study of distal tubular potassium and sodium transport in rat nephron. Am. J. Physiol.

Paracellular ion flux across epithelia occurs through selective and regulated pores in tight junctions; this process is poorly understood. Mutations in the kinase WNK4 cause pseudohypoaldosteronism type II (PHAII), a disease featuring hypertension and hyperkalemia. Whereas WNK4 is known to regulate several transcellular transporters and channels involved in NaCl and K ؉ homeostasis, its localization to tight junctions suggests it might also regulate paracellular flux. We performed electrophysiology on mammalian kidney epithelia with inducible expression of various WNK4 constructs. Induction of wild-type WNK4 reduced transepithelial resistance by increasing absolute chloride permeability. PHAII-mutant WNK4 produced markedly larger effects, whereas kinase-mutant WNK4 had no effect. The electrochemical and pharmacologic properties of these effects indicate they are attributable to the paracellular pathway. The effects of WNK4 persist when induction is delayed until after tight-junction formation, demonstrating a dynamic effect. WNK4 did not alter the flux of uncharged solutes, or the expression or localization of selected tight-junction proteins. Transmission and freeze-fracture electron microscopy showed no effect of WNK4 on tight-junction structure. These findings implicate WNK signaling in the coordination of transcellular and paracellular flux to achieve NaCl and K ؉ homeostasis, explain PHAII pathophysiology, and suggest that modifiers of WNK signaling may be potent antihypertensive agents.protein serine-threonine kinases ͉ ion transport

Oxygen consumption (QO2) and net K+ transport were studied in rabbit proximal tubule suspensions to define the early effects of cisplatin on proximal tubule function. Cisplatin caused dose-dependent inhibition of QO2, which was delayed in onset. The concentration of cisplatin required for inhibition decreased as the duration of exposure was increased [40-min exposure, threshold concentration of 10(-4) M, inhibitor constant (Ki) of 10(-3) M; 4-h exposure, threshold concentration of 3 X 10(-5) M, Ki of 10(-4) M]. Both ouabain-sensitive and ouabain-insensitive QO2 were reduced, indicating inhibition of all adenosinetriphosphatases, including Na(+)- K(+)-ATPase activity. There was a parallel fall in ouabain-sensitive net K+ transport and cytosolic K+ content, confirming the latter observation. Na(+)-K(+)-ATPase activity was unchanged in cell membranes prepared by hypotonic lysis from cisplatin-treated tubules, indicating an indirect cytosol-dependent mechanism of enzyme inhibition. Nystatin-stimulated QO2 was reduced in cisplatin-treated tubules, excluding inhibition of Na+ entry as the mechanism of injury and suggesting mitochondrial injury. The latter was confirmed by measurement of carbonylcyanide-m-chlorophenylhydrazone (CCCP)-uncoupled QO2 in intact cells and ADP-stimulated (state 3) QO2 in digitonin-permeabilized tubules. Furthermore, by maximally stimulating mitochondrial respiration with CCCP and nystatin, it was possible to demonstrate mitochondrial injury at a time when basal QO2 and K+ transport were apparently normal. These data suggest that mitochondrial injury is a central event in cisplatin toxicity to the proximal tubule.

Micropuncture studies were made in the rat on the relationship between net water and net sodium movement across proximal tubular epithelium during strong osmotic diuresis induced by intravenous infusion of a 20% mannitol solution. Results obtained in the mannitol series were compared with data obtained in rats during infusion of 0.9% saline. Collected tubular fluid and appropriate plasma samples were analyzed for C14 inulin or creatinine, chloride and radiosodium. The latter was injected intravenously several hours prior to tubular puncture. Net water reabsorption across the proximal tubule was less than under nondiuretic conditions. The concentration of radiosodium and of chloride in proximal tubular fluid was found to be below that of plasma in osmotic diuresis. Much steeper gradients for these ions were found across distal tubular epithelium indicating that parts other than the proximal convolution participate in the establishment of the over-all concentration gradients. Since net sodium movement occurred against an electrochemical potential gradient during osmotic diuresis, this study provides evidence for active transport of sodium ions across the proximal tubular wall.

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