SUMMARYPotassium transport was measured in equine red blood cells, using 8fRb+ influx as a convenient assay. A significant component of volume-and pH-sensitive K+-Cl-cotransport to the overall K+ flux was observed in all blood samples studied, although fluxes were variable between animals, and within individuals when measured at intervals over a period of weeks. The aryloxyacetic acid [(dihydroindenyl)oxy]alkanoic acid (DIOA), at a final concentration of 100jUM, inhibited most (>95 %) of the Cl--dependent K+ flux, and DIOA sensitivity was therefore used to define the activity of the K+-Cl-cotransporter. K+-Cl-cotransport was also sensitive to protein phosphatase inhibition with calyculin A or okadaic acid, with inhibition constants of 9 + 1 nm for calyculin and about 100 nm for okadaic acid. Peak fluxes were observed at an extemal pH of 6-7-7 0, with inhibition at higher and lower values. Volumesensitive K+ fluxes assayed in autologous plasma, controlled for osmolality, pH and potassium concentration, were significantly lower (28 + 8 % of control values, n = 6) than those measured in saline. This inhibition was mimicked by the culture medium RPMI, but disappeared following dialysis of the plasma. Phosphate (5 6 mM) inhibited volume-sensitive K+ fluxes by 48 + 2 %, n = 3; no significant effect was observed by increasing external magnesium concentrations to 0.5 or 2 mm. Thus, inhibition by RPMI, but not that by plasma, may be due to phosphate. Finally, volume-and pH-sensitive K+ fluxes were sensitive to oxygen tension and were abolished reversibly by equilibrating solutions with nitrogen, as opposed to air. Use of solutions equilibrated with different values of PO, may account for some of the variability in equine red blood cell KCI fluxes. The importance of these observations to equine red blood cell homeostasis and haemodynamics is discussed.

A sudden decrease in external medium osmolality (90 mosmol/kg) causes an immediate swelling of trout erythrocytes, followed by a regulatory volume decrease (RVD) due to activation of both a KCl cotransporter and a taurine transport pathway. Here, we determined how trout red cells respond when they are exposed to a gradual and slow decrease in medium osmolality (80 mosmol/kg at a rate of 0.7 mosmol/kg per min). Erythrocytes were unable to regulate their volume efficiently when swollen gradually and it increased continuously throughout the experimental period (120 min). As long as volume was increased slowly by 15-25%, regulatory pathways remained essentially inactivated, erythrocytes losing no significant amount of intracellular osmotically active solutes. Above this swelling threshold, a response was triggered but the quantity of solutes lost via the regulatory pathways was still not sufficient to counterbalance the continuous entry of water due to the slow and gradual decrease in medium tonicity.

1. Pressure induced a 4-to 5-fold stimulation of the residual (i.e. oubain-bumetanide insensitive) s6Rb+ influx across the human red cell membrane. This enhancement showed a broad pH. dependence with a maximum stimulation around pHo 7.2. At atmospheric pressure, the protein kinase inhibitors staurosporine and chelerythrine stimulated a normally silent component of 86Rb+ influx in a dose-dependent manner with a half-maximum stimulatory concentration at about 550 nm and 140 /M, respectively. The component stimulated by staurosporine was entirely Cl-dependent, but part of the chelerythrine effect was Cl-independent.3. Staurosporine (3/M), chelerythrine (200 #M) and N-ethylmaleimide (1 mM) stimulated further the increased residual 86RbV influx in cells at high pressure.4. The serine/threonine protein phosphatase inhibitors okadaic acid, cantharidin and calyculin A inhibited the stimulatory pressure effect in a dose-dependent manner with halfmaximum inhibitory concentrations of 70 nM, 2-5 /M and 3-3 nm, respectively. In contrast, deltamethrin, a specific protein phosphatase type 2B inhibitor, did not affect the stimulation by pressure, up to a concentration of 10 uM.