Dual control of pupal diapause by cyclic nucleotides in the bertha armyworm,Mamestra configurata Wlk.

Treatment of diapausing pupae of M. configurata with dibutyryl cyclic AMP or 8-(4-chlorophenylthio) cyclic AMP (CPT cyclic AMP) reduced the incidence of eclosion to zero compared to about 15% for controls, whereas treatment with cyclic GMP increased eclosion to more than 90%. Treatment with the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) resulted in a high incidence (79.8%) of eclosion, but treatment with dibutyryl cyclic AMP + IMBX or CPT cyclic AMP + IBMX gave low incidences (<9.1%) of eclosion. Other methylxanthines (theophylline, 8-phenyltheophylline, caffeine) and papaverine had relatively little effect on eclosion even at high doses.Treatment of post-diapause pupae with dibutyryl cyclic AMP or CPT cyclic AMP resulted in a low incidence (<5.0%) of eclosion compared to 98.8% eclosion in controls. Suppression of eclosion was more effective if dibutyryl cyclic AMP was given within the first 2 days of pupal-adult development at 20°C and became less effective as development progressed, indicating that dibutyryl cyclic AMP inhibits endocrine events initiating development rather than inhibiting subsequent metamorphic development. Treatment of post-diapausing pupae with cyclic GMP, IBMX, other methylxanthines or papaverine did not affect eclosion. These results are consistent with a dual control of pupal diapause in M. configurata by cyclic nucleotides, with cyclic AMP acting to maintain diapause and cyclic GMP acting to terminate it.     

Inactivation of cyclic GMP-dependent protein kinase by N alpha-tosyl-L-lysine chloromethylketone

Cyclic GMP-dependent protein kinase from bovine lung and cyclic AMP-dependent protein kinase from bovine heart are inactivated by N^α-tosyl-L-lysine chloromethylketone (TLCK) in the presence of cyclic GMP and cyclic AMP, respectively. The inactivation of both protein kinases is pseudo-first order, suggesting the rate limiting step is beyond the binding of TLCK. Cyclic GMP-dependent protein kinase is inactivated less than $\text{1}\text{4}$ as rapidly as cyclic AMP-dependent protein kinase, although it shows a higher apparent affinity for TLCK. Cyclic AMP stimulated the rate of inactivation of cyclic AMP-dependent protein kinase 10-fold but cyclic GMP stimulated the rate of inactivation of cyclic GMP-dependent protein kinase only 1.5-fold. The rate of inactivation of cyclic GMP-dependent protein kinase by TLCK is sufficiently rapid (half-time of about 30 min at 37°C with 2 mM TLCK) to account for the effects of TLCK on cell growth observed by others.     

Variation of the levels of cyclic AMP and cyclic GMP during development of the insect Ceratitis capitata.

Changes in the levels of adenosine 3′,5′-monophosphate (cyclic AMP) and guanosine 3′,5′-monophosphate (cyclic GMP) during development were studied in the Dipterous Ceratitis capitata. The developmental patterns were different to each other. Cyclic AMP showed a sharp maximum in the larval stage to decrease afterwards during adult development. Changes of cyclic GMP exhibited an opposite pattern, although its levels were always higher than those of cyclic AMP.     

A partial characterization of the cyclic nucleotide phosphodiesterases of Drosophila melanogaster

The cyclic nucleotide phosphodiesterases in crude homogenate, soluble material, and particulate preparations of adult Drosophila melanogaster flies, hydrolyze cyclic AMP with nonlinear kinetics. Cyclic GMP is hydrolyzed by the phosphodiesterases in crude homogenate and soluble material with linear kinetics. Physical separation techniques of gel filtration, velocity sedimentation, and ion-exchange chromatography reveal that Drosophila soluble fraction contains two major forms of cyclic nucleotide phosphodiesterase. Form I hydrolyzes both cyclic AMP and cyclic GMP. Inhibition experiments suggest that the hydrolysis of both cyclic nucleotides by Form I occurs at a single active site. The K_m's for hydrolysis of both substrates are about 4 μm. This form has a molecular weight of about 168,000 as estimated by gel nitration. Form II cyclic nucleotide phosphodiesterase is specific for cyclic AMP as substrate. Gel filtration indicates that this form has a molecular weight of about 68,000. The K_m for cyclic AMP is about 2 μm.     

Brain cyclic AMP levels and the initiation of adult development in the Cecropia silkmoth.

A six-fold increase in the level of brain cyclic AMP is observed in chilled Cecropia pharate adults within 24 hr of transfer from the cold to room temperature. This increase is not observed in pupae chilled for a period insufficient to allow initiation of adult development, nor after injury to diapausing pupae. Other tissues show a variable and minor response during initiation. Injected dibutyryl cAMP will cause initiation in insufficiently chilled pupae, but not in dauer pupae. The possible relationship of this rise in cAMP to the process of initiation is discussed.     

Interaction of cyclic nucleotides and ecdysterone in breaking the pupal diapause of the bertha armyworm, Mamestra configurata WLK.

Cyclic GMP and cyclic AMP have distinct and opposite effects upon the action of ecdysterone in diapausing pupae of the Bertha armyworm, $\text{Mamestra}$$\text{configurata}$. Cyclic GMP enhanced the effectiveness of suboptimal doses of ecdysterone in breaking diapause; the amount of cyclic GMP required to lower the ED50 of ecdysterone by half was 80 μg/g. Dibutyryl cyclic GMP had no apparent effect on the action of ecdysterone over a wide dose range (0.07 – 70 μg/g). On the other hand, cyclic AMP and dibutyryl cyclic AMP effectively blocked the diapause-breaking action of ecdysterone when administered simultaneously with the steroid hormone. The amount of cyclic AMP required to reduce the incidence of diapause termination from 100% to 50% was 60 μg/g; for dibutyryl cyclic AMP the amount required was only 14 μg/g. No cyclic nucleotide tested in the study could by itself break the pupal diapause of $\text{M.}$$\text{configurata}$. The concept that cyclic GMP and cyclic AMP provide at least different if not opposing regulatory influences in certain insect systems is discussed briefly in the light of these observations.     

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.     

Characterization of particulate cyclic nucleotide phosphodiesterases of rat kidney.

Particulate cyclic nucleotide phosphodiesterases of rat kidney display some distinct kinetic and regulatory properties. Only a small portion (5–10%) of the total homogenate low K_m cyclic AMP phosphodiesterase activity (measured with concentrations of cyclic AMP less than l μm) is tightly associated with kidney membranes. Cyclic GMP phosphodiesterase activity (measured with 0.25–200 μm cyclic GMP) is readily detectable in these fractionated and washed membranes. Low concentrations of cyclic GMP stimulated the hydrolysis of cyclic AMP (K_a ～- 0.5 μM), an effect not noted in most other membrane systems. High concentrations of cyclic GMP (K_i ～- 450 μM) and cyclic AMP (K_i ～- 150 μM) inhibited the hydrolysis of each other noncompetitively. Solubilization of membrane bound activities by sonication or Sarkosyl L markedly alters enzyme kinetic properties and the responses to cyclic nucleotides and sulfhydryl reagents. Incubation of membrane fractions with dithiothreitol (5 mm) or storage of the membranes at 4 °C results in a change in extrapolated kinetic constants for cyclic AMP hydrolysis and an increase in the rate of denaturation at 45 °C. Our findings raise the possibility that regulation of membrane-bound cyclic nucleotide phosphodiesterase activity involves interactions with cyclic nucleotides themselves, as well as oxidation and reduction of disulfide bonds and membrane-enzyme interactions.     

Two types of cyclic GMP binding site associated with the cyclic AMP-dependent protein kinase from lymphocytes

Low- and high-affinity binding sites for cyclic GMP were found to be associated with the cyclic AMP-dependent protein kinase (ATP: protein phosphotransferase, EC 2.7.1.37) from human tonsillar lymphocytes, but neither of them was identical with the cyclic AMP binding site.The enzyme activated by cyclic GMP phosphorylated the same site of calf thymus H2b histone as the cyclic AMP activated enzyme; however, more complex kinetics of activation were found with cyclic GMP.Two classes of cyclic GMP binding site were demonstrated by kinetic analysis of cyclic [³H]GMP binding in the enzyme preparations eluted by 0.1 M potassium phosphate (pH 7.0) from DEAE cellulose. The high-affinity cyclic GMP binding site (K_d about 44 · 10^?8 M belonged to some complex form of the protein kinase, as evidenced by the mutual inhibition of cyclic AMP binding and high affinity cyclic GMP binding. However, the high-affinity cyclic GMP binding site disappeared on Sephadex G-100 gel chromatography of the enzyme preparation, whereas the cyclic AMP binding activity was recovered quantitively as separate fractions. The low-affinity cyclic GMP binding site (K_d 2–5 · 10^?6 M) was demonstrated by the inhibitory effect of 10^?5 M cyclic GMP on cyclic AMP binding in each cyclic AMP binding fraction obtained by gel chromatography. However, cyclic AMP did not inhibit the binding of cyclic GMP to the low-affinity binding site.     

Fluctuations in cyclic adenosine 3':5'-monophosphate and cyclic guanosine 3':5'-monophosphate during the mitotic cycle of the acellular slime mould Physarum polycephalum.

Cyclic adenosine 3′:5′-monophosphate (cyclic AMP) and cyclic guanosine 3′:5′-monophosphate (cyclic GMP) have been determined at half-hourly intervals throughout the mitotic cycle of Physarum polycephalum. Cyclic AMP was constant at 1pmole/mg protein throughout except for a transient peak of 17pmoles/mg protein in the last quarter of G2. Cyclic GMP was more variable (2–4pmole/mg protein) rising to 9.5pmole/mg protein during the 3 hour S period and to 7pmole/mg protein during the last hour of G2. The significance of these changes is discussed.     

Ecdysteroids and diapause in pupae of Heliothis punctiger

Most pupae of H. punctiger enter diapause when reared at 19°C, 12L:12D. When pharate pupae were treated for only 12 hr at 28°C about 50% developed at 19°C. The proportion of non-diapausing pupae increased as the temperature at which the pharate pupal stage was spent increased.The quantity of injected 20-hydroxyecdysone necessary to promote development in diapausing pupae varied from about 1 μg g^?1 soon after pupation to about 4 μg g^?1 after 50 days. It fell somewhat after 150 days.Removing brains from non-diapausing pupae showed that the brain secreted its hormone at the time of pupation (or just before). However, if the pupae were kept at 19°C development did not occur unless the brain remained in situ for at least 20 hr at 28°C. Implanting brains from non-diapausing pupae into diapausing ones had no measurable effect.These results may be explained by postulating that the prothoracic gland is ‘activated’ by exposure to high temperature, but that it reverts to inactivity over a period at 19°C. The ‘active’ gland must then be stimulated by brain hormone for a long period to trigger secretion of its hormone, which results in development. Diapause is thus the result of the failure of the prothoracic gland to secrete.     

A direct, stimulating effect of cyclic GMP on purified phosphoribosyl pyrophosphate synthetase and its antagonism by cyclic AMP

The activity of phosphoribosyl pyrophosphate synthetase, purified from a line of rat hepatoma cells in continuous culture, is maximally stimulated (2–4 fold) by less than 10^?7M cyclic GMP. Half maximal stimulation occurs at 2 × 10^?9M. Cyclic GMP stimulates phosphoribosyl pyrophosphate synthetase by decreasing the Km of the enzyme for ATP from 50 μM to 10 μM without affecting the Vmax; it has no effect on the Km for ribose 5-phosphate, the other substrate. Cyclic AMP alone has no effect on the enzyme activity, but at micromolar concentrations it antagonizes the stimulation by cyclic GMP. GMP, GDP, and GTP do not stimulate enzyme activity; and AMP and ADP at micromolar concentrations do not antagonize the effect of cyclic GMP.There is no detectable cyclic nucleotide-activated protein kinase in the enzyme preparation. Cyclic GMP significantly stabilizes the enzyme to heat inactivation. We conclude that cyclic GMP binds directly to the enzyme in an allosteric fashion, causing it to have an increased affinity for one of its substrates, and that cyclic AMP directly antagonizes this effect.     

Signal transduction for Schistocerca gregaria ion transport peptide is mediated via both cyclic AMP and cyclic GMP

The second messengers involved in the signal transduction for Schistocerca gregaria, ion transport peptide (Schgr-ITP) that regulates ion and fluid transport across the ileum of the desert locust S. gregaria, were measured using competitive enzyme-linked immunosorbent assays (ELISAs). Synthetic Schgr-ITP elevates intracellular levels of both cyclic AMP and cyclic GMP, measured over a 15 min period in the presence of 3-isobutyl-1-methylxanthine, in a dose-dependent manner. Furthermore, crude corpora cardiaca (CC) extracts elevate intracellular cyclic AMP levels 2-fold greater than Schgr-ITP, suggesting that factors present in the CC, other than Schgr-ITP, also act via this second messenger. These results suggest that the interaction of Schgr-ITP with two separate receptors, most likely a G-protein coupled receptor and a membrane bound guanylate cyclase, elevates intracellular levels of cyclic AMP and cyclic GMP to regulate ion and fluid transport across the locust ileum. Cyclic AMP stimulates Cl⁻, K⁺ and Na⁺ reabsorption, whereas secretion of H⁺ into the lumen of the ileum is most likely mediated via cyclic GMP. Cyclic GMP also stimulates Cl⁻ uptake in a similar manner to cyclic AMP. The measurement of tissue (central nervous system) levels of Schgr-ITP using an indirect ELISA confirms that the peptide is only present in the locust brain and the CC. The amounts present are greatest in the CC, where the peptide is presumably stored for release into the hemolymph when locusts feed.     

Effects of isoproterenol,theophylline and carbachol on cyclic nucleotide levels and relaxation of bovine tracheal smooth muscle

The effect of theophylline and isoproterenol on bovine tracheal smooth muscle tension and cyclic AMP levels was investigated. Concentrations of isoproterenol (4 × 10^?6 M) and theophylline (10 mM) that relaxed carbachol-contracted tracheal muscle by 85–95% did not significantly elevate control levels of cyclic AMP. In the absence of carbachol, several-fold increases in cyclic AMP were caused by isoproterenol although no elevations by theophylline were measurable. However, when isoproterenol and theophylline were administered together, theophylline potentiated the rise in cyclic AMP caused by isoproterenol. Phosphodiesterase studies in tracheal muscle showed the presence of a high and a low K_m enzyme which were inhibited by theophylline. Cyclic GMP levels were elevated in muscles contracted by carbachol as well as in carbachol-contracted muscles that had been relaxed by theophylline. In non-tension studies, in which the tracheal muscle was not under isometric tension, carbachol or theophylline alone increased cyclic GMP and together they synergistically elevated cyclic GMP. Atropine blocked the elevation caused by carbachol but not that caused by theophylline. In contrast to theophylline, isoproterenol did not elevate cyclic GMP, and in carbachol-contracted muscles that had been relaxed by isoproterenol, cyclic GMP levels were no different from control. Also, in non-tension studies, isoproterenol decreased basal cyclic GMP and antagonized the increase in cyclic GMP due to carbachol.The results indicate that whole-tissue levels of cyclic AMP and cyclic GMP do not correlate with the state of tracheal smooth muscle tension. Cyclic GMP levels do not clearly correlate with either contraction or relaxation. The inhibition by carbachol of increases in cyclic AMP due to isoproterenol and the inhibition by isoproterenol of increases in cyclic GMP due to carbachol provide evidence for a reciprocal cholinergic-adrenergic antagonism of cyclic AMP and cyclic GMP levels. The antagonism did not appear to be due to either cyclic nucleotide affecting the elevation of the other since the levels of both cyclic nucleotides were depressed.     

Fetal and postnatal development of cyclic GMP phosphodiesterase activity in the rat brain

Cyclic GMP concentration and cyclic GMP phosphodiesterase activity were studied in rat mothers and fetuses at 17, 19 and 21 days of intrauterine life and 0, 1, 4, 10, 15,20, 30 and 45 days after birth. During this developmental period, the increase in cyclic GMP concentration was discrete and the value in 15-day-old rats was already similar to the adult level. Cyclic GMP phosphodiesterase activity increased from 17- to 19-day fetuses and was significantly reduced in 21-day fetuses, neonates, and 1-day-old rats. This reduction may be a result of fetal endocrine preparation for parturition. During postnatal development, cyclic GMP phosphodiesterase activity increased in a parallel way in the limbic system, corpora striata, cerebral hemispheres, and diencephalon, reaching maximal level between 20 and 30 days after birth, and then decreasing to the adult value. The highest activity was found in corpora striata and the lowest in diencephalon. Cerebellar cyclic GMP phosphodiesterase activity was very high in the 4-day-old rat (257% of adult value) and diminished significantly in the 10-day-old rat with no subsequent changes. Kinetic analysis of the enzyme during postnatal forebrain development showed an increase in both the V_max and the apparent K_m. A decrease in the enzyme's V_max was observed only in the cerebellum.The importance of cyclic GMP phosphodiesterase regulation of cyclic GMP concentrations in the brain during development is discussed.     

Levels of cyclic nucleotides in mouse regional brain following 300 ms microwave inactivation.

Abstract— The uniformity and speed of inactivation of mouse brain adenylate cyclase, guanylate cyclase and cyclic nucleotide phosphodiesterase were measured after 6 kW microwave irradiation (MWR). Inactivation of enzymes was uniform throughout the brain during heating and 100% loss of activity was evident after 300 ms. MWR. For comparison of effects of inactivation times on levels of cyclic nucleotides measured in regional brain areas, cyclic AMP and cyclic GMP were estimated after 1.5 kW MWR requiring 4 s of heating and 6 kW MWR requiring 300 ms. Except for corpus striatum, uniformly lower levels of cyclic AMP were measured following 300 ms vs. 4s MWR . There was no change in cyclic GMP levels in regional brain areas after 4s vs. 300 ms MWR . Cyclic AMP and cyclic GMP were measured from the same regional brain tissue samples after 300 ms and ratios calculated. The finding of much lower cyclic AMP:cyclic GMP ratios than had previously been reported suggests that slow inactivation times provide for the measurement of regional brain cyclic nucleotide values which are not consistent with the in-vivo state.     

The cyclic nucleotide phosphodiesterases of spinach chloroplasts and microsomes

Cyclic nucleotide phosphodiesterase was extracted from intact chloroplasts and partially purified. Peak 1_c activity from Sephadex G-200 was resolved by electrophoresis into two major bands (MWs 1.87 × 10⁵ and 3.7 × 10⁵). Both also possessed acid phosphatase, ribonuclease, nucleotidase and ATPase. The chloroplast peak 1_c cyclic nueleotide phosphodiesterase was located in the envelope. Peak 1_m cyclic nucleotide phosphodiesterase obtained from the microsomal fraction had a MW of 2.63 × 10⁵. Electrophoresis separated 1_m into two bands of cyclic nucleotide phosphodiesterase activity (MWs 2.63 × 10⁵ and 1.28 × 10⁵). Both contain ATPase, ribonuclease, nucleotidase, but not acid phosphatase. Peak 1_c has high activity towards 3′:5′-cyclic AMP and 3′:5′-cyclic GMP but little towards 2′:3′-cyclic nucleotides. Peak 1_m showed most activity towards 2′:3′-cyclic AMP, 2′:3′-cyclic GMP and 2′:3′-cyclic CMP with little activity towards 3′:5′-cyclic nucleotides. With 1_c, 3′:5′-cyclic AMP and 3′:5′-cyclic GMP exhibit mixed-type inhibition towards one another. The 2′:3′-cyclic AMP phosphodiesterase 1_m was competitively inhibited by 2′:3′-cyclic GMP. p-Chloromercuribenzoate inhibits 1_c but not 1_m. Electrophoresis after dissociation indicates that 1_c and 1_m are both enzyme complexes. After dissociation, the 1_c complex but not that of 1_m could be reassociated. The ribonuclease of the 1_m complex hydrolyses RNA to yield 2′:3′-cyclic nucleotides as the main products. These results are compatible with the 1_c cyclic nucleotide phosphodiesterase complex being involved in the metabolism of 3′:5′-cyclic AMP, and the 1_m complex being concerned with RNA catabolism.     

A kinetic analysis of the cyclic nucleotide phosphodiesterase regulation by the endogenous protein activator. A study of rat brain and frog sympathetic chain.

The effect of the endogenous protein activator on the kinetic characteristics of a highly purified, activator-deficient rat brain phosphodiesterase (EC 3.1.4.-) of a highly purified, activator-deficient rat brain phosphodiesterase (EC 3.1.4-) was studied. This enzyme preparation has only a high Km for cyclic AMP and a low Km for cyclic GMP. In the presence of 20 muM Ca2+, saturating concentrations of the activator decreased the Km of this enzyme for cyclic AMP from 350 muM to about 80 muM, without changing the V. The phosphodiesterase activator did not change the Km of phosphodiesterase for cyclic GMP; however, amoderate increase of V was seen. The activator lacks species specificity; the activator isolated from the bullfrog sympathetic chain produced the same qualitative and comparable quantitative changes in the kinetic properties of the purified rat brain phosphodiesterase. Cyclic GMP is a potent competitive inhibitor of the phosphodiesterase activation by the activator (Ki=1.8 muM), using cyclic AMP as a substrate. Cyclic AMP inhibits slightly the hydrolysis of cyclic GMP by phosphodiesterase in the presence of activator (Ki=155 muM) only.     

Purification and succinylation of cyclic GMP from large volume samples and radioimmunoassay of succinyl cyclic GMP.

Improved procedures for isolation of cyclic GMP and cyclic AMP and radioimmunoassay of cyclic GMP with succinylation are described. Procedures involved include modified chromatography on alumina and succinylation of cyclic GMP followed by purification of succinyl cyclic GMP on a Dowex AG 1×8 column. These procedures are convenient and applicable to any volume up to 50 ml of tissue extracts and especially for isotonic incubation mixtures. This assay system is sensitive to 6 femtomoles of cyclic GMP/tube. On radioimmunoassay, free and antibody bound [¹²⁵I]-labeled cyclic GMP are separated by Millipore filtration. Cyclic GMP levels in several tissue samples were determined in order to show the applicability of the procedures.     

The Influence of Hypoxia on the Concentrations of Cyclic Nucleotides in the Rat Brain

Abstract: In order to study the influence of hypoxia on cyclic nucleotides in the brain, we reduced arterial Po, for 15–30 min in lightly anaesthetised and artificially ventilated rats to obtain values ranging from about 45 to about 10 mm Hg. In an additional group (arterial Po₂ 18–22 mm Hg), the tissue hypoxia was aggravated by moderate arterial hypotension (mean arterial blood pressure about 80 mm Hg). In all animals, electrocortical activity was recorded. Cyclic GMP concentrations in cerebral cortex were unchanged in all groups but one. In that group, in which tissue hypoxia was severe enough to induce a suppression-burst EEG pattern and a measurable reduction in the adenylate energy charge, cyclic GMP concentrations were slightly increased ( p < 0.05). Cyclic AMP concentrations remained unaltered at all degrees of hypoxia studied. It is concluded that changes in cyclic nucleotides in brain tissue occur first at such severe degrees of hypoxia of the duration studied that function and metabolism are profoundly altered.