Effect of low temperatures on compound 48/80-induced intracellular Ca2+ changes and exocytosis of rat peritoneal mast cells.

Abstract

It has been well documented that compound 48/80-induced exocytosis of mast cells is accompanied by changes in intracellular Ca2+ concentration ([Ca2+]i) showing a biphasic pattern: an initial phase which constitutes an abrupt increase, followed by a plateau phase. The former is caused by Ca2+ release from intracellular Ca2+ stores, and the latter is the result of secondary Ca2+ influx. Low temperatures lead to the inhibition of exocytosis, but the precise mechanism remains unclear. The present study aims to reveal whether [Ca2+]i changes are affected by the environmental temperature. To this end, we developed a novel imaging method to record [Ca2+]i changes and exocytotic processes simultaneously. Rat peritoneal mast cells were loaded by Indo-1/AM or Fluo-3/AM for measuring [Ca2+]i, and the exocytosed granule matrices were stained by sulforhodamine-B. Cells were stimulated by compound 48/80, and [Ca2+]i changes and exocytosis were recorded by means of a real-time confocal microscope. At 37 degrees C, [Ca2+]i changes in stimulated mast cells showed a sustained plateau phase. Granule discharge was observed at the cell surface, and, in addition, most of the intracellular granule matrices were involved in compound exocytosis. The granule discharge and compound exocytosis proceeded over a period of a few minutes. At 4 degrees C, the plateau phase of [Ca2+]i changes declined rapidly, although the initial phase was not suppressed. Granule discharge occurred at the cell surface, but compound exocytosis ceased within a few minutes. These findings indicate that a low temperature inhibits compound exocytosis which can be caused by Ca2+ influx. The present imaging method represents a powerful tool for investigating the stimulus-secretion coupling of mast cells.