CHANGES IN TYPE-VI ADENYLYL-CYCLASE ISOFORM EXPRESSION CORRELATE WITH A DECREASED CAPACITY FOR CAMP GENERATION IN THE AGING VENTRICLE

We investigated the developmental regulation of the beta-adrenergic receptor-G(s)-adenylyl cyclase pathway in myocardial membranes from fetal, neonatal, adult, and mature adult rats by measuring the density of the beta-adrenergic receptor and the activities of the stimulatory guanine nucleotide-binding protein G(s) and the adenylyl cyclase enzyme. Total beta-adrenergic receptor content (in femtomoles per milligram protein) was greatest in the fetal (124.4+/-20.5 fmol/mg) and neonatal (122.3+/-16.1 fmol/mg) stages and gradually decreased in the adult (90.9+/-8.0 fmol/mg) and mature adult (70.0+/-9.6 fmol/mg) stages. An equivalent pattern was seen for adenylyl cyclase activity: the basal activity of the effector enzyme or that measured in the presence of 0.1 mmol/L isoproterenol with 0.1 mmol/L Gpp(NH)p, 10 mmol/L NaF, or 0.05 mmol/L forskolin was greater in the fetus and the neonate than in the adult and the mature adult. These data suggested that decreased stimulation of the catalytic unit by G(s) could be the underlying cause of diminished adenylyl cyclase activity with aging. However, quantification of G(s) by reconstitution into S49 cyc- membranes (in picomoles cAMP per microgram for 10 minutes) demonstrated no significant decrease during development from fetus (1.55+/-0.1 pmol/mug) to neonate (1.9+/-0.5 pmol/mug) and subsequent aging to adult (2.6+/-0.2 pmol/mug) and mature adult (1.9+/-0.2 pmol/mug). When Northern blot analysis was used to characterize the relative amounts of mRNA coding for G(salpha), no significant differences were seen among the developmental stages studied. Unlike G(salpha), the inhibitory protein G(ialsph2) was subject to developmental regulation; however, Concomitant evaluation of G(ialpha2) levels both by ADP-ribosylation with pertussis toxin and by immunoblotting showed the greatest amount in the fetus (0.82+/-0.07 pmol/mg) and the neonate (0.51+/-0.1 pmol/mg). The adult and mature adult both contained 0.09+/-0.02 pmol/mg. Also, the steady-state G(ialpha2), mRNA level paralleled the amount of protein. Similarly, the level of type V mRNA coding for adenylyl cyclase was greater in the mature adult than in the fetal and neonatal stages. In contrast, the level of type VI adenylyl cyclase mRNA decreased with age, paralleling the decline in the functional activity of adenylyl cyclase with age. Taken together, these data suggest that the age-related changes in the activity of the myocardial beta-adrenergic receptor-G(s)-adenylyl cyclase pathway is not primarily regulated by alteration in the level of G(s) or G(i) but by the density of beta-adrenergic receptors and by the activity of the catalyst adenylyl cyclase. In particular, steady-state mRNA measurements show that a decrease in the content of the type VI isoform of adenylyl cyclase correlates with a decrease in catalytic activity with age.