Caveolar remodeling is a critical mechanotransduction mechanism of the stretch-induced L-type Ca²⁺ channel activation in vascular myocytes

Activation of L-type voltage-dependent Ca²⁺ channels (VDCC_L) by membrane stretch contributes to many biological responses such as myogenic contraction of arteries. However, mechanism for the stretch-induced VDCC_L activation is unclear. In this study, we examined the hypothesis that caveolar remodeling and its related signaling cascade contribute to the stretch-induced activation of VDCC_L in rat mesenteric arterial smooth muscle cells. The VDCC_L currents were recorded with nystatin-perforated or with conventional whole-cell patch-clamp technique. Hypotonic (~230 mOsm) swelling-induced membrane stretch reversibly increased the VDCC_L currents. Electron microscope and confocal imaging analysis revealed that both hypotonic swelling and cholesterol depletion by methyl-β-cychlodextrin (MβCD) similarly disrupted the caveolae structure and translocated caveolin-1 (Cav-1) from membrane to cytosolic space. Accordingly, MβCD also increased VDCC_L currents. Moreover, subsequent hypotonic swelling after MβCD treatment failed to increase the VDCC_L currents further. Western blotting experiments revealed that hypotonic swelling phosphorylated Cav-1 and JNK. Inhibitors of tyrosine kinases (genistein) and JNK (SP00125) prevented the swelling-induced facilitation of VDCC_L currents. Knockdown of Cav-1 by small interfering RNA blocked both the VDCC_L current facilitation by stretch and the related phosphorylation of JNK. Taken together, the results suggest that membrane stretch is transduced to the facilitation of VDCC_L currents via caveolar structure-dependent tyrosine phosphorylation of Cav-1 and subsequent activation of JNK in rat mesenteric arterial myocytes.