Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, C58

Oral Communications

Role of Rho-kinase and Src-Family Kinases in carbachol-induced contraction of isolated rat bronchioles.

N. Irechukwu1, G. A. Knock1, J. P. Ward1

1. King's College London, London, United Kingdom.

  • Effects of inhibitors of Rho-kinase and Src-family kinases on carbachol contraction in rat bronchioles

Asthma is associated with remodelling of airway smooth muscle and hyper-responsiveness to constrictor stimuli. Potential therapeutic targets for the treatment of asthma are Rho-kinase and its principle activator RhoA. They have been implicated in both remodelling and constrictor hyper-responsiveness due to their involvement in smooth muscle migration, proliferation and constriction, the latter primarily through inhibition of myosin phosphatase (Ca2+-sensitization). The signalling pathways that control activity of RhoA/Rho-kinase however, such as the Src-family of tyrosine kinases (SrcFK), are largely unexplored. We studied carbachol-induced contraction in isolated wire myograph-mounted rat bronchioles (1260± 68µm), bathed in bicarbonate-buffered physiological salt solution, pH 7.4 at 37°C, and investigated the effects of inhibitors of Rho-Kinase (Y27632, 10µM) and SrcFK (PP2, 30µM) on these contractile responses. Carbachol (0.001-100µM) caused a concentration-dependent constriction (EC50 -5.84 ± 0.06 Log[M], n=18]), which was reproducible upon repetition (both EC50 and max unchanged, P>0.05, n=6). Y27632 significantly increased the EC50 (from -5.81 ± 0.09 to -5.32 ± 0.03, P<0.001, paired t-test, n=6) and decreased the maximal contractile response (by 17.3 ± 6.5%, p<0.05, paired t-test, n=6) (fig a). PP2 also significantly increased the EC50 (from -5.91 ± 0.10 to -5.64 ± 0.10, P<0.01, paired t-test, n=6) and decreased the maximal contractile response (by 10.1 ± 2.4%, p<0.01 paired t-test, n=6) (fig b). Carbachol also caused constriction in α-toxin permeabilised rat bronchioles with [Ca2+]i clamped at pCa 6.4 and in the presence of 10µM cyclopiazonic acid and 1µM GTP. This constriction was also inhibited by Y27632 (n =3). These results suggest the involvement of both Rho-kinase and SrcFK in contraction of rat bronchioles. We aim to further characterise these responses and determine whether SrcFK are acting upstream of Rho-kinase, as shown previously in pulmonary artery (1).

Where applicable, experiments conform with Society ethical requirements