Inhibition of calcium channels in rat central and peripheral neurons by omega-conotoxin MVIIC

Abstract

Inhibition of voltage-dependent calcium channels by omega-conotoxin MVIIC (omega-CTx-MVIIC) was studied in various types of rat neurons. When studied with 5 mM Ba2+ as charge carrier, omega-CTx-MVIIC block of N-type calcium channels in sympathetic neurons was potent, with half-block at 18 nM. Block of N-type channels had a rapid onset (tau approximately 1 sec at 1 microM omega-CTx-MVIIC) and quick reversibility (tau approximately 30 sec). The rate of block was proportional to toxin concentration, consistent with 1:1 binding of toxin to channels, with a rate constant (k on) of approximately 1 X 10(6) M-1. sec-1. Both potency and rate of block were reduced dramatically with increasing concentrations of extracellular Ba2+ omega-CTx-MVIIC also blocked P-type calcium channels in cerebellar Purkinje neurons, but both development and reversal of block were far slower than for N-type channels. The rate of block was proportional to toxin concentration, with k on -1.5 x 10(3) M-1. sec-1 at 5 mM Ba2+. From this value and an unblocking time constant of approximately 200 min, a dissociation constant of approximately 50 nM was estimated. Thus, block of P-type channels is potent but very slow. In hippocampal CA3 pyramidal neurons, omega-CTx-MVIIC blocked approximately 50% of the high-threshold calcium channel current; one component (approximately 20%) was blocked with the rapid kinetics expected for N-type channels, whereas the other component was blocked slowly. The component blocked slowly was reduced but not eliminated by preexposure to 200 nM or 1 microM omega-Aga-IVA.

Postsynaptic Calcium Transients Evoked by Activation of Individual Hippocampal Mossy Fiber Synapses

“…6 B, C); the effect was seen even at complex spines adjacent to the soma in which the block cannot be attributable to interference with action potential propagation. The widespectrum blocker of N-, P-, and Q-type VACCs, -conotoxin MVIIC (2 M) (McDonough et al, 1996), also substantially reduced the action potential-evoked Ca 2ϩ transients at complex spines (49.0 Ϯ 8% of baseline; n ϭ 4; p Ͻ 0.02), providing additional evidence that these channels are present and functional at complex spines.…”

β-Adrenergic Stimulation Selectively Inhibits Long-Lasting L-Type Calcium Channel Facilitation in Hippocampal Pyramidal Neurons

“…N-, P-, and Q-type channel currents were blocked by preincubating cells with -conotoxins GV IA and M V IIC (McCleskey et al, 1987;McDonough et al, 1996). Under these conditions the channel activity observed during delayed facilitation was blocked by the DHP antagonist nimodipine (1 M; n ϭ 4) (results not shown).…”

“…Acutely dissociated hippocampal CA1 neurons were obtained as described previously (McDonough et al, 1996). Briefly, Sprague Dawley rats (9 -14 d old) were anesthetized with halothane and decapitated.…”

“…Before recording, cells were incubated for 20 -30 min in a modified Krebs' solution containing (in mM): NaC l, 150; KC l, 5; C aC l 2 , 1; MgCl 2 , 1; H EPES, 10; and glucose, 5, pH 7.4, supplemented with -conotoxins M V IIC (5 M) and GV IA (1 M) to block P-, Q-, and N-type C a channels (McC leskey et al, 1987;McDonough et al, 1996). During recording, cells were superf used (15 ml /min) with an external solution containing (in mM): potassium aspartate, 125; KC l, 35; MgC l 2 , 5; H EPES(Na), 10; EGTA, 10; and C aC l 2 , 5.64 (to give an estimated free concentration of 60 nM) (Fabiato and Fabiato, 1979), pH 7.4, with C sOH.…”

“…L -type C a channels are prevalent in hippocampal pyramidal neurons, contributing ϳ30 -50% of total calcium current (Mintz et al, 1992;Swartz and Bean, 1992;McDonough et al, 1996). The majority of L -type channels are distributed on the surface of both the cell body and the proximal dendrites .…”