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© The Rockefeller University Press, 0021-9525/2000//839 $5.00
The Journal of Cell Biology, Volume 150, Number 4, , 2000 839-848

Differential Regulation of Granule-to-Granule and Granule-to-Plasma Membrane Fusion during Secretion from Rat Pituitary Lactotrophs

^a Department of Physiology and Biophysics, University of Colorado Medical School, Denver, Colorado 80220
^b Department of Biological Sciences, University of Denver, Denver, Colorado 80208
Department of Physiology and Biophysics, University of Colorado Medical School, 4200 East Ninth Avenue, Box C240, Denver, CO 80220.(303) 315-8110(303) 315-8946

bill.betz{at}uchsc.edu

We used fluorescence imaging of individual exocytic events together with electron microscopy to study the regulation of dense core granule-to-plasma membrane fusion and granule-to-granule fusion events that occur during secretion from rat pituitary lactotrophs. Stimulating secretion with elevated extracellular potassium, with the calcium ionophore ionomycin, or with thyrotropin releasing hormone or vasoactive intestinal polypeptide resulted in abundant exocytic structures. Approximately 67% of these structures consisted of multiple granules fused together sharing a single exocytic opening with the plasma membrane, i.e., compound exocytosis. For all of these stimulation conditions there appeared to be a finite number of plasma membrane fusion sites, 11 sites around each cellular equator. However, a granule could fuse directly with another granule that had already fused with the plasma membrane even before all plasma membrane sites were occupied. Granule-to-plasma membrane and granule-to-granule fusion events were subject to different regulations. Forskolin, which can elevate cAMP, increased the number of granule-to-granule fusion events without altering the number of granule-to-plasma membrane fusion events. In contrast, the phorbol ester PMA, which activates protein kinase C increased both granule-to-granule and granule-to-plasma membrane fusion events. These results provide a cellular mechanism that can account for the previously demonstrated potentiation of secretion from lactotrophs by cAMP- and PKC-dependent pathways.

Key Words: FM1-43 • fluorescence • homotypic fusion • compound exocytosis • phorbol ester

© 2000 The Rockefeller University Press

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