Evidence of Coordinated and Adjustable Osmolytes Movements Following Hyposmotic Swelling in Rainbow Trout Red Blood Cells

Abstract

BACKGROUND/AIMS: The osmolytes involved in the volume regulation of hyposmotically-swollen fish cells are well identified. However, if a coordination and adjustments of their fluxes are obvious, few studies have clearly illustrated these aspects. METHODS: Trout red blood cells volume variations were estimated from water contents obtained by a gravimetric method. Intracellular K+ and Na+ contents, and Cl- content of haemolysed cells were determined by photometry and colorimetry, respectively. The taurine contribution to cell volume regulation was calculated from the net changes of water, K+, Cl- and Na+ contents. The intracellular pH was calculated from the chloride distribution across the cells membranes according to the Donnan equilibrium. RESULTS: Cells responses to a rapid change (from 296 to 176 mOsm.kg-1) of the saline osmolality were examined in three conditions designed to not impact (Hypo. I) or to reduce the K+ (Hypo. II) and Cl- (Hypo. III) contributions to the volume regulation. Hypo. I condition caused an immediate increase in water content, followed by a 90 min. full regulation, concomitant with gradual lowering of K+ and Cl- contents and a surprising increase in Na+ content. Hypo. II and III conditions showed a partial and complete volume regulation, respectively. This was made possible by an increase in the taurine involvement. These experiments allowed to confirm that K+ and Cl- were released via KCl cotransport and by separate channels. The comparison of Hypo. I and III conditions led to the observation that the partially amiloride-sensitive Na+ influx is proportional to the taurine efflux; the latter being sustained mainly by a Na+/taurine cotransport. The Hypo. II condition was suitable for the (Na+/K+)ATPase activity inhibition. This effect could explain the observed lack of Na+ uptake, the consecutive depletion of intracellular taurine stock and the incomplete volume regulation. Finally, the results support the importance of taurine in pH control under Hypo. I (physiologic) condition. The alkalosis observed in Hypo. II and III conditions were the consequences of changes in the salines compositions, not of physiologic adjustments. CONCLUSION: The regulatory volume decrease process of trout RBCs is complex and adjustable through coordinated osmolytes movements. The obliged decrease in K+ and/or Cl- contributions stimulates taurine and Na+ pathways. This study highlights the importance of taurine as a compensatory variable in cell volume regulation and explains for the first time the significance of the Na+ uptake during this process