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Showing papers on "Cyclase published in 1971"


Journal ArticleDOI
TL;DR: It is concluded that guanyl nucleotides play a specific and obligatory role in the activation of adenyl cyclase by glucagon, and bind at sites, distinct from the glucagon binding sites, that appear to regulate both the response ofadenyl cycling to glucagon and the actions of fluoride ion on this system.

777 citations


Journal ArticleDOI
TL;DR: It is concluded that CT and certain prostaglandins produce small intestinal fluid secretion by increasing mucosal adenyl cyclase activity, thereby stimulating an active secretory process.
Abstract: The effects of several prostaglandins (PG) and a highly purified preparation of cholera enterotoxin (CT) on intestinal mucosal adenyl cyclase activity and the effect of CT on intestinal mucosal cyclic 3',5'-adenosine monophosphate concentration were determined in guinea pig and rabbit small intestine and were correlated with the effects of the same agents on ion transport. Adenyl cyclase activity, measured in a crude membrane fraction of the mucosa, was found at all levels of the small intestine with the highest activity per milligram protein in the duodenum. The prostaglandins, when added directly to the assay, increased adenyl cyclase activity; the greatest effect (2-fold increase) was obtained with PGE(1) (maximal effect at 0.03 mM) and PGE(2). The prostaglandins also increased short-circuit current (SCC) in isolated guinea pig ileal mucosa, with PGE(1) and PGE(2) again giving the greatest effects. The prior addition of theophylline (10 mM) reduced the subsequent SCC response to PGE(1) and vice versa. It was concluded, therefore, that the SCC response to PGE(1), like the response to theophylline, represented active Cl secretion. CT increased adenyl cyclase activity in guinea pig and rabbit ileal mucosa when preincubated with the mucosa from 1 to 2.5 hr in vitro or for 2.5 hr in vivo but not when added directly to the assay. The increments in activity caused by PGE(1) and NaF were the same in CT-treated and control mucosa. Cyclic 3',5'-AMP concentration in rabbit ileal mucosa was increased 3.5-fold after a 2 hr preincubation with CT in vitro. Phosphodiesterase activity in the crude membrane fraction of the mucosa was unaffected by either CT or PGE(1). A variety of other agents including insulin, glucagon, parathormone, thyroid-stimulating hormone, L-thyroxine, thyrocalcitonin, vasopressin, and epinephrine all failed to change adenyl cyclase activity. It is concluded that CT and certain prostaglandins produce small intestinal fluid secretion by increasing mucosal adenyl cyclase activity, thereby stimulating an active secretory process.

573 citations


Journal ArticleDOI
TL;DR: The first in a now-classic series of five articles in which Rodbell concluded that GTP was likely the active biological factor in separating glucagon, a hormone that can act to increase blood glucose levels, from the cell's receptor, had important implications for the treatment of various disorders and diseases.

486 citations


Journal ArticleDOI
TL;DR: In this article, Rodbell established that the glucagon receptor is involved in the activation of the enzyme adenyl cyclase and that intracellular signaling is achieved through specific GTP-binding heterotrimeric G-proteins.

475 citations


Journal ArticleDOI
TL;DR: Plasma membranes prepared from rat livers treated with digitonin or phospholipase A under conditions which result in substantial loss of glucagon- Stimulated adenyl cyclase activity but no loss of fluoride-stimulated activity are thought to reflect extensive modification of the structures responsible for hormone sensitivity without destruction of the catalytic component of the adeny cyclase system.

458 citations


Journal ArticleDOI
24 Dec 1971-Science
TL;DR: The results suggest that the physiological effects of dopamine in the ganglion, and possibly elsewhere in the nervous system, may be mediated by stimulating the synthesis of adenosine 3',5'-monophosphate.
Abstract: An adenyl cyclase activated by low concentrations of dopamine has been found in the mammalian superior cervical sympathetic ganglion The existence of this enzyme may account for the increased amount of adenosine 3',5' monophosphate associated with synaptic activity in the ganglion The results suggest that the physiological effects of dopamine in the ganglion, and possibly elsewhere in the nervous system, may be mediated by stimulating the synthesis of adenosine 3',5'-monophosphate

396 citations


Journal ArticleDOI
TL;DR: Results are consistent with an initial action of PHA at this level, and the early changes in cyclic AMP concentration appear to precede previously reported metabolic changes in PHA-stimulated cells.
Abstract: We have studied cyclic adenosine 3′,5′-monophosphate (cyclic AMP) concentrations in human peripheral blood lymphocytes after stimulation with phytohemagglutinin (PHA), isoproterenol, prostaglandins, and aminophylline. Purified lymphocytes were obtained by nylon fiber chromatography, and low speed centrifugation to remove platelets. Cyclic AMP levels were determined by a highly sensitive radioimmunoassay. At concentrations of 0.1-1.0 mmoles/liter isoproterenol and aminophylline produced moderate increases in cyclic AMP concentrations, whereas prostaglandins produced marked elevations. High concentrations of PHA produced 25-300% increases in cyclic AMP levels, alterations being demonstrated within 1-2 min. The early changes in cyclic AMP concentration appear to precede previously reported metabolic changes in PHA-stimulated cells. After 6 hr cyclic AMP levels in PHA-stimulated cells had usually fallen to the levels of control cells. After 24 hr the level in PHA-stimulated cells was characteristically below that of the control cells. Adenyl cyclase, the enzyme which converts ATP to cyclic AMP, was measured in lymphocyte homogenates. Adenyl cyclase activity was rapidly stimulated by fluoride, isoproterenol, prostaglandins, and PHA. Since adenyl cyclase is characteristically localized in external cell membranes, our results are consistent with an initial action of PHA at this level.

295 citations


Journal ArticleDOI
TL;DR: A method is described for separating 3′,5′-nucleoside monophosphates from other nucleotides on columns of aluminum oxide and an α-32P-labeled nucleoside triphosphate provides a specific precursor for use in assaying cyclase enzymes.

257 citations


Journal ArticleDOI
TL;DR: In vitro studies parallel in vitro studies with cerebellar slices which show an increase in cyclic AMP levels with NE, as well as histamine, which point to a possible transsynaptic modulation of adenyl cyclase activity.

242 citations


Journal ArticleDOI
TL;DR: A rapid, simple, one-step procedure is described for the separation of cyclic AMP from ATP, ADP, AMP, and P 1 based on selective adsorption of all nucleotides except cyclicAMP on hydrous aluminum oxide.

231 citations


Journal ArticleDOI
TL;DR: The present findings suggest that glucagon and fluoride ion activate adenyl cyclase by different mechanisms, suggesting that these agents react through different molecular entities in this complex enzyme system.

Journal ArticleDOI
TL;DR: Phosphorylation of the synaptic plasma membrane fractions indicated that some membrane proteins were at least comparable to and possibly better than the histones in effectiveness as substrate for the enzyme.

Journal ArticleDOI
TL;DR: It is concluded that the thyrotropin "receptor" and the adenyl cyclase (F- activation) are distinct entities, and it is suggested that prostaglandin E1 uses a different receptor than thyrotopin.

Journal ArticleDOI
TL;DR: The addition of phosphatidylinositol totally restored the norepinephrine activation of the solubilized adenylate cyclase, whereas, phosphatodylserine and phosph atidylethanolamine did not.

Journal ArticleDOI
TL;DR: Sodium fluoride-induced adenyl cyclase activity of rat and rabbit cerebral cortex homogenates is inhibited by a wide range of concentrations of lithium, suggesting that the psychosedative effects of lithium may be related to the inhibition of brain adenYL cyclase.

Journal ArticleDOI
TL;DR: It was concluded that adenyl cyclase exists in its membranous environment in a state of inhibition and treatment with detergents or freezing of the tissue could lead to nonspecific changes in membrane structure which cause reduction of the inhibition and increased catalytic activity.

Journal ArticleDOI
TL;DR: It is suggested that cyclic AMP is an intermediate in the light- and dark-induced changes in sodium permeability in the photoreceptor cell.
Abstract: Tomita has shown by electrophysiological measurements that the photoreceptors of the vertebrate retina are depolarized (excited) by darkness and hyperpolarized (inhibited) by light. Excitation is accompanied by an increase, and inhibition by a decrease, in the sodiumion permeability of the receptor cell. The retinal-rod outer segments of the frog contain an adenyl cyclase that is active in darkness and inactivated by light. This cyclase has a specific activity ten-times higher than the activity in previously described tissues. It is suggested that cyclic AMP is an intermediate in the light- and dark-induced changes in sodium permeability in the photoreceptor cell.

Journal Article
TL;DR: It is proposed that contact inhibition of growth may be mediated by activation of adenyl cyclase in the cell membranes with subsequent rise in cellular cAMP, and higher level in the nontumorigenic cultures.
Abstract: Summary Adenosine 3′,5′-cyclic monophosphate (cAMP) and cyclic nucleotide phosphodiesterase were assayed in strain L cells from the 2nd to the 6th day of incubation. cAMP increases markedly when the culture reaches confluency on the 4th day of incubation. The phosphodiesterase increased in a similar but less dramatic manner. cAMP and cyclic nucleotide phosphodiesterase were measured in two nontumorigenic cell lines and five tumorigenic cell lines at confluency. The level of cAMP is higher in the nontumorigenic cultures and is related to the population-doubling time of the cells. It is proposed that contact inhibition of growth may be mediated by activation of adenyl cyclase in the cell membranes with subsequent rise in cellular cAMP.

Journal ArticleDOI
TL;DR: It was concluded that the stimulatory effect of F- was due to an increase in Vmax, there being no significant effect on the Km for substrate (MgATP) or on the apparent Ka for Mg++.

Journal Article
TL;DR: PGE1 and isoproterenol at maximally effective doses produced no additive effect on cyclic AMP synthesis, suggesting that both drugs stimulate different receptors, but activate the same adenyl cyclase enzyme.
Abstract: The effects of drugs on adenyl cyclase activity in isolated human leukocytes were examined in two ways: 1) conversion of radioactive adenosine triphosphate to cyclic adenosine monophosphate (AMP) by broken cell preparations; and 2) accumulation of radioactive cyclic AMP within intact leukoytes after preincubation with radioactive adenine. Synthesis of cyclic AMP was increased by prostaglandin-E1(PGE1) and selected catecholamines, but not by glucagon or adrenocorticotropin. Responses to catecholamines were characteristic of a beta adrenergic receptor, both in order of potency or agonists [isoproterenol = epinephrine > norepinephrine > phenylephrine (= 0)] and in being competitively inhibited by beta adrenergic antagonists (propranolol and MJ-1999) but not by alpha antagonists (phentolamine and Dibenamine). The effect of PGE1 was not blocked by propranolol. PGE1 and isoproterenol at maximally effective doses produced no additive effect on cyclic AMP synthesis, suggesting that both drugs stimulate different receptors, but activate the same adenyl cyclase enzyme. The maximum effect of isoproterenol is only about 15% of that produced by PGE1; the possibility that isoproterenol stimulates adenyl cyclase in a fraction of the leukocytes (or specific types of leukocytes) has not been ruled out.

Journal ArticleDOI
TL;DR: It is proposed that Ca(++) inhibits the core enzyme directly since at the minimally inhibitory concentration (0.1 mm), CaCl(2) reduced to an equivalent extent basal activity, the response to graded doses of vasopressin and theresponse to maximal stimulatory concentrations of NaF.
Abstract: Adenyl cyclase activity was assayed in crude homogenates of the renal cortex, medulla, and papilla of the golden hamster. The specific activity (moles C-AMP/unit of time per mg protein of tissue) of the enzyme under basal conditions, was greatest in papilla, somewhat lower in medulla, and least in cortex. On an absolute scale, the sensitivity to vasopressin was greater in the medullary and papillary than in the cortical homogenates. In addition, at concentrations of 0.1-1.0 mm, CaCl(2) inhibited the enzyme in the order papilla > medulla > cortex. These results imply the existence of distinct differences in the composition of the adenyl cyclase-receptor complex in various parts of the kidney. We proposed that Ca(++) inhibits the core enzyme directly since at the minimally inhibitory concentration (0.1 mm), CaCl(2) reduced to an equivalent extent (a) basal activity, (b) the response to graded doses of vasopressin (0.5 to 50.0 mU/ml) and (c) the response to maximal stimulatory concentrations of NaF (10 mm). Prostaglandin E(1) (PGE(1) = 10(-7)m) had no effect on either basal adenyl-cyclase activity or the response to 10 mm NaF in medullary and papillary homogenates. 7-Oxa-13-prostynoic acid (10(-4)m) similarly had no effect under basal conditions or on stimulation with NaF in medullary homogenates. Both fatty acids, however, inhibited the enzymic response to vasopressin, particularly at low concentrations of the peptide. The straight-chain fatty acid, 11-eicosanoic acid (10(-7)m), was inactive on basal activity or on the response to vasopressin. The possibility that PGE(1) modifies the coupling mechanism between the core enzyme and the hormone-specific receptor is discussed.

Journal ArticleDOI
TL;DR: It is demonstrated that the addition of phosphatidyl-serine to a solubilized preparation freed of detergent, completely restores the glucagon responsiveness of adenyl cyclase.

Journal ArticleDOI
TL;DR: During the first ten minutes after the beginning of a continuous exposure of rat thymocyte populations (maintained in vitro) to epinephrine, there is an increase in the cellular concentration of cyclic AMP.
Abstract: : During the first ten minutes after the beginning of a continuous exposure of rat thymocyte populations (maintained in vitro) to epinephrine, there is an increase in the cellular concentration of cyclic AMP. The hormone also increases the activity of a crude preparation of the thymocyte's cyclic AMP-forming enzyme, adenyl cyclase. Between 30 and 45 minutes after the beginning of exposure to epinephrine, an additional part of the cell population begins to synthesize deoxyribonucleic acid (DNA). These changes are finally followed two to four hours later by an increase of the flow of cells into mitosis. Since cyclic AMP itself is known to stimulate both the initiation of DNA synthesis and thymocyte proliferation, and since the mitogenic action of epinephrine is shown to be potentiated by caffeine and inhibited by imidazole, it is concluded that the mitogenic action of this hormone is mediated by the cyclic nucleotide. (Author)

Journal ArticleDOI
TL;DR: The cyclase of the tumor as well as of the adrenals was stimulated by adrenocorticotropic hormone (ACTH) over similar concentration ranges, and the tumor enzyme was also stimulated by epinephrine, norepinephrine, and thyroid-stimulating hormone (TSH).
Abstract: Properties of adenyl cyclase of normal adrenals and of a corticosterone-producing adrenal cancer of the rat have been compared. Enzyme activity was found in all particulate fractions of both tissues. The cyclase of the tumor as well as of the adrenals was stimulated by adrenocorticotropic hormone (ACTH) over similar concentration ranges. Unexpectedly, the tumor enzyme was also stimulated by epinephrine, norepinephrine, and thyroid-stimulating hormone (TSH). These hormones produced a dose-related effect over a concentration span that was comparable with that for ACTH. The tumor cyclase was not responsive to angiotensin Il, vasopressin, glucagon, insulin, growth hormone, parathyroid hormone, and thyrocalcitonin. ACTH was the only hormonal preparation that stimulated normal adrenal cyclase. These findings are compatible either with the possibility that the adenyl cyclase receptor of the tumor has undergone structural alteration with a consequent loss of specificity for ACTH or with the possibility that the tumor possesses several cyclase regulatory receptors.

Journal ArticleDOI
TL;DR: Adenylate cyclase of homogenates or lysates of mouse and rat lymphoid tissues was activated by the addition of fluoride ion, epinephrine, or norepinephrine, but not by any of several other hormones.
Abstract: Adenylate cyclase of homogenates or lysates of mouse and rat lymphoid tissues was activated by the addition of fluoride ion, epinephrine, or norepinephrine, but not by any of several other hormones. The catecholamine stimulation was characterized as β-adrenergic. This activity was localized in the small lymphoid cells, was greater in thymic than in splenic or mesenteric node cells, and also was greater in mouse than in rat cells. Catecholamine-stimulated activity of mouse thymocytes remained constant from the 17-19th day of fetal development to 5 weeks after birth; it subsequently decreased to about one-half of the activity by 7-8 weeks. Similar decreases with age occurred in mouse spleen and rat thymus. In contrast, the glucagon-stimulated activity of rat liver increased during a similar period. Hypophysectomy of rats at 3 weeks did not influence the amount of cyclase activity of thymocytes assayed at 7 weeks. When intact mouse thymocytes were first incubated in a culture medium at 37°C with epinephrine or norepinephrine, a second addition of catecholamine after cell lysis no longer stimulated the enzymes. This loss of stimulation was selective, since basal activity and stimulation by fluoride ion were not affected. Incubation of intact cells with phytohemagglutinin markedly decreased the activity of lysates, whether assayed in the presence or absence of catecholamine or fluoride. In contrast, phytohemagglutinin added directly to the assay had no effect. No alternations occurred in adenylate cyclase activity as a result of the incubation of a 1:1 mixture of thymocytes from two strains of mice selected for the capacity of the cells to produce a mixed lymphocyte response.


Journal ArticleDOI
TL;DR: With most lines, adenyl cyclase activity remained constant during culture for 2 to 3 months, however, in rat embryo fibroblasts and SV40 (3T3) cells, the fluoride-stimulated enzyme levels fell with successive passages.

Journal ArticleDOI
TL;DR: Results in the inner medulla are consistent with the hypothesis that PGE(1) can compete with vasopressin for adenyl cyclase-binding sites, but the findings in the outer medulla suggest the situation is more complex.
Abstract: A B S T R A C T Vasopressin increased adenyl cyclase activity in homogenates of both inner and outer renal medulla of the rat. It also increased the concentration of cyclic 3',5'-adenosine monophosphate (AMP) in slices of both inner and outer medulla but not in renal cortex. In the inner medulla, a concentration of prostaglandin E1 (PGE1), which was ineffective by itself significantly reduced the stimulation of adenyl cyclase activity and cyclic AMP concentration induced by vasopressin. These results are consistent with the hypothesis that PGE1 can compete with vasopressin for adenyl cyclase-binding sites. However, the findings in the outer medulla suggest the situation is more complex. Although 10 M PGE1 had no effect by itself and inhibited the vasopressin-induced elevation of cyclic AMP, larger amounts of PGE1 increased both adenyl cyclase activity and cyclic AMP levels. The maximum effect on the latter parameter was at least 6 times as great as that of maximum amounts of vasopressin.

Journal ArticleDOI
TL;DR: The evidence for hormone receptors in relation to the adenyl cyclase system is reviewed, a detailed description of the interaction of ACTH with its physiological receptors in adrenal cortical tissue is presented, and the ACTH system is used as a model to speculate further on the nature and role of hormone receptors.
Abstract: Since the original demonstration that epinephrine stimulates the adenyl cyclase system of liver cells,l numerous hormonal substances have been shown to act by altering the activity of cell membrane-bound adenyl cyclase in their respective target tissues.2 Two general phenomena related to the interaction of hormones and adenyl cyclase have become clear. First, the enzyme is very widespread, occurring in almost all mammalian tissues examined except erythrocytes. Second, in any given tissue only a very limited number of hormones (often just one) influence the activity of the enzyme. This latter phenomenon is referred to as “hormone specificity.” Thus, only TSH stimulates thyroid adenyl cyclase,:’ only ACTH stimulates adrenal adenyl c y ~ l a s e , ~ and so forth. The specific cell membrane structures responsible for this great hormone specificity in each tissue have been referred to as receptors.5 Whereas the catalytic function of converting ATP to cyclic AMP resides in the adenyl cyclase enzyme, the discriminatory function of permitting only a narrow range of hormones to interact with the enzyme resides in the receptors. The exact relationship of these “catalytic” and receptor functions of the adenyl cyclase system has been the subject of much speculation.5 In fact, it is only recently that physicochemical proof of the existence of these receptors has been obtained. The purpose of this paper is to (1) review briefly the evidence for hormone receptors in relation to the adenyl cyclase system, (2) present a detailed description of the interaction of ACTH with its physiological receptors in adrenal cortical tissue, and (3 ) use the ACTH system as a model to speculate further on the nature and role of hormone receptors.

Journal ArticleDOI
TL;DR: Combined maximal doses of histamine and norepinephrine produced completely additive effects on the activation of adenyl cyclase, which suggests that at least two separate adenYL cyclase systems are present in the heart, each responsive to one of these hormones.
Abstract: A B S T R A C T Histamine has positive inotropic and chronotropic effects on the heart which are not abolished by beta adrenergic-blocking agents. Since the positive inotropic and chronotropic effects of other hormones on the heart are thought to be mediated by cyclic 3',5'-AMP, we examined the effect of histamine on adenyl cyclase in particulate preparations of guinea pig, cat, and human myocardium. Histamine at the peak of its dose-response curve, 3 X 10'moles/liter, produced approximately a 300% increase in cyclic 3',5'-AMP accumulation in the guinea pig, 60% in the cat, and 90% in the human heart particles. Half-maximal activity for the histamine mediated activation of adenyl cyclase in the guinea pig was 9 X 10-6moles/liter, almost identical with that observed for norepinephrine in the same preparation. DL-Propranolol, 1 X 10'moles/liter, did not abolish the activation of adenyl cyclase produced by histamine but did abolish the activation produced by norepinephrine. In contrast, diphenhydramine hydrochloride, Benadryl, 8 X 105moles/liter, abolished the activation of adenyl cyclase by histamine but not that produced by norepinephrine. These data suggest that there are at least two receptor sites in guinea pig heart mediating the activation of adenyl cyclase, one responsive to histamine, the other to norepinephrine. In addition, combined maximal doses of histamine and norepinephrine produced completely additive effects on the activation of adenyl cyclase, which suggests that at least two separate adenyl cyclase systems are present in the heart, each responsive to one of these hormones. However, definitive proof would require physical separation of the two enzymes.