The effect of a Rho–kinase inhibitor (Y-27632) on the relaxation of thromboxane a₂ mimetic (U-46619) induced contraction in the presence and absence of the endothelium

Abstract

Background: These experiments investigated mechanisms responsible for the slow relaxation of aortic rings following contraction elicited by the thromboxane mimetic, U46619. In this study we hypothesized that blocking only the Rho-kinase pathway with the Rho-kinase inhibitor (Y-27632) will shorten the extended relaxation time induced by U46619 and that production of NO contributes to the relaxation of blood vessels treated with the Rho kinase inhibitor.

Methodology: Aortic rings were obtained from a euthanized rabbit and placed in heated organ baths and contractile responses to the thromboxane mimetic, U46619 measured. After removal of the endothelium or inhibition of NO production, we subjected the U46619 induced vessels to the Rho kinase inhibitor (Y-27632), so as to determine if Y-27632 causes relaxation of U46619 induced vessels by acting through NO, which is the relaxing factor in the endothelium. Vessels exposed to U46619 relaxed at a significantly slower rate compared to other agonists such as phenylephrine (1.0 uM) or high KCl solutions (60 uM).

Result: Inhibition of Rho-kinase significantly reduced the relaxation time; i.e., the rate of relaxation was higher (0.13 ± 0.07 g/min) in the presence of a Rho kinase inhibitor (Y-27632; 1 uM) compared to vehicle-treated vessels (0.02 ± 0.01 g/min) (P< 0.05). We then investigated whether the enhanced rate of relaxation following inhibition of Rho kinase was dependent on the release of NO from the endothelium. Vessels treated with an inhibitor of NO production (L-NAME) or vessels where the endothelium was mechanically removed showed the same response to inhibition of Rho-kinase as vessels treated with the vehicle of L-NAME or vessels in which the endothelium was not denuded.

Conclusion: We conclude that Rho kinase plays an essential role in sustaining the contractile phase of vessels treated with the TxA2 mimetic U46619. The faster rate of relaxation of vessels following inhibition of Rho-kinase does not involve release of NO from the endothelium.

Keywords: Thromboxane A₂ mimetic (U46619), Rho-kinase inhibitor (Y-27632), Vascular Smooth Muscle (VSM), Endothelium and Nitric oxide (NO)s