Increase In Cyclic AMP Levels Research Articles

Positive Inotropic/Vasodilator Agents

Beta-adrenergic sympathomimetic agents such as dobutamine and dopamine, and phosphodiesterase inhibitors such as amrinone, milrinone, and enoxamone, exert a direct positive inotropic effect upon the myocardium by causing an increase in cyclic AMP levels. The phosphodiesterase inhibitors also exert a substantial direct vasodilator effect. Both the sympathomimetic agents and the phosphodiesterase inhibitors can be of value in the acute, short-term management of myocardial failure. At present, the use of these agents for long-term therapy of congestive heart failure is unproven, and remains investigational.

Age-dependence of octopaminergic modulation of flight muscle in the locust

1. The age-dependence of the modulatory actions of octopamine on the dorsal longitudinal flight muscle of the locust,Schistocerca gregaria, is described. 2. The maximal levels of octopaminergic potentiation of the amplitude and of the rates of contraction and relaxation of twitch tension occurred in the late 5th instar hopper and in the adult at the day of moulting. After 1 day of adult life the effects on twitch amplitude and contraction rate had declined to basal levels which were maintained for up to 28 days. The effect on the relaxation rate declined progressively up to day 7 of adult life before reaching a maintained level. Parallel effects were observed for the changes mediated by stimulation of the dorsal unpaired median modulatory octopaminergic neurone (DUMDL) to this muscle. 3. The octopamine-induced changes in the physiological parameters of twitch tension exhibited motor unit specificity as did changes in the octopamine-mediated increases in cyclic AMP levels. 4. Studies on the age-dependence of the modulatory effects of exogenously applied cyclic nucleotides suggest that age-dependent changes occur at multiple sites in the response cascade between the activation of octopamine receptors and the observed physiological responses.

Details of Mode and Mechanism of Action of Denopamine, a New Orally Active Cardiotonic Agent with Affinity for β1-Receptors

We investigated the detailed mode and mechanism of action of denopamine in canine right ventricular muscle. When administered in single doses, denopamine produced a positive inotropic effect (PIE) and increased cyclic AMP levels. Concentration--response curves for both variables were sigmoid. The maximum PIE of denopamine was almost the same as that produced by isoproterenol. However, the maximum increase in cyclic AMP caused by denopamine was approximately 65% of that attained with isoproterenol. When administered cumulatively, concentration--PIE curves for denopamine were bell-shaped, ascending at 10(-7) to 10(-6) M, reaching a maximum at approximately 3 X 10(-6) M which was approximately 75% of that attained with single administrations, and descending at higher concentrations. The bell-shaped curve, which was computer-fitted, suggested that denopamine behaves as an agonist with pD2 of 6.12 and as an antagonist with pD2 of 4.50. The PIE of denopamine was antagonized by atenolol (pA2 = 7.66) but not by ICI 118,551 (less than 10(-7) M), indicating that denopamine is a selective beta 1-receptor agonist. The PIE of denopamine was augmented by 3-isobutyl-1-methyl-xanthine. The PIE and increase in cyclic AMP level caused by 3 X 10(-6) M denopamine were abolished by 10(-6) M atenolol or 3 X 10(-6) M carbachol. From these results we concluded that the PIE of denopamine is derived from the mechanism of action as a selective beta 1-receptor partial agonist. We suggested the close coupling of the beta 1-receptor-adenylate cyclase-cyclic AMP system to positive inotropy.

Involvement of a plasma membrane phosphodiesterase in the negative control of cyclic AMP levels by vasopressin in rat hepatocytes

Vasopressin has been shown previously to lower the glucagon-induced increase of cyclic AMP levels in isolated rat hepatocytes by way of an enhanced phosphodiesterase (EC 3.1.4.17) activity. Five phosphodiesterase inhibitors were tested for their ability to prevent vasopressin from lowering cyclic AMP levels in intact hepatocytes and for their inhibitory effect in vitro on soluble and particulate phosphodiesterase activities partially purified from hepatocytes. Three soluble activities have been separated by DEAE-cellulose chromatography: a phosphodiesterase hydrolyzing both cyclic AMP and cyclic GMP, a form stimulated by cyclic GMP and a cyclic AMP-specific activity. The absence of any statistically significant correlation between the in vivo (in intact cells) and the in vitro (on isolated phosphodiesterases) potencies of the inhibitors does not support a role for the cytosolic phosphodiesterases in mediating the vasopressin-induced decrease in cyclic AMP levels. No statistically significant correlation was observed between the inhibition of the vasopressin effect on cyclic AMP accumulation and the inhibition of phosphodiesterase activity either associated with the native plasma membranes or solubilized from these membranes with 0.4 M NaCl. In contrast, a statistically significant correlation was observed between the degree of inhibition of the vasopressin effect in the intact cells and the degree of inhibition of the intrinsic phosphodiesterase still associated with the plasma membranes after high-salt treatment. These data indicate that a phosphodiesterase activity integral to the plasma membrane is very likely involved in the negative control of cyclic AMP levels by vasopressin.

Inhibition of cardiac slow action potentials by 8-bromo-cyclic GMP occurs independent of changes in cyclic AMP levels.

Cyclic GMP inhibits the slow inward Ca current of cardiac cells. This effect could be due to a cyclic GMP-mediated phosphorylation of the Ca channel (or some protein modifying Ca channel activity), or alternatively, to enhanced degradation of cyclic AMP owing to stimulation of a phosphodiesterase by cyclic GMP. To test the latter possibility, we examined the effect of extracellular 8-bromo-cyclic GMP on cyclic AMP levels in guinea pig papillary muscles, in parallel with electrophysiological experiments. Isoproterenol (10(-6) M) significantly increased the cyclic AMP levels and induced Ca-dependent slow action potentials. Superfusion with 8-bromo-cyclic GMP (10(-3) M) inhibited the slow action potentials induced by isoproterenol. However, muscles superfused with 8-bromo-cyclic GMP had cyclic AMP levels identical to those of muscles superfused with isoproterenol alone. Similarly, 8-bromo-cyclic GMP had no effect on the increase in cyclic AMP levels of muscles treated with forskolin (10(-6) M) or histamine (10(-6) M). We conclude that the inhibitory effect of cyclic GMP on slow Ca channels in guinea pig ventricular cells is not due to a decrease in the cyclic AMP levels. We hypothesize that a cyclic GMP-mediated phosphorylation is the most likely explanation for the Ca channel inhibition observed in this preparation.

Effects of SK&F 94120, an inhibitor of cyclic nucleotide phosphodiesterase type III, on human platelets

Elevation of cyclic AMP concentrations in platelets inhibits agonist-induced responses. Pharmacological interventions which could increase the levels of platelet cyclic AMP include activation of the synthesis of cyclic AMP or inhibition of its breakdown. In this study we have investigated the effects of SK&F 94120 on human platelet phosphodiesterase (PDE) activities separated by ion-exchange chromatography, and studied the effects of this agent on platelet responses caused by the agonists collagen, U44069 and ADP. Four PDE activities were identified from human platelet preparations. The PDE activities found comprised a cyclic GMP selective PDE, a Ca 2+/calmodulin stimulated PDE, a cyclic GMP stimulated PDE and a “low K m ” PDE activity called PDE III by analogy with activities described in other tissues. SK&F 94120 was found selectively to inhibit the “low K m ” PDE III activity with an ic 50 of 10.8 μM, which is consistent with the effects of this compound on cardiac ventricle PDE activities. Exposure of human platelets to SK&F 94120 produced concentration dependent increases in cyclic AMP, showing that inhibition of PDE III activity alone can cause an increase in the level of platelet cyclic AMP. SK&F 94120 also caused an inhibition of platelet responses to collagen, U44069 and ADP. However, SK&F 94120 was much less effective as an inhibitor of aggregation induced by ADP ( ic 50 > 100 μM) than by collagen ( ic 50 = 24.1 μ M ) or by U44069 ( ic 50 = 1.7 μ M ). Isobutylmethylxanthine (IBMX), a non-selective PDE inhibitor, was less effective than SK&F 94120 as an inhibitor of platelet responses for the same measured increase in cyclic AMP levels. M&B 22948 and rolipram, inhibitors of PDE I and PDE IV respectively, had no significant effect on platelet responses. These data suggest that selective inhibition of PDE III is the primary mechanism of action of SK&F 94120 as an inhibitor of agonist-induced platelet responses, and that increased cyclic AMP in the pool controlled by PDE III has important consequences on platelet responses. Moreover, these data suggest that some form of compartmentalization of cyclic AMP and/or PDE activity exists in human platelets.

A new heat-labile cytolethal distending toxin (CLDT) produced by Campylobacter spp.

A new heat-labile toxin cytolethal to CHO, Vero, HeLa, and HEp-2 cells and negative in Y-1 cells has been demonstrated in culture filtrates of many strains of Campylobacter spp. This new toxin was termed a cytolethal distending toxin (CLDT) to reflect the progressive cell distention and eventual cytotoxicity observed in all sensitive tissue cells. CLDT was distinct from previously reported cytotoxins and cholera-like enterotoxin produced by some Campylobacter spp. Since CHO elongation induced by either the Campylobacter enterotoxin or CLDT could not be differentiated after 24 h incubation, continuation of the assay for 96 h was essential for observation of CLDT-associated progressive morphological changes and cytolethal events. Specific assay conditions were required for demonstration of CLDT in Vero, HeLa, and HEp-2 cells. A 31 -fold increase in cyclic AMP levels was observed in CHO cells exposed for 24 h to CLDT of catalase negative or weak positive Campylobacter. CLDT was detected in culture filtrates from strains of Campylobacter jejuni, C. coli, C. fetus subsp. fetus, C. laridis and catalase negative or weak positive Campylobacter. Of 718 strains investigated from both human and animal isolations, 295 (41%) were found to produce this toxin. Campylobacter CLDT was negative in adult rabbit ligated Meal loops, suckling mouse and rabbit skin tests. Hemorrhagic responses were observed in rat ligated ileal loop tests of CLDT-positive cultures. The new CLDT toxin could only be neutralized by homologous rabbit antitoxin, was trypsin-sensitive, non-dialyzable and over 30000 in molecular weight. CLDT-producing strains were observed in many serogroups and biotypes of Campylobacter spp. The strains tested originated from many countries and no clear association of toxigenicity with serotype or biotype could be established.

Role of Cyclic AMP During Histamine Release. Histamine Release is not Directly Related to Increase in Cyclic AMP Levels in Rat Mast Cells Activated by Concanavalin A, Anti-IgE, Antigen, Prostaglandin D2and Isoproterenol

Activation of mast cells by bridging of IgE-receptors or concanavalin A (Con A) results in a rapid initial rise and fall in cyclic AMP (cAMP) levels followed by a second rise in cAMP levels and histamine release (Sullivan, T. et al. (1976) J. Immunol. 117, 713-716; Lewis, R.A. et al. (1979) J. Immunol. 123, 1663-1668; Ishizaka, T. et al. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 6812-6816). trans-4-Guanidinomethylcyclohexanecarboxylic acid 4-tert-butylphenyl ester (GMCHA-OPhBut), a strong trypsin inhibitor and an anti-allergic agent (Muramatu, M. et al. (1982) Hoppe-Seyler's Z. Physiol. Chem. 363, 203-211; Takei, M. et al. Agents Actions, in press), strongly and dose-dependently inhibited the initial and second rises in cAMP levels, and release of histamine from rat mast cells by Con A, anti-IgE and antigen. Addition of GMCHA-OPhBut after the initial rise in cAMP inhibited the second rise in cAMP and histamine release. These results suggested a possible participation of a trypsin-like proteinase, probably pH 7 tryptase present in rat mast cells, in the activation of adenylate cyclase by the above secretagogues, and the initial rise in cAMP was not directly related to the latter events. The second rise in cAMP is induced by prostaglandin D2 (PGD2), a metabolic product of arachidonic acid. PGD2 elevated the cAMP levels in mast cells whereas no histamine was secreted. GMCHA-OPhBut did not inhibit the increase in cAMP by PGD2. Therefore, the strong inhibitory effect of GMCHA-OPhBut on the second rise in cAMP might depend on the inhibition of an earlier process than the activation of adenylate cyclase by PGD2.(ABSTRACT TRUNCATED AT 250 WORDS)

Thrombin-induced abnormal platelet activation in spontaneously hypertensive rats is linked with phosphoinositides turnover and phosphorylation of 47,000 and 20,000 dalton proteins

We have shown earlier that abnormal platelet aggregation in spontaneously hypertensive rats (SHR) is not caused by prostaglandins. In this study platelets from SHR and normotensive (Wistar Kyoto, WKY) rats were used to examine the role of phosphoinositides and phosphorylation of 47,000 and 20,000 Dalton proteins in abnormal platelet activation in hypertension. Thrombin (0.05 U/ml) induced a rapid decrease in (32P)-PO4 labelled phosphatidylinositol-4,5-bisphosphate (PIP2), phosphatidylinositol-4-phosphate (PIP) and phosphatidylinositol (PI) in washed rat platelets. However, significantly greater loss of PIP2 and PI was seen in SHR platelets than in WKY platelets. For example the level of PIP2 declined by 32% in SHR platelets and only by 13% in WKY platelets at five seconds of incubation with thrombin. The loss of PI was similar in SHR and WKY platelets for the first five seconds of incubation with thrombin. However, by 15 seconds SHR platelets showed a significantly greater loss (24%) in PI than in WKY platelets (8%). Thrombin induced a 14% and 18% decrease in PIP at three seconds in WKY and SHR platelets respectively. In SHR platelets PIP level returned to the baseline in five seconds and then rose to 20% above the baseline by 30 seconds. In contrast PIP level in WKY platelets slowly reached the basal value by 30 seconds. Thrombin also produced a two- to three-fold greater accumulation of (32P)-phosphatidic acid (PA) in SHR platelets than in WKY platelets. Thrombin (0.05 U/ml) induced rapid phosphorylation of 47,000 Dalton (P47) and 20,000 Dalton (P20) proteins in both WKY and SHR platelets. Thrombin induced a four-fold greater increase in phosphorylation of P47 in SHR platelets than in WKY platelets in the first five seconds. Thrombin produced significantly greater increase in phosphorylation of P20 in SHR platelets (34% and 41%) than in WKY platelets (18% and 28%) at 5 and 15 seconds. Phosphorylation of P20 was followed by dephosphorylation in both WKY and SHR platelets. Aspirin (500 uM) did not affect phosphorylation of either P47 or P20 in SHR or WKY platelets. In other experiments prostaglandin El (0.5 uM), which stimulates adenylate cyclase via a guanine nucleotide regulatory protein termed Gs, caused an eighteen-fold increase in cyclic AMP level in SHR platelets as compared to a six-fold increase in WKY platelets. These data lead us to suggest that increased turnover of phosphoinositides and increased phosphorylation of P47 and P20 are involved in abnormal platelet activation in SHR platelets.

Effects of stimulants of electrogenic ion transport on cyclic AMP and cyclic GMP levels in locust rectum

AbstractA variety of agents stimulate the chloride‐dependent short‐circuit current (Isc) across isolated locust recta. In this study, the role of cyclic nucleotides in mediating this stimulation was examined by measuring levels of cyclic AMP and cyclic GMP in tissues exposed to these agents. After bathing recta in saline, there is a decline in Isc, and this is accompanied by a fall in cellular levels of cyclic AMP and cyclic GMP. Stimulants of Isc across locust recta, corpus cardiacum (CC) extracts containing chloride transport‐stimulating hormone (CTSH) and forskolin, also elevate cellular levels of cyclic AMP. Between 5 and 10 min after addition of CC extract, there is an increase in cyclic AMP levels. During this same time period, the Isc rises at its maximal rate. The Isc remains elevated for at least another 20 min, even though tissue cyclic‐AMP levels decline rapidly by 15 min. These results suggest that stimulation of Isc is induced by a rise in cyclic AMP, but the cessation of stimulated active ion transport is controlled by other biochemical processes. In contrast to the relatively rapid rise in cyclic‐AMP levels, a 60‐min exposure to CC extract was required to elicit a rise in cyclic GMP, and therefore it may not be involved in the early stages of Isc stimulation.

Effects of phosphodiesterase inhibitors on normal and chemically‐skinned isolated airway smooth muscle

1. The effects of three phosphodiesterase inhibitors (papaverine, isobutyl methyl xanthine (IBMX) and SKF 94120) were examined on tension responses and cyclic nucleotide content (both cyclic AMP and cyclic GMP) of normal and Triton X-100 skinned isolated trachealis of the guinea-pig. 2. The three inhibitors were approximately equipotent in eliciting concentration-dependent relaxation of histamine-induced contractions of the trachealis. 3. Papaverine-induced relaxation was associated with concentration-related increases in the levels of both cyclic nucleotides. 4. IBMX at low concentrations (1 mumol l-1) produced significant relaxation (36%) of histamine-contracted trachealis without changing cyclic nucleotide levels. At a ten fold higher concentration IBMX-induced relaxation (95%) was associated with a selective increase in tissue cyclic GMP levels. Only at the highest concentration tested (100 mumol l-1) did IBMX increase cyclic AMP levels significantly. 5. SKF 94120 (1 mumol l-1) elicited a 23% relaxation of the contracted trachealis without altering the tissue content of either cyclic nucleotide. At the two higher concentrations tested (10 and 100 mumol l-1), SKF 94120-induced relaxation was accompanied by a selective increase in the levels of cyclic AMP. 6. In the skinned trachealis Ca2+ (10 and 20 mumol l-1)-induced contractions were significantly inhibited by the calmodulin antagonist calmidazolium (10 mumol l-1) and by cyclic AMP (10 mumol l-1), the catalytic subunit of cyclic AMP-dependent protein kinase (0.1 mumol l-1) and cyclic GMP (10 mumol l-1). 7. Papaverine (100 mumol l-1) significantly inhibited (31 +/- 6%) the Ca2+-induced contractions of the skinned trachealis. Both IBMX and SKF 94120 were without effect. It is concluded that cyclic nucleotide-dependent mechanisms have an inhibitory action on the biochemical processes that lead to contraction of the guinea-pig trachealis. The results suggest that a functional sarcoplasmic reticular and/or plasma membrane is essential for the expression of IBMXand SKF 94120-induced relaxation. This is not the case for papaverine. The results also highlight the fact that significant relaxant responses of airway smooth muscle can be produced by phosphodiesterase- inhibiting drugs without concomitant elevations in tissue cyclic nucleotide content.

Regulation of fructose 2,6-biphosphate levels in Neurospora crassa

Both wild type and cr-1 mutant (adenylate cyclase and cyclic AMP-deficient) strains of Neurospora crassa contain fructose 2,6-biphosphate at levels of 2t nmol/g dry tissue weight. This level decreases by about 50% in both strains upon depriving the cells of carbon or nitrogen sources for 3 h. An increase in cyclic AMP levels produced by addition of lysine to nitrogen-starved cells produced no increase in fructose 2,6-biphosphate levels. Both strains respond to short-term addition of salicylate, acetate, or 2,4-dinitrophenol with an increase in fructose 2,6-biphosphate. Thus, the above-described regulation of fructose 2,6-biphosphate levels is cyclic AMP-independent. A suspension of the wild type produces a transient increase of fructose 2,6-biphosphate in response to administration of glucose, whereas the mutant strain does not respond unless it is fed exogenous cyclic AMP. Substitution of acetate for sucrose as a sole carbon source for growth leads to a differential decrease in fructose 2,6-biphosphate levels between the two strains: the wild type strain has 63% and the cr-1 mutant strain has 37% of the levels of fructose 2,6-biphosphate on acetate as compared to sucrose-grown controls. This may be the basis for an advantage of cr-1 over wild type in growth on acetate. Thus, although most regulation of fructose 2,6-biphosphate is cyclic AMP-independent, the levels can be regulated by a combination of carbon source and cyclic AMP levels.

Parathyroid hormone-induced changes in cyclic nucleotide levels during relaxation of the rat aorta

Parathyroid hormone is a potent vasodilator in vivo and relaxes vascular tissue in vitro . Since parathyroid hormone action in kidney and bone is thought to be mediated by stimulation of cellular cyclic AMP production, the present study was designed to monitor changes in cyclic AMP and cyclic GMP in vascular tissue during relaxation by parathyroid hormone. Rabbit aortic strips were quick-frozen at various times after exposure to parathyroid hormone and the percent relaxation and cyclic nucleotide levels were determined. Cyclic AMP concentrations were elevated about 3-fold within 30 seconds after treatment with hormone. This corresponded to a 10% relaxation of the norepinephrine-contracted tissue. After five minutes, cyclic AMP was still elevated 2-fold above basal and the relaxation response was maximal (36%). The cyclic AMP and relaxation responses to parathyroid hormone were markedly potentiated by forskolin or methylisobutylxanthine. Parathyroid hormone produced a small but significant increase in cyclic GMP concentrations only at early time points whereas sodium nitroprusside substantially increased cyclic GMP and relaxed the strips at all times studied. The increase in cyclic AMP levels after exposure to parathyroid hormone occurred prior to or coincident with the onset of relaxation of the aortic strips. These findings are supportive of the hypothesis that the vascular actions of parathyroid hormone involve cyclic AMP.

In vitro effect of prostaglandins on the accumulation of cyclic AMP in the avian oviduct

The effects of prostaglandins E 1, E 2, and F 2α (PGE 1, PGE 2, and PGF 2α, respectively) on cyclic AMP production in tissue samples from the uterus (U), uterovaginal sphincter (UVS), and vagina (V) of regularly laying hens as well as the effects of PGE 2 and forskolin on U and V of Japanese quail were studied. Both PGE 1 and PGE 2 enhanced cyclic AMP production in the hen oviductal segments, particularly in the V in a dose-related manner, PGE 1 being the more effective agonist. PGF 2α had no significant effect. Quail U appeared to be less responsive to PGE 2 which caused a 40% increase in cyclic AMP levels at the highest concentration tested (28 μ M), whereas forskolin at 10 μ M increased cyclic AMP production in quail U and V by 7- and 20-fold, respectively. It is suggested that the primary function of PGE 2 and PGE 1 is to relax the terminal portion of the oviduct, allowing the expulsion of the egg under the contractile influence of PGF 2α and other oxytocic factors.

Phenyliminoimidazolidine Derivatives Activate Both Octopamine1 and Octopamine2 Receptor Subtypes in Locust Skeletal Muscle

ABSTRACT The actions of the phenyliminoimidazolidine derivatives, NC7 and NC5 on octopamine receptors in the extensor-tibiae neuromuscular preparation of the locust are described. The derivatives activate both OCTOPAMINE1 and OCTOPAMINE2 receptors in this preparation. They are not selective agonists of OCTOPAMINE2 receptors that activate adenylate cyclase. The relative potencies of NC5 and NC7 on OCTOPAMINE1 and OCTOP-AMINE2 receptors differ, suggesting that they may be useful in the development of selective agonists that can distinguish between the different pharmacological subclasses of octopamine receptor. The octopamine-mediated increases in cyclic AMP levels in this preparation can be separated into high-and low-affinity components. NC5 and NC7 appear to activate the high-affinity component of this response selectively.

Pertussis toxin stimulates cholecystokinin-induced cyclic AMP formation but is without effect on secretagogue-induced calcium mobilization in exocrine pancreas

The role of a pertussis toxin sensitive GTP-binding protein in mediating between cholecystokinin receptors and phosphatidylinositol 4,5-bisphosphate phosphodiesterase as well as in preventing cholecystokinin from increasing cellular cyclic AMP has been investigated using dispersed acini from rabbit pancreas. Pertussis toxin pretreatment (500 ng/ml, 2 h) did not affect cholecystokinin(octapeptide) (CCK-8) -induced increases in cytosolic free Ca 2+ as judged from changes in fluorescence obtained from quin2-loaded acini. Although pretreatment with pertussis toxin was also without effect on resting acinar cell cyclic AMP levels, adenylate cyclase activity was increased, since inhibition of cyclic AMP phosphodiesterase activity by isobutylmethylxanthine (IBMX) resulted in an additional increase in cyclic AMP levels in toxin-treated acini, indicating that acinar cell adenylate cyclase activity is under some tonic inhibitory control by the pertussis toxin-sensitive inhibitory GTP-binding protein (G i) of the adenylate cyclase system. CCK-8 gave an increase in cyclic AMP levels in both control (1.6-fold) and toxin-treated (2.3-fold) acini, leading to cyclic AMP levels in the toxin-treated acini 2-times as high as those in control acini. In the presence of IBMX, the cyclic AMP response to CCK-8 was again markedly enhanced in acini pretreated with the toxin (3.2- vs. 1.8-fold), resulting in cAMP levels in the toxin-treated acini 3.7-times those in the absence of IBMX, 2.5-times those in control acini in the presence of IBMX and 7.0-times those in control acini in the absence of IBMX. Neither the pretreatment with pertussis toxin, nor the presence of IBMX alone, nor the combination had an effect on basal amylase secretion. However, all three treatments potentiated the stimulatory effect of CCK-8 on amylase secretion and the amount of potentiation was proportional to the cyclic AMP levels reached. Our findings suggest that in the intact pancreatic acinar cell G i inhibition of the catalytic subunit of the adenylate cyclase may largely be responsible for preventing cholecystokinin from increasing cellular cyclic AMP. They moreover show that cyclic AMP is a modulatory agent in rabbit pancreatic enzyme secretion, not able to stimulate secretion itself, but potentiating effects mediated by the phosphatidylinositol-calcium pathway.

Modulation of T cells by cyclic AMP in murine influenza virus infection. I. In vitro inhibition of cytotoxic T lymphocytes by exogenous cyclic AMP analogues and by agents which increase intracellular cyclic AMP concentrations

The effects of 8-bromo-cyclic AMP and cyclic AMP agonists (cholera toxin plus hydrocortisone and prostaglandin E2 plus 3-isobutyl-1-methylxanthine) on the cytotoxic activity of T cells generated during murine influenza virus infection have been examined. Treatment of influenza A/WSN virus primed primary immune spleen cells and their cultured secondary effector T cells from Balb/c mice with these agonists resulted in increases in cyclic AMP levels. These agents also had a marked inhibitory effect on the cytotoxic activity of primary immune spleen cells, but a much weaker inhibitory effect on that of secondary effector T cells. No significant effect of 8-bromo-cyclic GMP on the cytotoxic activity of these two cell types was observed. The cytotoxic activity of H-2-restricted, virus-specific T cells generated either in vivo (primary) or in vitro (secondary) was not significantly inhibited by treatment with histamine, whereas that alloreactive killer T cells generated in vivo was markedly inhibited. These results suggest that the actions of H-2-restricted, virus-specific T cells may not be regulated by histamine during viral or inflammatory processes.

Purified podophyllotoxin (CPH-86) inhibits lymphocyte proliferation but augments macrophage proliferation

Purified podophyllotoxin (CPH-86) is an inhibitor of microtubular aggregation used in the treatment of cancer, psoriasis and rheumatoid arthritis. To better understand its immunopharmacology we examined its effects on human lymphocytes and monocytes and guinea pig macrophages. CPH-86 inhibits mitogen-induced human lymphocyte proliferation and macrophage growth factor-stimulated macrophage proliferation with ID50s of approximately 10 −7M. The effect of CPH-86 on lymphocytes in conjunction with mitogen is nonlethal, evident during the early but not the late phases of proliferation, and associated with early increases in cyclic AMP levels. In contrast to these obviously inhibitory effects, CPH-86 (10 −7M) alone induces IL-1 by human monocytes and, with mitogen, it induces IL-2 production by human lymphocytes. It directly stimulates macrophage pproliferation and potentiates the effects of low doses of macrophage growth factor to do so. The latter effects may be mediated by colony stimulating factor production. The effects of CPH-86 are not mediated by inhibition of prostaglandin synthesis. The stimulation of monokine and lymphokine production by CPH-86 may represent positive features of its action and may be immunotherapeutic.

Fatty acid control of cyclic AMP levels inNeurospora crassa

Several saturated, monosaturated, and polyunsaturated fatty acids produce rapid increases in cyclic AMP levels in the fungusNeurospora crassa when added to the growth medium at 10–50 μM. The time courses of cyclic AMP increase resembled those previously shown to be induced by other agents, reaching peak cyclic AMP levels at about 2 min after fatty acid addition. These fatty acids had little or no influence on adenylate cyclase fromNeurospora crassa in vitro. On the basis of previous evidence that uncouplers of oxidative phosphorylation increase cyclic AMP levels and that fatty acids can act as uncouplers, we suggest that the fatty acids in vivo may act to increase cyclic AMP levels by acting as uncouplers of oxidative phosphorylation. In agreement with this suggestion, two fatty acids were shown to produce decreased ATP-ADP ratios inNeurospora at concentrations producing cyclic AMP increases.

Effects of prostaglandin E 1, Isoproterenol and forskolin on cyclic AMP levels and tension in rabbit aortic rings

The role of cyclic AMP in the control of vascular smooth muscle tone was studied by monitoring the effects of prostaglandin E 1 (PGE 1), isoproterenol and forskolin on cyclic AMP levels and tension in rabbit aortic rings. PGE 1, isoproterenol and forskolin all increased cyclic AMP levels in rabbit aortic rings. Isoproterenol and forskolin relaxed phenylephrine-contracted aortic rings, but PGE 1 contracted the rings in the presence or absence of phenylephrine. Isoproterenol relaxed these PGE 1-contracted aortic rings without further change in total cyclic AMP levels, which were already elevated by the PGE 1 alone. Pretreatment with forskolin potentiated the effects of PGE 1 on cyclic AMP levels. PGE 1 caused contradictions in muscles partially relaxed by forskolin, even though very large increases in cyclic AMP levels (30 fold) were produced by PGE 1 in the presence of forskolin. Isoproterenol was able to relax these forskolin-treated, PGE 1-contracted muscles with no further increase in cyclic AMP levels. Thus, there does not appear to be a good correlation between total tissue levels of cyclic AMP and tension in these experiments. Our results suggest that, if cyclic AMP is responsible for relaxation of smooth muscle, some form of functional compartmentalization of cyclic AMP must exist in this tissue.