Coincident microvillar actin bundle disruption and perinuclear actin sequestration in anoxic proximal tubule

The present studies investigated acute disruption of microvillar actin cyto skeleton and actin association with other cytoskeletal components in ATP-de pleted rabbit proximal tubular cells. Video-enhanced differential-interfere nce contrast microscopy and confocal microscopy were used to follow the fat e of F-actin during the disruption of microvilli. Within individual cells, all microvilli collapsed simultaneously. Microvillar actin filaments underw ent a parallel decrease in length. Using a sequential cytoskeletal extracti on protocol and electron microscopy, we revealed in the present studies the coincident sequestration of a distinct, perinuclear pool of actin that was primarily absent in control cells. Actin sequestration progressed in a dur ation-dependent manner, occurring as early as 15 min of anoxia when cellula r ATP dropped to <5% of control level. Phalloidin staining and depolymeriza tion treatment showed the majority (>90%) of this sequestered actin to be F -actin. A microvillar actin bundling protein villin was also sequestered in the same perinuclear complex of anoxic proximal tubules. In conclusion, th e present results demonstrate a coincident microvillar actin bundle disrupt ion and the perinuclear sequestration of F-actin in ATP-depleted proximal t ubular cells.