Different roles attributed to Cav1 channel subtypes in spontaneous action potential firing and fine tuning of exocytosis in mouse chromaffin cells

Abstract

J. Neurochem. (2011) 116, 105–121. Abstract This study examines the Cav1 isoforms expressed in mouse chromaffin cells and compares their biophysical properties and roles played in cell excitability and exocytosis. Using immunocytochemical and electrophysiological techniques in mice lacking the Cav1.3α1 subunit (Cav1.3−/−) or the high sensitivity of Cav1.2α1 subunits to dihydropyridines, Cav1.2 and Cav1.3 channels were identified as the only Cav1 channel subtypes expressed in mouse chromaffin cells. Cav1.3 channels were activated at more negative membrane potentials and inactivated more slowly than Cav1.2 channels. Cav1 channels, mainly Cav1.2, control cell excitability by functional coupling to BK channels, revealed by nifedipine blockade of BK channels in wild type (WT) and Cav1.3−/− cells (53% and 35%, respectively), and by the identical change in the shape of the spontaneous action potentials elicited by the dihydropyridine in both strains of mice. Cav1.2 channels also play a major role in spontaneous action potential firing, supported by the following evidence: (i) a similar percentage of WT and Cav1.3−/− cells fired spontaneous action potentials; (ii) firing frequency did not vary between WT and Cav1.3−/− cells; (iii) mostly Cav1.2 channels contributed to the inward current preceding the action potential threshold; and (iv) in the presence of tetrodotoxin, WT or Cav1.3−/− cells exhibited spontaneous oscillatory activity, which was fully abolished by nifedipine perfusion. Finally, Cav1.2 and Cav1.3 channels were essential for controlling the exocytotic process at potentials above and below −10 mV, respectively. Our data reveal the key yet differential roles of Cav1.2 and Cav1.3 channels in mediating action potential firing and exocytotic events in the neuroendocrine chromaffin cell.

Native α6β4∗ nicotinic receptors control exocytosis in human chromaffin cells of the adrenal gland

“…Experiments were started 48 h after plating to allow recovery of the nicotinic receptor after collagenase treatment (26). The protocol for mouse and rat chromaffin cell cultures was performed as described previously (27).…”

Atypical Ca2+ currents in chromaffin cells from SHR and WKY rat strains result from the deficient expression of a splice variant of the α1D Ca2+ channel

“…T-type channels do not inactivate in a Ca 2ϩ -dependent manner, as its rate of inactivation is not altered by changing the current-carrying species (26). Conversely, the smaller amplitude of the non-inactivating, sustained component in WKY/ SHR CCs compared with Wistar CCs could be due to a diminished expression of P/Q-, N-, or L-type Ca 2ϩ channels, although the percentages of each Ca 2ϩ channel subtype in WKY/SHR CCs are similar to those reported in other rodent species (9,12 Ϫ/Ϫ knockout (KO) mice, it has been reported that the number of spontaneously active CCs decreases by Ͼ50% (24), while another study reports no significant difference in spontaneous firing (34). This discrepancy has been explained by the use of a higher KCl concentration in the latter study, which, by producing a mild depolarization (ϳ6 mV), caused an approximately threefold increase of firing frequency (21).…”

“…The fact that spontaneous firing persists, despite the deletion of the Ca V 1.3 ␣ 1 -subunit, is explained by a parallel increase in the expression of other ␣ 1 -subunits, since total I Ca is not altered (24,34). Also, secretion elicited by depolarizing pulses apparently is not altered in Ca V 1.3 Ϫ/Ϫ KO mice compared with WT (34 non-inactivating Ca V 1.3 42 channels apparently is not compensated by overexpression of other sustained I Ca .…”

Selectivity of Action of Pregabalin on Ca2+ Channels but Not on Fusion Pore, Exocytotic Machinery, or Mitochondria in Chromaffin Cells of the Adrenal Gland

“…The mice were housed in the animal facility of the Universidad Autónoma de Madrid (UAM), Madrid Registry number ES-280790000097. The procedure of isolation and culture of cells was performed as reported previously (Pérez-Alvarez et al, 2011). The study protocol for the use of human chromaffin cells was approved by the Ethics Committees of the Hospital Ramón y Cajal and Universidad Autónoma de Madrid.…”