Decrease In Adenylate Cyclase Activity Research Articles

Heavy isotope labeling study of the turnover of forskolin-stimulated adenylate cyclase in BC3H1 cell line.

We have used the method of heavy isotope labeling to study the metabolic turnover of adenylate cyclase in a nonfusing muscle cell line, the BC3H1 cells. These cells contains an adenylate cyclase coupled to beta-adrenergic receptors and highly stimulated by forskolin, a potent activator of the enzyme. After transfer of the cells from normal medium to heavy medium (a medium containing heavy labeled amino acids, 2H, 13C, 15N), heavy isotope-labeled adenylate cyclase molecules progressively replace pre-existing light molecules. In sucrose gradient differential sedimentation, after a 5-day switch in heavy medium, the enzyme exhibited a higher mass (s = 8.40 +/- 0.03 S, n = 13) compared to the control enzyme (s = 7.40 +/- 0.04 S, n = 36). Indeed, the increase in the sedimentation coefficient of the heavy molecules was due to the synthesis of new molecules of adenylate cyclase labeled with heavy isotope amino acids since in the presence of cycloheximide, an inhibitor of protein synthesis, no change in the sedimentation pattern of the forskolin-stimulated adenylate cyclase occurred. After incorporation of heavy isotope amino acids in the adenylate cyclase molecules, the kinetics parameters of the enzyme (i.e. Km for ATP and EC50 for Mn2+ or Mg2+) did not change. However, adenylate cyclase from cells incubated with heavy medium exhibits an activity about 2-fold lower than control (cells in light medium). After switching the cells to the heavy medium, the decrease of the activity of the enzyme occurred during the first 24 h and thereafter remained at a steady state for at least 4 days. In contrast, 24 h after the switch, the sedimentation coefficient of forskolin-stimulated adenylate cyclase was progressively shifted to a higher value indicating that the heavy isotope-labeled enzyme replaced the pre-existing light form of the molecule. These observations show that the rapid decrease in adenylate cyclase activity and the synthesis of heavy adenylate cyclase molecules are two separate events. The relative amounts of heavy and light components of forskolin-stimulated adenylate cyclase obtained in sucrose gradient differential sedimentation were determined as a function of time beginning 24 h after the transfer into the heavy medium. The decrease of the pre-existing light form could be represented by simple first order kinetics with a half-time of 40 h. This result suggests that the metabolic renewal of forskolin-stimulated adenylate cyclase is comparable to that of most plasma membrane proteins.

WR-2721 inhibits parathyroid adenylate cyclase.

WR-2721 [S-2-(3-aminopropylamino)ethylphosphorothioic acid] is a chemoprotective and radioprotective agent that has been shown to lower serum calcium in dogs and in humans. This is secondary both to impaired release of Ca2+ from bone and diminished secretion of parathyroid hormone (PTH) from parathyroid glands. Because cAMP plays a role in the regulation of PTH secretion and WR-2721 has been shown to lower cAMP levels in radiated mouse spleen, we investigated the effects of WR-2721 on cAMP production in dispersed bovine parathyroid cells. Additionally, we studied the adenylate cyclase in plasma membranes from normal bovine parathyroid glands after exposure to WR-2721. With parathyroid cells incubated at 0.5 mM Ca2+, addition of WR-2721 in concentrations ranging from 0.02 to 2.0 mM resulted in a progressive decrease in intracellular cAMP (42-50%, respectively). In plasma membranes of bovine parathyroid cells a dose-dependent decrease in adenylate cyclase activity was noted. Inhibition of the cyclase was seen over a wide range of Mg2+ concentrations (2.5-40 mM). WR-2721 inhibited both basal and NaF, Gpp(NH)p, forskolin, and pertussin toxin-stimulated adenylate cyclase. These data suggest that WR-2721 inhibits the activity of parathyroid adenylate cyclase.

Antipain microinjection prevents progesterone to inhibit adenyl cyclase in Xenopus oocytes

Microinjection of antipain, an inhibitor of thiol and Ca 2+-dependent proteases, in immature Xenopus oocytes inhibited meiotic maturation induced by progesterone, but not by transfer of cytoplasm taken from maturing oocytes. Oocytes could be released from antipain inhibition by increasing progesterone concentration. α - 32P -ATP was microinjected to study adenylcyclase in ovo . As already reported, neosynthesis of cAMP was decreased following progesterone application. This decrease was not observed, or it was considerably reduced, in oocytes previously injected with antipain. In amphibian, full-grown ovarian oocytes are arrested at first meiotic prophase, and have a large nucleus known as the germinal vesicle. Progesterone induces the production of a cytoplasmic maturation-promoting factor (MPF), which itself triggers germinal vesicle breakdown (GVBD), and subsequent events of meiotic maturation ( Masui and Markert, 1971; Gerhart et al., 1984). A considerable body of evidences support the view that release from prophase block is due to inactivation of a cAMP-dependent protein kinase (reviewed by Maller, 1983). On the other hand, progesterone has been shown to induce a transient decrease in cAMP level ( Speaker and Butcher, 1977; Schorderet-Slatkine et al., 1982; Cicirelli et al., 1985), and this initial drop of cAMP, along with a number of studies indicating a decrease in adenylate cyclase activity ( Mulner et al., 1979; Baltus et al., 1981; Sadler and Maller, 1981; Finidori-Lepicard et al., 1981; Jordana et al., 1981), provided key support to the theory that an early drop in cAMP led to the dephosphorylation of a hypothetical protein which initiates maturation. It has been reported that several antiproteases of microbial origin, including antipain, could inhibit meiotic maturation induced by progesterone but not by the direct transfer of MPF into prophase-arrested oocytes ( Guerrier et al., 1977). Since the exact mechanism according to which progesterone inhibits adenylate cyclase activity remains unknown, this finding led us to investigate whether inhibition of adenylate cyclase following progesterone addition was also observed in antipain-injected oocytes. This communication presents the results of this study.

Human prepro atrial natriuretic factors 26–55, 56–92, and 104–123 increase renal guanylate cyclase activity

Human prepro atrial natriuretic factor 26–55, 56–92, and 104–123 as well as human atrial natriuretic factor (4–28) in the present investigation increased renal cortical and medullary cyclic GMP levels and maximally enhanced particulate guanylate cyclase activity [E.C. 4.6.1.2] two-fold in whole kidney homogenates, renal cortical and medullary membranes, and in isolated distal nephrons at their 1 μM concentrations. Dose-response relationships revealed that the half maximal [ED 50] activation of guanylate cyclase was at their 10 nM concentrations in rat, rabbit, and dog kidneys. Both human atrial natriuretic factor and the prepro factors decreased adenylate cyclase activity. These results suggest that prepro factors 26–55, 56–92, 104–123 may also be functionally active.

Impairment in Coupled Beta-Adrenergic Receptor and Adenylate Cyclase System during Bleomycin-Induced Lung Fibrosis in Hamsters

The purpose of this study was to evaluate the coupled beta-adrenergic receptor (BAR) and adenylate cyclase (AC) system of the lung during the course of the bleomycin-(Bleo) induced pulmonary fibrosis in hamsters. The BAR population, dissociation constants (Kd), AC activity, and its sensitivity to various stimulators were studied at 2, 4, 7, 14, and 21 days after intratracheal administration of either 1 unit of Bleo or an equivalent volume of saline. The BAR population in the lungs of Bleo-treated animals did not differ from control at the early times, but it was significantly reduced to 5.9 X 10(3) fmol and 3.6 X 10(3) fmol from the control values of 1.1 X 10(4) fmol and 1.5 X 10(4) fmol per lung at 14 and 21 days after treatment, respectively. The Kd values for control hamster lung ranged from 2.5 X 10(-11) M to 3.7 X 10(-11) M, and for Bleo-treated hamster lung, from 2.7 X 10(-11) M to 4.8 X 10(-11) M. The Kd at the earliest time, 2 days after treatment, did not differ significantly from the Kd values at the subsequent times in control, while for Bleo-treated hamster lung, the Kd values at 7, 14, and 21 days were significantly higher than the Kd at 2 days after treatment. The Kd values for Bleo-treated hamster lung were also significantly higher than control at 14 and 21 days. The AC activity of the lung in Bleo-treated hamster was significantly reduced to 67%, 40%, 38%, and 50% of their respective controls in response to H2O (basal), GTP (10(-4) M), GTP + isoproterenol (10(-4) M each), and NaF (10 mM) at 21 days after treatment. The extent of AC stimulation in Bleo-treated hamster lung in response to various stimulators was generally less than that of saline control. Reductions in the BAR population and increased Kd values in Bleo-treated hamster lung were attributed to its fibrogenic ability and not to nutritional deficiency, which may partly be accountable for decreased AC activity of the lung in these animals. However, there were qualitative differences in the lung AC activity between Bleo-treated and nutritionally deprived hamsters, since the enzyme from the latter group was generally more responsive to stimulators than the enzyme from the former group. It was concluded from the findings of this study that an impairment in the coupled BAR and AC system of the lung may be partly responsible for the fibrogenic ability of bleomycin.

Calmodulin mediates the stimulatory effect of 3,5,3'-triiodothyronine on adenylate cyclase activity in rat thymocyte plasma membranes.

We have previously demonstrated that T3 increases adenylate cyclase activity in rat thymocyte plasma membranes by a mechanism that is calcium dependent. In the present studies we have examined whether calmodulin participates in this response to T3. Initial experiments provided evidence that calmodulin is involved in regulating the activity of the guanyl nucleotide-dependent adenylate cyclase in this tissue. Thus, compared with findings in freshly prepared membranes, dialysis of thymocyte plasma membrane preparations for 20 h at 4 C decreased their calmodulin concentrations from an initial value of approximately 600 ng/100 micrograms protein by about 50% and decreased adenylate cyclase activity by approximately 80%. Although storage of the same preparations under comparable conditions resulted in no change in calmodulin content, it caused an approximate 30% decrease in adenylate cyclase activity. Addition of calmodulin had no effect on adenylate cyclase activity in fresh or stored membrane preparations, but produced a dose-dependent increase in enzyme activity in preparations that had been dialyzed. Further, when added to freshly isolated membranes, three calmodulin antagonists, trifluoperazine, calmidazolium, and calmodulin antibodies, all produced a concentration-dependent inhibition of adenylate cyclase activity, and this was completely reversed in all cases by the addition of high concentrations of exogenous calmodulin. The stimulation of guanyl nucleotide-dependent adenylate cyclase activity that T3 induced in fresh membrane preparations was present in membranes that had been stored, but was absent in those that had been dialyzed. In addition, the response to T3 in fresh membranes was inhibited or abolished by all three calmodulin antagonists. Both in dialyzed membranes and in the presence of antagonists, the response to T3 was restored by the addition of exogenous calmodulin. We conclude that calmodulin influences the activity of the guanyl nucleotide-dependent adenylate cyclase in rat thymocytes and ultimately mediates the stimulation of enzyme activity that T3 produces. Such mediation very likely explains the calcium-dependent nature of the stimulatory effect of T3 on thymocyte adenylate cyclase activity.

Inhibition of spermatogenesis, motility and metabolic enzymes of sperm by gossypol.

Available data about gossypol a potential male antifertility agent explain its inhibition of spermatogenesis and possible mechanism of action. Spermatogenesis in rats has been inhibited by gossypol with no adverse effects on Leydig cells or Sertoli cells. The degree of damage to rat testes seminiferous epithelium was dose and duration related. Opinion varies regarding the site and mechanism of gossypols antifertility action. Early studies suggested an inhibition of the development of the spermatozoal nucleus and acrosome indicating a site of action for gossypol at the level of the testes but not the epididymis. Other studies did not confirm spermatozoal head defects but rather showed lesions in the spermatozoa tail in the epididymis. This discrepancy may indicate differences in spermatozoa response to gossypol with various mature stages. Other studies with rats and guinea pigs indicated an inhibitory action of gossypol preventing epididymal spermatozoa from attaining maturation. No consensus about the nature or site of gossypols antifertility action has been reached. In men the 1st effect of gossypol is a rapid loss of spermatozoa motility followed by a reduction of spermatozoa count. This decreased motility resulted from a reduction in energy supply through inhibition of glycolysis and the respiratory chain and the uncoupling of oxidative phosphorylation and reduction in ATP synthesis. The influence of gossypol on the tubulin-dynein system has recently been investigated in sea urchins. Gossypol has also been shown to inhibit the activity of several enzymes involved in energy metabolism including several dehydrogenases. Gossypol also decreased adenylate cyclase activity and inhibited fructolysis in human spermatozoa. Oral administration of gossypol at 12-30 mg/kg/day for 5-12 weeks induced infertility in rats hamsters guinea pigs dogs monkeys and men.

Effect of pertussis toxin on the heart muscarinic-cholinergic receptors and their function

Administration of pertussis toxin to rats induced a significant increase in heart rate that was evident as soon as 24 hours after the administration of the toxin and that persisted for at least 15 days. Electrical stimulation of the vagus decreased dramatically the heart rate of control animals but was unable to do it so in rats treated with pertussis toxin. In cardiac membranes muscarinic agonists decreased adenylate cyclase activity (≈20–25%); no effect was observed in membranes obtained from toxin-treated animals. Agonist displacement of antagonist binding ( [ 3H] Quinuclidinyl benzilate) indicated that treatment with pertussis toxin decreased the proportion of receptors in the high affinity state for agonists. All these data suggest that blockade of the parasympathetic tone plays a key role in the induction of tachycardia by pertussis toxin.

Relationship between ethanol-induced alterations in fluorescence anisotropy and adenylate cyclase activity

The effects of butanol, ethanol, and ketamine on adenylate cyclase activity and fluorescence anisotropy were determined in membranes prepared from L6 cells. The experiments were designed to test the hypothesis that the effects of ethanol on adenylate cyclase activity are a consequence of ethanol-induced changes in bulk membrane order. Butanol and ethanol elicited concentration-dependent increases in adenylate cyclase activity and caused decreases in the fluorescence anisotropy of diphenylhexatriene. Butanol was more potent than ethanol in reducing fluorescence anisotropy, and it elicited a greater reduction in fluorescence anisotropy than did ethanol. Butanol was also more potent than ethanol in activating adenylate cyclase, but the highest concentration of butanol used caused a smaller increase in enzyme activity than did the highest concentration of ethanol. When the percent change in adenylate cyclase activity was plotted against the percent change in fluorescence anisotropy at each concentration of alcohol, the increase in isoproterenol-stimulated adenylate cyclase activity per unit change in fluorescence polarization was greater with ethanol than with butanol. Ketamine decreased fluorescence anisotropy but, unlike the alcohols, ketamine caused a decrease in adenylate cyclase activity. A reduction in assay temperature attenuated both the ethanol-induced activation of adenylate cyclase activity and the ethanol-induced reduction in fluorescence anisotropy. Although the data are consistent with the theory that ethanol acts upon a hydrophobic region of the membrane to enhance adenylate cyclase activity, activation of the enzyme does not appear to be a consequence of a decrease in bulk membrane order.

Effects of aluminum on bovine parathyroid adenylate cyclase.

It is known that the secretion of PTH is often impaired in association with aluminum (Al3+) accumulation in patients with renal failure. The mechanisms involved remain ill defined. Since adenylate cyclase plays a role in the regulation of PTH secretion, these studies examine the effects of Al3+ on parathyroid adenylate cyclase. In membranes from normal bovine parathyroid glands, basal adenylate cyclase activity, in the presence of 0.2 mM ATP and 20 mM Mg2+, increased by 22% as Al3+ was raised from 0-10 microM. Higher Al3+ concentrations caused a progressive decrease in adenylate cyclase activity, reaching 68% inhibition of control activity at 2 mM Al3+. Since adenylate cyclase activation is influenced by the interaction of multiple sites within the adenylate cyclase complex, the nature of the inhibition by Al3+ was explored by examining the interaction of Al3+ with substrate ATP and with Mg2+, an allosteric activating metal ion. In the presence of 20 mM Mg2+, Al3+ concentrations of 1-2 mM resulted in noncompetitive inhibition with respect to ATP [decrease in maximum velocity (Vmax) from 4176 in the absence of Al3+ to 1106 pmol cAMP/mg protein X 15 min; Michaelis Menten constant (Km) for ATP was unchanged]. In contrast, at fixed ATP (0.2 mM), 0.5 mM resulted in competitive inhibition of adenylate cyclase with respect to Mg2+, whereas at higher Al3+ concentrations the inhibition was noncompetitive. When Mg2+ was replaced by Mn2+ (enzyme activity reflects the activity of the catalytic unit), the inhibitory effect of Al3+ on adenylate cyclase activity was abolished. These data suggest that the inhibition of parathyroid adenylate cyclase by Al3+ occurs at the level of the allosteric metal activating site. These data provide a potential mechanism for the inhibition of PTH secretion by Al3+.

The functional activity of adult mouse Ley dig cells in monolayer culture: effect of lipoproteins, pregnenolone and cholera-toxin

Further studies were carried out on purified mouse Leydig cells to determine why they lose their hormone responsiveness after several days in monolayer culture. The effects of cholera-toxin on cyclic AMP and testosterone production were examined. It was found that cyclic AMP production could still be maximally stimulated by cholera-toxin after 4 days in culture when response to luteinizing hormone (LH) has declined. Testosterone production was, however, not maintained. Because this decline in testosterone production may have been due to the lack of a suitable substrate after several days in culture, cells were cultured initially in the presence of exogenous pregnenolone and low-density lipoproteins (LDL). Both substances were found to enhance basal and LH-stimulated testosterone production and to extend responsiveness of the cells until at least day 4, but by day 7 response was lost. Cells were then cultured in the presence of rat and human LDL and HDL and in both cases LDL was found to enhance consistently testosterone production, but HDL was much less effective. Scanning and transmission electron micrographs showed that changes in cell shape occurred during culture, but indicated that the cells were not depleted of lipid droplets by the end of culture or after LH stimulation. It is concluded that the eventual decline in testosterone synthesis is not due to lack of substrate, although the addition of exogenous substrate does extend the period of responsiveness. Nor is it due to a decrease in adenylate cyclase activity. At least part of the lesion is caused by a decrease in the enzymes required for the conversion of pregnenolone to testosterone.

The effect of growth conditions on adenylate cyclase activity and virulence-related properties of Bordetella pertussis.

Growth of Bordetella pertussis in Stainer & Scholte medium in which the NaCl had been replaced by one of several inorganic or organic salts resulted in a large decrease in adenylate cyclase activity, histamine-sensitizing activity and in the amounts of two cell-envelope polypeptides of Mr 28000 and 30000. Although some variation between strains was observed, there was never a case where one of these properties was lost independently of the others. Cultures in which these properties were lost had decreased amounts of extracellular cAMP when compared to NaCl-grown cultures. Adenylate cyclase activity was detected in three locations of B. pertussis cultures (extracellular, extracytoplasmic but cell-associated, and cytoplasmic). After growth in medium containing high concentrations of MgSO4, enzyme activity was decreased to a similar extent in all three locations.

Excitation-Contraction Coupling in Hypertrophied Myocardium

In hypertensive cardiac hypertrophy, inotropic responsiveness of alpha and beta adrenergic stimuli is reduced. We have previously shown that hearts from two-kidney, one-clip renal hypertensive rats (RHR) have increased beta-adrenergic receptor density and a defect in the guanine nucleotide regulatory protein, leading to decreased adenylate cyclase activity. In spontaneously hypertensive rats (SHR), beta-receptor density was decreased with no change in adenylate cyclase. In these present experiments, we have shown that the alpha 1-adrenergic receptor changes are in the opposite direction, decreasing in RHR and increasing in SHR. All these changes are reversible within 4 weeks following removal of the clipped kidney in RHR, at which time blood pressure and heart weight have also returned towards normal. Further studies on the excitation-contraction pathway have indicated that c-AMP-stimulated protein kinase is decreased in SHR with no changes seen in RHR. Subcutaneous infusion of epinephrine leads to some increase of cardiac mass associated with decreased beta-adrenergic receptors element and decreased adenylate cyclase activity. However, following angiotensin II infusion, even though hypertrophy is more pronounced, no changes in receptors or cyclase are detected. We conclude that different models of hypertensive cardiac hypertrophy associated with different biochemical defects in the adrenergic excitation response pathway, and that if some of these changes become irreversible, further cardiac deterioration and even heart failure may ensue.

The effect of age and cholesterol on the rat lung beta-adrenergic system

To assess the influence of membrane lipid composition on beta-adrenergic receptor number and adenylate cyclase activity in aging, we investigated the effect of cholesteryl hemisuccinate on these parameters in lung membranes of 3-, 12-, and 24-month-old CDF (F-344) rats. When cholesteryl hemisuccinate (0.5 mg/ml) was incubated with lung membranes, beta-adrenergic receptor density was increased by 70%. This effect was the same for each age group studied and indicated that the density of both basal and CHS-sensitive receptors is unaltered in rat lung with age. Forskolin, NaF, p[NH]ppG, and isoproteronol-stimulated adenylate cyclase activity is 30% lower in lung membranes from aged rats. Since enzyme activity is affected by the lipid environment and membrane composition often changes with age, we assessed adenylate cyclase activity following cholesteryl hemisuccinate incorporation. There was up to a 75% decrease in adenylate cyclase activity following cholesteryl hemisuccinate incorporation in lung membranes in each of the three age groups. In untreated membranes, there was no significant difference in cholesterol or lipid phosphate content with age. These data suggest that cholesterol content does not account for alterations in senescent rat lung adenylate cyclase activity.

Phosphorylated adenosine derivatives as low-affinity adenosine-receptor agonists. Methodological implications for the adenylate cyclase assay.

In cellular systems provided with activatory (Ra-site) receptors for adenosine, such as rat cerebral microvessels and rat liver plasma membranes, the adenosine-receptor antagonist 8-phenyltheophylline (10 microM) significantly decreased adenylate cyclase activity if ATP was the substrate and only if GTP was present. With dATP as substrate, adenylate cyclase activities in both preparations remained unaffected by 8-phenyltheophylline. In rat cerebral-cortical membranes, with inhibitory (Ri-site) receptors for adenosine, 8-phenyltheophylline significantly enhanced adenylate cyclase activity only in the presence of GTP and if ATP was the substrate. In rat cardiac ventricular membranes, which are devoid of any adenylate cyclase-coupled adenosine receptor, the methylxanthine had no GTP-dependent effect, irrespective of the substrate used. All assay systems contained sufficiently high amounts of adenosine deaminase (2.5 units/ml), since no endogenous adenosine, formed from ATP, was found chromatographically. In order to demonstrate a direct influence of phosphorylated adenosine derivatives on adenylate cyclase activity, we investigated AMP in a dATP assay system. AMP was verified chromatographically to remain reasonably stable under the adenylate cyclase assay conditions. In the microvessels, AMP increased enzyme activity in the range 0.03-1.0 mM, an effect competitively antagonized by 8-phenyltheophylline. In the cortical membranes, 0.1 mM-AMP inhibited adenylate cyclase, which was partially reversed by the methylxanthine. The presence of GTP was again necessary for all observations. In the ventricular membranes, AMP had no effect. Since the efficacy of adenosine-receptor agonists and, probably, that of other hormones on adenylate cyclase activity can be more efficiently measured with dATP as the enzyme substrate, this nucleotide seems preferable for adenylate cyclase measurements in systems susceptible to modulation by adenosine.

Impaired formation of beta-adrenergic receptor-nucleotide regulatory protein complexes in pseudohypoparathyroidism.

Decreased activity of the guanine nucleotide regulatory protein (N) of the adenylate cyclase system is present in cell membranes of some patients with pseudohypoparathyrodism (PHP-Ia) whereas others have normal activity of N (PHP-Ib). Low N activity in PHP-Ia results in a decrease in hormone (H)-stimulatable adenylate cyclase in various tissues, which might be due to decreased ability to form an agonist-specific high affinity complex composed of H, receptor (R), and N. To test this hypothesis, we compared beta-adrenergic agonist-specific binding properties in erythrocyte membranes from five patients with PHP-Ia (N = 45% of control), five patients with PHP-Ib (N = 97%), and five control subjects. Competition curves that were generated by increasing concentrations of the beta-agonist isoproterenol competing with [125I]pindolol were shallow (slope factors less than 1) and were computer fit to a two-state model with corresponding high and low affinity for the agonist. The agonist competition curves from the PHP-Ia patients were shifted significantly (P less than 0.02) to the right as a result of a significant (P less than 0.01) decrease in the percent of beta-adrenergic receptors in the high affinity state from 64 +/- 22% in PHP-Ib and 56 +/- 5% in controls to 10 +/- 8% in PHP-Ia. The agonist competition curves were computer fit to a "ternary complex" model for the two-step reaction: H + R + N in equilibrium HR + N in equilibrium HRN. The modeling was consistent with a 60% decrease in the functional concentration of N, and was in good agreement with the biochemically determined decrease in erythrocyte N protein activity. These in vitro findings in erythrocytes taken together with the recent observations that in vivo isoproterenol-stimulated adenylate cyclase activity is decreased in patients with PHP (Carlson, H. E., and A. S. Brickman, 1983, J. Clin. Endocrinol. Metab. 56:1323-1326) are consistent with the notion that N is a bifunctional protein interacting with both R and the adenylate cyclase. It may be that in patients with PHP-Ia a single molecular and genetic defect accounts for both decreased HRN formation and decreased adenylate cyclase activity, whereas in PHP-Ib the biochemical lesion(s) appear not to affect HRN complex formation.

Effect of prostaglandin F2 alpha on the adenylate cyclase and phosphodiesterase activity of ovine corpora lutea.

The objective of the present study was to determine the temporal relationships among luteal adenylate cyclase activity, luteal phosphodiesterase activity, luteal progesterone concentration and plasma progesterone concentration during prostaglandin F2 alpha (PGF2 alpha)-induced luteolysis in ewes. Corpora lutea (CL) were removed from cycling ewes on d 9 (d 0 = first day of estrus) at 0, 2, 4, 6, 12 and 24 h (seven to eight ewes/group) after PGF2 alpha administration (im). Jugular blood samples were collected at the time of enucleation of CL and analyzed for progesterone. Plasma and luteal progesterone concentrations were decreased (P less than .05) by 4 and 12 h after PGF2 alpha injection, respectively. Basal adenylate cyclase, luteinizing hormone (LH)-activated adenylate cyclase, guanylylimidodiphosphate [Gpp(NH)p]-activated adenylate cyclase and LH plus Gpp(NH)p-activated adenylate cyclase activities were decreased (P less than .05) by 2 h after PGF2 alpha injection. The decrease in adenylate cyclase activity paralleled the decrease in plasma progesterone concentration over time. Luteinizing hormone stimulated (P less than .05) adenylate cyclase activity relative to basal activity at 0, 2, 12 and 24 h post-PGF2 alpha; whereas, Gpp(NH)p stimulated (P less than .01) adenylate cyclase activity relative to basal activity at each time point. In contrast to the decrease in adenylate cyclase activity, phosphodiesterase activity was increased (P less than .05) at 2 and 4 h post-PGF2 alpha. These results suggest that a decrease in adenylate cyclase activity coupled with an increase in phosphodiesterase activity may decrease the intracellular adenosine 3',5' cyclic monophosphate (cAMP) concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Decrease in adenylate cyclase activity in dog livers during endotoxic shock.

The effects of endotoxin administration on adenylate cyclase in dog liver plasma membranes were studied. The basal, fluoride-, guanine nucleotide-, isoproterenol-, and glucagon-stimulated adenylate cyclase activities were decreased by 61, 62, 69, 83, and 63%, respectively, 2 h after in vivo administration of endotoxin. Endotoxin (100 micrograms/ml) in vitro decreased the guanine nucleotide-, isoproterenol-, and glucagon-stimulated adenylate cyclase activities by 24, 25, and 21%, respectively. These data demonstrate that endotoxin administered in vivo or in vitro had an inhibitory effect on the adenylate cyclase enzyme system in dog liver plasma membranes. Because of the involvement of the adenylate cyclase-adenosine 3',5'-cyclic monophosphate (cAMP) system in the regulation of hepatic carbohydrate metabolism, the finding that endotoxin administration decreased adenylate cyclase activity in the liver should contribute to the understanding of the pathophysiology of altered hepatic glucose homeostasis during shock.

Effect of chronic ethanol ingestion on the cyclic AMP system of the upper gastrointestinal tract in the rat.

Chronic ethanol consumption significantly increases gastric adenylate cyclase (AC) activity (p less than 0.05) without influencing low Km 3',5'-cyclic adenosine monophosphate (cAMP) phosphodiesterase (PD) activity in the rat. On the other hand, in the duodenum and upper part of the jejunum, chronic ethanol feeding leads to a significant decrease of adenylate cyclase activity (p less than 0.02) and, again, does not affect low Km cAMP phosphodiesterase activity. In addition, the effect of various hormonal secretagoques on small intestinal adenylate cyclase activity was investigated. Prostaglandin I2 and D2, as well as glucagon, do not stimulate AC at all. However, small intestinal adenylate cyclase exhibits a lower sensitivity to prostaglandin E2 and vasoactive intestinal peptide (VIP), and a lower efficacy to VIP after chronic ethanol consumption when compared to controls. The decrease of both basal and stimulated AC activity following ethanol ingestion in the upper small intestine may be due to membrane alterations and tissue damage caused by ethanol. The ethanol-induced increase in gastric AC may be of relevance with respect to an increased acid secretion observed after alcohol administration.

Adenosine A1 receptors are associated with cerebellar granule cells.

The cerebellum of mouse appears to have only the adenosine A1 receptor, which decreases adenylate cyclase activity, and not the A2 receptor, which increases adenylate cyclase activity. The adenosine analog N6-(L-phenylisopropyl)adenosine (PIA), stimulates the A1 receptor in a membrane preparation and decreases basal adenylate cyclase activity by 40%. The EC50 for PIA is approximately 50 nM. To associate the A1 receptor with a cerebellar cell type, three different neurological mutant mouse strains were studied: staggerer (Purkinje and granule cell defect), nervous (Purkinje cell defect), and weaver (granule cell defect). PIA was unable to effect a maximal decrease in adenylate cyclase activity of membranes prepared from cerebella of the staggerer and weaver mice in comparison with the respective littermate control mice. In contrast, membranes from nervous mice and their littermates showed similar PIA dose-response curves. Moreover, the diminished PIA response observed in the weaver cerebellum, when compared with the control littermate, was not detected in the striatum. This suggests no overall brain defect in the adenosine A1 receptors coupled to adenylate cyclase of the weaver mouse. We conclude that a loss of granule cells coincides with an attenuated response to PIA, implying that the A1 receptors are associated with the granule cells of the cerebellum.