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doi: 10.1242/10.1242/jcs.00202

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Cell swelling increases intracellular calcium in Necturus erythrocytes

Department of Biology, Ripon College, Ripon, WI 54971, USA

^* Author for correspondence (e-mail: lightd{at}ripon.edu)

Accepted 27 September 2002

This study examined the role of Ca²⁺ in regulatory volume decrease by Necturus erythrocytes. Hypotonic shock (50% tonicity) stimulated an increase in cytosolic free Ca²⁺, detected using epi-fluorescence microscopy and the fluorescent Ca²⁺ indicator fluo-4-AM (10 µM). A similar increase in fluorescence did not occur under isosmotic conditions, unless cells were exposed to the Ca²⁺ ionophore A23187 (0.5 µM). In addition, a low Ca²⁺ medium (amphibian Ringer solution with 5 mM EGTA), hexokinase (2.5 U/ml, an ATP scavenger), suramin (100 µM, a P2 receptor antagonist) and gadolinium (10 µM, a stretch-activated channel blocker) each inhibited the swelling-induced increase in Ca²⁺. Consistent with these studies, a low Ca²⁺ Ringer solution increased osmotic fragility, whereas volume recovery following hypotonic shock (measured with a Coulter counter) was potentiated with A23187 (0.5 µM). By contrast, a low Ca²⁺ extracellular medium or buffering intracellular Ca²⁺ with BAPTA-AM (100 µM) reduced the rate of volume recovery following hypotonic challenge. Finally, a low Ca²⁺ extracellular Ringer solution inhibited whole-cell currents that are activated during cell swelling (measured with the whole-cell patch clamp technique). Our results are most consistent with hypotonic shock causing an increase in cytosolic free Ca²⁺, thereby stimulating subsequent volume decrease.

Key words: Fluo-4, EGTA, BAPTA, Cell volume regulation, Hexokinase, P2 receptor

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