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After Ca2+ influx, mitochondria can sequester Ca2+ and subsequently release it back into the cytosol. This form of Ca2+-induced Ca2+ release (CICR) prolongs Ca2+ signaling and can potentially mediate activity-dependent plasticity. As Ca2+ is required for its subsequent release, Ca2+ removal systems, like the plasma membrane Ca2+-ATPase (PMCA), could impact CICR. Here we examine such a role for the PMCA in the bag cell neurons of Aplysia californica. CICR is triggered in these neurons during an afterdischarge and is implicated in sustaining membrane excitability and peptide secretion. Somatic Ca2+ was measured from fura-PE3-loaded cultured bag cell neurons recorded under whole cell voltage clamp. Voltage-gated Ca2+ influx was elicited with a 5-Hz, 1-min train, which mimics the fast phase of the afterdischarge. PMCA inhibition with carboxyeosin or extracellular alkalization augmented the effectiveness of Ca2+ influx in eliciting mitochondrial CICR. A Ca2+ compartment model recapitulated these findings and indicated that disrupting PMCA-dependent Ca2+ removal increases CICR by enhancing mitochondrial Ca2+ loading. Indeed, carboxyeosin augmented train-evoked mitochondrial Ca2+ uptake. Consistent with their role on Ca2+ dynamics, cell labeling revealed that the PMCA and mitochondria overlap with Ca2+ entry sites. Finally, PMCA-dependent Ca2+ extrusion did not impact endoplasmic reticulum-dependent Ca2+ removal or release, despite the organelle residing near Ca2+ entry sites. Our results demonstrate that Ca2+ removal by the PMCA influences the propensity for stimulus-evoked CICR by adjusting the amount of Ca2+ available for mitochondrial Ca2+ uptake. This study highlights a mechanism by which the PMCA could impact activity-dependent plasticity in the bag cell neurons. [ABSTRACT FROM AUTHOR]

Although the contribution of Ca2+ buffering systems can vary between neuronal types and cellular compartments, it is unknown whether distinct Ca2+ sources within a neuron have different buffers. As individual Ca2+ sources can have separate functions, we propose that each is handled by unique systems. Using Aplysia californica bag cell neurons, which initiate reproduction through an afterdischarge involving multiple Ca2+-dependent processes, we investigated the role of endoplasmic reticulum (ER) and mitochondrial sequestration, as well as extrusion via the plasma membrane Ca2+-ATPase (PMCA) and Na+/Ca2+ exchanger, to the clearance of voltage-gated Ca2+ influx, Ca2+-induced Ca2+-release (CICR), and store-operated Ca2+ influx. Cultured bag cell neurons were filled with the Ca2+ indicator, fura-PE3, to image Ca2+ under whole-cell voltage clamp. A 5 Hz, 1 min train of depolarizing voltage steps elicited voltage-gated Ca2+ influx followed by EGTA-sensitive CICR from the mitochondria. A compartment model of Ca2+ indicated the effect of EGTA on CICR was due to buffering of released mitochondrial Ca2+ rather than uptake competition. Removal of voltage-gated Ca2+ influx was dominated by the mitochondria and PMCA, with no contribution from the Na+/Ca2+ exchanger or sarcoplasmic/endoplasmic Ca2+-ATPase (SERCA). In contrast, CICR recovery was slowed by eliminating the Na+/Ca2+ exchanger and PMCA. Last, store-operated influx, evoked by ER depletion, was removed by the SERCA and depended on the mitochondrial membrane potential. Our results demonstrate that distinct buffering systems are dedicated to particular Ca2+ sources. In general, this may represent a means to differentially regulate Ca2+-dependent processes, and for Aplysia, influence how reproductive behavior is triggered. [ABSTRACT FROM AUTHOR]