PORE, S.A.; SILVERMAN, L.N.; SPIVAK, A.; LIGHT, D.B.**; Lake Forest College; Lake Forest College; Lake Forest College; Lake Forest College: Regulated Volume Decrease in Alligator mississippiensis Erythrocytes

|We examined regulated volume decrease (RVD) in Alligator mississippiensis erythrocytes. Cell volume was measured electronically with a Coulter counter. Exposure of erythrocytes to a hypotonic (0.5X) Ringer caused cells to quickly swell, which was followed by two-phase recovery: a rapid volume decrease (first 20 minutes) and a slower volume decrease. Addition of the pore-former gramicidin (5 &microM, choline Ringer) potentiated volume recovery, indicating K⁺ efflux is rate-limiting. In contrast, the K⁺ channel antagonist quinine (10 mM) partially inhibited RVD (especially phase one), whereas 10 nM Ca²⁺ Ringer (buffered with EGTA) or Ca²⁺ ionophore A23187 (0.5 &microM) completely blocked RVD, even when Na⁺ was replaced with choline (N.B., substitution of choline for Na⁺ enhanced volume recovery, especially for phase two, indicating Na⁺ influx attenuates RVD). Further, quinine and A23187 had an additive effect during phase two, but not phase one. Finally, the Cl- transport inhibitor DIDS (0.1 mM) increased osmotic fragility and inhibited RVD (phase one), whereas the K⁺-Cl^– cotransport blockers bumetanide (0.1 mM) and furosemide (0.1 mM), and acetazolamide (10 mM) had no affect. Conclusions: regulated volume decrease in this cell type depends, at least in part, on K⁺ efflux, most likely through a Ca²⁺-activated channel, especially during phase one. Our results also indicate that although extracellular Ca²⁺ is necessary for RVD, high levels of intracellular Ca²⁺ block volume recovery, probably by inhibiting K⁺ efflux. (Supported by funds from Lake Forest College).