Abstract 17402: Cardiac Myosin Regulatory Light Chain Phosphorylation Regulates Inotropic Response Against Alpha1- Adrenergic Stimulation
Background: Phosphorylation of myosin regulatory light chain (MLC2) in cardiac muscle is indispensable for normal cardiac function and cardiac-specific myosin light chain kinase (cMLCK) plays a main role in maintaining the its phosphorylation level. Here, we identified the additional other isoform of cMLCK in various mouse strains. To elucidate the importance of MLC2 phosphorylation in cardiac performance, we examined the effects of MLC2 phosphorylation on developed tension, twitch tension parameters and the inotropic response against phenylephrine, alpha1-adrenergic receptor agonist, using the mice that never express the previously reported cMLCK in nature.
Methods and Results: Using the antibodies against cMLCK and nanoLC-MS/MS analysis, cMLCK with the molecular mass of 61 kDa (cMLCK-2), in addition to the previously reported 86 kDa cMLCK, was revealed to be expressed in mice heart. Among various mouse strains, C57BL/6N expressed only cMLCK-2 due to the point mutation in the cMLCK gene, mylk3, though its closest relative strain, C57BL/6J, expressed both isoforms. The level of MLC2 phosphorylation in C57BL/6N ventricular muscle was significantly lower than that in C57BL/6J. Compared with C57BL/6J, basal developed tension of left ventricular papillary muscles was significantly small and time to peak tension as well as half relaxation time were significantly prolonged in C57BL/6N. Phenylephrine increased concentration-dependently contractile force without changing MLC2 phosphorylation level and C57BL/6J revealed more significant positive inotropic response than C57BL/6N. The increase in the MLC2 phosphorylation by calyculin A, an inhibitor of protein phosphatases 1 and 2A, led to the enhanced positive inotropic response in C57BL/6N but not in C57BL/6J. There was a significant positive correlation between positive inotropic response against phenylephrine and MLC2 phosphorylation level ranging from 15% to 30%.
Conclusion: Our results suggest that there exists the other isoform of cMLCK in mice heart. In addition, the basal level of MLC2 phosphorylation regulates developed tension, twitch parameters and sensitivity against alpha1-adrenergic receptor stimulation.
- © 2013 by American Heart Association, Inc.