CONTRIBUTION OF ACTIN CYTOSKELETAL ALTERATIONS TO ATP DEPLETION AND CALCIUM-INDUCED PROXIMAL TUBULE CELL INJURY

The actin cytoskeleton of rabbit proximal tubules was assessed by deox yribonuclease (DNase) binding, sedimentability of detergent-insoluble actin, laser-scanning confocal microscopy, and ultrastructure during e xposure to hypoxia, antimycin, or antimycin plus ionomycin. One-third of total actin was DNase reactive in control cells prior to deliberate depolymerization, and a similar proportion was unsedimentable from de tergent lysates during 2.5 h at 100,000 g. Tubules injured by hypoxia or antimycin alone, without glycine, showed Ca2+-dependent pathology o f the cytoskeleton, consisting of increases in DNase-reactive actin, r edistribution of pelletable actin, and loss of microvilli concurrent w ith lethal membrane damage. In contrast, tubules similarly depleted of ATP and incubated with glycine showed no significant changes of DNase -reactive actin or actin sedimentability for up to 60 min, but, nevert heless, developed substantial loss of basal membrane-associated actin within 15 min and disruption of actin cores and clubbing of microvilli at durations >30 min. These structural changes that occurred in the p resence of glycine were not prevented by limiting Ca2+ availability or pH 6.9. Very rapid and extensive cytoskeletal disruption followed ant imycin-plus-ionomycin treatment. In this setting, glycine and pH 6.9 d ecreased lethal membrane damage but did not ameliorate pathology in th e cytoskeleton or microvilli; limiting Ca2+ availability partially pro tected the cytoskeleton but did not prevent lethal membrane damage. Th e data suggest that both ATP depletion-dependent but Ca2+-independent, as well as Ca2+-mediated, processes can disrupt the actin cytoskeleto n during acute proximal tubule cell injury; that both types of change occur, despite protection afforded by glycine and reduced pH against l ethal membrane damage; and that Ca2+-independent processes primarily a ccount for prelethal actin cytoskeletal alterations during simple ATP depletion of proximal tubule cells.