Nakamura M, Sunagawa M, Kosugi T, Sperelakis N
Department of Molecular and Cellular Physiology, College of Medicine, University of Cincinnati, Ohio 45267-0576, USA.
Am J Physiol Cell Physiol. 2000 Aug;279(2):C480-7. doi: 10.1152/ajpcell.2000.279.2.C480.
To clarify interactions between the cytoskeleton and activity of L-type Ca(2+) (Ca(L)) channels in vascular smooth muscle (VSM) cells, we investigated the effect of disruption of actin filaments and microtubules on the L-type Ca(2+) current [I(Ba(L))] of cultured VSM cells (A7r5 cell line) using whole cell voltage clamp. The cells were exposed to each disrupter for 1 h and then examined electrophysiologically and morphologically. Results of immunostaining using anti-alpha-actin and anti-alpha-tubulin antibodies showed that colchicine disrupted both actin filaments and microtubules, cytochalasin D disrupted only actin filaments, and nocodazole disrupted only microtubules. I(Ba(L)) was greatly reduced in cells that were exposed to colchicine or cytochalasin D but not to nocodazole. Colchicine even inhibited I(Ba(L)) by about 40% when the actin filaments were stabilized by phalloidin or when the cells were treated with phalloidin plus taxol to stabilize both cytoskeletal components. These results suggest that colchicine must also cause some inhibition of I(Ba(L)) due to another unknown mechanism, e.g., a direct block of Ca(L) channels. In summary, actin filament disruption of VSM cells inhibits Ca(L) channel activity, whereas disrupting the microtubules does not.
为阐明血管平滑肌（VSM）细胞中细胞骨架与L型Ca(2+)（Ca(L)）通道活性之间的相互作用，我们使用全细胞电压钳技术研究了肌动蛋白丝和微管破坏对培养的VSM细胞（A7r5细胞系）L型Ca(2+)电流[I(Ba(L))]的影响。将细胞暴露于每种破坏剂1小时，然后进行电生理和形态学检查。使用抗α-肌动蛋白和抗α-微管蛋白抗体进行免疫染色的结果表明，秋水仙碱破坏了肌动蛋白丝和微管，细胞松弛素D仅破坏了肌动蛋白丝，而诺考达唑仅破坏了微管。暴露于秋水仙碱或细胞松弛素D而非诺考达唑的细胞中，I(Ba(L))大幅降低。当用鬼笔环肽稳定肌动蛋白丝或用鬼笔环肽加紫杉醇处理细胞以稳定两种细胞骨架成分时，秋水仙碱甚至可将I(Ba(L))抑制约40%。这些结果表明，秋水仙碱还必须通过另一种未知机制对I(Ba(L))产生一些抑制作用，例如直接阻断Ca(L)通道。总之，VSM细胞的肌动蛋白丝破坏会抑制Ca(L)通道活性，而微管破坏则不会。
