Loss of plasma membrane structural support in ATP-depleted renal epithelia

Renal ischemia induces cytoskeletal alterations, membrane perturbations, including bleb formation, and ultimately membrane lysis. The mechanisms that underlie these alterations are largely unknown. Through the use of isolated rat renal proximal tubule fragments and calibrated micropipette techniques, two potential mechanisms for membrane bleb formation during ATP depletion were examined: 1) decreased cytoskeletal retention of the plasma membrane and 2) increased intracellular pressure. Under control conditions, the pressure required to pull the membrane from the underlying cellular matrix was 73 +/- 10 kdyn/cm2. After 30 min of ATP depletion, this pressure was diminished by >95% and blebs began to emerge from the basal membrane. The intracellular pressure within these blebbed cells was only 0.08 +/- 0.02 kdyn/cm2. These observations indicate that, during ATP depletion, the strength of membrane retention diminished until the relatively low intracellular pressure was capable of driving membrane bleb formation. Cytochalasin D, which disrupts the actin cytoskeleton, decreased the strength of membrane retention by 65 +/- 7%. This suggests that, during ATP depletion, alterations of the actin cytoskeleton may mediate the loss of membrane retention.