Date of Award

2-1991

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Medical Sciences

Supervisor

Richard J. Haslam

Abstract

In platelets, the activators of guanylyl cyclase used were sodium nitroprusside (SNP) and SIN-1 (the active metabolite of molsidomine). PGE$\sb1,$ a functional analogue of prostacyclin (PGI$\sb2),$ and adenosine were the activators of platelet adenylyl cyclase studied. Changes in cyclic ($\sp3$H) nucleotides were measured in platelets prelabelled with ($\sp3$H) guanine and ($\sp3$H) adenine. Incubation of labelled platelets with SNP or SIN-1 caused inhibitions of platelet function which were associated with large dose-dependent increases in ($\sp3$H) cGMP and 1.4 to 3.0-fold increases in ($\sp3$H) cAMP. However, addition of either SNP or SIN-1 with PGE$\sb1$ or adenosine, at concentrations of the latter that had little effect alone, caused much larger increases in ($\sp3$H) cAMP and greatly enhanced the inhibition of platelet function. Experiments with an adenylyl cyclase inhibitor, 2$\sp\prime,$5$\sp\prime$-dideoxyadenosine (DDA), suggested that these synergistic interactions depend on an enhanced accumulation of cAMP. Hemoglobin inhibited all nitrovasodilator-mediated effects. Cilostamide, a selective inhibitor of platelet cGMP-inhibited cAMP phosphodiesterase (cGI-PDE), had effects identical to those of SNP, suggesting that the actions of the latter depend on inhibition of the same enzyme. Also, the results obtained with M&B 22,948, a cGMP-selective PDE inhibitor, strongly suggest that the increases in platelet ($\sp3$H) cAMP caused by nitrovasodilators, in the presence or absence of activators of adenylyl cyclase, are mediated by the inhibition of cAMP breakdown of cGMP. To determine if a similar synergism is seen in vascular smooth muscle (VSM), the interactions between isoproterenol and either nitrovasodilators or cAMP-PDE inhibitors were studied. A fast and sensitive prelabelling method was established for the measurement of cyclic nucleotides in cultured vascular smooth muscle cells (VSMC) from rat aorta. Using this method, agonist-induced changes in the cyclic nucleotide levels of two different aortic VSMC lines were studied. (Abstract shortened by UMI.)


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