Crassostrea virginica hemocytes, separated from serum and suspended in a maintenance medium, were exposed to the ionophore A23187 + EDTA, A23187 + EGTA, and Escherichia coli + A23187 + EDTA. As controls, identically treated hemocytes were exposed to A23187, EDTA, EGTA, and nonenriched maintenance medium. The activity levels of acid phosphatase in the media associated with the experimental and control sets of hemocytes were assayed at five time intervals postexposure to the various additives. It was determined that A23187 + EDTA stimulated significantly higher levels of enzyme release, with an initial burst at 1 min or earlier, than A23187, EDTA, or nonenriched maintenance medium alone. It was also determined that A23187 + EGTA stimulated significantly less release of acid phosphatase than A23187 + EDTA, but more than A23187, EGTA, or nonenriched maintenance medium individually. In all of the combinations tested, exposure to A23187 + EDTA + E. coli stimulated the greatest release of acid phosphatase from oyster hemocytes. Atomic absorption analyses of the concentrations of Ca2+ and Zn2+ in the maintenance medium and cytosol of all categories of hemocytes revealed that there were rapid and high levels of efflux of Zn2+ from all three sets of experimental hemocytes, especially those exposed to A23187 + EDTA and A23187 + EDTA + E. coli, but only low levels of exocytosis of Ca2+, which were no different between experimental and control groups. This appears to be the first demonstration of the requirement of a chelator (EDTA) in an ionophore-stimulated release of a bioactive molecule from invertebrate cells and of a correlation between the efflux of Zn2+ with ionophore-stimulated secretion of an enzyme. © 1992.
