EXTRACELLULAR ACIDOSIS MINIMIZES ACTIN CYTOSKELETAL ALTERATIONS DURING ATP DEPLETION

Extracellular acidosis has been shown to be protective during ischemia in renal tubule cells. However, the mechanism of protection remains u nknown. Since ischemia leads to disruption and polymerization of the c ortical actin cytoskeleton, we hypothesized acidosis may better preser ve the actin cytoskeleton during ischemia. Therefore, the purpose of o ur studies was to examine the effect of pH on the integrity of the act in cytoskeleton during ATP depletion and ATP repletion. To do this, we used an in vitro model of reversible ATP depletion in LLC-PK1 cells a t extracellular pH values (pH(o)) of 6.9, 7.4, and 7.9. Immunofluoresc ent studies with rhodamine-phalloidin demonstrated more marked redistr ibution and clumping of cortical actin at pH, 7.9 and 7.4 vs. 6.9 afte r 90 min of chemical anoxia. After 15 min of ATP depletion, G-actin, q uantified by the deoxyribonuclease assay, decreased from 53.7 +/- 0.8 to 43.2 +/- 1.5 mu g/mg protein at pH(o) 6.9 vs. 37.6 +/- 1.8 mu g/mg protein at pH(o) 7.4 (P < 0.001). After 60 min, there still was signif icantly less conversion of G-actin to F-actin at pH(o) 6.9 vs. 7.4, wi th a decrease from 55.9 +/- 2.0 to 39.6 +/- 2.0 mu g/mg protein at 6.9 vs. 35.8 +/- 2.4 at 7.4 mu g/mg protein (P < 0.05). Furthermore, extr acellular acidosis during the phase of ATP repletion resulted in more rapid normalization of cellular G-actin levels (95 +/- 3% of control v s. 82 +/- 2% for pH 6.9 vs. 7.4, respectively, P < 0.01). Together, th ese findings indicate the actin cytoskeleton is better preserved in an acidic environment during ATP depletion. We postulate acidosis mainta ins cell integrity in part by stabilizing the actin cytoskeleton.