CA2+ RELEASE FROM SUBPLASMALEMMAL STORES AS A PRIMARY EVENT DURING EXOCYTOSIS IN PARAMECIUM CELLS

A correlated electrophysiological and light microscopic evaluation of trichocyst exocytosis was carried out with Paramecium cells which poss ess extensive cortical Ca stores with footlike links to the plasmalemm a. We used not only intra- but also extracellular recordings to accoun t for polar arrangement of ion channels (while trichocysts can be rele ased from all over the cell surface). With three widely different secr etagogues, aminoethyldextran (AED), veratridine and caffeine, similar anterior Na-in and posterior K-out currents (both known to be Ca2+-dep endent) were observed. Direct de- or hyperpolarization induced by curr ent injection failed to trigger exocytosis. For both, exocytotic membr ane fusion and secretagogue-induced membrane currents, sensitivity to or availability of Ca2+ appears to be different. Current responses to AED were blocked by W7 or trifluoperazine, while exocytosis remained u naffected. Reducing [Ca2+](o), to less than or equal to 0.16 mu M (i.e ., resting [Ca2+](i)) suppressed electrical membrane responses trigger ed with AED, while we had previously documented normal exocytotic memb rane fusion. From this we conclude that the primary effect of AED (as of caffeine) is the mobilization of Ca2+ from the subplasmalemmal pool s which not only activates exocytosis (abolished by iontophoretic EGTA injection) but secondarily also spatially segregated plasmalemmal Ca2 +-dependent ion channels (indicative of subplasmalemmal [Ca2+](i) incr ease, but irrelevant for Ca2+ mobilization). The Ca-45(2+) influx prev iously observed during AED triggering may serve to refill depleted sto res. Apart from the insensitivity of our system to depolarization, the mode of direct Ca2+ mobilization from stores by mechanical coupling t o the cell membrane (without previous Ca2+-influx from outside) closel y resembles the model currently discussed for skeletal muscle triads.