Calcium not strain regulates localization of α-myosin heavy chain mRNA in oriented cardiac myocytes

Abstract.

Our aim was to test a hypothesis that localization of the α-myosin heavy chain (α-MyHC) mRNA in oriented neonatal rat cardiomyocytes is regulated either by calcium, or by mechanical strain, or by both. Myocytes, grown on collagen aligned on stretchable silicone membranes, were elongated and had an increased length to width ratio (L/W) compared with randomly oriented myocytes grown on conventional substrata. Oriented cells were stretched by 10% in the longitudinal direction, in the transverse direction or passively unloaded for 6 h. As expected, shape changes followed these mechanical deformations. In situ hybridization was used to determine the localization of α-MyHC mRNA by quantitative analysis of optical density under various mechanical perturbations in myocytes that were either spontaneously beating or treated with verapamil (10 mM) to block influx of calcium. Unstretched, longitudinally stretched, and cells stretched transversely all had mRNA dispersed to their extremities. Verapamil treatment resulted in a perinuclear pattern of mRNA under all three mechanical perturbations. Additionally, mRNA distribution was examined in myocytes that were passively unloaded in the presence and absence of verapamil. Unloading myocytes with intact calcium cycling does not result in a perinuclear accumulation of mRNA. These data suggest that calcium is essential for α-MyHC mRNA distribution throughout the cell whereas stretch and alignment affect myocyte shape but have little effect on mRNA localization.