Dibutyryl Cyclic Guanosine Monophosphate Research Articles

Cyclic nucleotides and aminophylline produce different effects on nociceptive motor and sensory responses in the rat spinal cord.

The effect of intrathecal (i.t.) and systemic (i.p. and i.v.) administration of morphine, aminophylline, dibutyryl cyclic adenosine monophosphate (DBcAMP) and dibutyryl cyclic guanosine monophosphate (DBcGMP) on motor and sensory responses of the spinal nociceptive system was studied in rats. Motor responses were assessed in the tail-flick test performed on rats with an intact spinal cord, or as flexor reflex activity elicited in the electromyogram of the tibialis anterior muscle by supramaximal electrical stimulation of the sural nerve in rats in which the spinal cord was transected at the lower thoracic level. The sensory response consisted of activity in single ascending axons of the spinal cord evoked by electrical stimulation of afferent C fibres in spinal rats. Morphine (20 micrograms i.t. or 2 mg/kg i.p.) prolonged the tail-flick latency and aminophylline (25 mg/kg i.p. or 50 micrograms i.t.) prevented the antinociceptive effect of morphine. Aminophylline alone, administered by i.t. injection, reduced the tail-flick latency in a dose-dependent way. Morphine (2 mg/kg i.v. or 10 micrograms i.t.) reduced flexor reflex activity, and this reduction was abolished by aminophylline (25 mg/kg i.v. or 50 micrograms i.t.). Morphine (2 mg/kg i.v.) depressed spontaneous and evoked activity in single ascending axons responding to stimulation of afferent C fibres. This depressant effect of morphine was not abolished by aminophylline (50 micrograms i.t.); the depression was antagonized by naloxone (10 micrograms i.t.). DBcAMP (5 to 100 ng i.t.) dose-dependently prolonged the tail-flick latency. The antinociceptive effect of DBcAMP (50 ng i.t.) was prevented by aminophylline (50 micrograms i.t.) or naloxone (5 micrograms i.t.).(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanism by which dibutyryl cyclic guanosine monophosphate antagonizes the action of cholecystokinin.

Mechanism by which dibutyryl cyclic guanosine monophosphate antagonizes the action of cholecystokinin.

Copper Stimulates Amylase Release From Isolated Rat Pancreatic Acini Independent of Ca2+

Cu2+ and other metal divalent cations influencing the oxidation state of protein sulfhydryl groups were investigated for their effects on pancreatic function. The transition cations stimulated the release of amylase from isolated rat pancreatic acini with a potency order Cu2+ ≈ Cd2+ > Zn2+. Amylase release was also stimulated by the organic sulfhydryl oxidant, diamide. Cu2+-stimulated amylase release was maximal after a delay of 10 min, whereas the effects of cholecystokinin were maximal within 5 min. Cu2+-induced amylase release was only partially inhibited by dibutyryl cyclic guanosine monophosphate, and was not inhibited by atropine. Furthermore, the effect of Cu2+ was additive to that of cholecystokinin and carbamylcholine. Depletion of intracellular Ca2+ by pretreatment of acini with cholecystokinin in Ca2+-free medium (plus ethyleneglycol bis(β-aminoethylether)-N,N′-tetraacetic acid) had no effect on subsequent stimulation by Cu2+, although it abolished the response to cholecystokinin. Cu2+ induced only a small and delayed increase in 45Ca2+ efflux, whereas cholecystokinin induced an immediate large increase. It was concluded that Cu2+ has multiple effects on pancreatic amylase release. A major component is independent of known secretagogue receptors or intracellular Ca2+. It is suggested that changes in the oxidation state of protein sulfhydryl groups may be involved in pancreatic zymogen release.

The effect of dibutyril cyclic guanosine monophosphate, a competitive antagonist to cholecystokinin-pancreozymin, on gastric acid secretion in the isolated mouse stomach

Dibutyryl cyclic guanosine monophosphate (dbcGMP), a specific competitive inhibitor of the gastrin, cholecystokinin-pancreozymin (CCK-PZ) family of peptides in pancreas, gallbladder and ileum, had no effect on basal acid secretion in the isolated mouse stomach nor on secretion stimulated by bethanechol or histamine. Secretion evoked by low doses of pentagastrin were likewise unaffected by dbcGMP but responses to high doses of pentagastrin were augmented. CCK-PZ and glucagon each inhibited acid secretion evoked by pentagastrin. DbcGMP blocked CCK-PZ-mediated inhibition but was without effect on inhibition by glucagon. These observations suggest that in the gastric glands there exist two receptors with different affinities for gastrin and CCK-PZ which mediate excitation and inhibition respectively.

Cholinergic and Peptidergic Receptors on Isolated Human Antral Smooth Muscle Cells

Smooth muscle cells were isolated from segment so of human antrum obtained at operation. The presence, activity, and specificity of receptors to acetylcholine, the C-terminal octapeptide of cholecystokinin, methionine-enkephalin, and vasoactive intestinal peptide were examined. Muscarinic cholinergic receptors sensitive to low concentrations of atropine, cholecystokinin receptors sensitive to proglumide and dibutyryl cyclic guanosine monophosphate, enkephalin receptors sensitive to naloxone were demonstrated and found to mediate contraction. Vasoactive intestinal peptide receptors were demonstrated also and found to mediate relaxation. The stoichiometry and specificity of responses were closely similar to those previously found in isolated gastric smooth muscle cells of the guinea pig. The existence of high-affinity functional receptors on smooth muscle cells, together with the presence of the peptides in nerve terminals adjacent to these cells, makes it possible for these peptides to act as neurotransmitters.

Restoration by cyclic guanosine monophosphate and extracellular calcium of butylated hydroxyanisole-suppressed primary murine thymus-dependent antibody response.

The effect of dibutyryl cyclic guanosine monophosphate (dbc-GMP) on butylated hydroxyanisole (BHA)-induced suppression of the primary in vitro thymus-dependent antibody response of BDF1 mouse spleen cultures was studied. When added at 0 hr relative to antigen addition, 1 mg of dbc-GMP (8 mM) restored by greater than 70% the BHA-inhibited primary immunoglobulin (Ig)M plaque-forming cell (PFC) response to sheep erythrocytes (SRBC). The suppression was not reversed by the addition of 50 microgram of dibutryl cyclic adenosine monophosphate (dbc-AMP), which is known to reverse suppressor T-cell activity. The addition of 10 mM extracellular calcium (Ca2+) at the same time as antigen to BHA-inhibited cultures resulted in more than 80% restoration of the anti-SRBC PFC response. Quantitation of c-GMP by radioimmunoassay demonstrated that BHA lowered by 58% the c-GMP content of splenic lymphocytes and abrogated the ability of lipopolysaccharide of E. coli (LPS) to elevate c-GMP levels in splenic lymphocytes. The data suggest that BHA exerts its immunosuppressive effect on the primary in vitro antibody response by inhibiting guanylate cyclase activity and effectively lowering c-GMP levels; exogenous dbc-GMP and Ca2+ can freely reverse the immunosuppressive effect of BHA.

Effects of cyclic-nucleotide derivatives on the growth of human colonic carcinoma xenografts and on cell production in the rat colonic crypt epithelium.

Previous studies have shown that various amine hormones are able to influence the growth rate of human colorectal carcinomas propagated as xenografts in immune-deprived mice, and it is now well known that the effects of many amine and other hormones are mediated by cyclic nucleotides, acting as second messengers within cells. In the present study the influence of various derivatives of cyclic adenosine monophosphate and cyclic guanosine monophosphate on the growth of two different lines of colorectal cancer growing in immune-deprived mice, and on the cell production rate in the colonic crypt epithelium of the rat, was assessed. Growth of each tumour line, as well as crypt-cell production, was suppressed by treatment wit N6O2' dibutyryl and N6 monobutyryl derivatives of cyclic adenosine monophosphate. Dibutyryl cyclic guanosine monophosphate, on the other hand, was found to promote the growth of Tumour HXK4 and to promote crypt cell production, but to have no significant effect on Tumour HXM2.

Heavy metal modulation of lymphocyte activities—II. lead, an in vitro mediator of B-cell activation

Investigation of the immunopotentiating effect(s) of Pb 2+ on the humoral immune response has provided evidence that a heavy metal can alter the proliferation and differentiation of B-cells. Pb 2+ interacted with B-cells to enhance both their proliferation and their differentiation into sheep red blood cell (SRBC)-specific PFC. Preincubation of B-cells with Pb 2+ for only 1 h enhanced their activities. Enhancement of in vitro humoral immunity to SRBC was due to direct Pb 2+ activation of B-cells and enhancement of T-cell help. Furthermore, Pb 2+ and the cyclic nucleotides dibutyryl cyclic adenosine monophosphate (DBcAMP) or cyclic guanosine monophosphate (cGMP) worked in a synergistic manner to enhance the B-cell proliferation, and Pb 2+ and DBcAMP synergistically enhanced PFC development. Ni 2+, which had been shown to enhance PFC development in cultures of spleen cells in a manner equivalent to Pb 2+, did not enhance PFC development in B-cell cultures as well as Pb 2+ and did not produce synergistic effects with DBcAMP or cGMP. The possible mechanisms by which Pb 2+ enhances B-cell activities and the implications of these effects are discussed.

Inhibition of the action of cholecystokinin octapeptide on the guinea pig ileum myenteric plexus by dibutyryl cyclic guanosine monophosphate

Inhibition of the action of cholecystokinin octapeptide on the guinea pig ileum myenteric plexus by dibutyryl cyclic guanosine monophosphate

Cyclic nucleotide metabolism in mouse brain during seizures induced by bicuculline or dibutyryl cyclic guanosine monophosphate

In adult male C57 black mice, intracortical injection of bicuculline (10 μg) elevated cGMP concentrations and produced contralateral clonic seizures. After intraperitoneal injection, bicuculline (10 mg/kg) elevated cGMP concentrations at the earliest time at which paroxysmal EEG activity could be detected. Changes in cAMP concentration followed those events. Further elevations of cGMP concentrations (and now of cAMP as well) were observed during clonic and tonic seizures. Intracortical injections of dibutyryl cGMP produced behavioral and EEG seizures. These data suggest that cGMP plays an important role in bicuculline seizures.

Cholinergic agonists and dibutyryl cyclic guanosine monophosphate inhibit the norepinephrine-induced accumulation of cyclic adenosine monophosphate in the rat cerebral cortex

Addition of cholinergic agonists, namely carbamylcholine (carbachol), acetylcholine, eserine, eserine plus acetylcholine and eserine plus choline chloride, and dibutyryl cyclic guanosine moue-phosphate inhibited the norepinephrine-induced accumulation of cyclic adenosine monophosphate in incubated slices of rat cerebral cortex. Methacholine was ineffective and atropine did not overcome the action of carbachol on the stimulation of cyclic adenosine monophosphate synthesis by norepinephrine. Carbachol stimulated the production of cyclic guanosine monophosphate in the tissue slices white norepinephrine did not influence cyclic guanosine monophosphate levels to a significant degree. The data are in keeping with the ‘Yin-Yang’ hypothesis in which under particular situations the intracellular levels of cyclic guanosine monophosphate may modulate cyclic adenosine monophosphate concentrations.

Dibutyryl cyclic guanosine monophosphate (Bt 2cGMP) inhibits the action of cholecystokinin octapeptide (CCK8) on the guinea pig ileum longitudinal muscle

Dibutyryl cyclic guanosine monophosphate (Bt 2cGMP) inhibits the action of cholecystokinin octapeptide (CCK8) on the guinea pig ileum longitudinal muscle

Studies on the oxidation of muscle triglycerides.

To study factors which might influence oxidation of muscle triglycerides, pieces of rat diaphragms were prelabelled with palmitate-1-14C for one hour and the subsequent changes in tissue lipids and 14CO2 production were measured. Muscle lipids were separated on silicic acid columns and the amount and specific activities of triglycerides, phospholipids and free fatty acids were determined before and after a second hour of incubation in the absence of glucose and radioactive palmitate. Changes in these parameters strongly suggested that over 80% of the 14CO2 produced during the second hour of incubation was derived from diaphragm triglyceride. Therefore, changes in 14CO2 production under these conditions were an index of triglyceride oxidation. The following agents had no effect on this pathway: cosyntropin (Cortrosyn®), nicotinic acid, insulin, acetylcholine, carbamylcholine chloride (Carbachol®), thyrotropin stimulating hormone, glucagon, prostaglandin E1 prostaglandin E2, dexamethasone, growth hormone, adenosine, di-butyryl cyclic guanosine monophosphate, imidazole, norepinephrine, and methoxamine. 14CO2 production was decreased by epinephrine (via its β-adrenergic action), isoproterenol, isobutyl-methylxanthine, and dibutyryl cyclic adenosine monophosphate. However, this effect was due to enhanced glycogenosis which diluted out lipid-derived carbon with a carbohydrate source. Aminophylline and theophylline increased lipid oxidation to 14CO2. Further studies revealed that this effect occurred distal to lipolysis (i.e., on free fatty acid oxidation to CO2). Since isobutylmethylxanthine did not affect 14CO2 production from lipid, aminophylline may influence free fatty acid oxidation by a mechanism other than inhibition of phosphodiesterase. The inability to find agents which modify oxidation of triglycerides in rat diaphragm suggests that rapid mobilization of lipid is not important as an energy source for muscle.

Membrane Fluidity in Human and Mouse Chediak-Higashi Leukocytes

Polymorphonuclear leukocytes from humans and mice with the Chediak-Higashi syndrome were characterized by spin label electron spin resonance spectrometry. Our results suggest that cells from afflicted mice and humans have membranes more fluid than controls. Order parameters for a spin label that probes near the membrane surface were 0.652 for normals and 0.645 for two Chediak-Higashi patients. Cells from Chediak-Higashi mice showed similar differences, as did isolated plasma membrane fractions. An increased membrane fluidity was also detected with a spin label that probes deeper in the bilayer. In vitro treatment of Chediak-Higashi mouse cells with 0.01 M ascorbate increased the order parameter to normal levels. In vitro incubation of mouse Chediak-Higashi cells with glucose oxidase increased the order parameter, similar to the effect of ascorbate. This increase was abolished when catalase was added to the incubation medium. In vitro incubation with dibutyryl cyclic guanosine monophosphate (1 muM to 0.1 mM) did not normalize order parameters. These results indicate that fluidity of Chediak-Higashi cell membranes was affected by treatments expected to alter the oxidation: reduction potential of the environment but was not affected by treatments expected to alter the ratio of intracellular cyclic nucleotides. The latter treatment would affect microtubule assembly. Therefore, it appears that the membrane fluidity abnormalities as demonstrated by electron spin resonance and the earlier demonstrated microtubule dysfunctions characteristic of Chediak-Higashi cells are coexisting defects and are probably not directly related.

Gonadotropin stimulation of the expansion of cumulus oophori isolated from mice: General condition for expansion in vitro

AbstractGonadotropin Stimulation of the Expansion of Cumulus Oophori Isolated from Mice: General Conditions for Expansion In VitroThe hormonal stimulation of cumulus expansion (mucification) was studied using oocyte‐cumulus cell complexes isolated from Graafian follicles of C57BL/6J mice. Cumulus expansion was stimulated in vitro with human chorionic gonadotropin (HCG) and highly purified rat follicle stimulating hormone (FSH) in Whitten's medium containing fetal bovine serum (FBS). Highly purified rat luteinizing hormone failed to stimulate cumulus expansion. The 50% maximal stimulating concentration of FSH was found to be about 3–4 ng/ ml. Cumulus expansion was also stimulated by dibutyryl cyclic adenosine monophosphate (DBcAMP), but not by dibutyryl cyclic guanosine monophosphate or butyrate. Neither progesterone, testosterone, nor 17β‐estradiol stimulated cumulus expansion. Expanded cumuli were dispersed by hyaluronidase.The stimulation of cumulus expansion by FSH, HCG or DBcAMP required the presence of FBS. The active component of serum had a molecular weight greater than 10,000 daltons and did not require the association of a steroid for activity. Since DBcAMP does not stimulate cumulus expansion in the absence of FBS, it is suggested that the serum factor(s) is involved after the step of FSH stimulation of cAMP production by the complex.The cumulus does not expand if FSH is added to cultures after seven hours of spontaneous oocyte maturation, when the oocytes are at late metaphase I or early anaphase I of meiosis. Therefore, the changes which occur in the oocytecumulus cell complex during spontaneous oocyte maturation affect the ability of the complex to respond to FSH and bring about the expansion of the cumulus.

Effects of drugs affecting endogenous amines or cyclic nucleotides on ethanol withdrawal head twitches in mice.

1 Twenty-four hours after ethanol withdrawal, dependent mice exhibited frequent head twitching. Naive mice exhibited similar twitching 15 min after treatment with 5-hydroxytryptophan (5-HTP) or 6 h after alpha-methyl-p-tyrosine (AMPT). Ethanol lessened the incidence of head twitches induced by any of these treatments. 5-HTP and AMPT each increased the incidence of head twitches induced by withdrawal of ethanol from dependent mice. 2 Drugs that affect the amount or activity of endogenous amines or cyclic nucleotides modified the incidence of head twitches. Nearly all drugs acted in the same direction on twitching elicited by any of these three treatments. 3 The incidence was lessened by: (a) methysergide, methergoline, MA 1420, p-chlorophenylalanine and p-chloroamphetamine; (b) dopamine, noradrenaline, L-DOPA, amphetamine and apomorphine; (c) hyoscine and nicotine; and (d) adenosine triphosphate, dibutyryl cyclic adenosine-3',5'-monophosphate (db cyclic AMP) and prostaglandins E1 and E2. 4 The incidence was increased by: (a) acetylcholine, carbachol and physostigmine; and (b) guanosine triphosphate, dibutyryl cyclic guanosine monophosphate (db cyclic GMP), theophylline and 3-isobutyl-1-methyl-xanthine. 5 These findings suggest that head twitching induced by these three treatments arises from a common biochemical mechanism, which may ultimately be a change in favour of cyclic GMP of the balance between this nucleotide and cyclic AMP within appropriate neurones. This imbalance appears to be elicited or increased by 5-hydroxytryptamine and acetylcholine and to be decreased by dopamine, noradrenaline and E prostaglandins. 6 Neither actinomycin D nor cycloheximide, given during the induction of ethanol dependence, altered the incidence of head twitches after ethanol withdrawal.

An in vitro model of phagocytosis in bovine and human retinal pigment epithelium

We have developed an in vitro system for studying phagocytosis in retinal pigment epithelium (RPE). Explants of bovine and human RPE and choroid have been maintained in organ culture, and phagocytosis of 1·1 μm diameter latex microspheres by the RPE has been studied over 24-hr periods. After a latent period of about 8 hr latex was rapidly phagocytized from the apical surface of the cell by more than 90% of the cells in the explant. Each latex bead was surrounded by a membrane derived from apical plasma membrane. Morphometric analysis of organellar area in electron micrographs of phagocytizing vs. fresh RPE cells showed that microvillus membrane is internalized during phagocytosis. A web of microfilaments was apparent near the plasma membrane where initial engulfment occurs, and many microtubules surrounded latex phagosomes deeper in the cytoplasm. Lysosomes fused with many latex phagosomes, but the number of lysosomes in bovine RPE cultured 22 hr exceeded that of fresh RPE. This suggests that either more lysosomes are assembled during acute phagocytic challenge, or that degradation o phagolysosomes occurs less rapidly when inert particles are phagocytized. Phagocytosis by bovine RPE was not affected by continual light or darkness. Dibutyryl cyclic adenosine monophosphate enhanced the total incorporation of latex, but dibutyryl cyclic guanosine monophosphate so altered the shape of cells that phagocytosis could not be assessed. Phagocytosis by human RPE in vitro is reported for the first time. Explants from rod-rich and cone-rich areas of human retina did not differ significantly in their phagocytic ability. The potential use of this explants preparation in the study of various aspects of the phagocytic mechanism in RPE is discussed.

Inhibition of the noradrenergic induction of pineal N-acetyltransferase by dibutyryl cyclic guanosine monophosphate and by ionophore X-537A

Utilizing rat pineals in vitro, it can be demonstrated that dibutyryl cyclic GMP and the calcium ionophore X-537A are both able to reversibly inhibit the elevation of pineal N-acetyltransferase induced by the β-agonists norepinephrine or isoproterenol. These results are discussed in terms of the known hyposensitivy of the pineal gland to overstimulation by β-agonists.

Inhibition of (125I) albumin transport across the isolated rat mesentery by dibutyryl-cyclic guanosine monophosphate.

(1976). Inhibition of [125I] albumin transport across the isolated rat mesentery by dibutyryl-cyclic guanosine monophosphate (1 figure) Archives Internationales de Physiologie et de Biochimie: Vol. 84, No. 4, pp. 894-896.

Interaction between cyclic adenosine monophosphate and cyclic gunaosine monophosphate in guinea pig ventricular myocardium.

The purpose of this study was to examine the mechanisms underlying adrenergic-cholinergic antagonism in ventricular myocardium. Myocardial contractility, cyclic adenosine monophosphate (AMP) levels, and cyclic guanosine monophosphate (GMP) levels were measure in isolated guinea pig ventricles after treatment with various inotropic agents given alone and simultaneously with acetylcholine. Acetylcholine alone markedly elevated cyclic GMP levels but did not substantially change myocardial contractility. However, the same concentration of acetylcholine significantly attenuated the inotropic effect of isoproterenol and histamine, two drugs that may act by increasing myocardial levels of cyclic AMP. The decrease in the inotropic response to isoproterenol did not appear to be due to a decrease in the generation of cyclic AMP, because cyclic AMP levels were similar in hearts receiving isoproterenol alone and those receiving isoproterenol with acetylcholine. Dibutyryl cyclic GMP also antagonized the intropic action of isoproterenol. Acetylcholine did not alter the inotropic effects of ouabain, an agent that increases myocardial contractility without changing cyclic AMP levels. These results suggest that cyclic GMP mediates the antiadrenergic effects of acetylcholine by specifically antagonizing the inotropic actions of cyclic AMP.