Journal
JOURNAL OF PHYSIOLOGY-LONDON
Volume 527, Issue 1, Pages 95-107Publisher
WILEY
DOI: 10.1111/j.1469-7793.2000.00095.x
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1. The role of the cytoskeleton in leptin-induced activation of ATP-sensitive K+ (K-ATP) channels was examined in rat CRI-G1 insulin-secreting cells using patch clamp and fluorescence imaging techniques. 2. In whole cell recordings, dialysis with the actin filament stabiliser phalloidin (10 mu M) prevented K-ATP channel activation by leptin. 3. Application of the actin filament destabilising agents deoxyribonuclease type 1 (DNase 1; 50 mu g ml(-1)) or cytochalasin B (10 mu M) to intact cells or inside-out membrane patches also increased K-ATP channel activity in a phalloidin-dependent manner. 4. The anti-microtubule agents nocodazole (10 mu M) and colchicine (100 mu M) had no effect on K-ATP channel activity. 5. Fluorescence staining of the cells with rhodamine-conjugated phalloidin revealed rapid disassembly of actin filaments by cytochalasin B and leptin, the latter action being prevented by the phosphoinositide 3 (PI 3)-kinase inhibitor LY 294002. 6. Activation of K-ATP channels by the PI 3-kinase product phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P-3) was also prevented by phalloidin. This is consistent with the notion that leptin activates K-ATP channels in these cells by an increase in PtdIns(3,4,5)P-3 or a similar 3-phosphorylated phosphoinositol lipid, resulting in actin filament disruption.
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