P/Q Ca2+ channels are functionally coupled to exocytosis of the immediately releasable pool in mouse chromaffin cells

YANINA D. ´ ALVAREZ, LORENA I. IBA˜NEZ, OSVALDO D. UCHITEL, FERNANDO D. MARENGO

CELL CALCIUM.

Año: 2007 p. 1 - 1

0143-4160

Chromaffin cell exocytosis is triggered by Ca2+ entry through several voltage-dependent channel subtypes. Because it was postulated thatimmediately releasable vesicles are closely associated with Ca2+ channels, we wondered what channel types are specifically coupled to therelease of this pool. To study this question, cultured mouse chromaffin cell exocytosis was followed by patch-clamp membrane capacitancemeasurements. The immediately releasable pool was estimated using paired pulse stimulation, resulting in an upper limit of 31±3 fF forcontrol conditions (ICa: 25±2 pA/pF). The N-type channel blocker -conotoxin-GVIA affected neither ICa nor the immediately releasablepool exocytosis; although the L channel blocker nitrendipine decreased current by 50%, it did not reduce this pool significantly; and the Rchannel inhibitor SNX-482 significantly reduced the current but induced only a moderate decrease in the estimated IRP exocytosis. In contrast,the P/Q channel blocker -Agatoxin-IVA decreased ICa by 37% but strongly reduced the immediately releasable pool (upper limit: 6±1 fF).We used 1A subunit knockout mice to corroborate that P/Q Ca2+ channels were specifically linked to immediately releasable vesicles, andwe found that also in this preparation the exocytosis of this pool was severely decreased (6±1 fF). On the other hand, application of a strongstimulus that caused the fusion of most of releasable vesicles (3 min, 50mMK+) induced similar exocytosis for wild type and knockoutcells. Finally, whereas application of train stimulation on chromaffin cells derived from wild type mice provoked typical early synchronousand delayed asynchronous exocytosis components, the knockout derived cells presented a strongly depressed early exocytosis but showed aprominent delayed asynchronous component. These results demonstrate that P/Q are the dominant calcium channels associated to the releaseof immediately releasable pool in mouse chromaffin cells.