G-protein- and cAMP-dependent L-channel gating modulation: a manyfold system to control calcium entry in neurosecretory cells.

Voltage-gated Ca2+ channels are crucial to the control of Ca2+ entry in neurosecretory cells. In the chromaffin cells of adrenal medulla, paracrinally or autocrinally released neurotransmitters induce profound changes in Ca2+ channel gating and Ca2+-dependent events controlling catecholamine secretion and cell activity. The generally held view of these processes is that neurotransmitter-induced modulation of the most widely expressed Ca2+ channels in these cells (N-, P/Q- and L-type) follows two distinct pathways: a direct membrane-delimited Gi/o-protein-induced inhibition of N- and P/Q-type and a remote cAMP-mediated facilitation of L-channels. Both actions depend on voltage, although with remarkably different molecular and kinetic aspects. Recent findings, however, challenge this simple scheme and suggest that L-channels do not require strong pre-pulses to be recruited or facilitated. They are available during normal depolarizations and may be tonically inhibited by Gi/o proteins activated by the released neurotransmitters. Like the N- and P/Q-channels, this autocrine modulation is localized to membrane microareas. Unlike N- and P/Q-channels, however, the inhibition of L-channels is largely independent of voltage and develops in parallel with cAMP-mediated potentiation of channel gating. As L-channels play a crucial role in the control of catecholamine release in chromaffin cells, the two opposite modulations mediated by Gi/o proteins and cAMP may represent an effective way to broaden the dynamic range of Ca2+ signals controlling exocytosis. Here, we review the basic features of this novel L-type channel inhibition comparing it to the well-established forms of L-channel potentiation and voltage-dependent facilitation.

Synthesis and voltage-clamp studies of methyl 1,4-dihydro-2, 6-dimethyl-5-nitro-4-(benzofurazanyl)pyridine-3-carboxylate racemates and enantiomers and of their benzofuroxanyl analogues.

Racemic methyl 1,4-dihydro-2, 6-dimethyl-5-nitro-4-(benzofurazanyl)pyridine-3-carboxylates (+/-)-10 and (+/-)-11 and their benzofuroxanyl analogues (+/-)-12 and (+/-)-13 were prepared using a modified Hantzsch reaction that involved the condensation of nitroacetone with methyl 3-aminocrotonate and the appropriate aldehydes. The racemic mixtures were resolved into the corresponding enantiomers. Whole-cell voltage-clamp studies on L-type Ca2+ channels expressed in a rat insulinoma cell line (RINm5F) showed that all the dextrorotatory antipodes were effective agonists of L-type Ca2+ currents, while the levorotatory ones were weak Ca2+ entry blockers. The (+)-enantiomer of benzofurazan-5'-yl derivative 11 demonstrated unusual activity in that, in addition to producing a potentiation of L-type currents, it interfered with the voltage-dependent gating of L-type channels by producing a net delay of their activation at low voltages. This compound represents an interesting tool to probe L-type Ca2+ channel structure and function.