Vol. 285, Issue 2, 828-834, May 1998

Guanine Nucleotide-Binding Inhibitory Protein-Mediated Inhibition of Adenylyl Cyclase Is Enhanced in Spontaneously Hypertensive Rat Preglomerular Arteriolar Smooth Muscle Cells¹

Subhash J. Vyas, Rupa Mokkapatti, Raghvendra K. Dubey, Mala R. Chinoy and Edwin K. Jackson

Center for Clinical Pharmacology and Departments of Pharmacology (R.M., E.K.J.) and Medicine (S.J.V., R.K.D.), University of Pittsburgh School of Medicine, Pittsburgh and Section of Pediatric Surgery (M.R.C.), Division of Surgery, The Pennsylvania State University College of Medicine, The Milton S. Hershey Medical Center, Hershey, Pennsylvania

The purpose of our study was to determine whether G_i -mediated control over adenylyl cyclase in preglomerular arteriolar smooth muscle cells (PGASMC) is enhanced in the spontaneously hypertensive rat (SHR). PGASMC were cultured from preglomerular microvessels isolated from adult SHR (14-15 wk of age) and age-matched WKY rats. Confluent monolayers of cells in third passage were used for the experiments. cAMP released into the media (30 min) as well as cellular levels of cAMP were measured in the presence of a phosphodiesterase inhibitor, 1-isobutyl-3-methyl-xanthine (IBMX; 100 µM) and expressed as pmol/mg protein. Total (released + cellular) cAMP was significantly lower in SHR (14.19 ± 2.30 pmol/mg protein) as compared with WKY (28.3 ± 3.04 pmol/mg protein). Correspondingly, the released (4.6 ± 0.4 pmol/mg protein) as well as cellular (9.78 ± 2.18 pmol/mg protein) cAMP levels were also significantly lower in SHR when compared with WKY (8.85 ± 1.26 and 18.86 ± 2.0 pmol/mg protein, respectively). The steady-state levels of none of the G_i subunits, namely G_i1, G_i2 and G_i3, were higher in the SHR PGASMC. Pertussis toxin treatment (PTX; 100 ng/ml; 24 hr) caused complete ADP-ribosylation of G_i subunits in both WKY and SHR PGASMC. The same treatment of PTX also produced a significant increase in total cAMP in SHR, but not in WKY, such that the total cAMP levels after PTX treatment were not significantly different between the two strains. Interestingly, PTX significantly increased the released (20.26 ± 0.90 pmol/mg protein) but not the cellular (13.63 ± 1.63 pmol/mg protein) cAMP in SHR. Forskolin (1 µM) induced similar increases in total cAMP and isoproterenol (1 µM) caused greater increases in total cAMP in SHR cells compared with WKY cells. These data strongly suggest that in SHR PGASMC total adenylyl cyclase activity is not altered. Furthermore, steady-state expressions of G _i-1, G _i-2 and G _i-3 are not increased whereas G_i -mediated inhibition of adenylyl cyclase is augmented in SHR PGASMC.