Inhibition of RhoA Kinase but not Protein Kinase C can potentiate Blebbistatin‐induced relaxation of smooth muscle contraction.

Abstract

Smooth muscle (SM) contraction involves numerous receptors and second messengers in a complex spatiotemporal pattern of changes that regulate force production and maintenance. RhoA kinase (ROCK) and protein kinase C (PKC) are two primary participants in this complex regulatory scheme. Blebbistatin (a cell‐permeable selective and specific small molecule inhibitor of myosin II) has been reported to inhibit Pi release and thereby prevent strong binding of cross‐bridges. We found that during a K+‐induced contraction of rabbit circular fundic strips, blebbistatin inhibited the late, slow maintained force to a greater extent than the early, fast force. Pharmacological inhibition of ROCK (Y27632, H1152) had no effect on the early, fast contractile response but significantly inhibited the late, slow contractile response. In contrast, inhibition of PKC (GF 109203x) inhibited the early, fast and late, slow contractile responses. Combined blockade of ROCK and myosin II gave a greater inhibition of the late, slow contraction than blockade of ROCK or myosin II alone. This was not true for the combined blockade of PKC and myosin II. More selective inhibitors of specific PKC isoforms did not show an additive effect of specific PKC isotypes with blebbistatin in inhibiting the late slow contraction. Partial resistance to the inhibitory effect of blebbistatin suggests that the early, fast phase of SM contraction involves unique spatiotemporal regulation of SM myosin. The potentiation by ROCK inhibitors of the inhibitory effect of blebbistatin on the late, slow contractile response suggests that ROCK may regulate force‐maintenance, in part, by a mechanism supplemental to cross‐bridge activation.