds to calcium channels in the nerve terminals of NMJs. Therefore, the inhibitory effect of IgG anti-GM1 mAb on SMAPs may involve N-type and P/Q-type calcium channels in motor nerve terminals at the NMJ.

Modulation of calcium currents is eliminated after cleavage of a strategic component of the mammalian secretory apparatus

Adenosine inhibits neurotransmitter secretion from motor nerves by an effect on the secretory apparatus in amphibia. In contrast, the inhibitory effect of adenosine is associated with decreases in calcium currents at mouse motor nerve endings. To determine if the action of adenosine in the mouse is mediated thorough a direct effect on calcium channels or through the secretory machinery, the effects of cleavage of the SNARE proteins on the action of adenosine were examined. Cleavage of the SNARE syntaxin with botulinum toxin type C (Botx/C) prevented the inhibitory effect of adenosine on nerve terminal calcium currents. Cleavage of the other SNAREs (synaptobrevin with Botx/D or SNAP-25 with Botx/A) failed to affect the inhibitory action of adenosine. The results provide evidence for an intimate coupling of nerve terminal calcium channels with a plasma membrane component of the SNARE complex, such that modulation of calcium currents by a G-protein coupled receptor cannot occur when syntaxin is cleaved.

Biochemical properties and subcellular distribution of an N-type calcium hannel α1 subunit

A site-directed anti-peptide antibody, CNB-1, that recognizes the α1 subunit of rat brain class B calcium channels (rbB) immunoprecipitated 43% of the N-type calcium channels labeled by [ 125I]ω-conotoxin. CNB-1 recognized proteins of 240 and 210 kd, suggesting the presence of two size forms of this α1 subunit. Calcium channels recognized by CNB-1 were localized predominantly in dendrites; both dendritic shafts and punctate synaptic structures upon the dendrites were labeled. The large terminals of the mossy fibers of the dentate gyrus granule neurons were heavily labeled, suggesting that the punctate labeling pattern represents calcium channels in nerve terminals. The pattern of immunostaining was cell specific. The cell bodies of some pyramidal cells in layers II, III, and V of the dorsal cortex, Purkinje cells, and scattered cell bodies elsewhere in the brain were also labeled at a low level. The results define complementary distributions of N- and L-type calcium channels in dendrites, nerve terminals, and cell bodies of most central neurons and support distinct functional roles in calcium-dependent electrical activity, intracellular calcium regulation, and neurotransmitter release for these two channel types.

Specificity of Lambert-Eaton Myasthenic Syndrome immunoglobulin for nerve terminal calcium channels

Lambert-Eaton Myasthenic Syndrome (LEMS) is a presynaptic, neuromuscular disorder characterized by impaired nerve-evoked release of ACh. Repetitive nerve stimulation, which increases the probability of quantal release, improves the transmission defect. An autoantibody to Ca 2+ channels of presynaptic motor nerve terminals is thought to mediate the pathogenesis of this disease. The goal of the present study was to examine the specificity of LEMS autoantibodies for nerve terminal Ca 2+ channels as compared to other voltage-sensitive ion channels in nerve terminals, and to determine if non-specific membrane damage contributed to the pathogenesis of LEMS. The ion channel specificity of LEMS autoantibody was assessed by comparing the ability of acute application of IgG isolated from the plasma of a patient with LEMS to reduce depolarization-dependent uptake of 45Ca 2+ and 22Na + into or efflux of 86Rb + from rat forebrain synaptosomes. The clinical diagnosis of LEMS was confirmed electrophysiologically by treatment of mice for 30 days with plasma (1.5 ml/day) taken from this patient. Characteristic reduction of quantal content elicited at 1 Hz and facilitation at 20 Hz was observed in mice treated wtih LEMS plasma compared to those treated with control plasma. One s, K +-stimulated 45Ca 2+ uptake was inhibited36.5 ± 14.5%and44.5 ± 9.8% by acute application of 2 and 4 mg/ml LEMS IgG, respectively; IgG from patients with small cell carcinoma of the lung (SCC) had no effect on 45Ca 2+ entry. The same concentrations of LEMS IgG affected neither voltage-dependent uptake of 22Na + into veratridine-depolarized synaptosomes nor 86R + efflux from K +-depolarized synaptosomes. Lactate dehydrogenase release from synaptosomes incubated with varying concentrations of LEMS IgG was not increased over control suggesting that the reduction in 45Ca 2+ uptake was not a result of disruption of synaptosomal membrane integrity. Likewise, normal activation of Ca 2+ channels was not prevented by IgG-induced alterations in synaptosomal membrane potentials as the fluorescence of a membrane potential-sensitive carbocyanine dye (diS-C 2(5)) was unaltered following acute application of 2 mg/ml LEMS or control IgG. These results indicate that immunoglobulins isolated from a patient with LEMS alter presynaptic nerve terminals Ca 2+ channel activity. These deficits are not due to decreases in synaptosomal integrity nor to IgG-induced alterations in the synaptosomal membrane potential but appear to result from direct action on voltage-dependent Ca 2+ channels. The antibody from this patient appears to be specific for nerve terminal Ca 2+ channels as it did not affect ion flux through voltage-dependent Na + or K + channels.

Review of evidence for loss of motor nerve terminal calcium channels in Lambert-Eaton myasthenic syndrome.

Review of evidence for loss of motor nerve terminal calcium channels in Lambert-Eaton myasthenic syndrome.

Calcium-dependent depolarizations originating in lizard motor nerve terminals

Action potentials and afterpotentials were recorded via a microelectrode inserted into motor axons innervating the lizard ceratomandibularis muscle. Intra-axonal injection of Lucifer yellow dye indicated that these axons innervate multiple (at least 6-25) motor terminals. In 10 mM tetraethylammonium (TEA), the action potential was followed by a sequence of afterpotentials, whose amplitude and duration increased with increasing proximity to motor nerve terminals. In axons impaled within 1 mm of their most distal terminals, these afterpotentials included a depolarizing plateau (mean amplitude and duration: 32 mV, 24 msec) and a subsequent smaller depolarization that decayed over a time course of several hundred milliseconds. These depolarizing afterpotentials were Ca dependent: They increased with increasing bath [Ca] and were abolished by low [Ca]-high [Mg] solutions, by omega-conotoxin (GVIA, 1 microM), by addition of Cd (1 microM) or Mn (0.3-1 mM) to the bath, and by selective perfusion of Cd over the terminal region. In [Ca]-free solutions the afterpotentials were restored by selective perfusion of Ca over the terminal region but not by Ca applied to the more proximal nerve trunk. When Na influx was eliminated by 1-10 microM tetrodotoxin or by substitution of TEA for bath Na, passage of depolarizing current into the axon evoked prolonged depolarizing afterpotentials that were blocked by Mn. Bay K 8644 (0.1-1 microM), a dihydropyridine that prolongs the opening of certain calcium channels, enhanced mainly the slower component of the depolarizing afterpotential. Nimodipine (0.1-1 microM), a dihydropyridine that favors the closed state of some calcium channels, shortened the plateau phase of the depolarizing afterpotential. Another antagonist dihydropyridine, nitrendipine (0.1-1 microM), had little or no effect on the depolarizing afterpotential but did antagonize the actions of Bay K 8644. These results suggest that the intraaxonally recorded Ca-dependent afterpotentials are caused by electrotonic spread of depolarizations produced by calcium influx into that axon's terminals and that some motor nerve terminal calcium channels are sensitive to certain dihydropyridines.