Role of protein kinase C- or RhoA-induced Ca(2+) sensitization in stretch-induced myogenic tone.

OBJECTIVE It has been suggested that Ca(2+) sensitization mechanisms might contribute to myogenic tone. However, specific mechanisms have yet to be fully identified. Therefore, we investigated the role of protein kinase C (PKC)- or RhoA-induced Ca(2+) sensitization in myogenic tone of the rabbit basilar vessel. METHODS Myogenic tone was developed by stretch of rabbit basilar artery. Fura-2 Ca(2+) signals, contractile responses, PKC immunoblots, translocation of PKC and RhoA, and phosphorylation of myosin light chains were measured. RESULTS Stretch of the resting vessel evoked a myogenic contraction and an increase in the intracellular Ca(2+) concentration ([Ca(2+)](i)) only in the presence of extracellular Ca(2+). Stretch evoked greater contraction than high K(+) at a given [Ca(2+)](i). The stretch-induced increase in [Ca(2+)](i) and contractile force were inhibited by treatment of the tissue with nifedipine, a blocker of voltage-dependent Ca(2+) channel, but not with gadolinium, a blocker of stretch-activated cation channels. The PKC inhibitors, H-7 and calphostin C, and a RhoA-activated protein kinase (ROK) inhibitor, Y-27632, inhibited the stretch-induced myogenic tone without changing [Ca(2+)](i). Immunoblotting using isoform-specific antibodies showed the presence of PKCalpha and PKCepsilon in the rabbit basilar artery. PKCalpha, but not PKCepsilon, and RhoA were translocated from the cytosol to the cell membrane by stretch. Phosphorylation of the myosin light chains was increased by stretch and the increased phosphorylation was blocked by treatment of the tissue with H-7 and Y-27632, respectively. CONCLUSIONS Our results are consistent with important roles for PKC and RhoA in the generation of myogenic tone. Furthermore, enhanced phosphorylation of the myosin light chains by activation of PKCalpha and/or RhoA may be key mechanisms for the Ca(2+) sensitization associated myogenic tone in basilar vessels.