CYCLIC ADENOSINE 3'',5''-MONOPHOSPHATE IN GUINEA-PIG CEREBRAL CORTICAL SLICES: STUDIES ON THE ROLE OF ADENOSINE

Abstract— In guinea-pig cerebral cortical slices levels of cyclic AMP increase in response to adenosine to about 200pmol/mg protein within 10 min and stay at that level up to 30 min. In the absence of calcium ions and the presence of 1mm -EGTA in the Krebs-Ringer-bicarbonate medium the effect of adenosine is enhanced, cyclic AMP levels rise to about 600 pmol/mg protein within 30 min. In normal and calcium deficient media restimulation of cyclic AMP formation with adenosine is possible after a prior stimulation with adenosine. When slices are preincubated for various periods of time with histamine or adenosine before addition of the complementary agent i.e. adenosine or histamine cyclic AMP levels obtained are unaltered compared to levels seen when adenosine and histamine are added together. Slices which are rendered unresponsive to stimulation with histamine + noradrenaline by a prior incubation with these agents do not regain any response during a 100 min period of incubation in medium. The PDE inhibitors diazepam, SQ 66007 and isobutylmethylxanthine are capable of restoring the sensitivity of the slices to histamine + noradrenaline. This suggests an involvement of PDE in the unresponsive phase of the slices. Addition of adenosine to slices not affected by histamine + noradrenaline does reestablish the response of these slices to the neurohormones. A dose-response curve of adenosine for the interaction with histamine + noradrenaline yields an ED₅₀ of 16 μM using sensitive or desensitized slices. An adenosine concentration of only 7 μM is necessary to restore the original increase of cyclic AMP in response to histamine + noradrenaline to slices insensitive to the biogenic amines. The data are discussed in terms of a possible activation of PDE within cerebral cortical slices from guinea-pig. Adenosine may reverse this activation. The possibility of inactivation of adenylate cyclase during stimulation of cyclic AMP formation and the role of adenosine and PDE inhibitors in this process is being considered.     

ADENOSINE 3'',5''-MONOPHOSPHATE IN GUINEA PIG CEREBRAL CORTICAL SLICES: EFFECT OF BENZODIAZEPINES

Several benzodiazepines, diazepam, chlordiazepoxide, desmethyldiazepam, methyloxazepam and oxazepam, potentiate the accumulation of cyclic AMP elicited by histamine and histamine: noradrenaline in cerebral cortical slices of guinea pig. In addition, these drugs increase basal levels of cyclic AMP by about 100 per cent. When adenosine is used to stimulate cyclic AMP formation only diazepam, desmethyldiazepam and methyloxazepam are increasing cyclic AMP levels significantly over respective controls. The order of potency is: diazepam > desmethyldiazepam > methyloxazepam > oxazepam > chlordiazepoxide. Diazepam decreases the rate of degradation of cyclic AMP after removal of the stimulatory agents (histamine : noradrenaline). Dose response curves for diazepam under two stimulatory conditions are shown. A significant effect is obtained at 50 μm -diazepam and an ED₅₀ of 40 μm is calculated with histamine as the stimulatory agent. When cyclic AMP formation is elicited by histamine : noradrenaline a significant effect of diazepam is seen at 10 μm and an ED₅₀ of 16 μm is obtained. These results lend support to the hypothesis that the psychotropic action of the benzodiazepines may, at least in part, involve the cyclic AMP generating systems of the central nervous system.     

IN VIVO CHANGES OF CEREBRAL CYCLIC ADENOSINE 3'',5''-MONOPHOSPHATE INDUCED BY BIOGENIC AMINES: ASSOCIATION WITH PHOSPHORYLASE ACTIVATION

—The intravenous injection of adrenaline, isoprenaline and histamine to 4-6-day-old chicks resulted in a rapid increase in the cyclic AMP content of cerebral hemispheres that had been removed and frozen within 0·5 s using a freeze-blowing technique. Noradrenaline, dopamine, adenosine, 5-HT and acetylcholine did not significantly alter the nucleotide concentration in vivo. Addition of adrenaline, isoprenaline and histamine to incubated chick cerebral cortex slices also increased the cyclic AMP content of the tissue. Noradrenaline was considerably less potent than these amines and adenosine was ineffective. Low phosphorylase a levels (16 per cent of total activity) were observed in instantaneously frozen cerebral hemispheres of untreated chicks. The injection of adrenaline, isoprenaline and histamine resulted in a rapid conversion of phosphorylase b to a and a significant fall in tissue glycogen. Administration of noradrenaline was without effect on the relative forms of phosphorylase and also failed to influence cerebral glycogen. Phosphorylase activation was not observed in chick cerebral slices under conditions producing large increases in cyclic AMP. It is suggested that in vivo phosphorylase activation and subsequent glycogenolysis may occur, at least in part, in glia and that these cells may be damaged during preparation of cerebral slices. The results provide evidence of a metabolic role for cyclic AMP in cerebral tissue.     

FUNCTIONAL COMPARTMENTS OF ADENINE NUCLEOTIDES SERVING AS PRECURSORS OF ADENOSINE 3'',5''-MONOPHOSPHATE IN MOUSE CEREBRAL CORTEX

Cyclic AMP accumulates in cerebral cortical slices from the C57B1/6J mouse incubated with the following stimulatory agents: norepinephrine, adenosine, veratridine and adenosine-biogenic amine combinations. The results with slices labelled with radioactive adenine or adenosine provide evidence for the existence of distinct functional compartments of adenine nuclcotides which serve as precursors of cyclic AMP on stimulation with specific agents. Thus, in slices labelled with [¹⁴C]adenine or [³H]adenosine the ratio of [¹⁴C] to [³H]cyclic AMP was dependent on the stimulatory agent; with veratridinc the ratio was 1.4 while with adenosine the ratio was 3.0. In addition, a greater than 2-fold difference in the ratio of endogenous/radioactive cyclic AMP was observed in adenine or adenosine-labelled slices after incubation with veratridine, norepinephrine, adenosine or adenosine-amine combinations; the lowest ratios after stimulation with veratridine and the highest after adenosine or adenosine-amine combinations. The high ratio observed with adenosine was in part due to a quite marked incorporation of the stimulant, adenosine, into the accumulating cyclic AMP. Such distinct functional compartments of cyclic AMP precursors may represent different cell types and/or morphological entities within one cell type.     

ADENOSINE 3'',5''-MONOPHOSPHATE IN GUINEA PIG CEREBRAL CORTICAL SLICES: EFFECTS OF α- AND β-ADRENERGIC AGENTS, HISTAMINE, SEROTONIN AND ADENOSINE

—Norepinephrine and epinephrine, in combination with either adenosine or histamine, enhanced the accumulation of cyclic AMP in guinea pig cerebral cortical slices. Isoproterenol had only marginal effects under the same conditions. Studies with d- and l-norepinephrine and with the α- and β-adrenergic blocking agents, phenoxybenzamine, phentolamine, dihydroergokryptamine, propranolol and sotalol, indicated that the effect of catecholamines on cyclic AMP levels in this tissue was stereo-specific and was mediated primarily via interaction with a classical α-adrenergic receptor. Studies with the antihistaminics, diphenhydramine and pheniramine, and the antiserotonin agent, methysergide, indicated that guinea pig cerebral cortical slices contain receptors for histamine and serotonin, whose activation also stimulates an enhanced accumulation of cyclic AMP in the presence of adenosine.     

CYCLIC ADENOSINE 3'',5''-MONOPHOSPHATE FORMATION IN GUINEA-PIG BRAIN SLICES: EFFECT OF H1- AND H2-HISTAMINERGIC AGONISTS

—A variety of histamine analogs elicit accumulations of radioactive cyclic AMP in guinea-pig neocortical and hippocampal slices labelled during a prior incubation with [¹⁴C]adenine. The H₁agonist, 2-aminoethylthiazole, elicits accumulation of cyclic AMP in neocortical and hippocampal slices both in the absence or presence of adenosine. The presence of adenosine increases the maximum response to 2-aminoethylthiazole and decreases the EC₅₀ by nearly 10-fold. In the absence of adenosine the effects of 2-aminoethylthiazole are antagonized in hippocampal slices by both d-brompheniramine and metiamide, while in the presence of adenosine only d-brompheniramine is an effective antagonist. The H₂-agonist, 4-methylhistamine, elicits a somewhat smaller accumulation of cyclic AMP than does 2-aminoethylthiazole in both cortical and hippocampal slices. In the presence of adenosine the response to 4-methylhistamine is enhanced, but is markedly lower than that seen with the combination of adenosine and 2-aminoethylthiazole. The dose-response relationship for 4-methylhistamine in the presence of adenosine appears in hippocampal slices to consist of two components. The response to 4-methylhistamine in the absence of adenosine is blocked by metiamide, while in the presence of adenosine the response is partially blocked by both H₁ and H₂-antagonists. The accumulation of cyclic AMP elicited by histamine is greatly increased by adenosine but the EC₅₀ is not significantly decreased. The results suggest that (i) both H₁- and H₂-receptors regulate cyclic AMP-formation in the central nervous system, (ii) the synergism between adenosine and histamine is mediated primarily by interaction with H₁-receptors and (iii) that adenosine greatly increases the affinity of the H₁-receptors for both H₁ and H₂-agonists without affecting its affinity for histamine.     

The accumulation of cyclic AMP and cyclic GMP in guinea pig brain slices: Effect of calcium ions,norepinephrine and adenosine

The effect of Ca²⁺ and putative neurotransmitters on formation of cyclic AMP and cyclic GMP has been studied in incubated slices of brain tissue. Cyclic AMP levels in cerebellar slices after about 90 min of incubation ranged from 10 pmol/mg protein in rabbit, to 25 in guinea pig, to 50 in mouse and 200 in rat. Cyclic GMP levels in the same four species showed no correlation with cyclic AMP levels and were, respectively, 1.3, 20, 5 and 30 pmol/mg protein. The absence of calcium during the prolonged incubation of cerebellar slices had little effect on final levels of cyclic AMP, while markedly decreasing final levels of cyclic GMP. Reintroduction of Ca²⁺ resulted in a rapid increase in cerebellar levels of cyclic GMP which was most pronounced for guinea pig where levels increased nearly 7-fold within 5 min. Prolonged incubation of guinea pig cerebral cortical slices in calcium-free medium greatly elevated cyclic AMP levels apparently through enhanced formation of adenosine, while having little effect on final levels of cyclic GMP. Norepinephrine and adenosine elicited accumulations of cyclic AMP and cyclic GMP in both guinea pig cerebral cortical and cerebellar slices. Glutamate, γ-aminobutyrate, glycine, carbachol, and phenylephrine at concentrations of 1 mM or less had little or noe effect on cyclic nucleotide levels in guinea pig cerebellar slices. Prostaglandin E₁ and histamine slightly increased cerebellar levels of cyclic AMP. Isoproterenol increased both cyclic AMP and cyclic GMP. The accumulation of cyclic AMP and cyclic GMP elicited by norepinephrine in cerebellar slices appeared, baed on dose vs. response curves, agonist-antaganonist relationships and calcium dependency, to involve in both cases activation of a similar set of ß-adrenergic receptors. In cerebellar slices accumulations of cyclic AMP and cyclic GMP elicted by norepinephrine and by a depolarizing agent, veratridine, were strongly dependent on the presence of calcium. The stimulatory effects of adenosine on cyclic AMP and cyclic GMP formation were antagonized by theophylline. The lack of correlations between levels of cyclic AMP and cyclic GMP under the various conditions suggested independent activation of cyclic AMP- and cyclic GMP-generating systems in guinea pig cerebellar slices by interactions with Ca²⁺, norephinephrine and adenosine.     

REGULATION OF CYCLIC ADENOSINE 3'', 5''-MONOPHOSPHATE CONCENTRATION IN CHICK CEREBRAL HEMISPHERES DURING DEVELOPMENT

Abstract— Investigations have been carried out into developmental aspects of cyclic AMP metabolism and responsiveness to neurohormones in chick cerebral hemispheres. The in vivo cyclic AMP concentration, measured after freeze-blowing, was found to be highest in the embryonic brain, and changes in the cyclic nucleotide content produced by ischaemia increased with age. The magnitude of the in vivo increases in cyclic AMP produced by isoprenaline and by histamine decreased throughout the first postnatal month. The onset of isoprenaline- and histamine-induced cyclic AMP accumulation in brain slices occurred around 17 days embryonic age, reached a maximum at about 3 days post-hatch and fell to approx 50% of this response at 28 days of age. Adenosine stimulated cyclic AMP formation to a similar extent at all ages studied.
The activities of adenylate cyclase and cyclic AMP phosphodiesterase of hemisphere homogenates were found to reach maximum near the time of hatching. Since the overall pattern of responsiveness of the cerebral cyclic AMP system to neurohormones does not correlate with these variations in enzyme activities, it is suggested that changes occurring at the synaptic receptor level may explain the developmental variations observed.     

A MODULATING ROLE OF PITUITARY-ADRENAL AXIS IN CEREBRAL METABOLISM OF ADENOSINE 3'',5''-MONOPHOSPHATE

Abstract— The effect of adrenalectomy or hypophysectomy on the metabolism of adenosine 3',5'-monophosphate (cyclic AMP) in the cerebral cortex of male Wistar rats was investigated.
The bilateral removal of adrenal glands reduced significantly the activity of cerebral adenylate cyclase [EC 4.6.1.1]. whereas that of cyclic 3'.5'-nucleotide phosphodiesterase [EC 3.1.4.17] remained unchanged. The formation of cyclic AMP measured in cerebral cortical slices from adrenalectomized or hypophysectomized rats was also diminished. Decreases in the activity of adenylate cyclase and formation of cyclic AMP following adrenalectomy were antagonized by in vivo administration of dexamethasone or aldosterone, while those observed in hypophysectomized rats were restored by ACTH or dexamethasone. It is suggested that the pituitary adrenal axis has a modulating role in the metabolism of cerebral cyclic AMP, possibly by changing adenylate cyclase activity.     

ACCUMULATION OF CYCLIC ADENOSINE 3', 5'-MONOPHOSPHATE IN CEREBRAL CORTICAL SLICES FROM RAT AND MOUSE: STIMULATORY EFFECT OF α- AND β-ADRENERGIC AGENTS AND ADENOSINE

Abstract— Norepinephrine, epinephrine, isoproterenol, and adenosine elicit enhanced accumulations of cyclic AMP in incubated slices of rat cerebral cortex. Combinations of norepinephrine, epinephrine, isoproterenol, or histamine with adenosine have a greater than additive effect on cyclic AMP levels. The effects of isoproterenol appear to be mediated via a classical β-adrenergic receptor whereas the effects of norepinephrine appear due to interactions with both α- and β-adrenergic receptors. The presence of the phosphodiesterase inhibitor, isobutylmethylxanthine, potentiates the effects of the catecholamines and reveals a histamine-mediated increase in cyclic AMP levels. After an initial stimulation of cyclic AMP formation with norepinephrine, followed by washing of the slices, the cyclic AMP-generating system is unresponsive to norepinephrine but does respond to an adenosine-norepinephrine combination. In mouse cerebral cortical slices, catecholamines appear to elicit an accumulation of cyclic AMP primarily via interaction with a β-adrenergic receptor.     

EFFECT OF β-ADRENERGIC DRUGS ON ADENOSINE 3'',5''-MONOPHOSPHATE IN RABBIT VAGUS NERVE

Abstract— Cyclic AMP was found to accumulate in rabbit vagus nerve after stimulation of specific β-adrenoceptors. The increase in cyclic AMP content by either isoproterenol or epinephrine was inhibited by the β-adrenoceptor antagonists sotalol and propranolol. α-Adrenoceptor agonists and antagonists, indirect sympathomimetics and theophylline had no effect on the accumulation of cyclic AMP in vagus nerve. The cyclic AMP increase caused by either β-adrenoceptor agents or adenosine was found to have no effect on resting potentials, action potentials or on post-tetanic hyperpolarization.     

INFLUENCE OF DIVALENT CATIONS ON REGULATION OF CYCLIC GMP AND CYCLIC AMP LEVELS IN BRAIN TISSUE

—Depolarizing concentrations of K⁺ elevate levels of both adenosine 3′,5′monophosphate (cyclic AMP) and guanosine 3′,5′monophosphate (cyclic GMP) in incubated slices of mouse cerebellum. Calcium is an essential requirement for the K⁺ -induced accumulation of cyclic GMP. Barium and Sr²⁺, but not Mn²⁺ or Co²⁺, can substitute for Ca²⁺ in this process. Relatively high concentrations of Mg²⁺ inhibit the effect of Ca²⁺ on K⁺-induced accumulation of cyclic GMP. In contrast, depolarizing concentrations of K⁺ are capable of elevating cyclic AMP levels in brain slices suspended in media containing Mg²⁺ and no other divalent cations. High concentrations of Ca²⁺ (1 mm or greater) augment this Mg²⁺ -dependent, K⁺-induced accumulation of cyclic AMP, however. Strontium and Mn²⁺, but not Ba²⁺ or Co²⁺, can substitute for Ca²⁺ in this process, and high concentrations of Mg²⁺ are not inhibitory. The divalent cation ionophore, A-23187 (10 μm ), in the presence of extracellular Ca²⁺ elevates the level of cyclic GMP, but not cyclic AMP, in incubated mouse cerebellum slices. The results of this study indicate that intracellular Ca²⁺ concentration is a major factor regulating cyclic GMP levels in brain. In addition the present results suggest that, in brain tissue, depolarization-induced accumulation of cyclic GMP, but not cyclic AMP, is closely linked to some Ca²⁺-dependent mechanism(s) mediating release of intracellular substances.     

Accumulation of adenosine 3'',5''-monophosphate in incubated slices from discrete regions of squirrel monkey cerebral cortex: effect of norepinephrine, serotonin and adenosine

THE ROLE of adenosine 3′5′-monophosphate (cyclic AMP) in the central nervous system is as yet unknown. However, a variety of putative neurotransmitters such as norepinephrine, serotonin and histamine have been found to elicit enhanced accumulations of cyclic AMP in cerebral cortical slices from a variety of species (SHIMIZU and DALY, 1972; FORN and KRISHNA, 1971; SHIMIZU, TANAKA, SUZUKI and MATSUKADO, 1971; FUMAGALLI, BERNAREGGI, BERTI and TRABUCCHI, 1971). In addition, depolarization of membranes in cortical slices elicits a marked accumulation of cyclic AMP, probably mediated through the action of adenosine (KAKIUCHI, RALL and MCILWAIN, 1969; SHIMIZU. CREVELING and DALY, 1970). These results, obtained with grey matter from the entire cerebral cortex, suggest an intimate relationship between neuronal activity, release of putative neurotransmitters, and enhanced accumulation of cyclic AMP in brain tissue. Since innervation and electrical activity in the cerebral cortex differs among functionally distinct neocortical areas and in the histologically distinct limbic cortex, it appeared possible that these differences would also be manifest in the regional control of cyclic AMP levels. We now report that the response of the cyclic AMP generating system to putative neurotransmitters and adenosine does differ among discrete functional regions of the squirrel monkey cerebral cortex.     

THE EFFECT OF ELECTRICAL STIMULATION UPON THE ACCUMULATION OF ADENOSINE 3'',5''-PHOSPHATE IN ISOLATED CEREBRAL TISSUE

The application of electrical pulses to slices of guinea pig cerebral cortex led to an increase in the levels of adenosine 3′,5′-phosphate (cyclic 3′,5′-AMP) of more than 11-fold within 10 min. This effect of electrical pulses was severely reduced in the presence of theophylline. Cyclic 3′,5′-AMP accumulation in slices was increased in the presence of norepinephrine and histamine about 1·5-fold and six-fold, respectively; the effect of electrical pulses was augmented in the presence of maximal amounts of either amine. For these and other reasons, the accumulation of cyclic 3′,5′-AMP induced by electrical stimulation cannot be ascribed to the release and action of either histamine or norepinephrine.     

THE EFFECT OF NEONATAL THYROIDECTOMY ON THE DEVELOPMENT OF THE ADENOSINE 3'',5''-MONOPHOSPHATE SYSTEM IN THE RAT BRAIN

Abstract— The effect of neonatal thyroidectomy on the cyclic AMP system in the developing rat brain was examined. Administration of ¹³¹I at birth led to a 16 per cent reduction in brain weight and a 70 per cent reduction in body weight by 40 days of age. The level of cyclic AMP in the brain increased 5-fold between birth and 40 days of age and this increase was partially reduced by early thyroidectomy. A similar increase in the activity of adenyl cyclase and phosphodiesterase was observed during development, but thyroidectomy produced no detectable changes in the activity of either enzyme. The activity of the cyclic AMP-dependent protein kinase was already maximal at birth and also was unaffected by thyroidectomy.
Norepinephrine increased levels of cyclic AMP 4- to 5-fold in brain slices prepared from adult rats, but was without effect on slices prepared from newborn or 3-day-old rats. The response to norepinephrine in thyroidectomized rats did not differ from that in control rats at any of the ages examined. Our findings indicate that neonatal hypothyroidism does not deleteriously affect the development of the cyclic AMP system in the rat brain.     

Inhibition of 3',5'-nucleotide phosphodiesterase in guinea pig cerebral cortical slices

Several compounds have been tested for their activity as inhibitors of 3′,5′-nucleotide phosphodiesterase in brain cortical slices from guinea pig. SQ 20,009 (1-ethyl-4-isopropylidenehydrazino)-1H-pyrazolo (3,4-b)pyridine-5-carboxylate, ethylester, hydrochloride), a very potent inhibitor of 3′,5′-nucleotide phosphodiesterase from rat and rabbit brain shows only moderate activity as 3′,5′-nucleotide phosphodiesterase inhibitor when tested in brain slices. It enhances cyclic AMP accumulation only when slices are stimulated by histamine. It does not affect cyclic AMP levels when histamine/norepinephrine are used as stimuli of cyclic AMP formation and decreases the activity of adenosine as stimulant slightly. Ro 20–1724 (4-(3-butoxy-4-methoxy)-2-imidazolidinone) a potent inhibitor of canine cerebral cortex PDE activity effectively augments the increase in cyclic AMP under all stimulating conditions mentioned, as does to a somewhat smaller extent the more water soluble Ro 20–2926 (4-(3-ethoxy-ethoxy-4-methoxy)-2-imidazolidinone). Dose-response curves for Ro 20–1724 under three stimulating conditions of increased cyclic AMP formation (0.1 mm histamine, 0.1 mm histamine/0.1 mm norepinephrine, 0.1 mm adenosine) yield an ED₅₀ of about 20 μm in all instances. A significant increase over respective controls is seen even at 1 μm Ro 20–1724 (histamine/norepinephrine). The drugs may be useful as tools for studying the regulation of cyclic AMP levels in the central nervous system.     

A RADIOISOTOPIC METHOD FOR MEASURING THE FORMATION OF ADENOSINE 3'',5''-CYCLIC MONOPHOSPHATE IN INCUBATED SLICES OF BRAIN

Abstract— A simple and sensitive method for measuring the effect of neurohormones and other chemical agents on the formation of adenosine 3',5'-cyclic monophosphate (cyclic 3',5'-AMP) in incubated slices of brain was developed. The principle of the method depends on pulse-labelling of adenosine-5'-triphosphate in slices of brain with [8-¹⁴C]adenine, followed by incubation in a medium containing the test substance, separation by thin-layer chromatography of the cyclic nucleotide formed in the slices, and radioassay. The purity of the cyclic nucleotide was confirmed by chromatography in a variety of systems and by hydrolysis with a specific, bovine-heart phosphodiesterase. The method was used to study the effect of histamine, norepinephrine, and adenosine on the accumulation of adenosine 3',5'-cyclic monophosphate in incubated slices of brain.     

COMPARISON OF LEVELS OF ADENOSINE 3'',5''-MONOPHOSPHATE IN HIGHLY PURIFIED AND MIXED PRIMARY CULTURES OF NEURONS AND NON-NEURONAL CELLS FROM EMBRYONIC CHICK SYMPATHETIC GANGLIA

Primary cultures containing ≥99% neurons, ≥99% non-neuronal cells (glia), or both cell types were prepared from the sympathetic ganglia of 12-day chick embryos. Levels of cyclic AMP in the non-neuronal cells (～14 pmol/mg protein) were approximately 3-fold higher than levels in the neurons (～4 pmol/mg protein). Mixed cultures had concentrations of cyclic AMP which fell between the values measured for pure neuronal and pure non-neuronal cultures. The measured cyclic AMP values of mixed cultures were indistinguishable from values predicted by summing the expected contributions of the neurons and non-neuronal cells. Thus, contact between the neurons and non-neuronal cells in these mixed cultures did not appear to alter the level of cyclic AMP in either cell type. Neuronal-glial interactions, such as the specific neuronal stimulation of non-neuronal cell proliferation, occurred independently of any changes in the level of cyclic AMP in the mixed cultures. Cell density was varied in both pure and mixed cultures, and both cyclic AMP concentrations and amounts of [³H]thymidine incorporation into DNA were measured. The cyclic AMP content of the non-neuronal cells varied inversely with cell density. [³H]Thymidine incorporation was independent of cell density in both neuronal and non-neuronal cultures. Parallel density-dependent decreases in cyclic AMP concentration and [³H]thymidine incorporation were observed in mixed cultures as cell density was increased. The data suggest that there is no relationship between changes in rate of non-neuronal cell proliferation and cyclic AMP levels in these cultures.     

DEVELOPMENT AND CHARACTERISTICS OF THE HISTAMINE-INDUCED ACCUMULATION OF CYCLIC AMP IN THE RABBIT CEREBRAL CORTEX

Abstract— In slices of adult rabbit cerebral cortex histamine at 5 μM produced a detectable rise in adenosine 3',5'-monophosphate (cyclic AMP). A maximum (20-fold) increase was observed in response to 0–5 mM histamine, with higher concentrations being less effective. The antihistaminic agent, tripelennamine, inhibited the response to 50 μM histamine in a dose-related manner. No effect on basal levels of cyclic AMP was noted with the highest dose of tripelennamine. The cyclic AMP response to 50 μM histamine was sustained for up to 1 h of incubation whether the slices and included medium were assayed together or the slices were assayed separately, although after 60 min of incubation cyclic AMP levels were higher when the medium was included in the assay. During development of the rabbit cerebral cortex, the first detectable increase of cyclic AMP in response to histamine occurred at fetal day 25, and from day 28 to birth the response was a 4-to 5-fold increase. A maximal (10-fold) response was observed at 4–8 days postpartum and by 20 days of postnatal age the response had decreased to the adult levels.     

CATECHOLAMINE INDUCED INCREASE OF CYCLIC ADENOSINE 3'',5''-MONOPHOSPHATE IN RAT BRAIN IN VIVO

Abstract— Four catecholamines injected into the cerebral ventricles increased the content of cyclic adenosine 3',5'-monophosphate (cAMP) in vivo in the whole brain of rats. The highest rise (2.6-fold) was measured 2 min after an injection of 100 μg epinephrine. Isoproterenol and norepinephrine were less active and dopamine hardly increased the cAMP level. These results are compatible with the view that physiological actions of catecholamines in the nervous system may be mediated by an increase of CAMP.