Rho kinase is involved in the augmented contractile response to cooling mediated by 5-HT1 receptors in equine digital veins

H. Zerpa, S.R. Bailey, Y. Berhane, J. Elliott. Dept of Veterinary Basic Sciences, Royal Veterinary College, London NW1 0TU.

The equine digital vascular bed has an important physiological role in thermoregulation. 5-hydroxytryptamine (5-HT) is a potent vasoconstrictor of equine digital vessels (Bailey and Elliott, 1998). Cooling may enhance the response of cutaneous vessels to 5-HT and α2-adrenoceptor agonists, as seen in the mouse tail artery, Rho kinase is involved in the augmented contractile response mediated by α2-adrenoceptors (Bailey et al., 2004). The aim of this study was to establish the effect of the Rho kinase inhibitor, fasudil, on the cooling-enhanced response to 5-Carboxamidotryptamine (5-CT) in the isolated equine digital vein (EDV).Isometric tension response ofEDV to 5-CT was studied as previously described (Bailey and Elliott, 1998) using blood vessels harvested from adult mixed breed horses of either sex killed at an abattoir. Pairs of EDV rings from the same horse (n=6) were studied simultaneously. The contractile effect of cumulatively increasing concentrations of 5-CT (0.1 nM-10 µM) was assessed in the absence and presence of fasudil (1 µM) at temperatures of either 30°C (the peripheral hoof temperature of horses maintained in a thermo-neutral environment) or 22°C. Data are presented as the increase in tension expressed as a percentage of the maximal response to a depolarizing Krebs solution (DKS; KCL 118 mM). Cumulative concentration response curves (CRC) were fitted to a two-phase logistic equation using Origen 7.5 software. Statistical comparisons between groups were performed using paired Student’s t test.

asignificant response compared to 30°C. bsignificant effect of fasudil vs control (P<0.05). First phase response (P1). Second phase response (P2).

Cooling to 22°C significantly increased the maximum contractile response to 5-CT and shifted the response curve to the left. Fasudil had no effect on the contractile response at 30°C, but significantly abrogated the effect of cooling, decreasing the maximum response and increasing the EC50. This suggests that Rho kinase plays an important role in the enhanced contractile response to cooling, mediated by 5-HT1B/D receptors in EDV. Inhibition of myosin phosphatase by Rho/Rho kinase activation could be one of the mechanisms involved in the cooling-enhanced response in these vessels. Alternatively, Rho/Rho kinase activation by cooling may potentially lead to translocation of 5-HT1 receptors to the cell membrane, as demonstrated in the mouse tail artery for the α2C adrenoceptor subtype (Bailey et al., 2004).

Bailey SR, and Elliott J. (1998). Equine Vet. J. 30: 124-130.
Bailey SR, Eid AH, Mitra S, Flavahan S, Flavahan NA. (2004). Circ Res . 28:1367-74.

HZ was supported by CDCH-UCV and the Programme Al b an, the European Union Programme of Scholarships for Latin America, scholarship No. E05D054410VE.