Regulation of adenylate cyclase in cultured cardiocytes from neonatal rats by adenosine and other agonists

The presence of adenosine receptors coupled to adenylate cyclase in cultured cardiocytes from atria and ventricles from neonatal rats is demonstrated in these studies. N-Ethylcarboxamideadenosine (NECA), l-N⁶-phenylisopropyladenosine (PIA), and 2-chloroadenosine (2-cl-Ado) stimulated adenylate cyclase in a concentration-dependent manner in both cultured atrial and ventricular cells. The order of potency of stimulation was NECA > PIA > 2-cl-Ado. The stimulation of adenylate cyclase by NECA was enhanced by guanine nucleotides and was blocked by 3-isobutyl-1-methylxanthine in both these cells. Other agonists such as epinephrine, norepinephrine, dopamine, F^?, and forskolin were also able to stimulate adenylate cyclase, although the extent of stimulation by these agents was higher in ventricular than in atrial cells. The stimulation of adenylate cyclase by epinephrine and norepinephrine was inhibited by propranolol but not by phentolamine. On the other hand, phentolamine, propranolol, and haloperidol inhibited dopamine-stimulated adenylate cyclase activity to the same extent. Forskolin, at its maximal concentration, potentiated the stimulatory effect of epinephrine, norepinephrine, and dopamine on adenylate cyclase in both atrial and ventricular cardiocytes, but the interaction of NECA with epinephrine, norepinephrine, or dopamine was different in atrial and ventricular cells. The stimulation by an optimal concentration of NECA was additive with maximal stimulation by the catecholamines in atrial cells but not in ventricular cells. The data suggest the existence of adenosine “Ra” and catecholamine receptors in cultured atrial and ventricular cardiocytes. It can be postulated that adenosine in addition to its role as a potent vasodilator might regulate cardiac performance through its interaction with “Ra” receptors associated with adenylate cyclase. The difference in the mode of interaction of adenosine with catecholamines in atrial and ventricular cells suggests that the mechanism by which these agents activate adenylate cyclase may be different in these cells.     

The effect of alpha and beta adrenergic receptor stimulation on the adenylate cyclase activity of human adipocytes.

The effects of the mixed agonist epinephrine and the beta agonist isoproterenol, each alone and in combination with the alpha adrenergic blocker phentolamine and the beta blocker propranolol on the adenylate cyclase activity of human adipocyte membrane fragments were determined in a calcium free buffer. Neither phentolamine (10 muM) nor propranolol (32 muM) affected basal adenylate cyclase activity. Epinephrine (10 muM) stimulated adenylate cyclase activity and this effect was slightly enhanced by phentolamine. The combination of epinephrine plus propranolol depressed adenylate cyclase below the basal level. Isoproterenol (10 muM) markedly stimulated adenylate cyclase; the addition of phentolamine caused an equivocal further increase while the addition of propranolol depressed adenylate cyclase activity to, but not below, the basal level. These findings are consistent with the hypothesis that human adipocytes have both alpha and beta adrenergic receptors and that these receptors are associated with the cell membrane adenylate cyclase system.     

The regulation of adenylate cyclase activity in turkey erythrocyte membranes by forskolin

The mechanisms by which forskolin stimulates adenylate cyclase activity in turkey erythrocyte membranes and is influenced by manganese and Gpp(NH)p were studied. Forskolin-dependent adenylate cyclase activity in particulate turkey erythrocyte membranes is enhanced following preincubation of membranes with isoproterenol and GMP (cleared membranes). In contrast, solubilization of turkey erythrocyte membranes, previously cleared, renders them relatively refractory to forskolin but not to Gpp(NH)p. Whereas adenylate cyclase activity due to the simultaneous presence of forskolin and Mn2+ in particulate turkey erythrocyte membranes is additive, their copresence becomes synergistic after solubilization. The apparent Kact for forskolin activation of adenylate cyclase is not influenced by clearance or by the presence of Mn2+ in particulate turkey erythrocyte membranes. Following solubilization, the Vmax for forskolin-dependent adenylate cyclase activation determined in the presence of Mn2+ is also independent of clearance. Forskolin activation of turkey erythrocyte adenylate cyclase appears to be influenced at sites in addition to the catalytic unit.     

Catecholamine-stimulated GTPase activity in turkey erythrocyte membranes.

Determination of specific GTPase (EC 3.6.1.--) activity in turkey erythrocyte membranes was achieved using low concentration of GTP (0.25 muM), inhibition of nonspecific nucleoside triphosphatases by adenosine 5'(beta,gamma-imino-triphosphate (App(NH)p) and suppression of the transfer of gamma-32P from GTP to ADP with an ATP regeneration system. Under these conditions catacholamines caused a 30--70% increase in GTP hydrolysis. The stimulation of GTPase activity by catecholamines required the presence of Mg2+ or Mn2+. DIfferent batches of membranes revealed the following specific activities (pmol 32Pi/mg protein min): basal GTPase (determined in the absence of catecholamine), 6-- 11; catecholamine-stimulated TTPase, 3--7; and residual non-specific NTPase 3--5. The stimulation of GTPase activity by catecholamines fulfilled the stereospecific requirements of the beta-adrenergic receptor, and was inhibited by propranolol. The concentrations of DL-isoproterenol which half-maximally activated the GTPase and adenylate cyclase were 1 and 1.2 muM, respectively. The following findings indicate that the catecholamine-stimulated GTPase is independent of the catalytic production of cyclic AMP by the adenylate cyclase. Addition of cyclic AMP to the GTPase assay did not change the rate of GTP hydrolysis. Furthermore, treatment of the membrane with N-ethylmaleimide (MalNEt) at 0 degrees C which caused 98% inhibition of the adenylate cyclase, had no effect on the catecholamine-stimulated GTPase. The affinity and specificity for GTP in the GTPase reactions are similar to those previously reported for the stimulation of the adenylate cyclase. The apparent Km for GTP in the basal and the catecholamine-stimulated GTPase reaction was 0.1 muM. These GTPase activities were inhibited by ITP but not by CTP and UTP. It is proposed that a catecholamine-stimulated GTPase is a component of the turkey erythrocyte adenylate cyclase system.     

Role of phosphatidylinositol turnover in alpha1 and of adenylate cyclase inhibition in alpha2 effects of catecholamines

Ligand binding and pharmacological studies have indicated that alpha-adrenergic receptors can be divided into alpha₁ and alpha₂. We suggest that alpha₁ receptors mediate those metabolic effects of alpha catecholamines which involve phosphatidylinositol turnover and the release of bound intracellular Ca²⁺ as well as the entry of extracellular Ca²⁺. In contrast, alpha effects of catecholamines are due to non-specific inhibition of adenylate cyclase through a mechanism independent of Ca²⁺. A similar classification for the effects of both histamine and serotonin suggests that they have separate type 1 or alpha receptors for Ca²⁺ dynamics which are different from type 2 or beta receptors which regulate adenylate cyclase.There is a significant correlation between hormone effects on phosphatidylinositol turnover and elevation of intracellular Ca²⁺. The available data suggest that the turnover of membrane-bound phosphatidylinositol is involved in Ca²⁺ gating in rat hepatocytes, rat and hamster adipocytes and blowfly salivary glands. In hamster adipocytes adenylate cyclase activity is also inhibited by alpha₂ catecholamines through a Ca²⁺ independent mechanism.     

Regulation of Adenosine-Sensitive Adenylate Cyclase from Rat Brain Striatum

An adenosine-sensitive adenylate cyclase has been characterized from rat brain striatum. In whole homogenates as well as in particulate fractions, N⁶-phenylisopropyl adenosine (PIA), 2-chloroadenosine, and adenosine N′-oxide were equipotent in stimulating adenylate cyclase. Although GTP inhibited basal as well as PIA-stimulated activity of whole homogenates, the enzyme showed an absolute dependency on GTP for stimulation by PIA, dopamine, epinephrine, and norepinephrine in a particulate fraction derived from discontinuous sucrose gradient centrifugation. Adenosine exerts two effects on this adenylate cyclase, stimulation at low concentrations and inhibition at high concentrations, suggesting the presence of two adenosine binding sites. The stimulation of adenylate cyclase by PIA was dependent on the concentration of Mg^2-. The degree of stimulation by PIA was greater at a low concentration of Mg²⁺, which suggests that stimulation by PIA was accompanied by increasing the apparent affinity for Mg²⁺. Activation of adenylate cyclase by PIA was blocked by theophylline or 3-isobutyl- 1-methylxanthine (IBMX). The pH optimum for basal or (PIA + GTP)-stimulated activities was broad, with a peak between 8.5 and 9.5. In the presence of GTP, stimulation by an optimal concentration of PIA was additive, with maximal stimulation by the catecholamines. Phospholipase A₂ treatment at a concentration of 1 U/ml for 5 min completely abolished the stimulatory effect of dopamine, whereas PIA-stimulated activity remained unaltered. These data suggest that rat brain striatum either has a single adenylate cyclase, which is stimulated by catecholamines and adenosine by distinct mechanisms, or has different cyclase populations, stimulated by either adenosine or catecholamines.     

Beta-adrenergic receptors: stereospecificity and lack of affinity for catechols

Studies of adenylate cyclase activity in rat liver, heart and fat cell microsomal preparations and in turkey and rat erythrocyte ghosts indicate that β-adrenergic receptors exhibit very strict stereospecificity for (?)-catecholamines. (+)-Isomers of active catecholamines and inactive catechol compounds do not inhibit the β-adrenergic-mediated stimulation of adenylate cyclase and thus do not interact with specific receptors. However, very high concentrations (above 10^?4 M) of (?)- and (+)-isomers, as well as of biologically inactive non-catecholamine catechols (e.g., pyrocatechol, dihydroxymandelic acid), inhibit in a nonspecific manner the basal, hormone (catecholamine, glucagon)- and NaF-stimulated adenylate cyclase activity. Studies with propranolol suggest that the low activity (0.1 to 1%) of (+)-isomers of norepinephrine can be explained by contamination with the (?)-isomer. The activity of soterenol, a potent non-catechol β-adrenergic agonist, is uninfluenced by (+)-catecholamines or catechols. It is concluded that the binding of ³H-labeled catecholamines to a variety of cells, microsomes and membranes as described in various previous studies cannot represent specific receptor interactions. Binding to receptors must demonstrate strict stereospecificity and must not be affected by unrelated catechol substances.     

The effect of membrane preparation and cellular maturation on human erythrocyte adenylate cyclase

We found that adenylate cyclase activity of human erythrocytes is potentially labile during isolation of their plasmalemma. Addition of 1 mM EGTA to solution used to remove hemoglobin from lysed cells protected activity. Human erythrocyte adenylate cyclase is minimally activated by catecholamines, in the absence or presence of exogenous guanyl nucleotide, but substantially by 5′-guanylyl imidodiphosphate or sodium fluoride and concentration-dependently by Mg²⁺ or Mn²⁺. Basal catalytic activity is an age-dependent component of the human erythrocyte; 5′-guanylyl imidodiphosphate- or fluoride-activated activities decline with cellular maturation proportionally to the decrease in basal activity.     

Calcium and adenosine affect human sperm adenylate cyclase activity

The adenylate cyclase activity of human ejaculated spermatozoa in broken-cell preparations was investigated. In the presence of 5 mM metal cations and 0.1 mM ATP, the relative enzyme activity with Mn²⁺, Ca²⁺, Mg²⁺, Ba²⁺ was 1.00, 0.28, 0.22, and 0.03, respectively. Added Ca²⁺ appeared to activate the enzyme in the presence of Mn²⁺ or Mg²⁺. The human sperm adenylate cyclase was stimulated by ～ 2-fold by free Ca²⁺ (lmM) in the presence of Mg²⁺ (5 mM). If the GTP analogue, 5′-guanylyl imidophosphate (Gpp(NH)p) was added to the sperm homogenate in the presence of 200 μM ethylene-glycol-bis (β-aminoethylether) N,N′-tetraacetic acid (EGTA), the adenylate cyclase activity was increased by approximately 25%, but with the addition of 280 μM Ca²⁺ there was a decrease in enzyme activity. A similar response to low concentrations of Ca²⁺ was obtained after complementation of the sperm enzyme with the guanine nucleotide regulatory component from human erythrocytes, where the addition of 40 μM Gpp(NH)p, 200 μM EGTA, and Ca²⁺ (≤ 160 μM) stimulated the sperm enzyme ～ 3–4-fold, but the further addition of Ca²⁺ (280 μM, final) neutralized the stimulatory effect. The addition of adenosine, and the nucleotides 5′-AMP and 5′-ADP inhibited the enzyme, whereas guanine and 5′-GMP had no appreciable effect. Human follicular fluid and serum also had little direct effect on the sperm adenylate cyclase. These resuls suggest that Ca²⁺ might be an important physiological modulator of the human sperm adenylate cyclase.     

Calcium ion effects on guanylate cyclase of brain.

Soluble guanylate cyclase activity of brain is stimulated by Ca²⁺ in the presence of low concentrations of Mn²⁺. Unlike Ca²⁺ stimulation of adenylate cyclase, the effect does not depend upon interaction of guanylate cyclase with a specific high-affinity Ca²⁺-binding protein. In the presence of Mg²⁺, Ca²⁺ inhibits soluble guanylate cyclase as well as the particulate enzyme. The concept that stimulation of brain cells results in increased cyclic GMP concentration secondary to Ca²⁺ influx merits additional critical study.     

Effects of calcium on a parathyroid hormone-sensitive adenylate cyclase inhibitor

Inhibition of parathyroid hormone (PTH)-sensitive adenylate cyclase by {Nle-8, Nle-18, Tyr-34} bPTH-(3–34) amide was studied in thyroparathyroid-ectomized dogs. The inhibitory effect was shown to be markedly enhanced by the addition of calcium ions into the $\text{in}\text{,}\text{vitro}$ assay system. At 0.1 mM Ca²⁺, complete inhibition by the antagonist was obtained. Chelation of exogenous Ca²⁺ by EGTA eliminated the Ca²⁺-induced inhibition. Both the basal and hormone-stimulated activities were decreased in the presence of 0.1 mM Ca²⁺, whereas the addition of EGTA increased both activities. Our results suggest that Ca²⁺ modulates canine renal PTH-sensitive adenylate cyclase and its inhibition by substituted bPTH-(3–34).     

Adrenergic versus VIPergic control of cyclic AMP in human colonic crypts

The actions of catecholamines on VIP-induced cyclic AMP is studied in human colon. We show that: (1) Epinephrine in the 10(-7)-10(-3) M concentration range (ED50 = 11.10(-6) M) inhibits VIP-induced cyclic AMP rise in isolated colonic epithelial cells; the maximal inhibition reaches 30% of VIP effect; epinephrine alters the efficacy of the peptide and does not modify its potency; epinephrine also reduces the basal cyclic AMP level. (2) The inhibition is found with other alpha adrenergic agonists with the order of potencies epinephrine greater than norepinephrine greater than phenylephrine. Clonidine has a poor intrinsic activity but antagonizes the action of epinephrine. (3) The inhibition of VIP action by epinephrine is reversed by the alpha antagonists dihydroergotamine, phentolamine and the alpha 2 antagonist yohimbine, while unaffected by the beta antagonist propranolol and the alpha 1 antagonist prazosin, (4) Epinephrine inhibits VIP-stimulated adenylate cyclase activity in preparations of colonic plasma membranes. Thus catecholamines exert through an alpha 2 adrenoreceptor a negative control on basal and VIP-stimulated cyclic AMP formation in human colon. We suggest that colonic cyclic AMP metabolism undergoes a dual control: VIPergic, activator and adrenergic, inhibitor.     

The α1-adrenergic receptor in human erythrocyte membranes mediates interaction in vitro of epinephrine and thyroid hormone at the membrane Ca-ATPase

Membrane Ca²⁺-ATPase activity was stimulated in vitro separately by T₄ (10⁻¹⁰ M) and by epinephrine (10⁻⁶ M). In the presence of a fixed concentration of T₄, additions of 10⁻⁸ and 10⁻⁶ M epinephrine reduced the T₄ effect on the enzyme. β-Adrenergic blockade with propranolol (10⁻⁶ M) prevented stimulation by epinephrine of Ca²⁺-ATPase activity, but did not prevent the suppressive action of epinephrine on T₄-stimulable Ca²⁺-ATPase. In contrast α₁-adrenergic blockade with unlabelled prazosin restored the effect of T₄ on Ca²⁺-ATPase activity in the presence of epinephrine. Like propranolol, prazosin prevented enhancement of enzyme activity by epinephrine in the absence of thyroid hormone. Neither prazosin nor propranolol had any effect on the stimulations by T₄ of red cell Ca²⁺-ATPase in the absence of epinephrine. Analysis of radiolabelled prazosin binding to human red cell membranes revealed the presence of a single class of high-affinity binding sites (K_d, 1.2 × 10⁻⁸ M; B_max, 847 fmol/mg membrane protein). Thus, the human erythrocyte membrane contains α₁-radrenergic receptor sites that are capable of regulating Ca²⁺-ATPase activity.     

Activation by a calcium-binding protein of guanylate cyclase in Tetrahymena pyriformis.

A Ca²⁺-binding protein (TCBP), which was isolated from $\text{Tetrahymena pyriformis}$, enhanced about 20-fold particulate-bound guanylate cyclase activity in $\text{Tetrahymena}$ cells in the presence of a low concentration of Ca²⁺, while the adenylate cyclase activity was not increased. The enhancement was eliminated by ethylene glycol-bis (β-aminoethyl ether)-N,N′-tetraacetic acid. The enzyme activity was not stimulated by rabbit skeletal muscle troponin-C, the Ca²⁺-binding component of troponin, or other some proteins. In the presence of TCBP, stimulating effect of calcium ion on the enzyme activity was observed within the range of pCa 6.0 to 4.6, and was immediate and reversible.     

Renal parathyroid hormone-dependent adenylate cyclase activity after repletion of vitamin D-deficient rats with vitamin D-2

Rats fed a diet deficient in both vitamin D and Ca²⁺ exhibited a greater depression of the renal parathyroid hormone (PTH)-dependent adenylate cyclase than was observed in rats fed diets deficient in either vitamin D or calcium. Total serum Ca²⁺ was decreased from a control level of 11.2 mg/dl to 8.5 mg/dl in rats fed the diet deficient in calcium alone, and to 5.4 mg/dl in rats fed the diet deficient in vitamin D. Serum calcium was decreased further to 4.3 mg/dl in rats fed the diet deficient in both vitamin D and Ca²⁺. Serum immunoreactive PTH was significantly elevated over control levels when rats were fed the test diets; however, there were no significant differences between the elevated levels in the three experimental groups. Repletion of rats deficient in vitamin D only with a single oral dose of 3200 I.U. vitamin D-2 resulted in restoration of serum calcium to normal levels, a return of serum PTH to the control state, and an associated increase in PTH-dependent adenylate cyclase activity to the control level by 72 h. Repletion of rats deficient in both vitamin D and Ca²⁺ with the same dose of vitamin D-2 raised serum Ca²⁺ to 7.2 mg/dl by 72 h, but did not cause a reduction in circulating PTH, nor did it result in any significant improvement in the responsiveness of the membrane adenylate cyclase to PTH. These results suggest that elevated PTH is a factor in the down regulation of the PTH-dependent adenylate cyclase, but do not rule out a role for calcium as a regulatory factor.     

β-2-adrenergic stimulation of ornithine decarboxylase activity in porcine granulosa cells in vitro

Epinephrine, norepinephrine, and isoproterenol produced dose-dependent stimulation of ornithine decarboxylase (EC 4.1.1.7) activity in isolated porcine granulosa cells maintained under defined conditions in vitro. β- but not α-receptor-blocking agents prevented enzyme stimulation by catecholamines. Application of preferential β-1 and β-2-receptor antagonists and agonists localized the epinephrine effect to β-2-adrenergic mediation. Epinephrine action was enhanced by the phosphodiesterase inhibitor, 1-methyl-3-isobutyl-xanthine, but not by saturating concentrations of the cyclic AMP analogue, 8-bromocyclic AMP, of follicle-stimulating hormone, or of prostaglandin E₂. However, stimulation by epinephrine was additive to that of luteinizing hormone. Follicular fluid obtained from immature Graafian follicles contined concentrations of norepinephrine and epinephrine active in vitro.Thus, catecholamines may participate in the regulation of ornithine decarboxylase activity in the ovary. Catecholamine effects may be mediated by β-2-receptors linked to the adenylate cyclase system.     

Ca2+-dependent regulation of calmodulin binding and adenylate cyclase activation in bovine cerebellar membranes

The binding parameters of ¹²⁵I-labeled calmodulin to bovine cerebellar membranes have been determined and correlted with the activation of adenylate cyclase by calmodulin. In the presence of saturating levels of free Ca²⁺, calmodulin binds to a finite number of specific membrane sites with a dissociation constant (K_d) of 1.2 nM. Furthermore, Scatchard analysis reveals a second population of binding sites with a 100-fold lower affinity for calmodulin. The Ca²⁺-dependence of calmodulin binding and of adenylate cyclase activation varies with the amount of calmodulin present, as can be infered from the model of sequential equilibrium reactions which describes the activation of calmodulin-dependent enzymes. On the basis of this model, a quantitative analysis of the effect of free Ca²⁺ and of free calmodulin concentration on both binding and activation of adenylate cyclase was carried out. This analysis shows that both processes take place only when calmodulin is complexed with at least three Ca²⁺ atoms. The concentration of the active calmodulin ·Ca²⁺ species required for half-maximal activation of adenylate cyclase is very similar to the K_d of the high affinity binding sites on brain membranes. A Hill coefficient of approx. 1 was found for both processes indicating an absence of cooperativity. Phenothiazines and thioxanthenes antipsychotic agents inhibit calmodulin binding to membranes and calmodulin-dependent activation of adenylate cyclase with a similar order of potency. These results suggest that the Ca²⁺-dependent binding of calmodulin to specific high affinity sites on brain membranes regulates the activation of adenylate cyclase by calmodulin.     

Alanine and glutamine synthesis and release from skeletal muscle. IV. beta-Adrenergic inhibition of amino acid release.

Alanine and glutamine formation and release were studied using the intact epitrochlaris preparation of rat skeletal muscle. Epinephrine reduced the release of alanine and glutamine in a concentration-dependent manner. Measurable inhibition was observed at 10(-9) M epinephrine, and maximal inhibition was obtained at 10(-5) M. Norepinephrine also reduced alanine and glutamine formation and release but the concentration required for maximal inhibition was approximately 100-fold greater than for epinephrine. Isoproterenol (beta agonist), but not phenylephrine (alpha agonist), reproduced the effects of epinephrine, and propranolol (beta antagonist), but not phentolamine (alpha antagonist), blocked the effect of the catecholamine. N6,O2'-Dibutyryl adenosine 3':5'-monophosphate reproduced the effects of epinephrine and theophylline potentiated the effect of submaximal concentrations of the hormone. Glucagon and prostaglandin E2 had no observable effect on amino acid release. Insulin did not modify the inhibition of alanine and glutamine release produced by epinephrine. Alanine and glutamine formation from added precursor amino acids was unaffected by epinephrine or cyclic adenosine 3':5'-monophosphate. Epinephrine reduced alanine formation in muscles obtained from diabetic rats or animals treated with thyroxine or cortisone. These findings indicate that physiological levels of catecholamines reduce alanine and glutamine formation and release from skeletal muscle. This effect is mediated by a beta-adrenergic receptor and the adenylate cyclase system and can be accounted for by an inhibition of muscle protein degradation.     

Mechanism of adenylate cyclase activation by the rat lung cytoplasmic factors

Epinephrine, histamine and prostaglandin E₁ stimulated adenylate cyclase activity in lung membranes and their stimulation of the enzyme activity was completely blocked by propranolol, metiamide and indomethacin, respectively. A partially-purified activator from the adult rat lung also enhanced adenylate cyclase activity in membranes. However, stimulation of adenylate cyclase by the rat lung activator was not abolished by the above receptor antagonists. Further, epinephrine, NaF and Gpp(NH)p stimulated adenylate cyclase activity rather readily, whereas stimulation of the enzyme activity by the lung activator was evident after an initial lag phase of 10 min. Also, the lung activator produced additive activation of adenylate cyclase with epinephrine, NaF and Gpp(NH)p. These results indicate that the lung activator potentiates adenylate cyclase activity in membranes by a mechanism independent from those known for epinephrine, NaF and Gpp(NH)p. Incubation of lung membranes for 30 min at 40°C resulted in a loss of adenylate cyclase activation by NaF and Gpp(NH)p. Addition of the released proteins to the heat-treated membranes did not restore the enzyme response to these agonists. However, heat treatment of lung membranes in the presence of 2-mercaptoethanol or dithiothreitol prevented the loss of adenylate cyclase response to NaF and Gpp (NH)p. N-ethylmaleimide abolished adenylate cyclase activation by epinephrine, NaF, Gpp(NH)p and the lung activator. These results indicate that the sulfhydryl groups are important for adenylate cyclase function in rat lung membranes.Abbreviations Gpp(NH)p 5-Guanylimidodiphosphate     

Demonstration of adenylate cyclase activity in human red blood cell ghosts

The presence of adenylate cyclase (ATP pyrophosphate-lyase (cyclizing) EC 4.6.1.1) activity was demonstrated in human erythrocyte ghosts and was found to be around 3 pmol adenosine ′,5′-monophosphatase (cyclic AMP) · 2 h^?1 · mg^?1 protein. This enzymatic activity is strongly stimulated by NaF and 5′-guanylimidodiphosphate, is slightly stimulated by epinephrine, norephrine, soproterenol, and prostaglandin E, and is inhibited by calcium. The hormone stimulation is not potentiated by 5′-guanylylimidodiphosphate.