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© The Rockefeller University Press, 0021-9525/1997//597 $5.00
The Journal of Cell Biology, Volume 136, Number 3, , 1997 597-607

Article

Microdomain Ca2+ Activation during Exocytosis in Paramecium Cells. Superposition of Local Subplasmalemmal Calcium Store Activation by Local Ca2+ Influx



Christian Erxleben, Norbert Klauke, Matthias Flötenmeyer, Marie-Pierre Blanchard, Claudia Braun, and Helmut Plattner

Faculty of Biology, University of Konstanz, D-78434 Konstanz, Germany

In Paramecium tetraurelia, polyamine-triggered exocytosis is accompanied by the activation of Ca2+-activated currents across the cell membrane (Erxleben, C., and H. Plattner. 1994. J. Cell Biol. 127:935– 945). We now show by voltage clamp and extracellular recordings that the product of current x time (As) closely parallels the number of exocytotic events. We suggest that Ca2+ mobilization from subplasmalemmal storage compartments, covering almost the entire cell surface, is a key event. In fact, after local stimulation, Ca2+ imaging with high time resolution reveals rapid, transient, local signals even when extracellular Ca2+ is quenched to or below resting intracellular Ca2+ concentration ([Ca2+]e <= [Ca2+]i). Under these conditions, quenched-flow/freeze-fracture analysis shows that membrane fusion is only partially inhibited. Increasing [Ca2+]e alone, i.e., without secretagogue, causes rapid, strong cortical increase of [Ca2+]i but no exocytosis. In various cells, the ratio of maximal vs. minimal currents registered during maximal stimulation or single exocytotic events, respectively, correlate nicely with the number of Ca stores available. Since no quantal current steps could be observed, this is again compatible with the combined occurrence of Ca2+ mobilization from stores (providing close to threshold Ca2+ levels) and Ca2+ influx from the medium (which per se does not cause exocytosis). This implies that only the combination of Ca2+ flushes, primarily from internal and secondarily from external sources, can produce a signal triggering rapid, local exocytotic responses, as requested for Paramecium defense.

Abbreviations used in this paper: AED, aminoethyldextran; Ca2+e, extracellular calcium; [Ca2+]i, intracellular free Ca2+ concentration; CaGr-2, calcium green-2; CLSM, confocal laser scanning microscopy.

This paper is supported by SFB156 as well as DFG grants Pl78/11 and Pl78/12, all from the Deutsche Forschungsgemeinschaft (DFG).

Please address all correspondence to H. Plattner, Faculty of Biology, University of Konstanz, P.O. Box 5560, D-78434 Konstanz, Germany. Tel.: (49) 7531-88-2228. Fax: (49) 7531-88-2245.


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