Role of Cyclic AMP in Alpha-Adrenoceptor Mediated Vasorelaxation in Goat Arterial Strip

BACKGROUND: Alpha adrenergic activation is known to produce vasoconstriction. It has been shown in our lab that phenylephrine (PE), an alpha-adrenergic agonist could cause vasorelaxation in conditions with high nitric oxide (NO) and it requires alpha adrenergic activation as phentolamine, an α receptor antagonist blocked the vasorelaxant effect of PE/NO. In subsequent MD thesis, it was shown that such vasorelaxant effect is through α1D receptor. These experiments were done in spiral strip preparation. However, in further experiments it was seen that PE under normal NO environment also induced vasorelaxation in spiral strip. Thus, in this study we have studied two different preparations made from a small artery: transverse cylinder and longitudinal strip to study the effect of PE on circular and longitudinal smooth muscle respectively. The α receptor subtype involved in such PE induced reduction in tension was delineated and the role of second messenger, cAMP in such scenario was also studied. AIM OF THE STUDY: To delineate the role of cyclic Adenosine Mono Phosphate (cAMP) in phenylephrine (PE) induced vasorelaxation. OBJECTIVES: 1. To induce vasorelaxation in isolated goat arterial strip by phenylephrine. 2. To test if inhibition of basal nitric oxide (NO) by L-NNA (Nϖ-Nitro-L-arginine) could prevent phenylephrine induced vasorelaxation in longitudinal strip. 3. To test if phenylephrine induced vasorelaxation in longitudinal strip is preventable by an alpha adrenoceptor blocker, phentolamine. 4. To test if specific alpha receptor subtypes blockers such as RS 17053 (α1A blocker), chloroethylclonidine dihydrochloride (α1B blocker) and BMY 7378 dihydrochloride (α1D blocker) could prevent phenylephrine induced vasorelaxation in longitudinal strip. 5. To test if inhibition of adenylyl cyclase by MDL-12330A could prevent phenylephrine induced vasorelaxation in longitudinal strip. 6. If phenylephrine induced vasorelaxation in longitudinal strip was prevented by adenylyl cyclase inhibitor MDL-12330A, then to delineate the downstream pathway of cAMP by using protein kinase A (PKA) blocker, H-89 and exchange protein directly activated by cAMP (EPAC) blocker, ESI-09. METHODS: Isolated artery from fresh goat leg was made into two different preparations: transverse cylinder and longitudinal strip and one end was tied to organ bath of 20ml capacity perfused with mammalian ECF solution (@ 37o Celsius and gassed with carbogen), other end was connected to a force transducer which in turn was connected to powerlab data acquisition system for recording tension. Drugs were added to both organ bath and drug reservoir for continuous perfusion. The changes in tension were analysed using Igor pro software. RESULTS: Phenylephrine produces only contractile response on circular smooth muscle of blood vessels both under normal and high NO environment. However, it induced relaxation in longitudinal strip from small artery. Such PE induced relaxant response on longitudinal smooth muscle is dependent on NO and α-adrenergic activation. The relaxant response is mediated through α1D receptor and cAMP doesn’t have a role in such PE induced relaxant response on longitudinal smooth muscle. CONCLUSION: The effect of PE on circular smooth muscle of small artery is only contraction. However, its effect on longitudinal smooth muscle of small artery is relaxation. PE induced relaxation of longitudinal smooth muscle is mediated through α1D adrenoceptor and is independent of cAMP. The spiral strip preparation predominantly has the longitudinal smooth muscle component and therefore the results in the earlier thesis were similar to longitudinal strip preparation