MAST-CELL ACTIVATION INVOLVES PLASMA-MEMBRANE POTENTIAL-SENSITIVE ANDTHAPSIGARGIN-SENSITIVE INTRACELLULAR CALCIUM POOLS

The regulation and role of the intracellular Ca2+ pools were studied i n rat peritoneal mast cells. Cytosolic free calcium concentration ([Ca 2+](i)) was monitored in fura-2 loaded mast cells. In the presence of Ca2+ and K+, compound 48/80 induced a biphasic increase in [Ca2+](i) c omposed of a fast transient phase and an apparent sustained phase. The sustained phase was partially inhibited by the addition of Mn2+. DTPA , a cell-impermeant chelator of Mn2+, reversed this inhibition, sugges ting that a quenching of fura-2 fluorescence occurs in the extracellul ar medium. In the absence of extracellular Ca2+, the transient phase, but not the sustained one, could be preserved, provided that mast cell s were depolarized. The transient phase was completely abolished by th apsigargin, a microsomal Ca2+-ATPase inhibitor. Maximum histamine rele ase induced by either compound 48/80 or antigen was obtained in the ab sence of added Ca2+ only when mast cells were depolarized. These hista mine releases were inhibited by low doses (< 30 nM) of thapsigargin. T hapsigargin at higher doses induced histamine release which was unaffe cted by changing the plasma membrane potential, but was completely dep endent on extracellular Ca2+, showing that a Ca2+ influx is required f or thapsigargin-induced exocytosis. Together, these results suggest th at the mobilization of Ca2+ from thapsigargin sensitive-intracellular pools induced by compound 48/80 or antigen is sufficient to trigger hi stamine release. The modulation of these pools by the plasma membrane potential suggest their localization is close to the plasma membrane.