Mitochondrial Ca2+Activates a Cation Current inAplysiaBag Cell Neurons

Abstract

Ion channels may be gated by Ca2+entering from the extracellular space or released from intracellular stores—typically the endoplasmic reticulum. The present study examines how Ca2+impacts ion channels in the bag cell neurons of Aplysia californica. These neuroendocrine cells trigger ovulation through an afterdischarge involving Ca2+influx from Ca2+channels and Ca2+release from both the mitochondria and endoplasmic reticulum. Liberating mitochondrial Ca2+with the protonophore, carbonyl cyanide-4-trifluoromethoxyphenyl-hydrazone (FCCP), depolarized bag cell neurons, whereas depleting endoplasmic reticulum Ca2+with the Ca2+-ATPase inhibitor, cyclopiazonic acid, did not. In a concentration-dependent manner, FCCP elicited an inward current associated with an increase in conductance and a linear current/voltage relationship that reversed near −40 mV. The reversal potential was unaffected by changing intracellular Cl−, but left-shifted when extracellular Ca2+was removed and right-shifted when intracellular K+was decreased. Strong buffering of intracellular Ca2+decreased the current, although the response was not altered by blocking Ca2+-dependent proteases. Furthermore, fura imaging demonstrated that FCCP elevated intracellular Ca2+with a time course similar to the current itself. Inhibiting either the V-type H+-ATPase or the ATP synthetase failed to produce a current, ruling out acidic Ca2+stores or disruption of ATP production as mechanisms for the FCCP response. Similarly, any involvement of reactive oxygen species potentially produced by mitochondrial depolarization was mitigated by the fact that dialysis with xanthine/xanthine oxidase did not evoke an inward current. However, both the FCCP-induced current and Ca2+elevation were diminished by disabling the mitochondrial permeability transition pore with the alkylating agent, N-ethylmaleimide. The data suggest that mitochondrial Ca2+gates a voltage-independent, nonselective cation current with the potential to drive the afterdischarge and contribute to reproduction. Employing Ca2+from mitochondria, rather than the more common endoplasmic reticulum, represents a diversification of the mechanisms that influence neuronal activity.