Mechanisms of Cyclic AMP Regulation in Cerebral Anoxia and Their Relationship to Glycogenolysis

Abstract: Anoxia elevates levels of cyclic AMP and depresses levels of cyclic GMP in cerebral cortex of mice. Similar effects are also observed in other regions of the brain. Aminophylline inhibits accumulation of cyclic AMP about 50% in hippocampus and cerebellum, but not in cerebral cortex and striaturn; however, this effect requires high doses (²⁵⁰ mgikg). Pretreatment of animals with reserpine, which depletes brain stores of norepinephrine, dopamine, and serotonin, and also produces sedation and mild hypothermia, markedly inhibits accumulation of cyclic AMP in all regions of anoxic brain. Destruction of norepinephrine terminals by treatment of neonatal animals with ⁶-OH- dopamine, which does not sedate or produce hypothermia, has an effect on cyclic AMP levels similar to that of reserpine. None of the above treatments modifies the effect of anoxia on cyclic GMP levels. These data indicate that norepinephrine is a major regulator of cyclic AMP levels in anoxic brain and that adenosine and, perhaps, other unidentified substances have lesser roles in this process. In contrast, biogenic amines and adenosine appear to have no effect on cyclic GMP regulation in anoxic brain. Reserpine slows the activation of phosphorylase and the utilization of ATP, and slightly attenuates the breakdown of glycogen caused by anoxia, but has no effect on the changes in glucose, lactate, or phosphocreatine. In contrast, ⁶-OH-dopamine has no effect on any of these anoxiainduced changes. It is concluded that the effect of reserpine on phosphorylase, glycogen, and ATP is most likely related to the hypothermic and sedative effect of the drug, and that either cyclic AMP is not responsible for initiating glycogenolysis in anoxic brain or only a small rise in cyclic AMP levels is necessary for this process.     

Cyclic GMP and cyclic GMP-dependent protein kinase in brown adipose tissue of developing rats

In order to ascertain the possible involvement of cyclic GMP in the physiological regulation of the function and development of brown fat of the rat, we have determined its tissue concentration in vivo under a variety of conditions. The steady-state concentration of cyclic GMP in interscapular brown adipose tissue of late foetus was about 80 pmol per g fresh weight. The concentration gradually declined during the first 2 weeks after birth to reach 40 pmol/g fresh weight and then remained constant into adulthood. The cyclic GMP content of brown fat was decreased by chemical sympathectomy and was increased after complete acclimatization of the animals to the cold. The activity of cyclic GMP-dependent protein kinase was also highest in tissue from newborn and cold-acclimatized rats.Both acute cold stress and injection of norepinephrine resulted in a significant but temporary increase in the concentration of cyclic GMP in brown fat, which was followed by a depression of the concentration below values in untreated animals. The concentration of cyclic AMP showed similar pattern of changes. Injection of phenylephrine was followed by a pronounced increase in the cyclic GMP content of brown fat, with little effect upon cyclic AMP. Injection of isoproterenol raised the content of cyclic AMP but not that of cyclic GMP. The ability of norepinephrine and phenylephrine to increase the concentration of cyclic GMP was abolished by pre-treatment of the animals with phenoxybenzamine, but not by pre-treatment with propranolol. Conversely, propranolol but not phenoxybenzamine abolished the effects of norepinephrine on the cyclic AMP content of the tissue.Thus we have established the responsiveness of the cyclic GMP content of brown fat to physiological and pharmacological stimuli and have evidence of the possible participation by cyclic GMP in the α-adrenergic stimulation and in the regulation of proliferative processes in the tissue.     

2 types of chronic epileptogenesis in rabbits during kindling stimulation of the hippocampus

The development of convulsant readiness in rabbits during kindling electrical stimulation of the hippocamp was studied as was the dependence of the motor seizure pattern on the degree of epileptiform activity generalization in the CNS. The kindling electrical stimulation of the hippocamp gave rise to the formation in different rabbits of the two main types of afterdischarges. One of them was characterized by high-frequency and high-amplitude spikes (total duration 8-30 s) and the other one by continuous, rather long (50-100 s) hypersynchronous paroxysms. In the interictal period, the animals with the first type demonstrated the occurrence of spontaneous spikes (in all the brain structures under study) that sometimes progressed to a more or less prolonged seizure discharges. At the same time in the animals with the second type of afterdischarges the EEG in the interictal period was slightly different from normal. Despite this fact the seizures induced by electrical stimulation ran a milder course (short-term clonic seizures) in animals with the first type of afterdischarges as compared to those with the second type (long-term clonicotonic seizures). It is assumed that the severity of the motor seizure does not depend on the degree of epileptic activity generalization in the CNS.     

Heterogeneity of β-Adrenoceptors in Guinea-Pig Brain: Radioligand Binding and Cyclic Nucleotide Generation

Abstract: In this report, we have examined the radioligand binding and second messenger signalling characteristics of β-adrenoceptors in the guinea-pig brain. [¹²⁵I]lodocyanopindolol ([¹²⁵I]ICYP)-labelled sites in the cerebellum and cerebral cortex were of similar densities ( B _max 34 and 24 fmol·mg⁻¹) and affinities ( K _D 20 and 55 p M ), respectively. Analysis of competition for [¹²⁵I]ICYP binding in the cerebellum was compatible with the presence of a β₂-adrenoceptor. In this tissue, isoprenaline evoked a cyclic AMP stimulation, and also potentiated cyclic GMP accumulations evoked in the presence of a nitric oxide donor, consistent with mediation via a β₂-adrenoceptor. The [¹²⁵I]ICYP binding profile in the cerebral cortex did not comply with those previously described for β-adrenoceptor subtypes, and isoprenaline failed to alter significantly cyclic AMP accumulation in the cerebral cortex, hippocampus, or neostriatum, even in the presence of forskolin or a phosphodiesterase inhibitor. Isoprenaline was also without effect on cyclic GMP accumulation or phosphoinositide turnover in the cerebral cortex. These results suggest that the guinea-pig cerebellum expresses a functional β₂-adrenoceptor coupled to cyclic AMP generation, and potentiation of cyclic GMP accumulation. However, the guinea-pig cerebral cortex displays binding sites that exhibit β-adrenoceptor-like pharmacology but fail to show functional coupling to cyclic AMP, cyclic GMP, or phosphoinositide signalling systems.     

Cyclic Nucleotide Phosphodiesterase Activity in Bovine Brain Coated Vesicles

Cyclic AMP phosphodiesterase activity in bovine brain coated vesicles displayed a Km of approximately 22 microM for cyclic AMP, a Vmax of 3.2 nmol/min/mg protein, and a Hill coefficient of 1.5, suggesting positive cooperativity. The enzyme activity was stimulated by cyclic GMP with maximal indexes of stimulation ranging between 40 and 300%. Both basal and stimulated phosphodiesterase activities were immunotitrated with polyclonal antibodies against clathrin attached to heat-inactivated, formaldehyde-fixed Staphylococcus aureus cells. The main form of phosphodiesterase activity present in the immunoprecipitated brain coated vesicle preparation also is stimulated by cyclic GMP. The allosteric behavior was modulated by cyclic GMP. All of these properties are typical of type II or cyclic GMP-sensitive phosphodiesterases in addition to their calcium and calmodulin independence. Competition experiments with a series of phosphodiesterase inhibitors, papaverine, 1-methyl-3-isobutylxanthine, and theophylline, showed inhibition of cyclic AMP hydrolysis. Trifluoperazine was inactive at the highest concentration used, 100 microM. These compounds also inhibited the cyclic GMP-stimulated cyclic AMP hydrolysis with trifluoperazine practically inactive. At 5 microM cyclic AMP none of the inhibitors was seen to stimulate the cyclic AMP hydrolytic activity. The presence of an enzyme for the breakdown of cyclic nucleotides in brain coated vesicles may suggest a role for these second messengers in the in vivo functions of this organelle.     

See-Saw Signal Processing in Pinealocytes Involves Reciprocal Changes in the α1-Adrenergic Component of the Cyclic GMP Response and the β-Adrenergic Component of the Cyclic AMP Response

Pineal cyclic AMP and cyclic GMP are regulated by norepinephrine (NE) acting through alpha 1- and beta-adrenoceptors. beta-Adrenergic stimulation appears to be an absolute requirement and alpha 1-adrenergic activation amplifies beta-adrenergic stimulation of the cyclic AMP response 10-fold and the cyclic GMP response 100-fold, respectively. Chronic deprivation of adrenergic stimulation, due to exposure to constant light (LL) or by surgical denervation, enhances the cyclic AMP response and diminishes the cyclic GMP response as compared to control animals in a 10:14 light/dark (LD) cycle. This phenomenon is termed see-saw signal processing. In the current study we find these changes do not reflect shifts in the time course or Ka of these responses. Dose-response studies indicate the beta-adrenergic component of cyclic AMP stimulation is enhanced and the alpha 1-adrenergic component of cyclic GMP stimulation is diminished in LL pinealocytes. Several observations indicate these changes may reflect alterations in Ca2+-sensitive postreceptor mechanisms.     

Amygdala Kindling Modifies Extracellular Opioid Peptide Content in Rat Hippocampus Measured by Microdialysis

Abstract: Opioid peptide release in the hippocampus was shown to be increased immediately following amygdala kindling stimulation in freely moving rats using microdialysis combined with a universal opioid peptide radioimmunoassay (RIA). Extracellular opioid peptide levels were elevated (55% above basal levels) within the first 10 min after electrical stimulation-induced partial seizures in previously nonkindled animals. Fully kindled rats showed lower extracellular opioid peptide levels (40% reduction) during the interictal period [16 ± 2.1 days (mean ± SEM) after the last stage V seizure], in comparison with values obtained from the sham-kindled group under basal conditions. However, opioid peptide release in fully kindled rats increased above 152% of interictal levels within the first 20 min after onset of fully kindled seizures, attaining peak levels equal to that of the partial kindled group and returning to prestimulation conditions 40–60 min following the ictal events. The majority of the immunoreactive material recovered from the hippocampus within the first 20 min following partial and generalized kindled seizures coeluted with dynorphin-A (1–6), dynorphin-A (1–8), and Leu-enkephalin by HPLC/RIA analysis. It is proposed that the enhanced opioid peptide release in hippocampus induced by amygdala kindling stimulation might be associated with either enhanced excitability or seizure suppression as seizure susceptibility fluctuates. The reduced interictal opioid peptide levels may also underlie some interictal behavioral disturbances.     

Somatostatin release in the hippocampus in the kindling model of epilepsy: a microdialysis study

Somatostatin biosynthesis in the hippocampus is activated during and following kindling epileptogenesis. The aim of this study was to investigate whether this phenomenon is associated with enhanced somatostatin release in vivo. Experiments have been run in awake, freely moving rats, implanted with a bipolar electrode in the right amygdala (for kindling stimulation), and with a recording electrode and a microdialysis probe in the left hippocampus. Basal somatostatin-like immunoreactivity (-LI) release was significantly greater in kindled than naive rats. In naive rats, a 2-min perfusion with 100 mM K(+) did not affect behavior and EEG recordings and nonsignificantly increased somatostatin-LI release; a 10-min K(+) perfusion evoked numerous wet dog shakes, electrical seizures (class 0; latency congruent with 8 min, duration congruent with 8 min), and somatostatin-LI release ( congruent with 350% of basal); and a single kindling after-discharge (4 +/- 3-s duration in the hippocampus) also evoked somatostatin-LI release ( congruent with 200% of basal). In kindled rats, a 2-min 100 mM K(+) perfusion evoked hippocampal discharges in three of seven animals (latency congruent with 2 min, mean duration congruent with 1.5 min) and increased somatostatin-LI release ( congruent with 250% of basal); a 10-min K(+) perfusion evoked behavioral seizures (class 1 to 5, latency congruent with 4 min, mean duration congruent with 12 min) with numerous wet dog shakes and robust somatostatin-LI release ( congruent with 350% of basal); and a kindling stimulation evoked generalized seizures (class 4 or 5, 77 +/- 15-s duration in the hippocampus) with remarkable somatostatin-LI release ( congruent with 300% of basal). These data demonstrate that hippocampal somatostatin release is increased in the kindling model in vivo.     

Alterations in lipid and calcium metabolism associated with seizure activity in the postischemic brain

Transient ischemia is known to lead to a long-lasting depression of cerebral metabolic rate and blood flow and to an attenuated metabolic and circulatory response to physiological stimuli. However, the corresponding responses to induced seizures are retained, demonstrating preserved metabolic and circulatory capacity. The objective of the present study was to explore how a preceding period of ischemia (15 min) alters the release of free fatty acids (FFAs) and diacylglycerides (DAGs), the formation of cyclic nucleotides, and the influx/efflux of Ca(2+), following intense neuronal stimulation. For that purpose, seizure activity was induced with bicuculline for 30 s or 5 min at 6 h after the ischemia. Extracellular Ca(2+) concentration (Ca(2+)(e)) was recorded, and the tissue was frozen in situ for measurements of levels of FFAs, DAGs, and cyclic nucleotides. Six hours after ischemia, the FFA concentrations were normalized, but there was a lowering of the content of 20:4 in the DAG fraction. Cyclic AMP levels returned to normal values, but cyclic GMP content was reduced. Seizures induced in postischemic animals showed similar changes in Ca(2+)(e), as well as in levels of FFAs, DAGs, and cyclic nucleotides, as did seizures induced in nonischemic control animals, with the exception of an attenuated rise in 20:4 content in the DAG fraction. We conclude that, at least in the neocortex, seizure-induced phospholipid hydrolysis and cyclic cAMP/cyclic GMP formation are not altered by a preceding period of ischemia, nor is there a change in the influx/efflux of Ca(2+) during seizure discharge or in associated spreading depression.     

Pharmacology of excitatory amino acid receptors mediating the stimulation of rat cerebellar cyclic GMP levele invitro

L-glutamate and related amino acids produced dose-related increases in cyclic GMP concentrations in cerebellar slices from 8-day old rats. The effects of L-glutamate, L-aspartate, and N-methyl-D-aspartate were antagonised in part by glutamate diethylester, while D-α-aminoadipate and DL-α-aminosuberate were ineffective. Kainic acid, although producing an effective stimulation of cyclic GMP, failed to achieve the same maximal response as glutamate, suggesting different sites of action. Cyclic GMP levels were also increased, although to a lesser extent, by L-glutamate in slices from hippocampus, medulla, hypothalamus, cortex, striatum and spinal cord of young animals. In the adult, however, this response was no longer observed.     

C-Type Natriuretic Peptide Is a Potent Stimulator of Cyclic GMP Production in Cultured Mouse Astrocytes

The effect of C-type natriuretic peptide (CNP), a novel member of the natriuretic peptide family, on cyclic GMP (cGMP) generation was studied in primary cultures of mouse astrocytes. CNP stimulated cGMP production by mouse astrocytes in a dose-dependent fashion, with an EC50 of 32 nM and a maximal stimulatory concentration of greater than 1 microM, which induced a rise of cGMP level from a baseline of 1.0 +/- 0.1 pmol/mg of protein to 196.2 +/- 22.0 pmol/mg of protein. Compared with our previously reported atrial and brain natriuretic peptide-induced cGMP responses, CNP had a lower EC50 and was 10-20 times more efficacious in its maximal effect on cGMP stimulation. These data lend support to the concept of a significant role of CNP in neuromodulation/neurotransmission.     

Exogenous, but not endogenous, cyclic GMP reduces hepatic pyruvate kinase activity

We investigated the effects of exogenous cyclic GMP and stimulants of endogenous cyclic GMP accumulation on L-form (hepatic) pyruvate kinase (ATP: pyruvate 2-O-phosphotransferase, EC 2.7.1.40) activity in isolated rat hepatocytes. Exogenous cyclic GMP (200 muM) reduced pyruvate kinase activity, but was less potent than exogenous cyclic AMP (50 muM) (Ki congruent to 120 muM vs. 30 muM, respectively), had a slower onset of action (1.0 vs. 0.3 min, respectively) and a less rapid maximal effect (5.0 vs. 1.0 min, respectively). Similar results were noted with dibutyryl cyclic GMP or dibutyryl cyclic AMP. 1.0 muM acetylcholine increased cyclic GMP concentrations in isolated hepatocytes from 233 +/- 16 to 447 +/- 3 pmol/g cell protein (P less than 0.001), but did not alter pyruvate kinase activity. Similar results were noted with carbamylcholine, NaN3 or acetylcholine plus eserine sulfate. The results suggest a differential effect of exogenous vs. endogenous cyclic GMP on L-form pyruvate kinase activity, and question the physiological relevance of observations with exogenous cyclic GMP in this system.     

Regional sensitivity of cyclic AMP and cyclic GMP in rat brain to central cholinergic stimulation

We determined cyclic AMP and cyclic GMP levels in 18 regions of rat brain following administration of two different centrally active cholinergic agonists. Administration of oxotremorine (2 mg/Kg IP), a muscarinic agonist, 10 minutes prior to sacrifice by exposure to high power microwave irradiation resulted in significant increases in cyclic GMP in cerebellum, brainstem, hippocampus, midbrain, thalamus and septal region. Cyclic AMP levels were significantly elevated in substantia nigra, nucleus interpeduncularis, hypothalamus, brainstem, midbrain and in the pituitary where a greater than tenfold increase was observed. Levels of plasma prolactin and corticosterone did not differ in any of the groups examined, but growth hormone was significantly lower in animals exposed to oxotremorine. Physostigmine (0.5 mg/Kg) a cholinesterase inhibitor, administered IP also produced elevations in cyclic AMP and cyclic GMP in several of the brain regions examined. These results indicate that multiple regions of the brain are responsive to central cholinergic activation of not only cyclic GMP, but also cyclic AMP system.     

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.     

Cannabinoid Receptors and Modulation of Cyclic AMP Accumulation in the Rat Brain

The mechanism by which cannabinoid compounds produce their effects in the rat brain was evaluated in this investigation. Cannabinoid receptors, quantitated by [3H]CP-55,940 binding, were found in greatest abundance in the rat cortex, cerebellum, hippocampus, and striatum, with smaller but significant binding also found in the hypothalamus, brainstem, and spinal cord. Using rat brain slice preparations, we evaluated the effect of desacetyllevonantradol on basal and forskolin-stimulated cyclic AMP accumulation in the regions exhibiting the greatest cannabinoid receptor density. Desacetyllevonantradol (10 microM) reduced cyclic AMP levels in the hippocampus, frontal cortex, and striatum. In the cerebellum, however, the response to desacetyllevonantradol was biphasic with cyclic AMP accumulation being decreased at lower and increased at higher concentrations. Desacetyllevonantradol reduced cyclic AMP accumulation in isoproterenol-stimulated slices in the cortex and cerebellum, but not in the hippocampus. Cells that responded to vasoactive intestinal peptide with an increase in cyclic AMP accumulation in the hippocampus and cortex also responded to desacetyllevonantradol. The modulation of cyclic AMP accumulation by desacetyllevonantradol could be attenuated following stereotaxic implantation of pertussis toxin, supporting the involvement of a G protein in the cannabinoid response in the brain. However, other actions of cannabinoid compounds may also affect the cyclic AMP levels in brain slice preparations.     

Direct Demonstration of the Activation of UDP-N-Acetylgalactosamine: [GM3]N-Acetylgalactosaminyltransferase by Cyclic AMP

Treatment of NG108-15 cells in culture with the opiate peptide [D-Ala2,D-Leu5]enkephalin produces maximal inhibition of cyclic AMP synthesis in less than 15 min. The activity of [GM3]:N-acetylgalactosaminyltransferase is similarly inhibited, but maximal inhibition is not observed for at least 30 min following the addition of [D-Ala2,D-Leu5]enkephalin. Conversely, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine rapidly potentiates the intracellular accumulation of cyclic AMP and, in a more gradual fashion, increases [GM3]:N-acetylgalactosaminyltransferase activity. The reductions in the activity of [GM3]:N-acetylgalactosaminyltransferase that occur following treatment of NG108-15 cells with indomethacin argues for a direct role of cyclic AMP in the observed changed in [GM3]:N-acetylgalactosaminyltransferase activity. By adding low concentrations of cyclic AMP (but not cyclic GMP) to microsomes derived from neonatal rat brain, we were able to demonstrate a dose-dependent phosphorylation of membrane protein and subsequent doubling of [GM3]:N-acetylgalactosaminyltransferase activity.     

Cyclic AMP Concentrations in Rat Neocortex and Hippocampus During and Following Incomplete Ischemia: Effects of Central Noradrenergic Neurons, Prostaglandins, and Adenosine

The concentrations of cyclic AMP, noradrenaline, glycogen, glucose, lactate, pyruvate, labile phosphate compounds, and free fatty acids were investigated in the rat neocortex and hippocampus during and following cerebral ischemia. An incomplete ischemia of 5 and 15 min duration was induced by bilateral carotid clamping combined with hypotension. The postischemic events were studied after 5, 15, and 60 min of recirculation. Five minutes of ischemia did not significantly alter the neocortical or hippocampal concentrations of cyclic AMP. After 15 min of ischemia the neocortical levels decreased significantly below control values. In the recirculation period following ischemia a significant elevation of the cyclic AMP concentrations was observed. Following 5 min of recirculation after 5 min of ischemia the levels increased from 2.53 +/- 0.21 nmol X g-1 to 5.18 +/- 0.09 nmol X g-1 in the neocortex and from 2.14 +/- 0.16 nmol X g-1 to 3.52 +/- 0.35 nmol X g-1 in the hippocampus. Five minutes of recirculation following 15 min of ischemia led to a significant increase in the levels of cyclic AMP, to 12.86 +/- 1.43 nmol X g-1 in the neocortex to 5.58 +/- 0.57 nmol X g-1 in the hippocampus. With longer recirculation periods the cyclic AMP levels progressively decreased and were similar to control values after 60 min. Depletion of cortical noradrenaline by at least 95% was performed by injections of 6-hydroxydopamine into the ascending axon bundles from the locus ceruleus. The lesion did not significantly change the ischemic or post-ischemic neocortical and hippocampal levels of cyclic AMP, glycogen, or free fatty acids including arachidonic acid.(ABSTRACT TRUNCATED AT 250 WORDS)     

Change in levels of cyclic AMP and cyclic GMP during pregnancy and larval development of the tsetse fly, Glossina morsitans

Cyclic AMP and cyclic GMP levels change very little in response to feeding and mating, but during pregnancy and at parturition major changes can be detected in both the mother and larva. In both the female head and larva (whole body) cyclic AMP levels reach a peak at parturition. In the larval brain and ring gland cyclic AMP is at its lowest at parturition but rises sharply, reaching a peak 1.5 hr later at the time of pupariation . Though cyclic AMP levels in the head and thorax are consistently 10-60 times greater than levels of cyclic GMP, both the female abdomen and larva contain high concentrations of cyclic GMP with a ratio of cyclic AMP:cyclic GMP approaching 1:1. In the abdomen, the pattern of high cyclic GMP closely parallels the activity cycle of the female's milk gland.     

Activation of Tyrosine Hydroxylase in PC12 Cells by the Cyclic GMP and Cyclic AMP Second Messenger Systems

Tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis, is subject to regulation by a variety of agents. Previous workers have found that cyclic AMP-dependent protein kinase and calcium-stimulated protein kinases activate tyrosine hydroxylase. We wanted to determine whether cyclic GMP might also be involved in the regulation of tyrosine hydroxylase activity. We found that treatment of rat PC12 cells with sodium nitroprusside (an activator of guanylate cyclase), 8-bromocyclic GMP, forskolin (an activator of adenylate cyclase), and 8-bromocyclic AMP all produced an increase in tyrosine hydroxylase activity measured in vitro or an increased conversion of [14C]tyrosine to labeled catecholamine in situ. Sodium nitroprusside also increased the relative synthesis of cyclic GMP in these cells. In the presence of MgATP, both cyclic GMP and cyclic AMP increased tyrosine hydroxylase activity in PC12 cell extracts. The heat-stable cyclic AMP-dependent protein kinase inhibitor failed to attenuate the activation produced in the presence of cyclic GMP. It eliminated the activation produced in the presence of cyclic AMP. Sodium nitroprusside also increased tyrosine hydroxylase activity in vitro in rat corpus striatal synaptosomes and bovine adrenal chromaffin cells. In all cases, the cyclic AMP-dependent activation of tyrosine hydroxylase was greater than that of the cyclic GMP-dependent second messenger system. These results indicate that both cyclic GMP and cyclic AMP and their cognate protein kinases activate tyrosine hydroxylase activity in PC12 cells.