Effects Of Exogenous Cyclic AMP Research Articles

Effect of cAMP-dependent protein kinase inhibitor (H89) on kinematic parameters of ram epididymal sperm in vitro

The present study investigated the effects of exogenous cyclic AMP (cAMP)-dependent protein kinase inhibitor (H89) on kinematic parameters of epididymal ram sperm in vitro. After incubation of epididymal sperm samples for 45 and 90 min in the presence of H89 (0.5, 5, and 50 μM), motion parameters were evaluated by computer-aided sperm analysis. At 50 μM concentration of H89, there was a significant decrease in the fast progressive motility (at 45 and 90 min), and also most of the kinematical parameters such as curvilinear velocity, straight line velocity, average path velocity, mean angular degree (MAD), amplitude of lateral head displacement, beat cross-frequency, linearity, and straightness but not in wobble (WOB) at 45 min. After 90 min all kinematic parameters were significantly decreased at 50 μM concentration except MAD and WOB. In conclusion, high concentrations of exogenous cAMP-dependent protein kinase inhibitor (H89) decrease the motion parameters of ram epididymal sperm.

Ca 2+-dependent protein phosphorylation associated with microsomal fraction of rat pancreas

1. 1. Microsomes isolated from cat pancreas were incubated with [γ- 32P]ATP in the presence or absence of Ca 2+. Following fractionation of phosphoproteins by sodium dodecyl sulfate-polyacrylamide absence of Ca 2+. Following fractionation of phosphoproteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis a single microsomal protein with an apparent molecular mass of 77,000 dalton (77K) was found to be phosphorylated in a Ca 2+-dependent mechanism. 2. 2. Maximal phosphate incorporation into the 77K protein was observed at 10 −6mol/l[Ca 2+ and was 4-fold higher than in the absence of Ca 2+. 3. 3. The 77K phosphoprotein showed characteristic of a stable phosphoester rather than an acyl phosphate. 4. 4. Measurable phosphate incorporation into the 77K protein was noted 5 s following addition of [γ- 32P]ATP and reached maximum at 9–10th min. 5. 5. The lack of effect of exogenous cyclic AMP, cyclic AMP-dependent protein kinase, calmodulin, the calmodulin antagonist trifluoperazine, leupeptin and the suppression of phosphorylation by some phospholipid-interacting drugs suggested that the 77K protein is a substrate for cyclic AMP- and calmodulin-independent, Ca 2+-activated phospholipid-sensitive kinase activity. 6. 6. Centrifugation of the pancreatic homogenate in a ficoll-sucrose densitu gradient indicated that both the 77K protein and enzyme were associated in a fraction enriched in rough endoplasmic reticulum.

Effect of fenoterol on secretions of an isolated single submucosal gland from the trachea.

The mechanism of secretion of airway submucosal glands is not known. Although the existence of cyclic AMP in the submucosal gland has been reported, the effect of stimulation of the gland by exogenous cyclic AMP has been denied. We hypothesize that the negative effect of exogenous cyclic AMP is due to the modified influence of ciliated epithelium on the submucosal gland, because epithelium releases chemical mediators. We isolated a single submucosal gland from a cat trachea. We measured mucous glycoprotein labeled with [3H]-glucosamine from the isolated submucosal glands. With stimulation of dibutyryl cyclic AMP, a percent increase of the secretion above control was about 3 times in isolated glands. Methacholine and fenoterol secreted mucus largely in isolated glands. We suggest that cyclic AMP increased the secretion in the glands. The difference of quantity of secretions between methacholine and fenoterol has been investigated further.

Effects of Exogenous Cyclic AMP on Growth Characteristics and Radiation Response of Reuber H35 Hepatoma Cells

Reuber H35 rat hepatoma cells, clone KRC, were used to study the effect of cyclic AMP on radiation-induced cell death. Treatment of logarithmically growing cultures with 0.5 mM cAMP for 17 hr prior to irradiation resulted in a decreased cell survival. Similar results were obtained with cultures irradiated after treatment with Bt2cAMP. Treatment of H35 cells with cAMP or Bt2cAMP caused inhibition of their proliferation and resulted in an accumulation of cells in early S phase and a depletion of G2-phase cells. In synchronized cultures cells were relatively radioresistant during their S phase. In addition to single-dose treatment with X rays, the effect of Bt2cAMP on radiation-induced cell death was studied during fractionated irradiation with 2.5 Gy per day. This fractionated irradiation resulted in a dose-reduction factor of 1.6 at the 10% survival level and a 10-fold decrease in the surviving cell population due to the cooperative effects of Bt2cAMP on growth rate and radiation survival. The effect of cAMP on radiation-induced mitotic delay was also studied. It appeared that whereas cAMP had no effect on the progression of G2 cells into mitosis, it prevented cells from recovery from the X-ray mitotic delay in G2.

Effect of ptsI and ptsH mutations on initiation of transcription of the Escherichia coli lactose operon.

Mutations in the pts genes (which code for the enzyme I and HPr protein - the general components of the phosphoenolypyruvate-dependent phosphotransferase system) lead to decreases in enzyme-inducible synthesis at the level of transcription. The intracellular content of cyclic AMP in the ptsIH mutant was severely diminished, while the ptsH bacteria contain the same amounts of this nucleotide as the wild-type cells. Nevertheless expression of the lac operon was diminished in the ptsH as well as in the ptsIH mutant. The exogenous cyclic AMP did not prevent repression of beta-galactosidase synthesis in a delta cya ptsI mutant in a wide range of concentrations in the growth medium (from 0.05 mM to 5 mM). The combination of ptsI or ptsH mutations with rpoC1 (synthesis of thermosensitive beta' subunit of RNA polymerase) leads to greater disturbance of beta-galactosidase production at the nonpermissive temperature than demonstrated in the pts+ rpoC1 strain. The stimulatory effect of exogenous cyclic AMP was more pronounced in pts rpoC1 than in pts+ rpoC1 bacteria. The data presented confirm the hypothesis that pts mutations alter the function of CRP in initiation of transcription.

Cyclic AMP interferes with pattern formation in the cyanobacteriumAnabaena variabilis

The impressive array of regulatory roles of adenosine 3',5'-cyclic monophosphate (cyclic AMP) in heterotrophic organisms has no counterpart among autotrophs. Although cyclic AMP appears to be essential for differentiation of chloronema cells in the moss Funaria hygrometrica [1,2], little is known about the functions, if any, of this nucleotide in other plants [3.4] and eukaryotic algae [5] where it has been detected. We recently observed that cyclic AMP is synthesized by the nitrogen-fixing blue-green alga (cyanobacterium) Anabaena variabilis, and that intracellular concentrations increased 3 to 4-fold during nitrogen starvation [6]. These results have been corroborated by Francko and Wetzel [7], who also detected cyclic AMP in Synechococcus leopoliensis and Microcystis aeruginosa [8]. Cyanobacteria have also been shown to possess adenylate cyclase activity [9] and cyclic nucleotide phosphodiesterase activity [10]. To further evaluate the significance of cyclic AMP in A. variabilis, we have measured the effects of exogenous cyclic AMP on the differentiation of heterocysts, cells which are the site of nitrogen fixation in Anabaena (see [ 11] for review). Our results, showing that exogenous cyclic AMP specifically alters the pattern of heterocyst spacing in A. variabilis, provide further evidence for a possible regulatory function for cyclic AMP in this organism.

Regulation of development in Dictyostelium discoideum. IV. Effects of ions on the rate of differentiation and cellular response to cyclic AMP.

The effects of ionic environment on both the intrinsic rate of differentiation and the response to exogenous cyclic AMP in Dictyostelium discoideum have been examined. K+ specifically inhibits the rate of early development when present at concentrations > 15 mM. Na+ does not inhibit at concentrations up to 25 mM, and can partially overcome K+ inhibition. The maximum rate of development also depends upon the presence of adequate levels of extracellular Ca++. The effect of exogenous cyclic AMP on the rate of development is inhibited by the absence of Ca++, and/or the presence of high concentrations of K+. Under optimal ionic conditions, the only effect of exogenous cyclic AMP on early developments of K+. Under optimal ionic conditions, the only effect of exogenous cyclic AMP on early development is a specific inhibition. The implications of these results for current models of early developmental regulation are discussed.

PH-Dependence of water and solute transport in toad urinary bladder

Stimulation of urea and water transport by vasopressin (ADH) appears to occur via independent pathways. We examined the effects of altering serosal or mucosal bath pH on transport of water, urea, and sodium. Compared to bladders with a serosal bath pH of 7.4 to 8.0, reducing the serosal bath pH to 6.8 led to a 60% fall in ADH-stimulated osmotic water flow, without decreasing the permeability of urea. Raising the serosal pH to 9.5 had the opposite effect: urea permeability was inhibited by 40% without altering water flow. Exogenous cyclic AMP-stimulated water and urea permeabilities were not dissociated, but were changed in the same direction by alterations in serosal pH: serosal acidification enhanced the effect of exogenous cyclic AMP on both urea and water, whereas the cyclic AMP effect on both was diminished by serosal alkalinization. This was especially marked for urea, suggesting that an alteration in the urea response to cyclic AMP may be particularly important in defining vasopressin-stimulated urea permeability as the serosal bath pH is altered. Mucosal acidification increased short circuit current but decreased both the urea and water response to ADH and 8-bromo-cyclic AMP. The response to cyclic AMP was less consistent. Mucosal alkalinization did not cause significant changes in either basal or stimulated transport. The data demonstrate distinct and separable effects of bath pH alterations on each of the transport systems examined.

Relation between catecholamine‐induced cyclic AMP changes and hyperpolarization in isolated rat sympathetic ganglia

1. The effect of catecholamines on cyclic adenosine 3'5'-monophosphate (cyclic AMP) production in isolated rat superior cervical ganglia has been measured under experimental conditions in which they also produce ganglion hyperpolarization.2. (+/-)Isoprenaline (1 muM) increased cyclic AMP levels by 8-100 times after 15 min incubation at 25 degrees C. Half-maximal stimulation occurred at about 0.03 muM. This was due to stimulation of beta-receptors, since it was prevented by 1 muM-propranolol but not by 1 muM-phentolamine.3. The alpha-agonists phenylephrine (100 muM), dopamine (100 muM) and clonidine (1 muM) did not produce a detectable increase in ganglionic cyclic AMP. Dopamine (100 muM) was also ineffective at 37 degrees C in the presence of 10 mM-theophylline.4. Exogenous cyclic AMP (0.01-1 mM) hyperpolarized the ganglion. This effect was replicated by other adenosine compounds, most effectively by adenosine and by adenosine 5'-monophosphate, and was antagonized by theophylline. Dibutyryl cyclic AMP was weaker than cyclic AMP.5. Neither theophylline nor the non-xanthine phosphodiesterase inhibitor, Ro 20-1724, enhanced the hyperpolarizing actions of noradrenaline or dopamine.6. Since catecholamine-induced hyperpolarization of the isolated rat ganglion is induced via alpha-receptors, whereas cyclic AMP-production is induced via beta-receptors, it is concluded that cyclic AMP is unlikely to mediate the hyperpolarization. The effect of exogenous cyclic AMP may be due to an action on external adenosine-receptors.

Separation of mitogen-induced suppressor and helper cell activities during inhibition of interferon production by cyclic AMP

The effect of exogenous cyclic AMP on mitogen-induced suppression and enhancement of the in vitro plaque-forming cell (PFC) response and on mitogen induction of immune interferon (also called type II) in cultures was examined. Mitogen induction of immune interferon was quantitatively associated with mitogen-induced suppressor activity, and cyclic AMP blocked both the suppressor activity and the production of immune interferon in mouse (C57B1/6) spleen cell cultures. The evidence is as follows: (a) The concentrations of dibutyryl cyclic AMP that blocked T-cell mitogen (staphylococcal enterotoxin A) suppressor activity were the same as those that blocked mitogen induction of immune interferon. (b) The blocking action of dibutyryl cAMP on both the suppressor and interferon effects of mitogen was a function of the time of dibutyryl cAMP addition to cultures relative to mitogen addition. (c) A dramatic immunoenhancing effect of mitogen occurred in the presence of dibutyryl cAMP under conditions that blocked production of immune interferon. Specifically, mitogen-induced helper cell function is dramatically enhanced in the presence of dibutyryl cyclic AMP, if the mitogen is added to cultures 24 to 48 hr after SRBC and dibutyryl cyclic AMP. Dibutyryl cyclic GMP did not affect the mitogen- or cyclic AMP-induced effects under the conditions of our test system. Under the conditions described here, then, cyclic AMP appears to selectively block suppressor cell activity while allowing or aiding mitogen-induced helper cell activity. It is possible that the immune response is a reflection of the ratio of helper to suppressor activities in the system.

Cyclic AMP in action of antidiuretic hormone: effects of exogenous cyclic AMP and its new analogue

Exogenous cyclic 3',5'-adenosine monophosphate (cAMP) stimulates the effect of the antidiuretic hormone, vasopressin (VP), only in pharmacologic quantities and results have often been inconsistent. The present study examined the ability of a new analogue, 8-[p-Cl-phenylthio]cyclic 3',5'-adenosine monophosphate (C1-PheS-cAMP) to mimic the effect of VP, both biochemically (protein kinase activation) and functionally (hydrosomatic response of perfused collecting tubules) in mammalian kidney tissue. C1PheS-cAMP was found to be about 100 times as effective as cAMP both biochemically and functionally. The increased effectiveness of C1PheS-cAMP is probably is probably due to a greater permeability across the cell membrane and to the resistance of C1PheS-cAMP to enzymatic degradation, Cyclic AMP phosphodiesterase inhibition was observed with C1PheS-cAMP, but its contribution to overall effect was minor. C1PheS-cAMP was found to be more effective than exogenous vasopressin, an effect probably due primarily to its resistance to catabolism. The results provide further new evidence that cAMP and protein kinase are involved in the cellular action of vasopressin. C1PheS-cAMP proved to be a useful tool in the study of hormone action, especially in steps subsequent to cAMP generation.

The adenosine-like effect of exogenous cyclic AMP upon nucleotide and PP-ribose-P concentrations of cultured human lymphoblasts

FEBS LettersVolume 66, Issue 1 p. 102-106 Full-length articleFree Access The adenosine-like effect of exogenous cyclic AMP upon nucleotide and PP-ribose-P concentrations of cultured human lymphoblasts Floyd F. Snyder, Floyd F. Snyder Division or Rheumatology, Department of Medicine, University of California, San Diego, La Jolla, California, 92093 USASearch for more papers by this authorJ.Edwin Seegmiller, J.Edwin Seegmiller Division or Rheumatology, Department of Medicine, University of California, San Diego, La Jolla, California, 92093 USASearch for more papers by this author Floyd F. Snyder, Floyd F. Snyder Division or Rheumatology, Department of Medicine, University of California, San Diego, La Jolla, California, 92093 USASearch for more papers by this authorJ.Edwin Seegmiller, J.Edwin Seegmiller Division or Rheumatology, Department of Medicine, University of California, San Diego, La Jolla, California, 92093 USASearch for more papers by this author First published: July 01, 1976 https://doi.org/10.1016/0014-5793(76)80595-9Citations: 48AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL References 1 H. Hilz, A. Tarnowski, Biochem. Biophys. Res. Comm., 40, (1970), 973– 981. 2 E. Kaukel, V. Fuhrmann, H. Hilz, Biochem. Biophys. Res. Comm., 48, (1972), 1516– 1524. 3 H. Hilz, E. Kaukel, Mol. Cell. Biochem., 1, (1973), 229– 239. 4 S.S. Solomon, J.S. Brush, A.E. Kitabchi, Science, 169, (1970), 387– 388. 5 B.A. Roller, K. Hirai, V. Defendi, Biochim. Biophys. Acta, 366, (1972), 402– 410. 6 E. Kaukel, H. Hilz, Biochem. Biophys. Res. Comm., 46, (1972), 1011– 1018. 7 D.K. Granner, L. Sellers, A. Lee, C. Butters, L. Kutina, Arch. Biochem. Biophys., 169, (1975), 601– 615. 8 J. Taylor-Papadimitriou, T. Karemfyllis, A. Eukarpidou, G. Karamanlidou, J. Cell. Sci., 19, (1975), 305– 313. 9 W.D. Yushok, J. Biol. Chem., 246, (1971), 1607– 1617. 10 F.F. Snyder, J.F. Henderson, Can. J. Biochem., 51, (1973), 943– 948. 11 H. Green, T.-S. Chan, Science, 182, (1973), 836– 837. 12 E.R. Giblett, J.E. Anderson, F. Cohen, B. Pollara, H.J. Meuwissen, Lancet, 2, (1972), 1067– 1069. 13 J.E. Lever, G. Nuki, J.E. Seegmiller, Proc. Nat. Acad. Sci., 71, (1974), 2679– 2683. 14 K.H. Astrin, Ph. D. Thesis (1973), University of California San Diego 15 F.F. Snyder, J.F. Henderson, S.C. Kim, A.R.P. Paterson, L.W. Brox, Cancer Res., 33, (1973), 2425– 2430. 16 M. Smith, G.I. Drummond, H.G. Khorana, J. Amer. Chem. Soc., 83, (1961), 698– 706. 17 F.F. Snyder, J.F. Henderson, J. Biol. Chem., 248, (1973), 5899– 5904. 18 J.F. Henderson, M.K.Y. Khoo, J. Biol. Chem., 240, (1965), 2349– 2357. 19 A. Hershko, A. Razin, J. Mager, Biochim. Biophys. Acta, 184, (1969), 64– 76. 20 W.R.A. Osborne, N. Spencer, Biochem. J., 133, (1973), 117– 123. 21 R.P. Agarwal, S.M. Sagar, R.E. Parks Jr., Biochem. Pharmacol., 24, (1975), 693– 701. 22 D.E. Atkinson, Biochemistry, 7, (1968), 4030– 4034. 23 H.J. Schaeffer, C.F. Schwender, J. Med. Chem., 17, (1974), 6– 8. 24 C.M. Smith, L.J. Fontenelle, M. Lalanne, J.F. Henderson, Cancer Res., 34, (1974), 463– 467. 25 G. Tyrsted, A.C. Sartorelli, Biochim. Biophys. Acta, 155, (1968), 619– 622. Citing Literature Volume66, Issue1July 01, 1976Pages 102-106 ReferencesRelatedInformation

Stimulation of cartilage macromolecule synthesis by adenosine 3′,5′-monophosphate

The role of cyclic AMP in the regulation of cartilage macromolecule synthesis in vitro was studied in pelvic cartilage from 10–12 day chick embryos. Incubation of cartilages in medium containing 0.5 mM cyclic AMP resulted in a 30% inhibition of 35SO 4 −2, [ 3H]leucine and [ 3H]uridine incorporation into proteoglycan, total protein and RNA, respectively. Higher concentrations of cyclic AMP had no greater effects. In contrast, butyrylated cyclic AMP derivatives (0.5–5.0 mM) added to the incubation medium stimulated (50–100%) the incorporation of these radiolabeled precursors into cartilage macromolecules. Theophylline, in concentrations (0.1–0.5 mM) which raise intracellular cyclic AMP, also increases the incorporation of radiolabeled precursors into macromolecules. The data indicate that exogenous cyclic AMP and butyrylated cyclic AMP derivatives have paradoxical effects on cartilage macromolecule synthesis. Butyrylated cyclic AMP derivatives, not exogenous cyclic AMP, mimic the effects of intracellular cyclic AMP. Incubation of embryonic chicken cartilage with exogenous cyclic AMP results in the extracellular degradation of the cyclic AMP to adenosine. Adenosine (0.125 mM) inhibits precursor incorporation into cartilage macromolecules. The metabolism of exogenous cyclic AMP generates sufficient adenosine to account for the observed inhibitory effects of exogenous cyclic AMP on cartilage macromolecule synthesis. Butyrylated cyclic AMP derivatives are not degraded during incubation with cartilage. The data indicate that cartilage is a tissue in which the effect of cyclic AMP is to stimulate anabolic processes.

Effect of Exogenous Cyclic AMP and other Adenine Nucleotides on Neutrophil Chemotaxis and Motility

Cyclic AMP placed in the bottom of the modified Boyden chemotaxis chamber is weakly chemotactic for rabbit peritoneal neutrophils. Dibutyryl-cyclic AMP under similar conditions gives a biphasic response, being weakly chemotactic at low concentrations and inhibiting cell movement at higher concentrations. When mixed with the cells in the upper compartment of the chemotaxis chamber, cyclic AMP and dibutyryl-cyclic AMP enhance neutrophil spontaneous motility; on the other hand, dibutyryl-cyclic AMP mixed with the cells in the upper compartment consistently inhibits neutrophil chemotactic responsiveness, while cyclic AMP has a variable effect on chemotaxis. The effect of ADP and ATP was to inhibit chemotaxis and spontaneous motility. AMP is totally without effect.

Modulation of Stimulated Acid Secretion by Dibutyryl Cyclic Amp in the Dog Stomach

An in vivo canine gastric chamber preparation was used to determine effects of exogenous cyclic AMP and dibutyryl cyclic AMP on histamine-driven acid secretion and the role of blood flow in these responses. Histamine was infused intravenously at a submaximal rate 1.2 mu/kg-min) followed by the intraarterial infusion of either cyclic AMP (0.15 mg/kg-min) or dibutyryl cyclic AMP (0.15 mg/kg-min. Acid secretion and mucosal blood flow (aminopyrine clearance) were measured at 15-minute intervals, and electrical potential difference was measured continuously. Histamine alone decreased the potential difference from 82 mV to 48 mV within 60 minutes, increased aminopyrine clearance from 1.0 ml/min to 2.4 ml/min and increased acid output to 220 muEq/15 min. All values returned towards control following termination of histamine infusion. Dibutyryl cyclic AMP significantly increased histamine stimulated acid secretion to 350 muEq/15 min, but did not alter mucosal blood flow or electrical potential. Cyclic AMP did not change secretory, circulatory or electrical response to histamine. Therefore, dibutyryl cyclic AMP appears to increase histamine-stimulated acid secretion, and the effect is not mediated by increased mucosal blood flow.

Regulation of pancreatic acinar function: effects of cyclic AMP, dibutyryl cyclic AMP, and theophylline in vitro.

The role of cyclic AMP in the regulation of pancreatic acinar function has been assessed by measuring the effects of exogenous cyclic AMP, dibutyryl cyclic AMP, and theophylline on protein synthesis and amylase secretion. The rate at which slices of rat pancreas incorporated leucine into protein did not change as a consequence of treatment with either cyclic AMP or dibutyryl cyclic AMP, nor did the slices alter their rate of amylase secretion. Moreover, theophylline did not enhance the ability of submaximal doses of pancreozymin to stimulate amylase secretion or to suppress protein synthesis. These results fail to demonstrate that cyclic AMP regulates either the synthesis or secretion of pancreatic digestive enzymes but they do not rule out the possibility.

Enhancement of plasminogen activator by vasopressin and a drenaline: a role of cyclic AMP?

To evaluate the hypothesis that cyclic AMP is involved in the regulation of plasminogen activator level in plasma, the effect of exogenous cyclic AMP and of its dibutyryl derivative; of glucagon, lysine vasopressin and adrenaline (activators of adenyl cyclase) have been investigated in human volunteers on their intravenous infusion. Cyclic AMP, dibutyryl cyclic AMP and glucagon had no effect on plasminogen activator. The marked rise induced by adrenaline and lysine vasopressin was not potentiated by the simultaneous administration of theophylline, an inhibitor of phosphodiesterase. These results do not support the hypothesis that cyclic AMP is the final common pathway for the effect on plasminogen activator induced by lysine vasopressin and adrenaline. There is also at present no conclusive evidence that cyclic AMP is involved as a physiologic regulator of the fibrinolytic system.

Inhibition of marrow growth by cyclic AMP.

SummaryCyclic AMP, dibutyryl cyclic AMP, epinephrine, and theophylline inhibited growth of marrow colonies in vitro. The interpretation of this observation is uncertain since various adenine nucleotides had a similar effect and since several substances lowering intracellular cyclic AMP levels were unable to reverse the inhibitory effect of exogenous cyclic AMP.