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Showing papers in "Journal of Neurochemistry in 1980"


Journal ArticleDOI
TL;DR: This report provides evidence that the increase in glucose utilization evoked by functional activation in neural tissue is due to the opening of Na+ channels and activation of Na+, K+ -ATPase activity.
Abstract: Sokoloff later noted, "This report provides evidence that the increase in glucose utilization evoked by functional activation in neural tissue is due to the opening of Na+ channels and activation of Na+, K+ -ATPase activity."

583 citations


Journal ArticleDOI
TL;DR: The use of Sep‐Pak™ C18 reverse‐phase cartridges to adsorb gangliosides from aqueous solutions was studied and retained lipids can be eluted with methanol or chloroform‐methanol.
Abstract: The use of Sep-Pak C18 reverse-phase cartridges to adsorb gangliosides from aqueous solutions was studied When upper phases formed from chloroform-methanol tissue extracts or aqueous salt solutions containing gangliosides are rapidly passed through the cartridges, the lipids are retained and the non-lipid components can be washed through Gangliosides and other retained lipids can subsequently be eluted with methanol or chloroform-methanol

478 citations


Journal ArticleDOI
TL;DR: Levels were highest during the initial period of reperfusion, then decreased to approach control levels after 120 min, suggesting that postischemic accumulation of arachidonic acid metabolites could be blocked by prior administration of inhibitors of cyclooxygenase activity.
Abstract: Several of the cyclooxygenase products of arachidonic acid were measured in the cerebral hemispheres of gerbils subjected to transient interruption of the cerebral circulation. The levels of PGD2, PGF2 alpha, PGE2, TXB2, 13, 14-H2-15-keto-PGE2, and the stable nonenzymic product of prostacyclin, 6-keto-PGF1 alpha, were not altered at the end of a 5-min period of ischemia. However, the onset of reperfusion was accompanied by a rapid accumulation of these products. Levels were highest during the initial period of reperfusion, then decreased to approach control levels after 120 min. PGD2, PGF2 alpha, and PGE2 were the predominant metabolites detected. This postischemic accumulation of arachidonic acid metabolites could be blocked by prior administration of inhibitors of cyclooxygenase activity.

349 citations


Journal ArticleDOI
TL;DR: The possibility that the isolated enzyme represents a proteolytic complex composed of units with distinctly different activities was raised, as preliminary attempts to dissociate the enzyme into catalytic units of lower molecular weight were not successful and led to loss of activity.
Abstract: A highly purified preparation of a cation-sensitive neutral endopeptidase was obtained from bovine pituitaries. The enzyme constitutes almost 0.1% of the protein in bovine pituitary homogenates. Polyacrylamide gel electrophoresis of the enzyme showed a single protein band, and in gel filtration experiments on calibrated Sepharose 6B columns the enzyme eluted slightly ahead of thyroglobulin, suggesting an apparent molecular weight of about 700,000. Polyacrylamide gel electrophoresis in SDS-containing buffers indicated the presence of three major components with molecular weights ranging from about 24,000 to 28,000. The enzyme hydrolyzes bonds between hydro-phobic and small neutral amino acids in both model synthetic substrates and biologically active peptides such as substance P, LH-RH, and bradykinin. Peptide bonds in which the carbonyl group is contributed by a glutamyl or arginyl residue are also hydrolyzed, especially if they are preceded in the sequence by hydrophobic amino acids. Leupeptin exclusively inhibited enzymatic activity toward the arginine-containing substrates. This observation, together with the high molecular weight and broad specificity of the enzyme, raised the possibility that the isolated enzyme represents a proteolytic complex composed of units with distinctly different activities. Preliminary attempts to dissociate the enzyme into catalytic units of lower molecular weight were not successful and led to loss of activity.

298 citations


Journal ArticleDOI
TL;DR: The view that HA might have a neurotransmitter role in mammalian brain has been indirectly reinforced by the identification of histaminergic neurons in the CNS of the marine mollusk Aplysia californica (for a review, see Weinreich, 1977).
Abstract: For a long time the history of histamine (HA) has been confounded with that of autacoids, i.e., endogenous substances with potent biological effects but whose physiological function is only a matter of speculation. However, starting from about 1970, sufficient data, mostly of a biochemical nature, have been collected to justify its present classification in most reviews or textbooks at the top of the list of putative neurotransmitters in the brain. The main landmarks in this evolution are probably the development of neurochemical methods allowing for assessment of the possibilities that: synthesis and storage of HA occurs in neuronal pools, in which it is in a dynamic state; that selective lesions of neuronal tracts are followed by changes in HA and L-histidine decarboxylase (HD); and that HA receptors can be detected in brain by various approaches. Recently the view that HA might have a neurotransmitter role in mammalian brain has been indirectly reinforced by the identification of histaminergic neurons in the CNS of the marine mollusk Aplysia californica (for a review, see Weinreich, 1977). The short review will consider the. most significant neurochemical data obtained concerning the metabolism of the amine, the localisation of putative histaminergic pathways and the characterization of receptors. Other aspects not covered here can be found in recent reviews: behavioral and pharmacological aspects (Green et al . , 1978; Schwartz et al., 198Oa), neuroendocrinological aspects (Weiner and Ganong, 1978), and neurovegetative aspects (Owen, 1977).

294 citations


Journal ArticleDOI
TL;DR: In vitro data support the hypothesis that both superoxide radicals and lipid peroxidation are involved in the mechanism of polyunsaturated fatty acid‐induced brain edema.
Abstract: The involvement of superoxide free radicals and lipid peroxidation in brain swelling induced by free fatty acids has been studied in brain slices and homogenates. The polyunsaturated fatty acids linoleic acid (18:2), linolenic acid (18:3), arachidonic acid (20:4), and docosahexaenoic acid (22:6) caused brain swelling concomitant with increases in superoxide and membrane lipid peroxidation. Palmitic acid (16:0) and oleic acid (18:1) had no such effect. Furthermore, superoxide formation was stimulated by NADPH and scavenged by the addition of exogenous superoxide dismutase in cortical slice homogenates. These in vitro data support the hypothesis that both superoxide radicals and lipid peroxidation are involved in the mechanism of polyunsaturated fatty acid-induced brain edema.

270 citations


Journal ArticleDOI
Stig Rehncrona1, David Smith1, Björn Åkesson1, Eva Westerberg1, Bo K. Siesjö1 
TL;DR: Measurements of changes in brain fatty acid and phospholipid composition may be a useful tool to establishment of whether peroxidative damage is important in vivo in situations with a critically reduced oxygen supply.
Abstract: The occurrence of peroxidative damage, as distinguished from anaerobic damage, to brain fatty acids and phospholipids was characterized in vitro. Fe2+ and ascorbic acid were used to stimulate peroxidation in cortical homogenates from rat brain incubated with or without oxygen. Lipid peroxidation was established in samples incubated with oxygen by increased diene conjugation, accumulation of thiobarbituric acid-reactive material (TBAR) and of lipid-soluble fluorescent products. No peroxidation occurred in samples incubated in the absence of oxygen (100% N2). Lipid peroxidation was characterized by a selective loss of arachidonic acid and docosahexaenoic acid and by degradation of ethanolamine phosphoglyceride, while choline phosphoglyceride did not change. During the course of peroxidation there were parallel increases in products of lipid peroxidation concomitant with the decrease in polyenoic fatty acids. The maximal changes in diene conjugation and TBAR occurred earlier than the maximal changes in fluorescent material and fatty acids. It is concluded that measurements of changes in brain fatty acid and phospholipid composition may be a useful tool to establishment of whether peroxidative damage is important in vivo in situations with a critically reduced oxygen supply. Estimation of lipid-soluble fluorescence in vivo may also be useful, since it is considered to reflect the accumulation of stable end products of peroxidation.

258 citations


Journal ArticleDOI
TL;DR: The results suggest that frontal cortical catecholamine systems may serve to regulate various neurotransmitter mechanisms within the basal ganglia, including 5‐hydroxytryptamine and GABA.
Abstract: The effect of lesions of the catecholamine nerve terminals in the medial prefrontal cortex of the rat on neurotransmitter mechanisms within the basal ganglia has been investigated. Bilateral 6-hydroxydopamine lesions were stereotaxically placed in the dopamine-rich (DA) area of th frontal cortex. Animals were pretreated with desmethylimipramine to block the uptake of neurotoxin into noradrenergic (NA) terminals and to make it more selective for DA terminals. The lesion produced a selective reduction of both NA and DA from the medial prefrontal cortex, a result related to falls in tyrosine hydroxylase activity at this site. Lesioned animals showed enhanced DA turnover and utilisation in striatal and limbic regions. There was no change in subcortical tyrosine hydroxylase activity. In addition there were significant falls in other putative neurotransmitters within basal sites, including 5-hydroxytryptamine and GABA. Decreased activity of the neurotransmitter-synthesizing enzyme glutamate decarboxylase and choline acetyltransferase was also recorded in certain regions of the basal ganglia. The results suggest that frontal cortical catecholamine systems may serve to regulate various neurotransmitter mechanisms in the basal ganglia.

248 citations


Journal ArticleDOI
TL;DR: The studies described here demonstrate that primary cultures of adrenal medulla cells provide an excellent experimental system for obtaining more detailed information on stimulus‐secretion coupling and other functional aspects of the adrenalmedulla.
Abstract: The functional stability of primary cultures of adrenal medulla cells was investigated. Isolated cells were prepared by treatment of bovine adrenal glands with collagenase followed by purification on Percoll density gradients and were maintained in Dulbecco's medium containing 10% fetal calf serum. Within 12 h after plating on plastic culture dishes, the cells became firmly attached and exhibited good survival for periods of time up to 3 weeks, as indicated by their morphology using light and electron microscopy, by maintenance of their content of catecholamines, tyrosine hydroxylase, dopamine-beta-hydroxylase, and phenylethanolamine N-methyltransferase, and their ability to respond to secretagogues. During the first 10 days to 2 weeks in culture there was little or no change in any of these parameters. During the 3rd week there were progressive losses of catecholamine and enzyme activities and increased vacuolization of medullary cells. The cells synthesized protein and RNA with no apparent loss in activities over the period studied, but did not incorporate [3H]thymidine into PCA-precipitable material. The cells responded to secretagogues and secretory antagonists similarly to isolated perfused adrenal glands. The studies described here demonstrate that primary cultures of adrenal medulla cells provide an excellent experimental system for obtaining more detailed information on stimulus-secretion coupling and other functional aspects of the adrenal medulla.

241 citations


Journal ArticleDOI
TL;DR: Autoradiography revealed that the pyramidal neurons of the hippocampus and areas of the caudate and cortex failed to recover normal rates of protein synthesis even after 48 h.
Abstract: Regional protein synthesis was measured in rat brain at intervals up to 48 h following occlusion of the four major arteries to the brain for either 10 or 30 min. Four-vessel occlusions produces ischemia in the cerebral hemispheres and oligemia in the midbrain-diencephalon and brainstem. During the hour following 10 min of ischemia, protein synthesis, measured by incorporation of [14C]valine into protein, was inhibited in the cerebral cortex by 67%. Normal rates of protein synthesis were attained within 4 h of recirculation. In rats subjected to 30 min of ischemia, protein synthesis was inhibited by 83% during the first hour of recirculation in the cortex, caudate-putamen, and hippocampus. Recovery of protein synthesis in these regions was slow (25-48 h). The midbrain-diencephalon showed less inhibition, 67%, and faster recovery (by 12 h). Protein synthesis was unaffected in the brainstem. [14C]Autoradiography revealed that the pyramidal neurons of the hippocampus and areas of the caudate and cortex failed to recover normal rates of protein synthesis even after 48 h. The accumulation of TCA-soluble [14C]valine was enhanced (55-65%) in the cortex, caudate, and hippocampus after 30 min of ischemia; the increase persisted for 12 h. A smaller rise in [14C]valine content (30%) and more rapid normalization of valine accumulation (by 7 h) were observed in the midbrain-diencephalon; no changes were found in the brainstem. In the cortex, recovery was more rapid when the duration of ischemia was reduced. Thus, the degree of inhibition of protein synthesis, the accumulation of valine in the tissue, and the length of time required to reestablish normal values for these processes were dependent on both the severity and the duration of the ischemic insult. Restoration of normal rates of protein synthesis after ischemia was slow compared with the normalization of cerebral energy metabolites.

236 citations


Journal ArticleDOI
TL;DR: For example, alpha-Fluoromethylhistidine (α-FMH) has been used for in vitro and in vivo studies of brain histidine decarboxylase (HD) as discussed by the authors.
Abstract: alpha-Fluoromethylhistidine (alpha-FMH), a new potent inhibitor of histidine decarboxylase (HD), has been used for in vitro and in vivo studies of brain HD. Following a preincubation with (+)-alpha-FMH, brain HD activity was inhibited in a time-dependent and concentration-dependent manner. The enzyme activity was not restored by overnight dialysis against standard buffer. The (-) antimer of alpha-FMH was ineffective. When injected intraperitoneally in a single dose of 20 mg/kg, (+/-)-alpha-FMH induced a complete loss in HD activity in cerebral cortex and hypothalamus as well as in peripheral tissues, such as stomach. At a dosage of 100 mg/kg (+/-)-alpha-FMH did not alter histamine-N-methyltransferase, DOPA decarboxylase, and glutamate decarboxylase activities. The maximal decrease of HD activity occurred after 2 h in both cerebral cortex and hypothalamus, but the time course of the recovery of enzyme activity was slower in the cerebral cortex. The enzyme activity reached control value within 3 days in hypothalamus and was not fully restored after 4 days in cerebral cortex. Contrasting with the diminished HD activity, a substantial concentration of histamine remained present in five regions of mouse brain. Thus, alpha-FMH is a highly specific irreversible inhibitor of brain HD activity and its efficacy makes it useful to study the physiological role of brain histamine.

Journal ArticleDOI
TL;DR: The findings are discussed in relation to the hypothesis that the deleterious effects of ischemia are due to an increase in free radical production which in turn leads to increased lipid peroxidation.
Abstract: Thirty minutes of total cerebral ischemia (decapitation) decreased total glutathione (GSH + GSSG) by 7% but had no detectable effect on the concentration of oxidized glutathione (GSSG), reduced ascorbate, or total ascorbate, In a model of reversible, bilateral hemispheric ischemia (four-vessel occlusion) no changes in glutathione or ascorbate were detected after 30 min of ischemia. During 24 h of reperfusion following such an insult no detectable change in total ascorbate, reduced ascorbate, or oxidized glutathione was noted: however, total brain glutathione declined by 25%. The findings are discussed in relation to the hypothesis that the deleterious effects of ischemia are due to an increase in free radical production which in turn leads to increased lipid peroxidation.

Journal ArticleDOI
TL;DR: Measuring receptor binding in isolated membranes overcomes potentially artifactual sources of variations in drug potency and has permitted identification of several new subclasses of neurotransmitter receptors.
Abstract: Just as multiple forms or \"isoenzymes\" of particular enzymes exist, so there may be multiple receptors for an individual neurotransmitter. Some of these have been well known for many years; for instance, the distinction of muscarinic and nicotinic cholinergic receptors, aand P-adrenergic receptors, and , more recently, distinct 6,and P,adrenergic receptors. In different parts of the body histamine acts at distinct H, and H, receptor sites. All of these discriminations of receptor populations were based initially on pharmacological variations. During the past few years it has been possible to label neurotransmitter receptors by binding techniques (Snyder, 1978), an approach which affords more subtle molecular discriminations of receptor properties. In pharmacological experiments drugs may vary in potency because of differential bioavailability and metabolism, even in isolated organs, rather than distinct populations of receptors. Measuring receptor binding in isolated membranes overcomes potentially artifactual sources of variations in drug potency and has permitted identification of several new subclasses of neurotransmitter receptors.

Journal ArticleDOI
TL;DR: H2 O2 production by rat brain in vivo was observed with a method based on the measurement of brain catalase, and in studies in vitro, the production of H2 O 2 by ratbrain mitochondria with either dopamine or serotonin as substrate was confirmed.
Abstract: H2 O2 production by rat brain in vivo was observed with a method based on the measurement of brain catalase. The administration to the rat of 3-amino-1, 2, 4-triazole, an H2 O2- dependent inhibitor of catalase, caused progressive inhibition of brain catalase activity in both the supernatant and pellet fractions of homogenates of the striatum and prefrontal cortex. The prevention of catalase inhibition by prior administration of ethanol confirmed that catalase inhibition in vivo was dependent upon H2 O2. A significant portion of the catalase (30-33%) appeared in the supernatant fraction from a slow-speed homogenization procedure and was not significantly contaminated by either erythrocytes or capillaries. In the whole homogenate, less than 6% of the catalase activity was attributed to erythrocytes. Modification of intracellular monoamine oxidase activity by either pargyline or reserpine did not change the rate of inhibition of catalase by aminotriazole. A probable interpretation of these data is that H2 O2 generated by mitochondrial monoamine oxidase does not reach the catalase compartment; the catalase is contained in particles described by other investigators as the microperoxisomes of brain. In studies in vitro, the production of H2 O2 by rat brain mitochondria with either dopamine or serotonin as substrate was confirmed.

Journal ArticleDOI
TL;DR: Rat brain was found to enzymatically methylate phospholipids to form phosphatidylcholine with S‐adenosyl‐l‐methionine serving as the methyl donor and methyltransferase activity was localized in the microsomes and synaptosomes.
Abstract: Rat brain was found to enzymatically methylate phospholipids to form phosphatidylcholine with S-adenosyl-l-methionine serving as the methyl donor Methyltransferase activity was localized in the microsomes and synaptosomes In synaptosomes, at least two enzymes were found to be involved in the formation of phosphatidylcholine The first methyltransferase which catalyzes the methylation of phosphatidylethanolamine to form phosphatidyl-N-monomethylethanolamine was found to have a pH optimum of 75, a low Km for 5-adenosyl-l-methionine and a partial requirement for Mg2 Methyltransferase I is tightly bound to membranes The second methyltransferase (II) catalyzes the successive methylations of phosphatidyl-N-monomethylethanolamine to phosphatidyl-N, N-dimethylethanolamine and then to phosphatidylcholine In contrast to methyltransferase I, methyltransferase II has a pH optimum of 105, a high apparent Km for S-adenosyl-l-methionine and no requirement for Mg2 Methyltransferase II is easily solubilized by sonication The highest specific activity for both enzymes was found in the synaptosomal plasma membrane

Journal ArticleDOI
TL;DR: It is speculated that the inhibition of B‐50 protein kinase may be related to known modulatory effects of ACTH and related peptides on certain types of neurotransmission and behaviour.
Abstract: ACTH inhibits the phosphorylation of a rat brain membrane-bound protein (B-50). Both the protein kinase and the substrate protein could be extracted from the membranes by means of treatment with Triton X-100 in 75 mM-KCl. Using column chromatography over DEAE-cellulose and ammonium sulphate precipitation a protein fraction (ASP 55–80) enriched in endogenous B-50 phosphorylating activity was obtained. The time course of the endogenous phosphorylation of B-50 in this fraction showed a linear incorporation with time for at least 10 min and reached an estimated maximal incorporation of 0.65 mol P/mol B-50 after 60 min. The inhibition by ACTH1_24 of the B-50 protein kinase in ASP 55–80 was dose-dependent; the half-maximal effective concentration was 5 × 10−6 M, being 10 to 50 times lower as compared with intact synaptic plasma membranes (SPM). cAMP, cGMP and various endor-phins had no effect on the B-50 protein kinase. The B-50 protein kinase required both magnesium and calcium for optimal activity. Using two-dimensional electrophoresis on polyacrylamide slab gels the B-50 protein kinase and the B-50 protein could be identified and purified. The isoelectric point (IEP) of the kinase is 5.5 and the apparent molecular weight 70,000, whereas the IEP of the substrate protein B-50 is 4.5 and the apparent molecular weight 48,000. Amino acid analysis on microgram quantities of purified kinase and B-50 protein revealed basic/acidic amino acid ratios in agreement with the respective lEP's. It is speculated that the inhibition of B-50 protein kinase may be related to known modulatory effects of ACTH and related peptides on certain types of neurotransmission and behaviour.

Journal ArticleDOI
TL;DR: By affecting binding of agonists, but not antagonists, GTP may regulate opiate receptor interactions with their physiological effectors.
Abstract: Guanine nucleotides differentiate binding of tritium-labeled agonists and antagonists to rat brain membranes. In the absence of sodium, GTP (50 μM) decreased binding of [3H]-labeled agonists by 20–60% and [3H]-labeled antagonists by 0–20%. In the presence of 100 mM-NaCl, GTP had no effect on antagonist binding, but decreased agonist binding by 60–95%. GMP was less potent than either GTP or GDP in decreasing agonist binding. GTP (50 μM) reduced high-affinity [3H]dihydromorphine sites by 52% and low-affinity sites by 55%. Without sodium, GTP reduced high-affinity [3H]-naloxone sites by 36%; in the presence of 100 mM-NaCl, GTP had no effect on either high- or low-affinity [3H]naloxone sites. GTP increased the association rate of [3H]dihydromorphine twofold and the dissociation rate by fourfold, while having no effect on association or dissociation rates of the antagonist [3H]diprenorphine. The affinities of uniabeled antagonists in inhibiting [3H]-diprenorphine binding were not affected by GTP or sodium, but the affinities of agonists were reduced 40- 120-fold, with met- and leu-enkephalin affinities reduced by the greatest degree. GTP and sodium lowered [3H]dihydromorphine binding in an additive fashion, while divalent cations, especially manganese, reversed the effects of GTP on [3H]-labeled agonist binding by stimulating membrane-bound phosphatases that hydrolyze GTP to GMP and guanosine. These results suggest that by affecting binding of agonists, but not antagonists, GTP may regulate opiate receptor interactions with their physiological effectors.

Journal ArticleDOI
TL;DR: The observed decrease in tissue GSH concentration mainly reflects a decrease in the glutathione pool size, due to an imbalance between breakdown and synthesis secondary to tissue energy failure, and the results fail to support the hypothesis that peroxidative damage occurs during or following brain ischemia.
Abstract: The influence of complete and pronounced incomplete cerebral ischemia on cortical concentrations of reduced (GSH) and oxidized (GSSG) glutathione was studied in lightly anaesthetized (70% N2 O) rats. GSH was extracted with HCl-methanol-perchloric acid and GSSG with trichloroacetic acid in the presence of N-ethylmaleimide and measured fluorometrically, giving normal concentrations in cortical tissue of about 2 and 0.01 μmol.g−1 respectively. Reversible complete ischemia was induced by increasing the intracranial pressure to above the systolic blood pressure by infusing mock CSF into the cisterna magna. Reversible pronounced incomplete ischemia was induced by bilateral carotid artery clamping combined with hypovolemic hypotension. Whether complete or incomplete, a 30-min ischemic period caused a similar decrease in cortical GSH concentration (to about 90% of control) without any concomitant accumulation of GSSG in the tissue (or in CSF). Prolongation of the ischemic period (complete ischemia) to maximally 120 min caused an almost linear decrease of the tissue glutathione concentration to 45% of the preischemic value. During subsequent recirculation following a 30 min period of either complete or pronounced incomplete ischemia, there was a further decrease in cortical GSH concentrations without a reciprocal increase in GSSG concentrations. Lipid peroxidation (verified by determination of malondialdehyde production) induced in brain cortical tissue in vitro caused oxidation of tissue GSH with accumulation of GSSG. As the observed decrease in GSH during brain ischemia in vivo was not accompanied by any reciprocal increase in GSSG the results fail to support the hypothesis that peroxidative damage occurs during or following brain ischemia. The finding of an unchanged GSSG concentration does, however, not exclude the possibility of an increased turnover rate in the glutathione reductase reaction. It is concluded that the observed decrease in tissue GSH concentration mainly reflects a decrease in the glutathione pool size, due to an imbalance between breakdown and synthesis secondary to tissue energy failure.

Journal ArticleDOI
TL;DR: The effects of acute and chronic ethanol ingestion on the binding of the inhibitory neurotransmitter 3H‐γ‐aminobutyric acid (GABA) was investigated in inbred strains of mice, suggesting the possible involvement of γ‐aminoic acid synaptic function in the neuropharmacological effects of ethanol.
Abstract: The effects of acute and chronic ethanol ingestion on the binding of the inhibitory neurotransmitter 3H-γ-aminobutyric acid (GABA) was investigated in inbred strains of mice. Acute effects were analyzed in DBA/2J (alcohol avoiding) and C57BL/6J (alcohol preferring) mice, while chronic effects were analyzed in C57 mice. Acute ethanol (4 g/kg) produced a significant increase in the binding capacity of the low-affinity γ-aminobutyric acid binding site in both C57 (12%) and DBA (23%) mice, without significantly altering other binding parameters. The enhanced binding capacity may be responsible for the behavioral depression associated with ethanol use. Chronically treated C57 mice exhibited a decrease in the density of low-affinity GABA receptor sites. These results suggest the possible involvement of γ-aminobutyric acid synaptic function in the neuropharmacological effects of ethanol. The possibility that chronic ethanol administration may result in adaptation of γ-aminobutyric acid receptors to the continuous presence of ethanol-induced central nervous system depression is discussed.

Journal ArticleDOI
TL;DR: Kinetic studies indicated that the Vmax of MAO B was increased in aging rat brain, whereas the Km was unaltered, and age‐related changes in total protein, MAO A, andMAO B were thus independent of each other.
Abstract: The activity of monoamine oxidase (MAO) was assayed in the brains of young adult (7-8 weeks old) and aging (95-104 weeks old) male Sprague-Dawley rats. When expressed per milligramme of tissue, total protein content was increased to a similar extent in whole brain and in all seven brain regions of the aging rat, whereas MAO A activity (assayed by using 5-hydroxytryptamine as substrate) was unchanged in whole brain but increased in the cerebellum, and fell in the brainstem, midbrain, hippocampus, and cortex; and MAO B activity (assayed by using beta-phenylethylamine as substrate) increased in whole brain and all regions, except the brainstem. Age-related changes in total protein, MAO A and MAO B were thus independent of each other. Kinetic studies indicated that the Vmax of MAO B was increased in aging rat brain, whereas the Km was unaltered. The increase in MAO B activity was restricted to the extrasynaptosomal mitochondrial fraction of the aging brain, whereas a reduction in MAO A activity was found in the intrasynaptosomal, but not the extrasynaptosomal mitochondrial fraction.

Journal ArticleDOI
TL;DR: The turnover rates of phenylethylamine and tryptamine have been estimated as their rates of accumulation after inhibition of monoamine oxidase by pargyline and were found to be 1.53 nmol/g/h and 0.24 nmol / g/h, respectively.
Abstract: The turnover rates of phenylethylamine and tryptamine have been estimated as their rates of accumulation after inhibition of monoamine oxidase by pargyline and were found to be 1.53 nmol/g/h and 0.24 nmol/g/h, respectively. Rate constants for the substrate activities of these amines towards monoamine oxidase were calculated as 100 h−1 and 150 h−1, respectively. Tryptamine was found to exhibit a biphasic response to increasing pargyline dosage, demonstrating its dual activity to type B and type A monoamine oxidase, for which two rate constants were obtained, kB=100 h−1 and kA=50 h−1.

Journal ArticleDOI
TL;DR: The development of the enzymes of aerobic glycolytic metabolism correlate with the onset of neurological competence in the two species, the guinea pig being a "precocial" species born neurologically competent and the rat being a “non‐precsocial” species Born neurologically immature.
Abstract: Key enzymes of ketone body metabolism (3-hydroxybutyrate dehydrogenase, 3-oxo-acid:CoA transferase, acetoacetyl-CoA thiolase) and glucose metabolism (hexokinase, lactate dehydrogenase, pyruvate dehydrogenase, citrate synthase) have been measured in the brains of foetal, neonatal, and adult guinea pigs and compared to those in the brains of neonatal and adult rats. The activities of the guinea pig brain ketone-body-metabolising enzymes remain relatively low in activity throughout the foetal and neonatal periods, with only slight increases occurring at birth. This contrasts with the rat brain, where three- to fourfold increases in activity occur during the suckling period (0-21 days post partum), followed by a corresponding decrease in the adult. The activities of the hexokinase (mitochondrial and cytosolic), pyruvate dehydrogenase, lactate dehydrogenase, and citrate synthase of guinea pig brain show marked increases in the last 10-15 days before birth, so that at birth the guinea pig possesses activities of these enzymes similar to the adult state. This contrasts with the rat brain where these enzymes develop during the late suckling period (10-15 days after birth). The development of the enzymes of aerobic glycolytic metabolism correlate with the onset of neurological competence in the two species, the guinea pig being a "precocial" species born neurologically competent and the rat being a "non-precocial" species born neurologically immature. The results are discussed with respect to the enzymatic activities required for the energy metabolism of a fully developed, neurologically competent mammalian brain and its relative sensitivity to hypoxia.

Journal ArticleDOI
TL;DR: The present results suggest that the anticonvulsant properties of the four catalytic inhibitors of GABA‐T tested are at least in part mediated through a direct influence on GABA receptors and uptake sites.
Abstract: Four catalytic inhibitors of GABA aminotransferase (gabaculine, γ-acetylenic GABA, γ-vinyl GABA, ethanolamine O-sulphate) as well as aminooxyacetic acid and valproate were studied for effects on neurochemical assays for GABA synthesis, receptor binding, uptake and metabolism in mouse and rat brain preparations. Gabaculine did not interfere with GABA synthesis as reflected by the activity of glutamate decarboxylase (GAD), it was only a weak inhibitor (IC50= 0.94 mM) of GABA receptor binding sites but was a moderately potent inhibitor of GABA uptake (IC50= 81 μM) and very potent (IC50= 1.8 μM) with respect to inhibition of the GABA-metabolizing enzyme GABA aminotransferase (GABA-T). γ-Acetylenic GABA was a weak inhibitor of GAD and GABA binding (IC50 > 1 mM), but virtually equipotent to inhibit uptake and metabolism of GABA (IC50 560 and 150 μM, respectively). This was very similar to γ-vinyl GABA, except that this drug did not decrease GAD activity. Ethanolamine O-sulphate was found to show virtually no inhibition of GAD and GABA uptake, but was a fairly potent inhibitor of GABA binding (IC50= 67 μM) and in this respect, 500 times more potent than as an inhibitor of GABA-T. Aminooxyacetic acid was a powerful inhibitor of both GAD and GABA-T (IC50 14 and 2.7 μM, respectively), but had very little affinity to receptor and uptake sites for GABA. Valproate showed no effects on GABA neurochemical assays which could be related to anticonvulsant action. The present results suggest that the anticonvulsant properties of the four catalytic inhibitors of GABA-T tested are at least in part mediated through a direct influence on GABA receptors and uptake sites.

Journal ArticleDOI
TL;DR: The direct demonstration of the NGF content of most end organs requires assays more sensitive than those currently available and the high levels of NGF produced by some organs are probably of some other physiological significance.
Abstract: CONCLUSION NGF proteins probably act as informational molecules transferred from end organs to the neurons of the sympathetic nervous system. The direct demonstration of the NGF content of most end organs requires assays more sensitive than those currently available. The high levels of NGF produced by some organs are probably of some other physiological significance.

Journal ArticleDOI
TL;DR: H2‐receptors are involved in the activation of adenylate cyclase of the capillary fraction of the guinea pig cerebral cortex, and this action elicits a twofold stimulation in the accumulation of cyclic AMP in this capillary fractions.
Abstract: A fraction enriched in capillaries has been prepared from the guinea pig cerebral cortex. The purity of this fraction was checked by light- and electron-microscopic examination and by its high enrichment in alkaline phosphatase and γ-glutamyl transpeptidase. In the capillary-rich fraction, the endogenous level of histamine was 1.9%’of that measured in the initial hornogenate. The histamine-synthesizing enzyme, I-histidine decarboxylase, and the metabolizing enzyme, histamine-N-methyltransferase, were barely detectable. In addition, histamine elicits a twofold stimulation in the accumulation of cyclic AMP in this capillary fraction with an EC50 of 5 γM. Agonists and antagonists of the two types of histamine receptors (H1 and H2) were used for the characterization of the receptors mediating this action: H2-receptor agonists were able to activate the adenylate cyclase with “relative potencies” similar to that found on typical H2-receptors, and cimetidine, a specific H2-receptor antagonist, competitively inhibited the response to histamine with a K1 value reflecting its interaction with a single population of H2-receptors. On the contrary, data obtained with H1-receptor agonists and antagonists reflect their interaction with H2-receptors rather than H1-receptors. Thus H2-receptors are involved in the activation of adenylate cyclase of the capillary fraction.

Journal ArticleDOI
TL;DR: The results indicate that α‐tubulin is an integral vesicle membrane protein, whereas most of the β sub‐unit is peripherally attached and can be easily dissociated from the vesicles membrane with EGTA.
Abstract: The major protein in isolated synaptic vesicles from bovine cerebral cortex has been compared to tubulin by sodium dodecyl sulphate-urea polyacrylamide gel electrophoresis, by two-dimensional gel electrophoresis, and by peptide mapping following limited proteolysis of the protein by Staphylococcus aureus protease. The results establish in purified synaptic vesicles the presence of tubulin, which is composed of the alpha and beta subunits. In the presence of ethyleneglycolbis)aminoethyl ether)-N,N'-tetraacetic acid (EGTA) or magnesium in the isolation buffers, the synaptic vesicles contained mainly the alpha-tubulin whereas the beta subunit was less abundant. Similarly, synaptosomal plasma membranes that were prepared in the presence of EGTA also contained more of alpha-tubulin than of the beta subunit. Non-ionic detergents such as Triton X-100 or Nonidet P-40 failed to solubilize the tubulin from the synaptic vesicles. Ionic detergents such as deoxycholate and sodium dodecyl sulphate solubilized all the vesicle proteins, including tubulin. The results indicate that alpha-tubulin is an integral vesicle membrane protein, whereas most of the beta subunit is peripherally attached and can be easily dissociated from the vesicle membrane with EGTA.

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TL;DR: The half‐life of the glucose content of brain was found to be significantly prolonged during anesthesia and to be significant and positively correlated with the plasma glucose concentration.
Abstract: A method has been developed for the measurement of the turnover rate constant or the half-life of the free glucose content of brain. It is based on an equation derived by the mathematical analysis of a kinetic model of the equilibration of the specific activity of the free glucose in brain with that of the plasma during an infusion of radioactive glucose. The method requires the measurement of the time course of the specific activity of glucose in the arterial plasma during an intravenous infusion of radioactive glucose for a period of 1 to 4 min and the specific activity of the free glucose in brain at the termination of the infusion. The turnover rate constant, or the half-life, is then calculated from these data by means of the operational equation of the method. The technique has been applied to conscious and anesthetized rats. In conscious rats the half-life of the free glucose content of brain was found to be 1.6 +/- 0.5 min (mean +/- S.D.) when the animals were killed by decapitation and 1.2 +/- 0.2 min (mean +/- S.D.) when they were killed by microwave irradiation; this difference is not statistically significant. In anesthetized rats, the half-life was found to be 2.6 +/- 0.8 min (mean +/- S.D.) in those killed by decapitation and 1.8 +/- 0.3 min (mean +/- S.D.) in those killed by microwave irradiation; this difference is statistically significant. The half-life of the glucose content of brain was found to be significantly prolonged during anesthesia and to be significantly and positively correlated with the plasma glucose concentration (r = 0.78; p < 0.001).

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TL;DR: Enrichment of alkaline phosphatase and ‐γ‐glutamyl transpeptidase activity in microvessel preparations supports the endothelial localization of these enzymes.
Abstract: Microvessels were isolated from rat brain using a double collagenase treatment which removed the endothelial basement membranes. The isolate was characterized by intact luminal and abluminal membranes and an absence of pericytes and astrocyte membranes. Minimal contamination by 5'-nucleotidase, an enzyme believed exclusively localized within the plasma membranes of neuroglia, established the purity of the isolated microvessels. Enrichment of alkaline phosphatase and gamma-glutamyl transpeptidase activity in microvessel preparations supports the endothelial localization of these enzymes.

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TL;DR: The results suggest that lithium administration causes selective changes in brain neurotransmitter receptor systems and that the net result of these changes may be a decrease in GABAergic and serotoninergic activity.
Abstract: Chronic, but not acute, consumption of lithium leads to a significant decrease in serotonin and GABA receptor binding in selected regions of the rat brain, with no changes noted in P-adrenergic or cholinergic muscarinic receptor binding. In addition, the concentration of β-methoxytyramine, a dopamine metabolite, in the corpus striatum was increased in the animals treated chronically with lithium, suggesting a possible enhancement in dopamine release, or inhibition of uptake, in this brain area. In contrast, chronic consumption of rubidium had no effect on any of the parameters studied. The results suggest that lithium administration causes selective changes in brain neurotransmitter receptor systems and that the net result of these changes may be a decrease in GABAergic and serotoninergic activity. The fact that these alterktions are noted only after chronic administration suggests that they may be related to the therapeutic action of lithium in the prophylactic treatment of recurrent manic- depressive psychosis.

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TL;DR: An aqueous extract derived from selected intraocular tissues of 15‐day chick embryos contains a soluble macromolecular agent which is capable of ensuring the survival of 8-day chick embryonic ciliary ganglionic neurons in monolayer culture.
Abstract: An aqueous extract derived from selected intraocular tissues of 15-day chick embryos contains a soluble macromolecular agent which is capable of ensuring the survival of 8-day chick embryonic ciliary ganglionic neurons in monolayer culture. When this ciliary neuronotrophic factor (CNTF) was concentrated using ultrafiltration and subjected to Sephadex G100 and G200 chromatography, activity was detected in most of the eluted fractions. A peak of the most active fractions was eluted in a region corresponding to a molecular weight of 35-40 X 10(3) and contained about 20-30% of the applied protein. CNTF activity bound readily to DE-52 cellulose resin at neutral pH and was eluted with NaCl in a narrow region containing about 20-40% of the applied protein. Gel electrophoretic staining profiles of the active DE52 fraction indicated considerable (but still only partial) simplification in protein composition. While significant CNTF activity losses were incurred in response to each of the above treatments, an active material could be conveniently generated in one working day in milligram amounts having a specific activity of 60,000 trophic units/mg protein. This trophic activity is in the same range as that of the only other known neuronotrophic factor, Nerve Growth Factor.