Calcium and regulated volume decrease in Atlantic salmon and American alligator erythrocytes

Abstract

A cell’s ability to regulate its volume is a property that has been conserved in a diversity of cell types; however, the mechanisms by which this compensation operates are not clearly understood. The goal of this study was to elucidate the role of Ca2+ in regulated volume recovery (RVD) in Atlantic salmon and American alligator erythrocytes. Mean cell volume was determined by electronic sizing using a Coulter counter, and changes in intracellular Ca2+ concentration were visualized using fluorescence microscopy and fluo-4. Hypotonic shock led to an increase in cytosolic Ca2+ in salmon. This increase resulted from Ca2+ influx because it was not observed in cells bathed in a hypotonic, low Ca2+ medium (10 nM, buffered with EGTA). Furthermore, cells of both species exposed to a low Ca2+ environment failed to recover from cell swelling, indicating that extracellular Ca2+ was needed for RVD. Interestingly, the Ca2+ ionophore A23187 inhibited RVD in Na-Ringer and when Na+ was replaced with NMDG+, indicating this inhibition was not due to Na+ influx. Further, although the Ca2+ channel antagonists gadolinium and lanthanum inhibited RVD in alligator, they enhanced volume recovery in salmon. Finally, pilot studies with hexokinase and ONO-RS-082 suggest that RVD in alligator depends on the activation of P2 receptors and the formation of arachidonic acid, respectively. In conclusion, our results indicate extracellular Ca2+ is necessary for RVD in both species. Future studies will examine how Ca2+ elicits volume recovery. (Supported by Lake Forest College.)