Q-type Ca 2+ channels are located closer to secretory sites than L-type channels: functional evidence in chromaffin cells

Abstract

This study uses a new strategy to investigate the hypothesis that, of the various Ca2+ channels expressed by a neurosecretory cell, a given channel subtype is coupled more tightly to the exocytotic apparatus than others. The approach is based on the prediction that the degree of inhibition of the secretory response by various Ca2+ channel blockers will differ at low (0.5 mM) and high (5 mM) extracellular Ca2+ concentrations ([Ca2+]o). So, at low [Ca2+]o the K+-evoked catecholamine release from superfused bovine chromaffin cells was depressed 60-70% by 2 microM omega-agatoxin IVA (P/Q-type Ca2+ channel blockade), by 3 microM omega-conotoxin MVIIC (N/P/Q-type Ca2+ channel blockade), or by 3 microM lubeluzole (N/P/Q-type Ca2+ channel blockade); in high [Ca2+]o these blockers inhibited the responses by only 20-35%. At 1-3 microM omega-conotoxin GVIA (N-type Ca2+ channel blockade) or 3 microM furnidipine (L-type Ca2+ channel blockade), secretion was inhibited by 30 and 50%, respectively; such inhibitory effects were similar in low or high [Ca2+]o. Combined furnidipine plus omega-conotoxin MVIIC, omega-agatoxin IVA or omega-conotoxin GVIA exhibited additive blocking effects at both Ca2+ concentrations. The results suggest that Q-type Ca2+ channels are coupled more tightly to exocytotic active sites, as compared to L-type channels. This hypothesis if founded in the fact that external Ca2+ that enters the cell through a Ca2+ channel located near to chromaffin vesicles will saturate the K+ secretory response at both [Ca2+]o, i.e. 0.5 mM and 5 mM. In contrast, Ca2+ ions entering through more distant channels will be sequestered by intracellular buffers and, thus, will not saturate the secretory machinery at lower [Ca2+]o.

L-type calcium channels in adrenal chromaffin cells: Role in pace-making and secretion

“…L-type channels predominate the secretion despite their contribution to the total current in single cell experiments is partial or very small, as in the case of BCCs [73,74]. A dominance of L-type channel-mediated secretion is thus reported in bovine [4,[75][76][77], mouse [78,79], rat [9,10,80,81] and cat chromaffin cells [82,83] more or less independently of whether intact adrenal glands or cultured chromaffin cells are used. The reason for this general finding is that during prolonged depolarization (far from physiological conditions) L-type channels are probably more favored because of their slower time-dependent inactivation and lower steady-state inactivation with respect to the other HVA channels (P/Q, N and R), which inactivate more rapidly and completely during prolonged stimuli.…”

Neuronal somatic ATP release triggers neuron–satellite glial cell communication in dorsal root ganglia

“…We (Fig. 2D) and others (7,10) have found that L-type Ca 2ϩ channels, which are known to be loosely associated with release machinery (34,35), mediate somatic exocytosis. In contrast, N-and P/Q types Ca 2ϩ channels, which are closely associated with fusion machinery, are involved in the exocytosis at synaptic terminals (34,36).…”

“…2D) and others (7,10) have found that L-type Ca 2ϩ channels, which are known to be loosely associated with release machinery (34,35), mediate somatic exocytosis. In contrast, N-and P/Q types Ca 2ϩ channels, which are closely associated with fusion machinery, are involved in the exocytosis at synaptic terminals (34,36). These observations also are consistent with our conclusion that the intracellular Ca 2ϩ levels required for somatic release is lower than those required for the release from most nerve terminals (4,36).…”

Fast exocytosis mediated by T- and L-type channels in chromaffin cells: distinct voltage-dependence but similar Ca2+-dependence

“…This slower exocytotic process is primarily controlled by Ca 2+ inXux through multiple isoforms of high-voltage activated Ca 2+ channels, which contribute to secretion proportionally to their density of expression and gating properties, with no speciWc requirements of Ca 2+ channels colocalization to the release sites (Kim et al 1995;Engisch and Nowycky 1996;Klingauf and Neher 1997;Lukyanetz and Neher 1999;Ulate et al 2000;Carabelli et al 2003;Chan et al 2005;Giancippoli et al 2006;Polo-Parada et al 2006). Alternatively, other groups suggest a preferential role of some channel subtype in close proximity to the secretory sites (Lopez et al 1994;Artalejo et al 1994;Lara et al 1998;O'Farrel and Marley 2000;Albillos et al 2000) and a further role of mitochondria in sequestering Ca 2+ entering through Ca 2+ channels (Ales et al 2005). In addition to this, the eYciency of stimulus-secretion coupling can be altered in many ways, some relevant examples are: (1) PKC activation, which enhances exocytosis (Gillis et al 1996), (2) rapid and sustained increase of intracellular cAMP, which potentiates Ltype channel densities and secretion at diVerent proportions , (3) transdiVerentiation of chromaYn cells with astrocyte-conditioned medium, which promotes the acquisition of a neuronal phenotype and accelerates the exocytotic kinetics, by a better coupling between secretory vesicles and Ca 2+ channels (Ardiles et al 2006) and (4) -adrenergic stimulation, which enhances the stimulus-secretion coupling of Ltype Ca 2+ channels by repeated action potential-like stimulations (Polo-Parada et al 2006).…”