The objectives of this proposal are to investigate the biochemical changes in vascular smooth muscle of spontaneously hypertensive rat (SHR). These studies are aimed to further our understanding of the mechanisms underlying increased vascular tension development in this model of hypertension. Experiments are designed to investigate the effect of calmodulin- and cAMP-dependent protein kinase mediated phosphorylation of sarcoplasmic reticulum (SR) and sarcolemma (SL) vesicles isolated from bovine carotid artery on Ca++ uptake and (Ca++, Mg++)-ATPase activity. Further, we will investigate whether calmodulin- and cAMP-dependent protein kinase phosphorylate the same protein at different sites or they phosphorylate different proteins of SR and SL membranes isolated from bovine carotid artery. We will investigate the effect of calmodulin- and cAMP-dependent protein kinase mediated phosphorylation of SR and SL vesicles isolated from vascular smooth muscle of SHR and WKY on Ca++ uptake and (Ca++, Mg++)-ATPase activity. The objective of these studies being to understand the molecular mechanisms underlying reduced Ca++ sequestration by microsomes isolated from vascular smooth muscle of SHR. Further, the hypothesis that increased tension in caudal artery strip of SHR is due to increased responsiveness of contractile proteins to Ca++ will be tested. To achieve this objective we will study in skinned caudal artery strips of SHR and WKY (i) contractile response to graded doses of Ca++, (ii) the effect of catalytic subunit of cAMP-dependent protein kinase on contractile responsiveness to Ca++. Experiments are also designed to test whether increased reactivity of caudal artery strips of SHR to norepinephrine and Ca++ is due to the increased myosin light chain phosphorylation. Estimations will be made of tension development in response to graded doses of norepinephrine using intact caudal artery strips and to graded doses of Ca++ using skinned caudal artery strips. The extent of myosin light chain phosphorylation will be quantitated after the steady-state tension development for each dose of NE or Ca++. The extent of myosin light chain phosphorylation will be correlated with the extent of tension generation in the caudal artery strips of WKY and SHR.
