Extracellular acidosis is cytoprotective in several models against anoxia/hypoxia and a variety of toxicants. The goal of this study was to determine the temporal relationships among toxicant exposure, the initiation of extracellular acidosis, C1 influx, Cl channel inhibition and the onset of cellular death in rabbit renal proximal tubule suspensions. Extracellular acidosis was produced by adding HCI or H2S04 to renal proximal tubule suspensions to decrease the extracellular buffer pH to 6.4 or by resuspending renal proximal tubules in a pH 6.4 buffer. The initiation of extracellular acidosis 15 mm after the mitochondnal inhibitors antimycin A or carbonyl cyanide p-(trifluoromethoxy)-phenylhydrazone addition, a time point in which adenosine triphosphate levels are depleted and intracellular K is de-creased, ameliorated lactate dehydrogenase release, a marker of necrotic cellular death. The initiation of extracellular acidosis 120 mm after the addition of the toxicants tetrafluoroethyl-L-cysteine or t-butyl hydroperoxide decreased lactate dehydro-genase release 120 mm later. Increased C1 influx is an impor-tant step during the late phase of toxicant-induced cellular injury. Therefore, we determined if extracellular acidosis cytoprotection was associated with inhibition of C1 influx and whether the C1 channel inhibitors indanyloxyacetic acid (1.0 mM), niflumic acid (1 00. tM) and 5-nitro-2-(3-phenylpropyiamino)-benzoic acid (100 pM) decreased CI influx and cellular death in renal proximal tubules exposed to antimycin A.

Indeed, all three Cl channel inhibitors significantly decreased 36Cl influx and cellular death. In contrast, extracellular acidosis did not decrease 36CI influx but did prevent lactate dehy-drogenase release. These results demonstrate that extracellular acidosis cytoprotection occurs dunng the late phase of cellular injury at a site distal to C1 influx. Furthermore, the Cl influx