Perimetric [Ca2+](i) rise and exocytosis detected by ultraviolet laser scanning confocal microscopy in rat peritoneal mast cells

It has been reported that in dialysed mast cells an increase in mean [Ca2+](i) is neither necessary nor sufficient for secretion; however, it is possible that juxtamembranal [Ca2+](i) may exceed the mean [Ca2+](i) before exocytosis. The present study was carried out to analyse spatial and temporal dynamics of [Ca2+](i) and concomitant exocytosis in intact rat peritoneal mast cells using UV laser scanning confocal microscopy. Stimulation with Compound 48/80 (16 mu M) increased mean [Ca2+](i), causing an initial rapid elevation from 40 to 200 nM, which lasted for 6 s and was followed by a delayed secondary increase to 600 nM. Exocytotic images were seen in the cell perimeter 16 s after the stimulation. Perimetric and nuclear [Ca2+](i) increased to several micromoles per litre, while that in the intermediate region remained low. In a Ca2+-deficient environment, Compound 48/80 still increased perimetric [Ca2+](i) to micromolar values and induced exocytosis. This study clearly indicates, for the first time, that a perimetric increase in [Ca2+](i) to micromolar levels precedes exocytosis, in intact, as opposed to permeabilized cells.