Evidence for myosin light chain kinase mediating noradrenaline‐evoked cation current in rabbit portal vein myocytes

1 The role of myosin light chain kinase (MLCK) in the activation of the noradrenaline‐evoked non‐selective cation current ( I cat ) was examined with the whole‐cell recording technique in single rabbit portal vein smooth muscle cells. 2 Intracellular dialysis with 5 μM MLCK (11–19) amide, a substrate‐specific peptide inhibitor of MLCK, markedly reduced the amplitude and rate of activation of noradrenaline‐evoked I cat . A similar result was obtained when the cells were dialysed with 10 μM AV25, which also inhibits MLCK by an action at the auto‐inhibitory domain of MLCK. 3 Inhibitors of binding of ATP to MLCK, wortmannin and synthetic naphthalenesulphonyl derivatives (ML‐7 and ML‐9), at micromolar concentrations, also reduced the amplitude of noradrenaline‐evoked I cat . 4 ML‐7 and ML‐9 (both at 5 μM) reduced the amplitude of I cat induced by both guanosine 5′‐ O ‐(3‐thiotriphosphate) (GTPγS) and 1‐oleoyl‐2‐acetyl‐ sn ‐glycerol (OAG). 5 MLCK (11–19) amide, AV25 and ML‐9 did not inhibit the noradrenaline‐evoked Ca 2+ ‐activated potassium current at a holding potential of 0 mV. In addition, MLCK (11–19) amide and AV25 did not reduce the non‐selective cation current induced by ATP in rabbit ear artery cells. 6 Intracellular dialysis with 2 μM Ca 2+ and 9 μM calmodulin activated I cat , which developed over a period of about 5 min. 7 Intracellular dialysis with the non‐hydrolysable analogue of ATP, 5′‐adenylylimidodiphosphate (AMP‐PNP), reduced the amplitude and rate of activation of noradrenaline‐evoked I cat . 8 The results indicate that MLCK mediates noradrenaline‐activated I cat in rabbit portal vein smooth muscle cells.