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We have studied the regulation of the K-Cl cotransporter KCC1 and its functional interaction with the Na-K-Cl cotransporter. K-Cl cotransporter activity was substantially activated in HEK-293 cells overexpressing KCC1 (KCC1-HEK) by hypotonic cell swelling, 50 mM external K, and pretreatment with N-ethylmaleimide (NEM). Bumetanide inhibited 86Rb efflux in KCC1-HEK cells after cell swelling [inhibition constant (Ki) approximately 190 microM] and pretreatment with NEM (Ki approximately 60 microM). Thus regulation of KCC1 is consistent with properties of the red cell K-Cl cotransporter. To investigate functional interactions between K-Cl and Na-K-Cl cotransporters, we studied the relationship between Na-K-Cl cotransporter activation and intracellular Cl concentration ([Cl]i). Without stimulation, KCC1-HEK cells had greater Na-K-Cl cotransporter activity than controls. Endogenous Na-K-Cl cotransporter of KCC1-HEK cells was activated <2-fold by low-Cl hypotonic prestimulation, compared with 10-fold activation in HEK-293 cells and >20-fold activation in cells overexpressing the Na-K-Cl cotransporter (NKCC1-HEK). KCC1-HEK cells had lower resting [Cl]i than HEK-293 cells; cell volume was not different among cell lines. We found a steep relationship between [Cl]i and Na-K-Cl cotransport activity within the physiological range, supporting a primary role for [Cl]i in activation of Na-K-Cl cotransport and in apical-basolateral cross talk in ion-transporting epithelia.

To gain better insight into the interaction between thermoregulation and osmoregulation, we examined the thermal sweating response to drinking in cell-dehydrated humans. Cell dehydration (CDH) was induced by infusion of a 3% NaCl solution, at 1.2 ml/kg, for 2 h; infusion of a 0.9% NaCl solution in a separate experiment served as a control (euhydrated condition, EH). After infusion, subjects were heated by immersion of the lower legs in 42 degrees C water at an ambient temperature of 28 degrees C for 90 min. Subjects drank 4.3 ml/kg of H2O (approximately 38 degrees C) at 60 min of heating. The 3% NaCl infusion increased plasma osmolality by 13.6 +/- 0.8 mosmol/kgH2O and plasma arginine vasopressin concentration ([AVP]) by 3.3 +/- 0.7 pg/ml. Neither variable was altered with 0.9% NaCl infusion. Before drinking, esophageal temperature (Tes) had increased by 0.91 +/- 0.08 degrees C in CDH and by 0.40 +/- 0.11 degrees C in EH. Local chest sweating rate (SRch) had increased by 0.67 +/- 0.08 and 0.63 +/- 0.07 mg.min-1.cm-2 in CDH and EH, respectively. Thus the change in SRch per unit rise in Tes was much lower in CDH than in EH. Drinking immediately increased SRch and reduced Tes in CDH, with a reduction in plasma [AVP] and thirst rating. Drinking did not change thermoregulatory and osmoregulatory responses in EH. These results suggest that the act of drinking itself eliminates, at least partially, an osmotic inhibitory input to the thermoregulatory center, as well as osmotic AVP secretion and thirst.

After a 7-h H2O and Na+ depletion period (DP), produced by intermittent light exercise (8 bouts) at 35 degrees C, we examined thirst and taste palatability responses to 10 different NaCl solutions during 23 h of rehydration (RH) at 25 degrees C. During DP, net H2O and Na+ loss were 27.2 +/- 2.9 ml/kg and 3.29 +/- 0.45 meq/kg, respectively. Plasma osmolality (POsm) and plasma Na+ concentration ([Na+]p) increased significantly during DP by 3.4 +/- 1.2 mosmol/kgH2O and 3.0 +/- 1.0 meq/kgH2O, respectively. Plasma volume (PV) decreased by 6.5 +/- 1.9%. Thirst rating, renal fractional reabsorption of H2O, and plasma arginine vasopressin concentration (PAVP) increased as POsm increased. This increased thirst was accompanied by increased palatability ratings to H2O. During RH, subjects drank deionized H2O ad libitum and ate a Na(+)-free diet for 23 h. POsm and [Na+]p returned to control levels within 1 h RH and remained at or below the control thereafter. PV remained reduced by approximately 5% throughout RH. The increased thirst and PAVP returned to their respective control levels within 1 h of RH as POsm decreased, but thirst rating increased against between 17 and 23 h of RH without increase in POsm or PAVP. Palatability ratings to a 1 M NaCl solution at and after 3 h RH and palatability ratings to 0.3 M at 17 and 23 h RH were significantly higher than control. Plasma aldosterone concentration (PAldo) increased after DP, decreased with drinking, and increased again between 6 and 23 h of RH, accompanied by a marked decrease in fractional Na+ excretion to < 0.07%. Thus both Na+ preference and thirst in humans are influenced by body fluid and electrolyte status. The increased Na+ palatability (Na+ appetite) was preceded by osmotically induced thirst, and accompanied by nonosmotically driven thirst [extracellular fluid (ECF) thirst] and increased PAldo. The "Na+ appetite" and "ECF thirst" along with increased renal Na+ retention could contribute to ECF volume regulation after thermally induced H2O and Na+ depletion.