Calcium-dependent inhibition of L, N, and P/Q Ca2+ channels in chromaffin cells: Role of mitochondria

Abstract

The hypothesis that the buffering of Ca2+ by mitochondria could affect the Ca2+-dependent inhibition of voltage-activated Ca2+ channel, (/Ca), was tested in voltage-clamped bovine adrenal chromaffin cells. The protonophore carbonyl cyanide m-chlorophenyl-hydrazone (CCCP), the blocker of the Ca2+ uniporter ruthenium red (RR), and a combination of oligomycin plus rotenone were used to interfere with mitochondrial Ca2+ buffering. In cells dialyzed with an EGTA-free solution, peak /Ca generated by 20 msec pulses to 0 or +10 mV, applied at 15 sec intervals, from a holding potential of -80 mV, decayed rapidly after superfusion of cells with 2 μM CCCP (τ = 16.7 ± 3 sec; n = 8). In cells dialyzed with 14 mM EGTA, CCCP did not provoke /Ca loss. Cell dialysis with 4 μM ruthenium red or cell superfusion with oligomycin (3 μM) plus rotenone (4 μM) also accelerated the decay of /Ca. After treatment with CCCP, decay of N- and P/Q-type Ca2+ channel currents occurred faster than that of L-type Ca2+ channel currents. These data are compatible with the idea that the elevation of the bulk cytosolic Ca2+ concentration, [Ca2+]c, causes the inhibition of L- and N- as well as P/Q-type Ca2+ channels expressed by bovine chromaffin cells. This [Ca2+]c signal appears to be tightly regulated by rapid Ca2+ uptake into mitochondria. Thus, it is plausible that mitochondria might efficiently regulate the activity of L, N, and P/Q Ca2+ channels under physiological stimulation conditions of the cell.