CALCIUM LOADING OF SECRETORY GRANULES IN STIMULATED NEUROHYPOPHYSEAL NERVE-ENDINGS

The total calcium content of secretory granules, CA(g), was evaluated in isolated neurohypophysial nerve endings. The Ca-g in the resting st ate, as measured by X-ray microanalysis, is relatively high with an av erage of 7.4+/-0.6 mmol/kg wet weight. Following a depolarizing potass ium challenge, a subpopulation of granules with even higher Ca-g could be detected, dispersed over a wider range of concentrations (up to 70 mmol/kg wet weight). After subsequent rinsing in physiological saline , CA(g) decreased to control values. This could have resulted from Ca2 + extrusion, or from preferential secretion of calcium-enriched granul es. Our data can be interpreted in favor of the second explanation sin ce no decrease in Ca-g was observed when secretion was blocked by a hy perosmotic saline. The effect of hyperosmotic conditions on isolated n erved endings was further studied by monitoring free cytoplasmic Ca2with the calcium-sensitive dye Fura-2 and by conventional electron mic roscopy. It was demonstrated that hyperosmotic treatment alone did not increase basal cytosolic Ca2+ concentrations but did significantly re duce the potassium-induced cytosolic rise in Ca2+. Electron microscopy of nerve endings in hyperosmotic conditions showed numerous exocytoti c figures at various stages. The observed changes in Ca-g are in accor d with a published hypothesis which proposes that intragranular calciu m is a significant variable in regulated secretion.