bluuebunny/crossref_metadata_2025 huggingface dataset:
Glycogen Metabolism in Bovine Adrenal Medulla
Authors: Millaruelo Ana , Sagarra M. Rosa , Miras‐Portugal M. Teresa
Published: October 2006

Abstract: Glycogen content was determined both in whole adrenal medullary tissue and in isolated adrenal chromaffin cells, in which it responds to glucose deprivation and restoration. [14C]glucose incorporation into glycogen in isolated adrenal chromaffin cells is increased by previous glucose deprivation (“fasting”). Total glycogen synthase activities are 452 ± 66 mU/g in whole tissue and 305 ± 108 mU/g in isolated cells. The Km of glycogen synthase for UDP‐glucose is 0.67 mM with 13 mm glucose‐6‐phosphate and 1 mm without this effector. The in vitro inactivation process of glycogen synthase a has been found to be mainly cyclic AMP‐dependent, but it also responds to Ca2+. Total glycogen phosphorylase activities are 8.69 ± 1.26 U/g in whole tissue and 2.38 ± 0.30 U/g in isolated cells. The requirements for interconversion in vitro of both glycogen synthase and phosphorylase suggest a system similar to that of other tissues. During incubation of isolated adrenal chromaffin cells with 5 mm‐glucose, phosphorylase a activity decreases and synthase a activity increases; these changes are more marked in “fasted” cells. Glycogen content and glycogen synthase and phosphorylase activities are higher in the adrenal medulla than in the brain, suggesting a greater metabolic role of glycogen in the adrenal medulla.
Cholinergic Modulation of the Release of [3H]Acetylcholine from Synaptosomes of the Myenteric Plexus
Authors: Briggs Clark A. , Cooper Jack R.
Published: October 2006

Abstract: The purpose of these experiments was to determine if cholinergic agents affected the release of acetylcholine (ACh) from a synaptosomal preparation of the guinea pig ileum myenteric plexus. The synaptosomal preparation was first incubated with the precursor [3H]choline; subsequently, release of the stored [3H]ACh was measured. The release was decreased by oxotremorine or exogenous ACh plus hexamethonium and increased by exogenous ACh plus atropine. The nicotinic agonist 1,1‐dimethyl‐4‐phenylpiperazinium (DMPP) evoked release that was inhibited by nicotinic antagonists or muscarinic agonists. Release was stimulated half‐maximally by approximately 2 μm‐ and maximally by 10 μm‐DMPP. Either in the absence of calcium or at 0°C, DMPP was without effect. The effect of 10 μm‐DMPP was brief, a significant stimulation occurring only within the first 2 min at 37°C. Tetrodotoxin also inhibited excitation by DMPP but not completely. Thus, the release of [3H]ACh appears to be presynaptically modulated, negatively by muscarinic agonists and positively by nicotinic agonists.
Influence of Severe Hypoglycemia on Mitochondrial and Plasma Membrane Function in Rat Brain
Authors: Agardh C.‐D. , Chapman A. G. , Pelligrino D. , Siesjö B. K.
Published: October 2006

Abstract: Previous experiments have shown that severe hypoglycemia disrupts cerebral energy state in spite of a maintained cerebral oxygen consumption, suggesting uncoupling of oxidative phosphorylation. Other studies have demonstrated that hypoglycemia leads to loss of cerebral cortical phospholipids and phospholipid‐bound fatty acids. The objective of the present study was, therefore, to study respiratory characteristics of brain mitochondria during severe hypoglycemia and to correlate respiratory activity to mitochondrial phospholipid composition. Mitochondria were isolated after 30 or 60 min of hypoglycemia with ceased EEG activity, and after a 90‐min recovery period, and their resting (state 4) and ADP‐stimulated (state 3) oxygen consumption rates and phospholipids and phospholipid‐bound fatty acid content were measured. After 30 min of hypoglycemia, state 3 respiration decreased without any increase in state 4 respiration or change in ADP/O ratio. This decrease, which occurred with glutamate plus malate—but not with succinate—as substrates, was partly reversed by addition of bovine serum albumin and KCI. Chemical analyses of isolated mitochondria did not reveal changes in their phospholipid or fatty acid content. The results thus failed to account for the dissociation of cerebral energy state and oxygen consumption. It is emphasized, though, that uncoupling may well occur in vivo due to accumulation of free fatty acids and “futile cycling” of K+ and Ca2+. After 60 min of hypoglycemia, a moderate decrease in state 3 respiration was observed also with succinate as substrate, and there was some decrease in ADP/O ratios in KCI‐containing media. However, the changes in ADP/O ratios were more conspicuous during recovery; in addition, state 4 respiration increased significantly. It is concluded that changes in mitochondrial function after 30 min of hypoglycemia are potentially reversible but that true mitochondrial failure develops in the recovery period following 60 min of hypoglycemia. This conclusion was corroborated by results demonstrating incomplete recovery of cerebral energy state. Since EEG and sensory evoked potentials return after 30 min but not after 60 min of hypoglycemia it seemed difficult to explain failure of return of electrophysiological function after 60 min of hypoglycemia solely by mitochondrial dysfunction; plasma membrane function was therefore assessed by measurements of extracellular potassium activity ([K+]e). The results showed that whereas [K+]e remained close to control in the recovery period following 30 min of hypoglycemia it rose progressively during recovery following 60 min of hypoglycemia. Possibly, inhibition of Na+ K+–activated ATPase could contribute to the permanent loss of spontaneous or evoked electrical activity.
Regional Blood‐Brain Barrier Permeability to Amino Acids After Portacaval Anastomosis
Authors: Mans Anke M. , Biebuyck Julien F. , Shelly Kirk , Hawkins Richard A.
Published: October 2006

Abstract: The influx of phenylalanine, tryptophan, leucine, and lysine across the blood‐brain barrier of individual brain structures was studied in rats 7–8 weeks after a portacaval shunt or sham operation. The method involved a brief infusion of labeled amino acid in tracer quantity and quantitative autoradiography. The clearance rates of phenylalanine, tryptophan, and leucine were increased in proportion to each other in every region examined, but not by the same factor. Tryptophan clearance increased the most (about 200%) and leucine the least (about 30%), compared with phenylalanine (about 80%). This was unexpected, as all three amino acids are believed to be transported by the same mechanism. The changes were most marked in several limbic structures and the reticular formation, whereas the hypothalamus was least affected. Plasma clearance of lysine was decreased in all areas by about 70%. Since the circulating lysine concentration was decreased by 13%, the actual rate of lysine influx was even more reduced. The results demonstrate specific alterations in two different amino acid transport systems. The resulting excess brain neutral amino acids, some of which are neurotransmitter precursors, as well as reduced basic amino acid availability, may be of etiological significance in hepatic encephalopathy.
Increased Levels of 3′,5′‐Cyclic Adenosine Monophosphate Induced by Cobaltous Ion or 3‐Isobutylmethylxanthine in the Incubated Mouse Retina: Evidence Concerning Location and Response to Ions and Light
Authors: Cohen Adolph I.
Published: October 2006

Abstract: Dark levels of 3′,5′‐cyclic adenosine monophosphate (cyclic AMP) of mouse retinas incubated in Earle's medium were elevated by 3‐isobutylmethylxanthine (IBMX) and/or Co2+ or Mn2+, but not by Cd2+, methylverapamil, or excess Mg2+ or Ca2+. Light reduced elevated dark levels of cyclic AMP in the presence of agents known to block the light modulation of postreceptoral neurons (aspartate, Co2+ high Mg2+), a finding consistent with a cyclic AMP metabolism in photoreceptors. Co2+‐elevated cyclic AMP levels were not less light‐sensitive than cyclic GMP levels. Ouabain substantially increased IBMX‐elevated cyclic AMP with a persistent light response, but reduced the dark action of Co2+. IBMX, but not Co2+, also increased cyclic AMP in receptorless (rd/rd) retinas; haloperidol partly reduced this IBMX effect. In normal retinas in Co2+ medium, progressively replacing Na+ by K+ (but not choline+) from 1–50 mm caused a progressive fall in dark, light‐sensitive cyclic AMP levels, but from 50 to 100 mm‐K+ there appeared haloperidol‐preventable increases in both the dark‐ and light‐insensitive levels of cyclic AMP. In IBMX‐aspartate medium a haloperidol‐preventable, light‐insensitive increase in cyclic AMP appeared from 20 mm‐K+ upwards. Haloperidol‐preventable increases in cyclic AMP as induced by high K+ required Co2+ in normal retinas, but not in receptorless retinas, and 5 mm‐Co2+ greatly increased the response to dopamine in receptorless retinas. The postdopaminergic neurons, which are 4th‐order neurons, may have become hyper‐sensitive to dopamine in receptorless retinas consequent to the absent signal from the 1st‐order photoreceptors, or directly, as an effect of the same gene underlying the dystrophy.
Brain Cortical Fatty Acids and Phospholipids During and Following Complete and Severe Incomplete Ischemia
Authors: Rehncrona S. , Westerberg E. , Åkesson B. , Siesjö B. K.
Published: October 2006

Abstract: To explore the possibility that peroxtdative degradation of brain tissue lipid constituents is an important mechanism of irreversible ischemic damage, we measured cortical fatty acids and phospholipids during reversible brain ischemia in the rat. Neither complete nor severe incomplete ischemia (5 and 30 min) caused any measurable breakdown of total or individual fatty acids or phospholipids. Except for a small (and reversible) decrease of inositol plus serine phosphoglycerides in the early postischemic period following 30 min of incomplete ischemia, there were no significant losses of fatty acids or phospholipids during recirculation. Since peroxidation, induced in brain cortical tissue in vitro, characteristically involves degradation of polyenoic fatty acids (arachidonic and docosahexaenoic acids) and of ethanolamine phosphoglycerides, the present in vivo results fail to support the hypothesis that peroxidation of membrane lipids is of primary importance for ischemic brain cell damage. Both complete and severe incomplete ischemia caused a similar increase in the tissue content of free fatty acids (FFA). Thus the FFA pool increased by about 10 times during a 30‐min ischemic period, to constitute 1 ‐ 2% of the total fatty acid pool. Since there was a relatively larger increase in polyenoic FFA (especially in arachidonic acid) than in saturated FFA, the release of FFA may be the result of activation of a phospholipase A2 unbalanced by reesterification. Increased levels of FFA persisted during the initial recirculation period, but a gradual normalization occurred and the ischemic changes were essentially reversed at 30 min after restoration of circulation. The pathophysiological implications of the changes in FFA are discussed with respect to mitochondrial dysfunction, formation of cellular edema and prostaglandin‐mediated deterioration of postischemic circulation.
[14C]Glucose Metabolism in Sympathetic Ganglia of Chicken Embryos and in Primary Cultures of Neurons and of Other Cells from These Ganglia
Authors: Larrabee Martin G.
Published: October 2006

Abstract: Metabolism of [1‐14C]glucose and [6‐14C]glucose was measured in sympathetic ganglia excised from chicken embryos 12–16 days old and in primary cultures of neurons or nonneurons prepared from these ganglia. Some metabolic rates tended to change with the tissue/medium ratio, so this variable had to be controlled. Less C‐6 than C‐1 of glucose was put out in CO2 by all three types of preparations, indicating operation of the hexosemonophosphate shunt. The C‐6/C‐1 ratio was greater for the neuronal cultures and for intact ganglia than for the nonneuronal cultures. The C‐6/C‐1 ratio for the neurons increased with the amount of tissue added to a given volume of incubation medium, in agreement with previous experiments on embryonic dorsal root ganglia (Larrabee, 1978). Per unit of protein, the output of C‐1 of glucose in CO2 was higher in both the neuronal and the nonneuronal cultures than in intact ganglia, whereas that of C‐6 was higher in the neuronal cultures and lower in the nonneuronal ones than in the ganglia. The rates of release in lactate of C‐1 and C‐6 of glucose were 3–5 times higher from both types of cultures than from intact ganglia. The average rates of incorporation of C‐1 and C‐6 of glucose into tissue constituents were lower in the cultures than in intact ganglia, significantly so for incorporation of C‐6 in the nonneuronal cultures.
GABA Autoreceptors Are Not Coupled to Benzodiazepine Receptors in Rat Cerebral Cortex
Authors: Brennan Michael J. W.
Published: October 2006

Abstract: Depolarization‐induced release of [3H]γ‐aminobutyric acid ([3H]‐GABA) from preloaded slices of rat cerebral cortex was inhibited by muscimol and THIP in a dose‐dependent fashion. This inhibition of release was prevented by the GABA antagonists bicuculline and picrotoxin. These results confirm previous reports postulating the existence of GABA autoreceptors on GABAergic terminals. Since benzodiazapines are known to facilitate postsynaptic GABA actions, the effect of flunitrazepam on the inhibition of GABA release mediated through the autoreceptors has been examined. At a concentration of 1 μm or 10 μm, flunitrazepam had no effect on the IC50 values for muscimol or THIP in inhibiting stimulated GABA release. It thus seems that GABA autoreceptors are not functionally coupled to benzodiazepine receptors in rat cerebral cortex.
Studies on the Biosynthesis of Intermediate Filament Proteins in the Rat CNS
Authors: Strocchi Paola , Dahl D. , Gilbert Jeffrey M.
Published: October 2006

Abstract: The biosynthesis of brain intermediate filament proteins [neurofilament proteins and glial fibrillary acidic protein (GFA)] was studied with cell‐free systems containing either rat spinal cord polysomes (free polysomes or rough microsomes) and rabbit reticulocyte factors or wheat germ homogenate containing spinal cord messenger RNA. The products of translation were isoated by immunoaffinity chromatography and then analyzed by two‐dimensional gel electrophoresis (2DGE) followed by fluorography. The free polysome population was found to synthesize two neurofilament proteins (MW 145K, p15.4, and MW 70K, pl 5.3) and three isomers of GFA (α, β, and γ) that differ in isoelectric point. Wheat germ homogenate containing messenger RNA extracted from free cord polysomes synthesized two proteins that comigrated with neurofilament protein standards at 145K 5.4 and 70K 5.3; these proteins were partially purified by neurofilament affinity chromatography. The wheat germ system also synthesized the α, β, and γ isomers of GFA as characterized by immunoaffinity chromatographic purification and comigration with standards in 2DGE analysis. Our data are consistent with the conclusion that synthesis of neurofilament proteins requires multiple messenger RNAs. Also, synthesis of intermediate filament proteins occurs in the free polysome population; detectable amounts of these proteins were not synthcsized by the rough microsomes.
Changes in Brain Glycogen During Slow‐Wave Sleep in the Rat
Authors: Karnovsky M. L. , Reich P. , Anchors J. M. , Burrows B. L.
Published: October 2006

Abstract: During slow‐wave sleep, rat brain glycogen increases within a few minutes to about 70% above waking levels. Upon awakening, the increment is lost within 2–5 min. After repeated episodes of sleep, brain glycogen levels are comparable to those observed after only a single episode of sleep. Liver glycogen is unaffected by slow‐wave sleep.
Phosphoinositide Metabolism in the Retina: Localization to Horizontal Cells and Regulation by Light and Divalent Cations
Authors: Anderson Robert E. , Maude Maureen B. , Kelleher Paula A. , Rayborn Mary E. , Hollyfield Joe G.
Published: October 2006

Abstract: Isolated retinas from Xenopus laevis incorporated greater amounts of [3H]inositol and 32Pi into phosphoinositides when incubated in light than did control retinas incubated in the dark. Inositol was primarily incorporated into phosphatidylinositol (83–86%), while phosphate labeled the polyphosphoinositides (72–79%). The incorporation of radioactive glycerol, serine, choline, or ethanolarnine into retinal lipids was unaffected by light. Following incubation with [3H]inositol, the cell type involved in the light response was identified by light and electron microscope autoradiography to be the horizontal cell. These results are consistent with a classic phosphatidylinositol effect in the retina. An interesting feature of this response is that the stimulus (light) is received in the photoreceptor cell and the effect is manifest in the horizontal cell.
Identification of Muscarinic Receptors in Rat Cerebral Cortical Microvessels
Authors: Grammas Paula , Diglio Clement A. , Marks Bernard H. , Giacomelli Filiberto , Wiener Joseph
Published: October 2006

Abstract: Microvessels isolated from rat cerebral cortex consist mainly of capillaries (>85%). Fresh, intact microvessel preparations have been analyzed by radioligand binding techniques for muscarinic receptors. Scatchard analysis of specific quinuclidinyl benzilate (QNB) binding indicates that microvessels possess a large number of muscarinic sites (914 fmol/mg protein) of high affinity (Kd= 0.034 nM). The association and dissociation rate constants (0.37 min−1 nM−1 and 0.0067 min−1, respectively) yield an equilibrium Kd of 0.018 nM. Displacement of [3H]QNB by muscarinic ligands and control substances is typical of muscarinic receptors. The results indicate that cerebral microvessels possess a large population of muscarinic receptors.
Effects of Vasoactive Intestinal Polypeptide, Monoamines, Prostaglandins, and 2‐Chloroadenosine on Adenylate Cyclase in Rat Cerebral Microvessels
Authors: Huang Minta , Rorstad O. P.
Published: October 2006

Abstract: Adenylate cyclase in microvessels isolated from rat cerebral cortex was stimulated by guanine nucleotides, catecholamines, prostaglandin E1, prostaglandin E2, and 2‐chloroadenosine. Catecholamine stimulation was mediated by interaction with β‐adrenergic receptors. The order of relative potency was: isoproterenol > epinephrine > norepinephrine. Activation of microvessel adenylate cyclase by prostaglandins E1 and E2 as well as by 2‐chloroadenosine was dose related. Twenty‐two peptides were tested for possible effects on the microvessel adenylate cyclase. Only vasoactive intestinal polypeptide (VIP) was stimulatory. No inhibitory action was observed. Activation by VIP required guanosine triphosphate and was dose dependent from 10 nM to μM (ED50= 0.1 μM). At 30°C, stimulation of adenylate cyclase by the peptide increased linearly with time for up to 15 min. The effect of VIP was not inhibited by phentolamine or propranolol, suggesting that its action was not elicited by interaction with α‐ or β‐adrenergic receptors. Activation achieved by VIP and isoproterenol, prostaglandin E1, or 2‐chloroadenosine was the sum of the individual stimulations, suggesting that receptors for VIP were distinct from those for isoproterenol, prostaglandin E1, and 2‐chloroadenosine.
Regional Cerebral Glucose Utilization in Rats with Portacaval Anastomosis
Authors: Mans Anke M. , Biebuyck Julien F. , Davis Donald W. , Bryan Robert M. , Hawkins Richard A.
Published: October 2006

Regional cerebral glucose utilization was measured using [2‐14C]glucose in rats with an end‐to‐side portacaval anastomosis. The experiments were conducted in two groups of rats 4 to 8 weeks after portacaval shunting was established. One group was paralyzed and given N2O:O2 (70:30), whereas the other was conscious, unstressed, and unaware of the experiment. In both groups the rate of glucose utilization was decreased in almost all brain structures by an average of 20% after portacaval shunting. The results showed definitively that cerebral energy metabolism was reduced at a time when there were no obvious neurological abnormalities.
Changes in Brain Levels of Acidic, Basic, and Neutral Amino Acids After Consumption of Single Meals Containing Various Proportions of Protein
Authors: Glaeser Bruce S. , Maher Timothy J. , Wurtman Richard J.
Published: October 2006

Abstract:Rats fasted overnight were allowed to consume single meals containing 0, 18, or 40% protein or continued to fast; after 2 h, brains and sera were taken and assayed for various amino acids. In general, serum levels of most amino acids were reduced by the 0% protein meal and elevated by the high‐protein meal when compared with those associated with fasting conditions. Exceptions were those not diminished by the 0% protein meal (tryptophan, methionine, proline) and those increased (alanine) or decreased (glycine) by all of the test meals. Amino acids exhibiting the broadest normal ranges (estimated by comparing their serum levels after 40% protein with those after 0% protein) were tyrosine, leucine, valine, isoleucine, and proline; serum lysine and histidine, two basic amino acids, also varied more than threefold. Brain levels of lysine, histidine, and some of the large neutral amino acids (LNAAs) also exhibited clear relationships to the protein content of the test meal: those of valine, leucine, and isoleucine were depressed by the 0% protein but increased (compared with 0% protein) when protein was added to the meal: brain tyrosine was increased by all of the test meals in proportion to their protein contents; tryptophan, phenylalanine, and glutamate were increased after the 0% protein meal but not by protein‐containing meals; brain lysine, histidine, and methionine were increased after the high‐protein meal, and brain alanine was increased slightly by all of the meals. For each of the LNAAs, significant correlations were observed between its brain level in any animal and the ratio of its serum concentration to the sum of the concentrations of its LNAA competitors (for blood‐brain barrier transport). For valine, tyrosine, lysine, and histidine, significant correlations were obtained between their brain and serum levels.
Neurone‐Specific Enolase and Creatine Phosphokinase Are Protein Components of Rat Brain Synaptic Plasma Membranes
Authors: Lim L. , Hall C. , Leung T. , Mahadevan L. , Whatley S.
Published: October 2006

Abstract: Neuron‐specific enolase and creatine phosphokinase were found, by 2‐dimensional gel analysis, in rat brain synaptic plasma membranes (SPM). The identity of these enzymes was confirmed by comigration with purified rat brain NSE and CPK and by peptide analysis. The specific enzymatic activities of enolase and creatine phosphokinase, as well as of pyruvate kinase, also present on the membranes, were comparable to those in the homogenates when these three enzymes were fully activated. In the SPM all three enzymes, particularly enolase, were partially cryptic in that enzymatic activities were very low unless the membranes were treated with Triton X‐100. They were resistant to both low‐salt and high‐salt extraction and to trypsin, except when Triton X‐100 was present. These results suggest that the enzymes are tightly bound protein components of the membrane and that they may constitute an assembly capable of generating ATP.
Respiration in Primary Cultured Cerebellar Granule Neurons and Cerebral Cortical Neurons
Authors: Jameson Nancy , Olson James , Nguyen Hung , Holtzman David
Published: October 2006

Abstract: Respiration was measured polarographically in primary cultures enriched with cerebellar granule neurons or cerebral cortical neurons. The basal respiratory rate, measured on the sixth day after culturing, was 12.00 natom equiv. O/mg protein/min for the cortical neurons and 12.70 natom equiv. O/mg protein/min for the granule neurons. Maximal stimulation by 2,4‐dinitrophenol produced a 20–40% increase over the basal rate for both neuronal types. Oligomycin inhibited neuronal basal respiration by 45%. These respiratory rates in neurons from primary culture are markedly lower than those measured in astrocytes grown under similar conditions.
The Significance of Mast Cells as a Source of Histamine in the Mouse Brain
Authors: Orr Edward L. , Pace Kelly R.
Published: October 2006

Abstract: Knowledge of the relative contributions of mast cells and neurons to the overall pool of histamine in the brain is a prerequisite to determining the significance and role of this amine in brain function. Consequently, we analyzed the levels of brain histamine in four genotypes (+/+, W/+, Wv/+, and WIWv) of WBB6F1 mice, whose numbers of brain‐associated mast cells vary in a genotypically specific manner. Although mast cell numbers ranged from a total absence of mast cells (W/Wv) to an average of about 500 mast cells/brain (W/+), no significant differences between genotypes were found in the quantities of histamine in whole brains, brain regions, or crude subcellular fractions. Thus, in this strain of mice, mast cells are not a significant source of histamine in the brain. This suggests that most of the histamine is of neuronal origin. Since neuronal histamine levels are maintained only by continued histidine decarboxylase activity, complete inhibition of this enzyme by α‐fluoromethylhistidine, a “suicide” inhibitor of histidine decarboxylase, would totally deplete W/Wv mice of brain histamine. This was not found to occur in the W/Wv mice, suggesting that neuronal stores of histamine can be maintained in the absence of histidine decarboxylase, or that an additional nonneuronal, non‐mast cell source of histamine exists in the W/Wv mouse brain.
Inositol Phospholipid Hydrolysis in Rat Cerebral Cortical Slices: II. Calcium Requirement
Authors: Kendall David A. , Nahorski Stefan R.
Published: October 2006

Abstract: The calcium requirement for agonist‐dependent breakdown of phosphatidylinositol and polyphos‐phoinositides has been examined in rat cerebral cortex. The omission of added Ca2+ from the incubation medium abolished [3H]inositol phosphate accumulation from prelabelled phospholipid induced by histamine, reduced that due to noradrenaline and 5‐hydroxytryptamine, but did not affect carbachol‐stimulated breakdown. EC50 values for agonists were unaltered in the absence of Ca2+. Removal of Ca2+ by preincubation with EGTA (0.5 mM) abolished all responses, but complete restoration was achieved by replacement of Ca2+. The EC50for Ca2+ for histamine‐stimulated [3H]inositol phosphate accumulation was 80 μM. Noradrenaline‐stimulated breakdown was antagonised by manganese (IC50 1.7 mM), but not by the calcium channel blockers nitrendipine or nimodipine (30 μM). The calcium ionophore A23187 stimulated phosphatidylinositol/polyphosphoinositide hydrolysis with an EC50 of 2 μM, and this response was blocked by EGTA. Omission of Ca2+ or preincubation with EGTA or Mn2+ (EC50= 230 μM) greatly enhanced the incorporation of [3H]inositol into phospholipids. The IC50for Ca2+ in inhibiting incorporation was 25 μM. The results show that different receptors mediating phosphatidylinositol/ polyphosphoinositide breakdown in rat cortex have quantitatively different Ca2+ requirements, and it is suggested that rigid opinions regarding phosphatidylinositol/ polyphosphoinositide breakdown as either cause or effect of calcium mobilisation in rat cortex are inappropriate.
Experimental Brain Ischemia: Neuron‐Specific Enolase Level in Cerebrospinal Fluid as an Index of Neuronal Damage
Authors: Steinberg R. , Gueniau C. , Scarna H. , Keller A. , Worcel M. , Pujol J. F.
Published: October 2006

Abstract: Levels of neuron‐specific enolase (NSE) were measured in rat CSF following occlusion of the four major arteries to the brain for 10, 20, or 30 min. In the CSF of rats submitted to 30 min of total ischemia, an up to ninefold increase of NSE level occurred within the first few hours and then slowly diminished. Significant levels were seen for as long as 8 days. Histological observations 3 days after ischemia showed neuronal loss as well as neu‐ ronal damage in several forebrain regions such as hippocampus, striatum, and thalamus. Ischemia was followed by transient decreases in exploration behavior and neurological states that were no longer visible 24 h later. After 10 or 20 min ischemia, NSE levels were increased to a lesser degree and fewer damaged neurons were observed. The positive correlation between duration of ischemia and amount of NSE release in CSF indicates that the measurement of NSE in the CSF is a sensitive and reliable index of neuronal lesions.
Biochemical and Immunochemical Studies on the GABAergic System in the Rat Fallopian Tube and Ovary
Authors: Apud J. A. , Tappaz M. L. , Celotti F. , Negri‐Cesi P. , Masotto C. , Racagni G.
Published: October 2006

Abstract: γ‐Aminobutyric acid (GABA) and glutamic acid decarboxylase (GAD) activities were measured in the ovary and the Fallopian tube of rats and compared with brain values. GABA levels in the Fallopian tube were about twice as high as in the brain, while in the ovary they represented only about 5% of the amino acid content of the CNS. In vitro decarboxylation of glutamate, measured via CO2 formation, occurred both in the Fallopian tube and in the ovary. These two organs contained, respectively, 10% and 1% of brain GAD activity. However, the actual formation of GABA from glutamate in a high‐speed supernatant was detectable only in the Fallopian tube, where it represented about 5% of brain GAD activity. In contrast with the enzyme present in ovary, liver, anterior pituitary, and kidney, that in the Fallopian tube was quantitatively precipitated by a specific antiserum directed against rat neuronal GAD. Moreover, subcutaneous transplantation resulted in a quantitative decrease of both GABA levels and GAD activity in the Fallopian tube while no change occurred in the ovary, and vagus nerve section induced a 50% decrease of GAD activity in the Fallopian tube, although GABA levels were not significantly altered. The findings suggest an extrinsic GABAergic innervation in the rat Fallopian tube but not in the ovary.
S‐100 Modulates Ca2+‐Independent Phosphorylation of an Endogenous Protein (Mr= 19K) in Brain
Authors: Qi De‐Fang , Kuo J. F.
Published: October 2006

Abstract: A new brain enzyme (tentatively named protein kinase X), which catalyzes protamine phosphorylation modulated by S‐100, was reported recently. An endogenous substrate protein (Mr= 19K) for protein kinase × was isolated from brain by means of S‐100‐Sepharose 4B affinity chromatography. S‐100, but not calmodulin, promoted phosphorylation of the 19K Mr protein in a Ca2+‐independent manner, and this reaction was inhibited by gossypol. The substrate protein, localized in the particulate fraction, was present at a much higher level in brain from adult than neonatal rats (2‐day‐old), a developmental change similar to that seen for protein kinase X. It is suggested that a protein phosphorylation system modulated by S‐100 exists in brain, and that this process may be involved in regulation of certain neural functions.
Synthesis and Turnover of Cytoskeletal Proteins in Cultured Astrocytes
Authors: Chiu Fung‐Chow , Goldman James E.
Published: October 2006

Abstract: We previously reported that the cytoskeleton of rat astrocytes in primary culture contains vimentin, glial fibrillary acidic protein (GFAP), and actin. These proteins were found in a fraction insoluble in Triton X‐100 and thought to be assembled in filamentous structures. We now used primary astrocyte cultures to study the kinetics of synthesis and turnover of these cytoskeletal proteins. The intermediate filament proteins were among the most actively synthesized by astrocytes. High levels of synthesis were detectable by the third day of culture in the early log phase of growth, and the pattern of labeling at day 3 was similar to that at 14 days when the cultures had reached confluency. In short‐term incorporation experiments vimentin, GFAP, and actin in the Triton‐insoluble fraction were labeled within 5 min after exposure of the cultures to radioactive leucine. We did not detect any saturation of labeling for up to 6 h of incubation. The turnover of filament proteins studied by following the decay of radioactivity from prelabeled vimentin, GFAP, and cytoskeletal actin displayed biphasic decay kinetics for all three proteins. In the initial phase a fast‐decaying pool with a half‐life of 12–18 h contributed about 40% of the total activity in each protein. A major portion, about 60%, of each protein, however, decayed much more slowly, exhibiting a half‐life of about 8 days.
Rate Equations and Kinetics of Uptake of α‐Aminoisobutyric Acid and γ‐Aminobutyric Acid by Mouse Cerebrum Slices Incubated in Media Containing L(+)‐Lactate or a Mixture of Succinate, L‐Malate, and Pyruvate as the Energy Source
Authors: Cohen Stephen R.
Published: October 2006

Abstract: Influx of α‐aminoisobutyric acid (AIB) and γ‐aminobutyric acid (GABA) by mouse cerebrum slices incubated with L‐lactate or a mixture of succinate, Lmalate, and pyruvate (SMP) as the energy source follows the phenomenological rate equation for influx from pyruvate and glucose media: v =Vmax/(1 +Kt/S) +kuS, where v is rate and S is concentration of amino acid. There are two kinetically distinct, parallel components for concentrative uptake, one saturable, and one unsaturable. Rates are less with lactate than with pyruvate and still less with SMP (only GABA was studied), disproving the hypotheses that lower rates with pyruvate compared to glucose are due to an abnormal redox state in the tissue or to a Krebs cycle unbalanced by input at only one point. The carriers for AIB and GABA are qualitatively different. In lactate medium the capacity of each AIB carrier is unchanged but its affinity is reduced to one‐third. In lactate and SMP media the capacity of the saturable GABA carrier is diminished although its affinity is increased. Rates from these media with added glucose or a glucose analog confirm that amino acid and glucose fluxes are not coupled.
Determinants of Deoxyglucose Uptake in Cultured Astrocytes: The Role of the Sodium Pump
Authors: Brookes N. , Yarowsky P. J.
Published: October 2006

Abstract: Glucose utilization in primary cell cultures of mouse cerebral astrocytes was studied by measuring uptake of tracer concentrations of [3H]2‐deoxyglucose ([3H]2‐DG). The resting rate of glucose utilization, estimated at an extracellular K+ concentration ([K+]o) of 5.4 mM, was high (7.5 nmol glucose/mg protein/min) and was similar in morphologically undifferentiated and “differentiated” (dibutyryl cyclic AMP‐pretreated) cultures. Resting uptake of [3H]2‐DG was depressed by ouabain, by reducing [K+]o, and by cooling. These observations suggest that resting glucose utilization in astrocytes was dependent on sodium pump activity. Sodium pump‐dependent uptake in 2–3‐week‐old cultures was about 50% of total [3H]2‐DG uptake but this fraction declined with culture age from 1 to 5 weeks. Uptake was not affected by changes in extracellular bicarbonate concentration ([HCO3‐]o) in the range of 5–50 mM but was significantly reduced in bicarbonate‐free solution. At high [HCO3‐]o (50 mM) uptake was insensitive to pH (pH 6–8), whereas at low [HCO3‐]o (<5m M) uptake was markedly pH‐dependent. Elevation of [K+]o from 2.3 mM to 14.2–20 mM (corresponding to extremes of the physiological range of [K+]o) resulted in a 35–43% increase in [3H]2‐DG uptake that was not affected by culture age or by morphological differentiation. Our results indicate a high apparent rate of glucose utilization in astrocytes. This rate is dynamically responsive to changes in extracellular K+ concentration in the physiological range and is partially dependent on sodium pump activity.
Biphasic Liberation of Arachidonic and Stearic Acids During Cerebral Ischemia
Authors: Yasuda Hiroshi , Kishiro Kouji , Izumi Noriyoshi , Nakanishi Masato
Published: October 2006

Abstract: The mode of free fatty acid (FFA) liberation from the mouse brain during ischemia was investigated at various times after decapitation and under nizofenone treatment. Normal nonischemic brain FFAs consist mainly of palmitic acid (16:0), stearic acid (18:0), and oleic acid (18:1) with smaller amounts of arachidonic acid (20:4), docosahexaenoic acid (22:6), and others. Postde‐capitative ischemia induced a rapid, biphasic release of 20:4 after a short lag of <30 s. The first phase showed a rapid 6.4‐fold increase within 1 min of decapitation, followed by the second phase involving a slow release at less than one‐fifth the rate of the first phase and lasting for at least 10 min. A similar, but not so marked, biphasic liberation was observed with 18:0. However, all of the other fatty acids (16:0, 18:1, 22:6, and others) were released only in a single phase at a slow rate. The time course for the rapid and specific liberation of 20:4 coincided with the time course for the decrease in brain ATP concentration during ischemia. Pretreatment of the animals with nizofenone resulted in a marked suppression of both FFA liberation and ATP depletion during ischemia. This suppression was particularly noteworthy with 20:4 and 18:0. The present study indicates that there is a specific and rapid liberation of 20:4 and 18:0 in a very early stage of ischemia and that this liberation seems to depend on availability of ATP in the brain. The physiological role of this transient 20:4 liberation during ischemia is discussed.
Regional Differences in the Coupling of Muscarinic Receptors to Inositol Phospholipid Hydrolysis in Guinea Pig Brain
Authors: Fisher Stephen K. , Bartus Raymond T.
Published: October 2006

Abstract: The differential effects of muscarinic agents on inositol phospholipid hydroilysis and the role in this process of putative muscarinic receptor subtypes (M1 and M2) were investigated in three regions of guinea pig brain. Addition of the agonist oxotremorine‐M to slices of neostriatum, cerebral cortex, or hippocampus incubated in the presence of myo‐[2‐3H]inositol and Li+ resulted in a large accumulation of labeled inositol phosphates (733, 376, and 330% of control, respectively). In each tissue, the principal product formed was myo‐inositol 1‐phosphate (59–86%), with smaller amounts of glycerophosphoinositol and inositol bisphosphate. Only trace amounts of inositol trisphosphate could be detected. Regional differences were observed in the capacity of certain partial agonists to evoke inositol lipid hydrolysis, the most notable being that of bethanechol, which was four times more effective in the neostriatum than in either the cerebral cortex or hippocampus. In addition, the full agonists, oxotremorine‐M and carbamoylcholine, were more potent stimulators of inositol phosphate release in the neostriatum than in the cerebral cortex. The putative M1 selective agonist 4‐m‐chlorophenylcarbamoyloxy‐2‐butynyl trimethyl ammonium chloride had little stimulatory effect in any brain region, whereas the putative M1 selective antagonist pirenzepine blocked the enhanced release of inositol phosphates with high affinity in the cerebral cortex and hippocampus (Ki = 12.1 and 13.9 nM; “M1”) but with a lower affinity in the neostriatum (Ki = 160 nM; “M2”). In contrast to its differential effects on stimulated inositol lipid hydrolysis, no regional differences were observed in the capacity of pirenzepine to displace [3H]quinuclidinyl benzilate, a muscarinic antagonist, bound to membrane fractions. Atropine, an antagonist that does not discriminate between receptor subtypes, inhibited the enhanced release of inositol phosphates with similar affinities in the three regions (Ki = 0.40–0.60 nM). The results indicate that by measurement of inositol lipid hydrolysis, regional differences in muscarinic receptor coupling characteristics become evident. These differences, which are not readily detected by radioligand binding techniques, might be accounted for by either the presence of functionally distinct receptor subtypes, or alternatively, by regional variations in the efficiency of muscarinic receptor coupling to inositol lipid hydrolysis.
Resolution of Rat Brain Synaptic Phosphoprotein B‐50 into Multiple Forms by Two‐Dimensional Electrophoresis: Evidence for Multisite Phosphorylation
Authors: Zwiers H. , Verhaagen J. , van Dongen C. J. , de Graan P. N. E. , Gispen W. H.
Published: October 2006

Abstract:Phosphoprotein B‐50 was extracted from rat brain membranes by alkaline extraction and purified by ammonium sulphate precipitation and flat‐bed isoelectric focusing. The purified protein shows microheterogeneity upon isoelectric focusing in a narrow pH gradient (pH 3.5–5.0). As visualized by two‐dimensional gel electrophoresis, B‐50 resolved into four clearly separated forms which differ slightly in isoelectric point. The forms are in part mutually convertible by exhaustive phosphorylation (using protein kinase C) and dephosphorylation (usingEscherichia colialkaline phosphatase). Proteolysis withStaphylococcus aureusprotease yielded two radioactive peptides. Analysis of their molecular weights and the time course of their formation suggests that B‐50 was cleaved at only one specific site. Our data indicate the presence of more than one phosphorylatable site. The possibility that the heterogeneity of B‐50 was in part due to a glycoprotein nature of B‐50 was studied extensively. However, none of the six different methods used revealed the presence of glyco‐moieties in B‐50.
Dolichol in Human Brain: Regional and Developmental Aspects
Authors: Sakakihara Yoichi , Volpe Joseph J.
Published: October 2006

Abstract: Distinct regional differences in dolichol content were defined in human brain from 15 to 76 years of age. Concerning the regional distribution of dolichol, levels were: (1) higher in cortical gray matter than in subcortical white matter, (2) highest among cortical regions in temporal gray matter, (3) highest among all brain regions in thalamus, and (4) lowest among all brain regions in lower brain stem and spinal cord. The developmental changes in the contents of dolichol were found to be different among brain regions. For example, among regions with the highest levels of dolichol, in thalamus there was a six to sevenfold increase, but in parietal gray matter, only a 2.5‐fold increase. Regional and developmental changes in the proportions of the individual molecular species (isoprenologues) of dolichol were also observed. The findings indicate that the metabolism of dolichol is not uniform among regions of developing and aging human brain and may have implications for the role of dolichol in normal and diseased human brain.
Effects of Organic and Inorganic Lead on Synaptosomal Uptake, Release, and Receptor Binding of Glutamate in Young Rats
Authors: Regunathan S. , Sundaresan R.
Published: October 2006

Abstract: Chemical changes in central glutamate neurotransmission were assessed by measuring synaptic membrane receptor binding, the uptake and release by synaptosomes of glutamate in rats treated acutely with tetraethyl lead and chronically with lead acetate. The activity of Na+, K+‐ATPase in synaptosomes was measured to correlate with the changes in uptake/release studies. The affinity of receptor binding and uptake systems was significantly reduced although the number of receptor sites and the capacity of uptake systems were increased. The activity of Na+, K+‐ATPase was also found to be increased in synaptosomes. The changes were more marked in inorganic lead toxicity, and all three regions studied—cerebral cortex, cerebellum, and brainstem—showed significant alterations.
Streptozotocin‐Induced Diabetes Reduces Brain Serotonin Synthesis in Rats
Authors: Trulson Michael E. , Jacoby Jacob H. , MacKenzie Robert G.
Published: October 2006

Abstract: The rate of brain 5‐hydroxytryptamine (serotonin) synthesis and turnover in streptozotocin‐diabetic rats was assessed using three separate methods: the rate of 5‐hydroxytryptophan accumulation following decarboxylase inhibition with Ro 4–4602; the decline in 5‐hydroxyindoleacetic acid levels following monoamine oxidase inhibition with pargyline; and the rate of 5‐hydroxyindoleacetic acid accumulation following blockade of acid transport with probenecid. Each of the three methods revealed that 5‐hydroxytryptamine synthesis and turnover is decreased by 44–71% in diabetic rats with plasma glucose levels of between 500 and 600 mg%. In addition, the levels of free and bound plasma tryptophan were measured and the levels of the free amino acid were found to be the same in control and diabetic rats. Since diabetic rats exhibit a 40% decrease in brain tryptophan, the free tryptophan level in plasma does not predict brain tryptophan levels in diabetic rats. These data are discussed within the context of psychiatric disturbances experienced by diabetic patients.
Motor Activity Increases Tryptophan, 5‐Hydroxyindoleacetic Acid, and Homovanillic Acid in Ventricular Cerebrospinal Fluid of the Conscious Rat
Authors: Chaouloff F. , Laude D. , Guezennec Y. , Elghozi J. L.
Published: October 2006

Abstract: An investigation was made into the effects of running (I h at 20 m/min) on central serotonergic and dopaminergic metabolism in trained rats. Methodology involved continuous withdrawal of cerebrospinal fluid (CSF) from the third ventricle of conscious rats and measurements of tryptophan (TRP), 5‐hydroxyindoleacetic acid (5‐HIAA), and homovanillic acid (HVA) levels during a 2 h post‐exercise period. All three compounds were increased during the hour following exercise and returned to their basal values within an hour later. CSF flow rate was stable when metabolite levels were elevated. Brain determinations indicated that CSF metabolite variations only qualitatively paralleled brain changes. Indeed, post‐exercise TRP, 5‐HIAA, and HVA levels were increased to a greater extent in brain when compared to CSF. It is suggested that increased serotonergic and dopaminergic metabolism, caused by motor activity, may be involved in the behavioral effects of exercise.
Failure of Glucose and Branched‐Chain Amino Acids to Normalize Brain Glucose Use in Portacaval Shunted Rats
Authors: Mans Anke M. , Davis Donald W. , Biebuyck Julien F. , Hawkins Richard A.
Published: October 2006

Abstract Several abnormalities in brain and plasma amino acid concentrations caused by portacaval shunting in rats return toward normal after 4 days of intravenous infusion with either glucose or glucose with branched‐chain amino acids. To assess the effect of such treatment on brain energy metabolism, regional brain glucose use was measured using [14C]glucose and autoradiography, 5 weeks after portacaval shunting. In one experiment intravenous glucose or glucose with branched‐chain amino acids was given for 4 days. In a separate experiment the treatment was given orally for 2 weeks, and in addition to glucose use, brain monoamines and amino acids were measured. No other food was provided; the rats had free access to water. Normally fed shunted rats and sham‐operated rats served as controls. Both types of oral treatment lowered the high concentrations of tyrosine, phenylalanine, and glutamine in plasma and brain. Glucose without amino acids normalized brain tryptophan. Levels of brain norepinephrine, 5‐hydroxytryptamine (serotonin), and 5‐hydroxyindoleacetic acid were significantly raised after shunting. Treatment had no effect on norepinephrine but the glucose diet brought the indoles into the normal range. In contrast, neither intravenous nor oral treatment affected brain glucose use, which remained depressed by 25–30% in all brain areas examined.