This proposal examines whether lipoxgenase (LO) products of arachidonic acid metabolism mediate the vasoconstrictor effect of angiotensin II (AII). This notion is based on findings that LO products are obligatory mediators of AII-induced adrenal aldosterone secretion and their inhibition decreases the vascular pressor response to AII. Initial studies will focus on the dose response relation of several LO inhibitors on the attenuation of the in vitro contractile responses to AII, norepinephrine and KCI in isolated intact and de-endothelialized rat femoral artery strips and in the mesenteric and hind limb circulations. Simultaneous measurements by HPLC and RIA of the LO products 12- and 15-hydroxyeicosotetranoic acid (HETE) will be done on femoral arteries under the experimental conditions described above. Further studies will be examine the direct effect of HETEs on contractility by addition to femoral vessels alone and to arteries treated with AII and LO blockers. The above studies should show if LO pathway products are authentic metabolites in vascular tissue and if they have a role in mediating the vascular effects of AII. To elucidate the site of HETE production the ability of AII to stimulate HETEs in smooth muscle and endothelial cells will be studied in separate and combined tissue culture systems. AII cellular action involves changes in intracellular calcium (Cai++) and inositol triphosphate (IP3). The project will also examine in cultured vascular smooth muscle cells. (VSMC) whether 12- and 15-HETE stimulate Cai++ and IP3 and if LO blockade diminishes the AII-induced rise in Cai++. These intracellular calcium mobilization using several probes to elucidate mechanisms of HETE action on second messengers. These studies may introduce the exciting prospect of applying LO inhibitors to modulate tissue responses to AII in research and clinical medicine.
