atrial-natriuretic-factor and phenylalanyl-prolyl-arginine-chloromethyl-ketone

Thrombin and the atrial natriuretic peptide (ANP) possess a number of functionally antagonistic properties in vascular endothelial cells. Thus, regulatory interactions that modulate the activity of one or the other could have important sequelae with regard to cardiovascular homeostasis. Thrombin treatment effected a dose- and time-dependent reduction in ANP receptor activity (maximal 70% to 80% inhibition) in cultured bovine aortic endothelial cells. This resulted from a decrease in total receptor number as well as a modest reduction in the affinity of the receptor for its ligand. The inhibition was largely confined to the type C receptor population, in that thrombin had no effect on maximal type A receptor-linked cGMP accumulation. The protein kinase C-activating phorbol ester 12-O-tetradecanoylphorbol 13-acetate effected a similar reduction in binding activity; however, suppression of protein kinase C activity did not reverse the thrombin effect. Pretreatment of endothelial cells with cycloheximide did not completely prevent the thrombin-dependent inhibition, and thrombin did not effect a reduction in type C receptor mRNA levels, findings that argue for a postsynthetic inhibitory locus. The inhibition of receptor activity was effectively irreversible in that suspension of protein synthesis blocked the recovery of receptor density on the cell surface. Reduction in type C receptor density was accompanied by modest increases in the stability of ANP in the culture medium and enhancement of the cellular cGMP response to the peptide, particularly at low ligand concentrations. These findings demonstrate a potentially important interaction between these two agonist systems in regulating endothelial cell function within the vascular wall.

Topics: Amino Acid Chloromethyl Ketones; Animals; Antithrombins; Aorta; Atrial Natriuretic Factor; Cattle; Cells, Cultured; Cycloheximide; Dose-Response Relationship, Drug; Down-Regulation; Endothelium, Vascular; Enzyme Inhibitors; Protein Kinase C; Protein Synthesis Inhibitors; Receptors, Atrial Natriuretic Factor; Receptors, Thrombin; Staurosporine; Tetradecanoylphorbol Acetate; Thrombin; Time Factors