The MLCK‐mediated α 1 ‐adrenergic inotropic effect in atrial myocardium is negatively modulated by PKC ɛ signaling

1 The present study examined the role of myosin light chain kinase (MLCK), PKC isozymes, and inositol 1,4,5‐trisphosphate (IP 3 ) receptor in the positive inotropic effect of α 1 ‐adrenergic stimulation in atrial myocardium. 2 We measured inotropic effects of phenylephrine (0.3–300  μ M ) in isolated left atrial preparations (1 Hz, 37°C, 1.8 m M Ca 2+ , 0.3  μ M nadolol) from male 8‐week FVB mice ( n =200). Phenylephrine concentration‐dependently increased force of contraction from 1.5±0.1 to 2.8±0.1 m N (mean±s.e.m., n =42), which was associated with increased MLC‐2a phosphorylation at serine 21 and 22 by 67% and translocation of PKC ɛ but not PKC α to membrane (+30%) and myofilament (+50%) fractions. 3 MLCK inhibition using ML‐7 or wortmannin right‐shifted the concentration–response curve of phenylephrine, reducing its inotropic effect at 10  μ M by 73% and 81%, respectively. 4 The compound KIE1‐1 (500 n M ), an intracellularly acting PKC ɛ translocation inhibitor peptide, prevented PKC ɛ translocation and augmented the maximal inotropic effect of phenylephrine by 40%. In contrast, inhibition of Ca 2+ ‐dependent PKC translocation (KIC1‐1, 500 n M ) had no effect. Chelerythrine, a PKC inhibitor, decreased basal force without changing the inotropic effect of phenylephrine. 5 The IP 3 receptor blocker 2‐APB (2 and 20  μ M ) concentration‐dependently decreased basal force, but did not affect the concentration–response curve of phenylephrine. 6 These results indicate that activation of MLCK is required for the positive inotropic effect of α 1 ‐adrenergic stimulation, that the Ca 2+ ‐independent PKC ɛ negatively modulates this effect, and that PKC α and IP 3 receptor activation is not involved.British Journal of Pharmacology (2006) 148 , 991–1000. doi: 10.1038/sj.bjp.0706803