G-Protein Inhibition of N- and P/Q-Type Calcium Channels: Distinctive Elementary Mechanisms and Their Functional Impact

Abstract

Voltage-dependent G-protein inhibition of presynaptic Ca 2ϩ channels is a key mechanism for regulating synaptic efficacy. G-protein ␤␥ subunits produce such inhibition by binding to and shifting channel opening patterns from high to low open probability regimes, known respectively as "willing" and "reluctant" modes of gating. Recent macroscopic electrophysiological data hint that only N-type, but not P/Q-type channels can open in the reluctant mode, a distinction that could enrich the dimensions of synaptic modulation arising from channel inhibition. Here, using high-resolution single-channel recording of recombinant channels, we directly distinguished this core contrast in the prevalence of reluctant openings. Single, inhibited N-type channels manifested relatively infrequent openings of submillisecond duration (reluctant openings), which differed sharply from the high-frequency, millisecond gating events characteristic of uninhibited channels. By contrast, inhibited P/Q-type channels were electrically silent at the single-channel level. The functional impact of the differing inhibitory mechanisms was revealed in macroscopic Ca 2ϩ currents evoked with neuronal action potential waveforms (APWs). Fitting with a change in the manner of opening, inhibition of such N-type currents produced both decreased current amplitude and temporally advanced waveform, effects that would not only reduce synaptic efficacy, but also influence the timing of synaptic transmission. On the other hand, inhibition of P/Q-type currents evoked by APWs showed diminished amplitude without shape alteration, as expected from a simple reduction in the number of functional channels. Variable expression of N-and P/Q-type channels at spatially distinct synapses therefore offers the potential for custom regulation of both synaptic efficacy and synchrony, by G-protein inhibition.

Differential Gating and Recruitment of P/Q-, N-, and R-Type Ca2+Channels in Hippocampal Mossy Fiber Boutons

“…Presynaptic N-type channels showed a slightly, but not significantly, more positive midpoint potential of activation curve, but almost identical activation and deactivation kinetics than P/Q-type channels. These findings are consistent with the slight differences between native and recombinant P/Q-and N-type channels reported previously (Wheeler et al, 1996;Colecraft et al, 2000). In contrast, presynaptic R-type channels in MFBs showed a markedly lower activation threshold and a shallower steady-state activation curve than P/Q-or N-type channels, consistent with previous data for recombinant channels (Bourinet et al, 1996).…”

Domain III regulates N-type (CaV2.2) calcium channel closing kinetics

“…Since L-channel deactivation is not affected by roscovitine, we used L-N chimeric channels to demonstrate that the N-channel domain III is required for roscovitineinduced slowed deactivation. Our kinetic and pharmacological evidence supports domain III as a primary controller of cal-cium channel deactivation and suggests that this domain may play a dominant role in generating the high-P O gating characteristic of N-type channels (Colecraft et al 2001;Lee and Elmslie 1999).…”

“…4). The steeper voltage dependence of N-channel Deact likely reflects the voltage dependence of the N-channel open state (Colecraft et al 2001;Lee and Elmslie 1999).…”

“…L-type calcium channels typically gate with brief open times and a low open probability (P o ) (maximum Յ 0.2), while N-type and P/Q-type channels gate with high P o (maximum ϭ 0.8 -0.9) and long open times (Colecraft et al 2001;Elmslie 1997;Forti and Pietrobon 1993;Hess et al 1984;Lee and Elmslie 1999;Marks and Jones 1992). In addition, L-channels show a voltageindependent open state (Marks and Jones 1992), while the N-channel open state is voltage dependent (Colecraft et al 2001;Lee and Elmslie 1999). One possible explanation for these gating differences is domain "specialization," which has been proposed for voltage-dependent sodium channels (Chen et al 1996;Horn et al 2000).…”

G-Proteins Modulate Cumulative Inactivation of N-Type (CaV2.2) Calcium Channels

“…Arguably, the effect of G␤␥ on Ca 2ϩ entry is more closely mimicked by Cgtx than by Cd 2ϩ . Although G␤␥-bound "reluctant" channels display lowprobability openings with submillisecond open times, the most noticeable effect of G␤␥ is to increase the latency to first channel opening (Carabelli et al, 1996;Patil et al, 1996;Lee and Elmslie, 2000;Colecraft et al, 2001). In the context of a brief APW, the major effect will be to decrease the number of Ca-channels that open, with only small effects on Ca 2ϩ flux per se.…”