Corticotropin releasing factor receptor-mediated stimulation of adenylate cyclase activity in the rat brain

Corticotropin releasing factor (CRF)-stimulated adenylate cyclase activity and receptor binding were examined in rat brain homogenates using a potent synthetic CRF analog--[D-Tyr3,D-Pro4,Nle18,21,alpha-helical]CRF3-41 (alpha-hel CRF3-41). Binding of alpha-hel CRF3-41 in the rat brain was saturable, reversible, of high affinity and exhibited relevant peptide specificity. This analog also stimulated adenylate cyclase activity of various brain regions; the greatest magnitude of stimulation was in the cerebral cortex followed by the septum, cerebellum and thalamus. Adenylate cyclase stimulation in the cerebral cortex was concentration-dependent with an ED50 of 2.5 +/- 0.4 nM for alpha-hel CRF3-41 and an ED50 of 16 +/- 2 nM for ovine and rat CRF. Maximal stimulation was comparable for all peptides. Agonist-stimulated adenylate cyclase activity was competitively blocked by the CRF antagonists. The inactive CRF analog, ovine CRF1-39, at concentrations less than 1 microM, did not significantly stimulate adenylate cyclase. Adrenalectomy, which has been reported to modulate CRF receptor number and CRF-stimulated adenylate cyclase activity in the anterior pituitary, had no effect on CRF receptor binding or CRF-stimulated adenylate cyclase activity in the cerebral cortex. These results suggest that, as in the anterior pituitary, at least some of the physiological responses mediated by CRF receptors in the brain utilize the cyclic nucleotide regulatory pathway as a post-receptor mechanism.     

The resolution of dopamine and beta 1- and beta 2-adrenergic-sensitive adenylate cyclase activities in homogenates of cat cerebellum, hippocampus and cerebral cortex.

The stimulation of adenylate cyclase by dopamine and various β-adrenergic agonists has been investigated in homogenates from 3 areas of cat brain: the cerebral cortex, cerebellum and hippocampus. The purpose of the study was to determine whether the β-adrenergic receptors coupled to adenylate cyclase could be classified as either β₁ and β₂ subtypes in the different regions studied.

The stimulation of adenylate cyclase by the β-adrenergic agonist, (−)isoproterenol (5 × 10⁻⁶M), was completely blocked by the specific β-adrenergic antagonist, (−)alprenolol (10⁻⁵ M), but not by the dopaminergic antagonist, fluphenazine (10⁻⁵ M), whereas the stimulation of adenylate cyclase by (−)epinephrine (10⁻⁴ M) was blocked to varying extents by these two drugs in each of the 3 regions studied. The (−)epinephrine effect was always blocked in the combined presence of (−)alprenolol and fluphenazine. The adenylate cyclase stimulation by (−)epinephrine which is not blocked by (−)alprenolol was due to interaction of (−)epinephrine with a dopaminergic-sensitive adenylate cyclase which has been characterized in cerebral cortex, hippocampus and cerebellum.

Regional differences in the affinity of β-adrenergic-sensitive adenylate cyclase for various agonists were investigated in the presence of fluphenazine (10⁻⁵ M). In the cerebellum the potency order was (±)protokylol> (±)hydroxybenzylisoproterenol> (±)isoproterenol> (−)epinephrine> (±)salbutamol> (−)norepinephrine, indicating the presence of a β₂-adrenergic receptor. In the cerebral cortex the potency order was (−)isoproterenol> (±)protokylol> (±)hydroxybenzylisoproterenol> (−)epinephrine= (−)norepinephrine((±)salbutamol being inactive). A similar pattern was found in the hippocampus indicating the presence of a β₁-adrenergic receptor in these two regions. (±)Salbutamol was a partial agonist in the cerebellum and a competitive antagonist in the cerebral cortex.

The ratio of the antagonist potencies of (±)practolol and (±)butoxamine preferential β₁- and β₂-adrenergic antagonists respectively, to block the stimulation of adenylate cyclase was 25 in the cerebellum, compared to 0.5 in the cerebral cortex and 1.6 in the hippocampus. These results confirm the presence of a β₂ subtype of receptor coupled to adenylate cyclase in the former and β₁ subtypes in the latter two regions. The comparison between the affinities of a series of β-adrenergic agonists and antagonists for the β-adrenergic receptors coupled with an adenylate cyclase in cerebral cortex and cerebellum with their affinities for well characterized β₂-adrenergic receptors in lung and β₁-adrenergic receptor in heart substantiated this conclusion.     

The resolution of dopamine and β1- and β2-adrenergic-sensitive adenylate cyclase activities in homogenates of cat cerebellum, hippocampus and cerebral cortex

The stimulation of adenylate cyclase by dopamine and various β-adrenergic agonists has been investigated in homogenates from 3 areas of cat brain: the cerebral cortex, cerebellum and hippocampus. The purpose of the study was to determine whether the β-adrenergic receptors coupled to adenylate cyclase could be classified as either β₁ and β₂ subtypes in the different regions studied.The stimulation of adenylate cyclase by the β-adrenergic agonist, (−)isoproterenol (5 × 10⁻⁶M), was completely blocked by the specific β-adrenergic antagonist, (−)alprenolol (10⁻⁵ M), but not by the dopaminergic antagonist, fluphenazine (10⁻⁵ M), whereas the stimulation of adenylate cyclase by (−)epinephrine (10⁻⁴ M) was blocked to varying extents by these two drugs in each of the 3 regions studied. The (−)epinephrine effect was always blocked in the combined presence of (−)alprenolol and fluphenazine. The adenylate cyclase stimulation by (−)epinephrine which is not blocked by (−)alprenolol was due to interaction of (−)epinephrine with a dopaminergic-sensitive adenylate cyclase which has been characterized in cerebral cortex, hippocampus and cerebellum.Regional differences in the affinity of β-adrenergic-sensitive adenylate cyclase for various agonists were investigated in the presence of fluphenazine (10⁻⁵ M). In the cerebellum the potency order was (±)protokylol> (±)hydroxybenzylisoproterenol> (±)isoproterenol> (−)epinephrine> (±)salbutamol> (−)norepinephrine, indicating the presence of a β₂-adrenergic receptor. In the cerebral cortex the potency order was (−)isoproterenol> (±)protokylol> (±)hydroxybenzylisoproterenol> (−)epinephrine= (−)norepinephrine((±)salbutamol being inactive). A similar pattern was found in the hippocampus indicating the presence of a β₁-adrenergic receptor in these two regions. (±)Salbutamol was a partial agonist in the cerebellum and a competitive antagonist in the cerebral cortex.The ratio of the antagonist potencies of (±)practolol and (±)butoxamine preferential β₁- and β₂-adrenergic antagonists respectively, to block the stimulation of adenylate cyclase was 25 in the cerebellum, compared to 0.5 in the cerebral cortex and 1.6 in the hippocampus. These results confirm the presence of a β₂ subtype of receptor coupled to adenylate cyclase in the former and β₁ subtypes in the latter two regions. The comparison between the affinities of a series of β-adrenergic agonists and antagonists for the β-adrenergic receptors coupled with an adenylate cyclase in cerebral cortex and cerebellum with their affinities for well characterized β₂-adrenergic receptors in lung and β₁-adrenergic receptor in heart substantiated this conclusion.     

Increased corticotropin-releasing factor receptors in rat cerebral cortex following chronic atropine treatment

Rats were treated chronically with atropine (14 days, 20 mg/kg/day, s.c.) and corticotropin-releasing factor (CRF) receptors and CRF-mediated adenylate cyclase activity were measured in discrete brain regions. Chronic atropine treatment produced significant increases in muscarinic cholinergic receptors in the frontoparietal cortex (30% increase) and hippocampus (20% increase). No significant changes in the concentration of [¹²⁵I]Tyr^o-rat CRF binding sites were observed in olfactory bulb, cerebellum, striatum and hippocampus. In contrast, there was a significant and selective increase (35%) in CRF receptors in the frontoparietal cortex of atropine-treated rats. However, no significant corresponding changes in the V_max or EC₅₀ of CRF-stimulated adenylate cyclase activity accompanied the upregulation of CRF receptors in the cerebral cortex. These results demonstrate that (1) CRF receptors in rat brain are subject to receptor regulation, (2) the upregulation of CRF receptors occurs as a consequence of chronic muscarinic cholinergic receptor blockade, and (3) this interaction between acetylcholine and CRF may be limited to the cerebral cortex.     

Characterization of corticotropin-releasing factor receptor-mediated adenylate cyclase activity in the rat central nervous system

We report here that corticotropin-releasing factor (CRF) stimulates adenylate cyclase activity in the rat central nervous system (CNS). In frontoparietal cortex homogenates, the stimulation by CRF was dependent on time, temperature, tissue protein concentration, and guanine nucleotides. The rank order of potency for CRF analogs and fragments in stimulating adenylate cyclase activity [(Nle21,38) rat CRF greater than rat CRF approximately equal to acetyl ovine CRF (4-41) approximately equal to alpha helical ovine CRF greater than ovine CRF much greater than ovine CRF (1-39) approximately equal to ovine CRF (7-41)] was consistent with their affinities for CRF receptors in the brain and their relative potencies in stimulating pituitary adrenocorticotropic hormone secretion in vitro. The putative CRF receptor antagonist, alpha helical ovine CRF (9-41), did not stimulate adenosine 3',5'-cyclic monophosphate (cAMP) production but was able to attenuate the stimulation by various concentrations of rat CRF. The regional distribution of 125I-Tyr(o)-ovine CRF binding (olfactory bulb greater than frontoparietal cortex approximately equal to cerebellum greater than hypothalamus greater than striatum greater than or equal to midbrain greater than hippocampus greater than or equal to spinal cord) did not correspond with the regional degree of CRF receptor-mediated stimulation of adenylate cyclase (frontoparietal cortex greater than olfactory bulb greater than or equal to cerebellum greater than midbrain greater than or equal to hippocampus greater than striatum greater than or equal to hypothalamus greater than spinal cord). In addition, marked differences were observed in the ability of forskolin to potentiate CRF-stimulated cAMP production in the various brain areas examined. In summary, these data demonstrate that at least one of the second-messenger systems mediating the effects of CRF in the CNS involves stimulation of cAMP production and provides further support for a neurotransmitter role for this neuropeptide in the brain. Significant differences in the regulation of CRF-stimulated cAMP production and the disparity between CRF receptor number and receptor-mediated adenylate cyclase activity in discrete regions of the rat CNS suggest that some populations of CRF receptors in the brain may be functionally coupled to alternative signal transduction mechanisms.     

Brain forskolin binding in mice dependent on and tolerant to ethanol

Chronic ethanol ingestion by mice was previously shown to result in decreased activation of adenylate cyclase by guanine nucleotides and beta-adrenergic agonists, and in the loss of the high affinity beta-adrenergic agonist binding site in frontal cortex and hippocampus but not in cerebellum. These results indicate a regional specificity of ethanol's actions on beta-adrenergic receptors, the guanine nucleotide binding protein (Gs) and/or adenylate cyclase. To further detail the anatomical specificity of the effects of ethanol ingestion on receptor-coupled adenylate cyclase (AC) systems we have quantified the binding of [3H]forskolin to brain sections of control and ethanol-fed mice. High-affinity forskolin binding, thought to represent the complex of the alpha-subunit of Gs (as) and AC, was decreased in several brain areas including frontal cortex and hippocampus, but not in cerebellum, nucleus accumbens and certain other brain areas of ethanol-fed mice. Guanine nucleotides, such as Gpp(NH)p, generally enhanced forskolin binding in control animals. In ethanol-fed mice, however, Gpp(NH)p failed to enhance forskolin binding in most brain regions. These findings suggest that chronic ethanol ingestion may decrease the amount or function of as-AC in certain brain regions. Moreover, the regulation of the formation of this complex in different brain regions may affect responses to ethanol ingestion in mice.     

Age-related changes in bindings of second messengers in the rat brain

Age-related alterations in bindings of major second messengers in the brain were studied in 3-week- and 6-, 12-, 18- and 24-month-old Fisher 344 rats using receptor autoradiography. [³H]Phorbol 12,13-dibutyrate (PDBu) and [3H]forskolin were used to label protein kinase C (PKC) and adenylate cyclase, respectively. In immature rats (3-week-old), [³H]PDBu binding showed a significant decrease only in the cerebellum as compared to adult rats (6-month-old), whereas [³H]forskolin binding exhibited a significant reduction in the neocortex, nucleus accumbens, thalamus and substantia nigra. In aged rats, [³H]PDBu binding showed no significant change in all brain areas. In contrast, [³H]forskolin binding showed a conspicuous reduction in various brain areas in 18-month-old rats as compared to adult animals. The age-related reduction was especially observed in the cerebral cortex, hippocampal CA3 pyramidal cell layer, dentate gyrus, thalamus and molecular layer of cerebellum of 24-month-old rats. The results indicate that adenylate cyclase system in the rat brain is more susceptible to aging processes than phosphoinositide cycle system. Furthermore, our data demonstrate that the change in the adenylate cyclase system is more pronounced than that in the phosphoinositide cycle system in immature rat brain. These findings suggest that the adenylate cyclase system is primarily affected in aging processes and this may lead to age-related neurological deficits.     

Beta-adrenergic receptor binding in brain of alcoholics

The binding of agonists and antagonists to beta-adrenergic receptors in brain tissue obtained postmortem in nonalcoholic controls and matched intoxicated and sober alcoholics was measured to assess the state of the receptors and their coupling to adenylate cyclase. Binding of antagonist, iodocyanopindolol, to cerebral cortical and cerebellar membrane preparations was not different in alcoholics compared to that in controls, suggesting that the number of beta-adrenergic receptors was not affected by chronic ethanol ingestion. Agonist binding data, however, indicated the loss of the high-affinity agonist binding state of the beta-adrenergic receptor, representing the receptor-guanine nucleotide binding protein (Gs) complex. Such changes were observed in cerebral cortex but not in cerebellum of intoxicated alcoholics. These data suggest that cerebral cortical beta-adrenergic receptors are uncoupled from adenylate cyclase in these subjects. In cerebral cortical and cerebellar membranes of sober alcoholics both the high- and low-affinity agonist binding sites were observed. These findings are similar to those seen in animal studies and suggest that the effect of chronic ethanol ingestion on beta-adrenergic receptor-adenylate cyclase coupling is brain region specific and reversible with abstinence. Ethanol-induced changes in the coupling of receptors to adenylate cyclase may contribute to the physiological and behavioral manifestations of alcohol abuse.     

Effect of thyroid deficiency on the regional development of glutaminase, a glutamatergic neuron marker, in the rat brain

The effect of thyroid deficiency on the activity of phosphate-activated glutaminase (the marker for glutamatergic neurons) was studied in different parts of the rat brain at ages 5, 10, 15 and 25 days, and at day 130 following 102 days of rehabilitation. The brain regions investigated were the cerebral cortex, basal forebrain, hippocampus and cerebellum. During normal development, the activity of glutaminase increased relatively earlier in the cerebral cortex and hippocampus than in the cerebellum, while the absolute value reached a much higher level in the hippocampus than in other brain regions. In the basal forebrain, the developmental pattern of glutaminase was bimodal, and the rise in enzyme activity after 15 days coincided with the decrease in the cerebral cortex. These regional developmental changes in glutaminase activity correlated well with known information on the formation of glutamatergic cells and pathways in the brain. Neonatal thyroid deficiency had little effect on the developmental patterns of enzyme activity, the exception being a transient decrease in 10-day-old hypothyroid hippocampus. The present results, together with previous findings, indicate that the effect of thyroid hormone on neural maturation is cell-type specific and the glutamatergic neurons are not the main targets of thyroid hormone action.     

Inhibition of dopamine release by melatonin: regional distribution in the rat brain

Dopamine release evoked by electrical field stimulation of slices from various regions of rat brain was assessed in the presence of 10⁻¹⁰–10⁻⁵ M melatonin. Inhibition of dopamine release by melatonin was observed in the ventral hippocampus, medulla pons, preoptic area and median and posterior hypothalamus. No inhibitory effect of melatonin on dopamine release was observed in the cerebral cortex, cerebellum, dorsal hippocampus and striatum. Equal concentrations of melatonin were needed to produce half-maximal inhibition in all the regions affected. The results indicate that the brain sites for the inhibitory effect of melatonin on dopamine neurosecretion overlap the sites reportedly involved in its modulation of neuroendocrine functions.     

Unilateral cortical application of interleukin-1beta (IL1beta) induces asymmetry in fos, IL1beta and nerve growth factor immunoreactivity: implications for sleep regulation

Unilateral injection of interleukin-1 beta (IL1beta) into the somatosensory cortex enhances EEG slow wave activity ipsilaterally during non-rapid eye movement sleep [Yasuda, T., Yoshida, H., Garcia-Garcia, F., Kay, D., Krueger, J.M., 2005. Interleukin-1beta has a role in cerebral cortical state-dependent electroencephalographic slow-wave activity. Sleep 28, 177-184]. We show that a similar unilateral microinjection of IL1beta (10 ng) into layer VI or onto the surface of the primary somatosensory cortex induced increases in the neuronal activity marker, Fos, relative to the contralateral side that received saline or heat-inactivated IL1beta. When IL1beta was microinjected into layer VI, increases in Fos-immunoreactive nuclei were evident in layers II, III and VI of the somatosensory cortex and connected cortical regions, such as the endopiriform, secondary somatosensory, piriform and prefrontal cortex. Asymmetrical increases in Fos were also observed in subcortical regions, such as the reticular thalamus, which receives a main cortical projection, and hypothalamic regions implicated in sleep regulation, such as the ventrolateral preoptic area and dorsal median preoptic nucleus. Fos activation was not observed in many other brain regions. In the reticular thalamus and somatosensory cortex, the number of IL1beta-immunoreactive glial cells increased. Further, the number of NGF-immunoreactive cells in the primary somatosensory cortex and magnocellular preoptic nucleus increased on the IL1beta-injected side. These results are consistent with the hypothesis that sleep is initiated within the cortex after the local activation of specific cytokines and that whole organism sleep is coordinated via cortical connections with the subcortical sites.     

Alterations in Cyclic AMP Concentration and Adenylate Cyclase Activity in Specific Brain Areas of Rats with Inherited Hypothalamic Diabetes Insipidus (Brattleboro Rats)

The concentration of cyclic AMP (cAMP) and the activity of sodium-fluoride-stimulated adenylate cyclase was measured in 29 microdissected brain areas of homozygous Brattleboro rats and their Long-Evans control rats. In ten of the investigated brain areas a decreased cAMP level was measured in Brattleboro rats. It was particularly decreased in the supraoptic nucleus, cingulate and parietal cortex, hippocampus, habenula and organum vasculosum laminae terminalis. Significantly lower cAMP levels were also found in the periventricular nucleus, bed nucleus of the stria terminalis, area postrema and locus coeruleus. An increased cAMP concentration was detected only in the subcommissural organ of Brattleboro rats. In most brain areas, where cAMP was decreased, sodium fluoride-stimulated adenylate cyclase activity was significantly increased (supraoptic nucleus, parietal cortex, periventricular nucleus, bed nucleus of the stria terminalis, locus coeruleus) or unchanged (hippocampus, habenula, organum vasculosum laminae terminalis). The coincidence of alterations in cAMP concentration and adenylate cyclase activity in brain areas of Brattleboro rats with relatively dense vasopressinergic innervation and/or vasopressin receptor population in control rats, suggests an influence of brain vasopressin on the cAMP-adenylate cyclase second messenger system.     

β-Adrenergic receptor binding in brain of alcoholics

The binding of agonists and antagonists to β-adrenergic receptors in brain tissue obtained postmortem in nonalcoholic controls and matched intoxicated and sober alcoholics was measured to assess the state of the receptors and their coupling to adenylate cyclase. Binding of antagonist, iodocyanopindolol, to cerebral cortical and cerebellar membrane preparations was not different in alcoholics compared to that in controls, suggesting that the number of β-adrenergic receptors was not affected by chronic ethanol ingestion. Agonist binding data, however, indicated the loss of the high-affinity agonist binding state of the β-adrenergic receptor, representing the receptor-guanine nucleotide binding protein (G_s) complex. Such changes were observed in cerebral cortex but not in cerebellum of intoxicated alcoholics. These data suggest that cerebral cortical β-adrenergic receptors are uncoupled from adenylate cyclase in these subjects. In cerebral cortical and cerebellar membranes of sober alcoholics both the high- and low-affinity agonist binding sites were observed. These findings are similar to those seen in animal studies and suggest that the effect of chronic ethanol ingestion on β-adrenergic receptor-adenylate cyclase coupling is brain region specific and reversible with abstinence. Ethanol-induced changes in the coupling of receptors to adenylate cyclase may contribute to the physiological and behavioral manifestations of alcohol abuse.     

Vasoactive intestinal polypeptide (VIP) inhibits potassium-induced release of cholecystokinin (CCK) from rat caudato-putamen but not from cerebral cortex

CCK release elicited by 40 mM potassium from slices of rat caudato-putamen (cp) was inhibited by VIP. The effect of VIP was maximal at 10(-7) M. VIP does not inhibit CCK release from cerebral cortex at either 10(-7) or 10(-6) M. VIP is known to elevate levels of cAMP in rat brain. VIP inhibition of CCK release appears to be independent of activation of adenylate cyclase because treatment of cp slices with forskolin (2 X 10(-6) to 10(-4) M) does not mimic the inhibitory action of VIP.     

The role of adenylate cyclase in relaxation of brain arteries: studies with forskolin

The natural product, forskolin, which stimulates adenylate cyclase by a direct, non-receptor-mediated mechanism, was studied for its effect on the tension of isolated brain arteries and adenylate cyclase activity of cerebral arteries. Helical strips of bovine and porcine basilar arteries and bovine middle cerebral arteries, which had been precontracted with prostaglandin F2 alpha (PGF2 alpha) or KCl, relaxed potently to administration of forskolin with ED50 values, ranging from 22 to 69 nM. Incubation of forskolin with a broken cell preparation of bovine cerebral arteries resulted in an efficacious stimulation of adenylate cyclase, approximating 5 times basal activity at a forskolin concentration of 1 microM. The metal salts nickel chloride and manganese chloride decreased the potency of vasorelaxation by vasoactive intestinal peptide (VIP), which stimulates adenylate cyclase via the VIP receptor. In contrast, nickel chloride had little effect on vasorelaxation by forskolin. The endogenous nucleoside, adenosine, which acts via the adenosine receptor and adenylate cyclase, relaxed bovine basilar and middle cerebral arteries with ED50 values ranging from 0.26 to 0.94 microM. The data presented support a role for adenylate cyclase in mediating vasodilation of brain blood vessels.     

Age-related changes of the nitric oxide system in the rat brain

This work examines the age-related changes of the NO pathway in the central nervous system (CNS), analyzing nitric oxide synthase (NOS) isoform expression, the level of nitrotyrosine-modified proteins, and the NOS activity in the cerebral cortex, decorticated brain (basal ganglia, thalamus, hypothalamus, tegtum and tegmentum) and cerebellum of young, adult and aged rats. Our data demonstrate that the different NOS isoforms are not uniformly expressed across the CNS. In this sense, the nNOS and eNOS isoenzymes are expressed mainly in the cerebellum and decorticated brain, respectively, while the iNOS isoenzyme shows the highest level in cerebellum. Concerning age, in the cerebral cortex nNOS significantly increased its expression only in adult animals; meanwhile, in the cerebellum the eNOS expression decreased whereas iNOS increased in adult and aged rats. No age-related changes in any isoform were found in decorticated brain. NOS activity, determined by nitrate plus nitrite quantification, registered the highest levels in the cerebellum, where the significant increase detected with aging was probably related to iNOS activity. The number of nitrotyrosine-modified immunoreactive bands differed among regions; thus, the highest number was detected in the decorticated brain while the cerebellum showed the least number of bands. Finally, bulk protein nitration increased in cerebral cortex only in adult animal. No changes were found in the decorticated brain, and the decrease detected in the cerebellum of aged animals was not significant. According to these results, the NO pathway is differently modified with age in the three CNS regions analyzed.     

Postnatal development of regional binding of corticotropin-releasing factor and adenylate cyclase activity in the rat brain.

1. Corticotropin-releasing factor (CRF) plays a major role in the endocrine, autonomic and behavioral responses to stress. The distribution of CRF and CRF receptors in hypothalamic and extra-hypothalamic brain regions is consistent with its stress-related functions. 2. In most brain regions, CRF acts primarily, if not exclusively, through activation of the adenylate cyclase systems. 3. While previous studies have demonstrated the prenatal presence of CRF receptors, in the early postnatal period the abundance of CRF receptors relative to the magnitude of CRF-stimulated cAMP production suggests that CRF receptors are not fully linked to adenylate cyclase. 4. Because of our interest in the possible involvement of CRF signal transduction in the development of the neonatal stress response, we have examined postnatal development of CRF receptors in relation to adenylate cyclase activity in the rat. 5. CRF binding decreased significantly in the hippocampus and striatum from postnatal days 7-21. Basal adenylate cyclase activity peaked in the second-third week of postnatal life in each brain region. Preliminary studies suggest that early stress can alter the maturation of second messenger systems in the frontal cortex.     

Region-specific causal mechanism in the effects of ammonia on cerebral glucose metabolism in the rat brain

Ammonia, which is considered to be the main agent responsible for hepatic encephalopathy, inhibits oxidative glucose metabolism in the brain. However, the effects of ammonia on cerebral glucose metabolism in different brain regions remains unclear. To clarify this issue, we added ammonia directly to fresh rat brain slices and measured its effects on glucose metabolism. Dynamic positron autoradiography with [¹⁸F]2-fluoro-2-deoxy-d-glucose and 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium (WST-1) colorimetric assay revealed that ammonia significantly increased the cerebral glucose metabolic rate and depressed mitochondrial function, as compared to the unloaded control in each of the brain regions examined (cerebral cortex, striatum, and cerebellum), reflecting increased glycolysis that compensates for the decrease in aerobic metabolism. Pre-treatment with (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801), a N-methyl-d-aspartate (NMDA) receptor antagonist, significantly attenuated these changes induced by ammonia in cerebellum, but not in cerebral cortex or striatum. The addition of ammonia induced an increase in cyclic guanosine monophosphate (cGMP) levels in cerebellum, but not in cerebral cortex or striatum, reflecting the activation of the NMDA receptor-nitric oxide-cGMP pathway. These results suggested that NMDA receptor activation is responsible for the impairment of glucose metabolism induced by ammonia specifically in cerebellum.     

Effects of GTP on hormone-stimulated adenylate cyclase activity in cerebral cortex, striatum, and hippocampus from rats treated chronically with lithium

The effects of lithium on guanosine triphosphate (GTP) stimulated adenylate cyclase activity and hormone-induced GTP activation of the enzyme have been studied in three regions of the rat brain. Chronic treatment with lithium, giving a serum lithium level of 0.71 +/- 24 mmol/L, reduced isoprenaline-induced GTP stimulation of adenylate cyclase activity in cortical membranes at concentrations of GTP up to 2 microM. No effect of lithium was observed at higher concentrations of GTP. The enzyme activity stimulated by GTP alone was unaltered by lithium ex vivo. In striatal membranes, lithium ex vivo decreased both dopamine-induced GTP activation of adenylate cyclase and GTP-stimulated adenylate cyclase activity at concentrations of GTP below 2 microM. No effects of lithium ex vivo were found in striatum at 2 microM GTP and above. In hippocampal membranes, lithium ex vivo did not influence either serotonin-induced GTP stimulation of the adenylate cyclase or GTP-stimulated enzyme activity at low levels of GTP. However, at 50 microM GTP, lithium ex vivo enhanced serotonin-stimulated enzyme activity. The present results suggest that lithium ex vivo decreases neurotransmitter activation of the cortical beta-adrenergic adenylate cyclase by influencing the mechanisms by which receptor agonists enhance the GTP stimulation of the adenylate cyclase. Furthermore, lithium ex vivo exerts a region-specific action on the brain adenylate cyclases, but in the brain regions studied, an effect of lithium on N-protein level might be of significance for the action of lithium ex vivo on neurotransmitter activation.     

Induction of oxidative stress and inhibition of superoxide dismutase expression in rat cerebral cortex and cerebellum by PTU-induced hypothyroidism and its reversal by curcumin

The present study was carried out to elucidate the effectiveness of curcumin in ameliorating the expression of superoxide dismutase (SOD) in cerebral cortex and cerebellum of rat brain under 6-propyl-2-thiouracil (PTU)-induced hypothyroidism. Induction of hypothyroidism in adult rats by PTU resulted in augmentation of lipid peroxidation (LPx), an index of oxidative stress in cerebellum but not in cerebral cortex. Curcumin-supplementation to PTU-treated (hypothyroid) rats showed significant reduction in the level of LPx in both the regions of brain. The decreased translated products (SOD1 and SOD2) and the unchanged activity of SOD in cerebral cortex of PTU-treated rats were increased on supplementation of curcumin to the hypothyroid rats. Declined translated products of SOD1 and SOD2 in cerebellum of PTU-treated rats were alleviated on administration of curcumin to hypothyroid rats. On the other hand, the decreased activity of SOD in cerebellum of PTU-treated rats was further declined on administration of curcumin to the hypothyroid rats. Results of the present investigation indicate that curcumin differentially modulates the expression of superoxide dismutase in rat brain cortex and cerebellum under PTU-induced hypothyroidism.