bluuebunny/crossref_metadata_2025 huggingface dataset:
DISTRIBUTION AND PROPERTIES OF ANGIOTENSIN CONVERTING ENZYME OF RAT BRAIN
Authors: Yang H.‐Y. T. , Neff N. H.
Published: October 2006

Abstract— Angiotensin converting enzyme of rat brain was studied using Hip‐His‐Leu as substrate. The highest specific activity of the enzyme was associated with the microsomal fraction. The specific activity of the microsomal enzyme in several regions of the rat brain varied significantly. For example, the specific activities of the striatal and pituitary enzymes were about 10‐fold greater than that of the cerebral cortical enzyme. The enzyme required chloride ion; moreover, activity was inhibited in the presence of disodium EDTA or O‐phenanthroline, effects suggesting that the converting enzyme of brain is a metalloprotein. SQ‐20881, a nonapeptide that inhibits converting enzyme in peripheral tissue, was a potent inhibitor of the enzyme of brain. In addition to Hip‐His‐Leu, the microsomal fraction was capable of liberating C terminal dipeptides from angiotensin I, Hip‐Gly‐Gly and Z‐Gly‐ Gly‐Val. The broad substrate specificity of the enzyme suggests that, in addition to the possible contribution of the enzyme to the brain renin‐angiotensin system, other naturally occurring peptides might also be substrates for the enzyme.
THE EFFECT OF HYPERCAPNIA UPON INTRACELLULAR pH IN THE BRAIN, EVALUATED BY THE BICARBONATE‐ CARBONIC ACID METHOD AND FROM THE CREATINE PHOSPHOKINASE EQUILIBRIUM
Authors: Siesjö B. K. , Folbergrová Jaroslava , MacMillan V.
Published: October 2006

Abstract— Abstract‐Intracellular pH in the brain was evaluated by the bicarbonate‐carbonic acid method and from the creatine phosphokinase equilibrium, in rats exposed to 6–40 % CO2 for 45 min. There was a very good agreement between the two methods, indicating that the creatine phosphokinase equilibrium in vivo shows the pH dependence predicted from previous in vitro studies. The stepwise increase in the tissue CO2 tension from 45 to 265 mm Hg resulted in a lowering of the intracellular pH from 7.04 to 6.68. The regulation of intracellular pH in hypercapnia was far better than that which can be predicted from physicochemical buffering alone, and calculations indicate that the intracellular buffer base concentration increased by more than 10 mequiv./kg at the maximal Pco2 values encountered.
Increased monoamine oxidase activity following repeated electroshock seizures
Authors: Pryor G. T. , Scott M. K. , Peache SUSAN
Published: October 2006

We Have recently reported that chronic electroconvulsive seizures (ECS) lead to a long‐lasting increase (up to 6 weeks) in the activity of monoamine oxidase (MAO) in the brain of rats (Pryor and Otis, 1970). Other investigators have shown increased levels of norepinephrine (NE) and 5‐HT, more rapid clearance of intracisternally‐injected [3H]HNE (Kety, Javoy, Thierry, Julou and Glowinski, 1967), and increased tyrosine hydroxylase activity (Musacchio, Julou, Kety and Glowinski, 1969) at 24 h after a series of two ECS daily for 7 days. Together, these data suggest a sustained activation of the 5‐HT and/or NE systems following chronic ECS. We now report the results of a series of experiments in which some of the potential variables that may be involved in the MAO response were investigated and that indicate some biochemical specificity of the response. In these experiments, succinate dehydrogenase (SDH) activity was also assayed to assess possible nonspecific effects of repeated seizures on mitochondrial metabolism and catechol O‐methyltransferase (COMT) activity was determined to see if this extracellular degradative enzyme for NE was affected in the same way as MAO located intracellularly in the mitochondria.
THE INHIBITION OF PHENYLALANINE AND TYROSINE HYDROXYLASES BY HIGH OXYGEN LEVELS
Authors: Fisher D. B. , Kaufman S.
Published: October 2006

Abstract— The Km for oxygen for rat liver phenylalanine hydroxylase depended on the structure of the reduced pterin cofactor. When the synthetic cofactor, 6,7‐dimethyltetrahydropterin, was employed, the apparent Km for oxygen was 20%. When the natural cofactor, tetrahydrobiopterin, was used, the apparent Km for oxygen was 0.35 %. Substrate inhibition (40 per cent inhibition at 43% oxygen) was observed with the natural cofactor but not with the synthetic cofactor. Oxygen also caused substrate inhibition with bovine adrenal medulla and brain tyrosine hydroxylases. The inhibition was more dramatic in the presence of the natural cofactor than with the synthetic cofactor. Substrate inhibition by oxygen of brain tyrosine hydroxylase may explain the lowered brain levels of norepinephrine and dopamine observed after treatment of animals with hyperbaric oxygen.
FORMATION OF GABA FROM PUTRESCINE IN THE BRAIN OF FISH (SALMO IRIDEUS GIBB.)
Authors: Seiler N. , Al‐Therib M. J. , Kataoka K.
Published: October 2006

Abstract— It was demonstrated after intraperitoneal and intracerebral injections of [1,4‐14C]‐putrescine.2 HCl that GABA is formed in vivo in the trout brain via a pathway in which glutamic acid is not an intermediate. Intraperitoneal and intracerebral injections of both thiosemicarbazide and 3‐mercaptopropionic acid had no measurable effects on GABA concentration, transformation of glutamic acid into GABA in vivo, or on glutamate de‐carboxylase activity in the brain within the first 3 h after the application of the inhibitors. Only a small decrease in concentration of pyridoxal phosphate was noticed in the fish brain after thiosemicarbazide administration. The relatively high concentrations of pyridoxal phosphate in the trout brain may be one of the reasons for the ineffectiveness of thiosemicarbazide in inhibiting glutamate decarboxylase in vivo. After intracerebral injections of [1‐14C]GABA, a half‐life of 7 h was determined for GABA. The slow turnover rate of GABA in trout brain, which can be assumed from this observation, may give a further explanation of the ineffectiveness of the glutamate decarboxylase inhibitors in lowering the GABA content ot fish brain within a few hours.
GANGLIOSIDE COMPOSITION AND CONTENT OF RAT‐BRAIN SUBCELLULAR FRACTIONS
Authors: Avrova N. F. , Yu. Chenykaeva E. , Obukhova E. L.
Published: October 2006

Abstract—The composition and content of gangliosides from rat‐brain microsomal, synaptosomal, mitochondrial and myelin fractions were studied. Outer membranes of synaptosomes were also isolated, separated into subfractions and investigated. Of all the fractions studied the outer membranes of synaptosomes are richest in gangliosides, in one of their sub‐fractions the concentration of gangliosides per mg of protein is five times higher than in the homogenate. Microsomes are rich in gangliosides as well, but to a lesser degree, whereas the mitochondrial fraction contains considerably smaller amounts of gangliosides per mg of protein than does the homogenate. The ganglioside pattern of outer membranes of synaptosomes and of their subfractions is somewhat different from that of the homogenate; the outer membranes contain approximately one‐third less monosialogangliosides. On the contrary a very high content of monosialogangliosides is characteristic of the ganglioside pattern of the myelin fraction. In this fraction monosialoganglioside GMI(nomenclature of Svennerholm, 1963) constitutes 60–63 per cent of ganglioside sialic acid, or 75–80 molar per cent of gangliosides, the content of di‐ and trisialogangliosides being much lower than in other fractions. Fatty acid and long chain base composition of gangliosides from synaptosomal and microsomal fractions and homogenate is very similar, almost identical. In gangliosides from myelin fractions the relaitve content of palmitic and monoenoic acids is higher and that of arachinic acid and C20‐sphingosine—lower than in other fractions studied. The difference in ganglioside composition of synaptosomes and their outer membranes and on the other hand of myelin appears to reflect the difference in ganglioside composition of neuronal and oligodendroglial plasma membranes.
PURIFICATION AND SOME PROPERTIES OF CHOLINE ACETYLTRANSFERASE (EC 2.3.1.6) FROM BOVINE BRAIN
Authors: Chao L.‐P. , Wolfgram F.
Published: October 2006

Abstract— Choline acetyltransferase (ChAc) has been isolated and highly purified from the caudate nuclei of bovine brains. The procedure involved: (1) making an acetone‐ and chloroform‐insoluble powder from the tissue; (2) treating the powder with aqueous buffer and chromatographing the extractable soluble proteins on an organomercurial‐sepharose column; (3) removing impurities by passage through columns of DEAE‐cellulose and hydroxyapatite; and (4) separation of the heme‐containing protein from ChAc by denaturing the former with a mixture of chloroform and n‐butanol. The purified ChAc was essentially homogeneous as judged by polyacrylamide gel electrophoresis and exhibited a pH optimum at about pH 7. The partially purified ChAc dissociated into two non‐identical subunits when chromatographed with a dilute buffer on Bio‐gel A. It did not dissociate when a more concentrated buffer was used. The purified ChAc dissociated on the Bio‐gel A even in the presence of a high salt concentration. The dissociation was accompanied by a great loss of enzymatic activity, and we concluded that the presence of other proteins tends to prevent the dissociation of ChAc on gel filtration.
PROTEIN COMPOSITION OF MYELIN OF THE PERIPHERAL NERVOUS SYSTEM
Authors: Greenfield S. , Brostoff S. , Eylar E. H. , Morell P.
Published: October 2006

Abstract— Myelin was purified from the peripheral nervous system (PNS) of several species. The protein composition of these preparations was examined by discontinuous polyacrylamide gel electrophoresis in buffers containing sodium lauryl sulphate. Proteins characteristic of all samples include, in order of increasing mobility: a series of high molecular weight proteins, the major peripheral nerve protein (P0), two uncharacterized proteins, and two basic proteins (P1 and P2). Quantitative results, obtained by densitometry of gels stained with Fast Green showed differences in protein distribution, both between species, and from different types of nerves obtained from the same animal. The relative amounts of P1 and P2 proteins were the most variable; e.g. myelin from guinea‐pig sciatic nerve had little or no P2 protein, whereas 15 per cent of the myelin protein of beef posterior intradural root was Pz protein. P0, P1 and P2 proteins from rabbit sciatic nerve and P0 and P2 proteins from beef dorsal and ventral intradural roots were purified and their amino acid compositions were determined. Our results indicated that the P1 protein is very similar in size and amino acid composition to the basic protein of central nervous system myelin, whereas the P0 and P2 proteins are unique to the PNS.
HIGH‐AFFINITY TRANSPORT AND PHOSPHORYLATION OF 2‐DEOXY‐d‐GLUCOSE IN SYNAPTOSOMES1,2
Authors: Diamond I. , Fishman R. A.
Published: October 2006

2‐Deoxy‐d‐glucose (2 DG) entered synaptosomes (from rat brain) by a high‐affinity, Na+‐independent glucose transport system with a Km, of 0.24 mM. 3‐O‐methyl‐glucose, D‐glucose, and phloretin were competitive inhibitors of 2‐DG transport with Ki's of 7 mM, 64 μM, and 0·75 μM, respectively. Insulin was without effect. 2‐DG uptake was also saturable at high substrate concentrations with an apparent low affinity Km, of 75 mM, where the Kl, for glucose was 17.5 mM. We are not certain whether the rate‐limiting step for the low‐affinity uptake system is attributable to transport or phosphorylation. However, the high‐affinity glucose transport system probably is a special property of neuronal cell membranes and could be useful in helping to distinguish separated neurons from glial cells.
OPTIMAL FREEZING CONDITIONS FOR CEREBRAL METABOLITES IN RATS
Authors: Pontén U. , Ratcheson R. A. , Salford L. G. , Siesjö B. K.
Published: October 2006

Abstract— Optimal freezing conditions for metabolites were evaluated in 250‐450 g rats. As a standard procedure, the brains were frozen in such a way that the blood pressure and arterial oxygenation were upheld during the freezing. The progression of the freezing front was determined by means of implanted thermocouples, and the interruption of the circulation by means of injections of carbon particles into the blood stream. The freezing gave rise to a rapid interruption of the circulation in the superficial cortical layer first reached by the freezing front well before the temperature reached 0°C. In deeper regions the progression of the freezing front was slower and interruption of the circulation occurred simultaneously with the freezing of the tissue. Measurements of labile cerebral metabolites, including phosphocreatine, ATP, ADP, AMP and lactate, failed to show signs of autolysis in the part of cortex which became unperfused at temperatures above zero. Since the energy state was identical in superficial cortical areas and in areas that did not freeze until after 40–90 s, it is concluded that the freezing technique gives optimal conditions for metabolites also in deep cerebral structures.

Decapitation of unanaesthetized animals gave rise to large autolytic changes in the cerebral cortex. In unanaesthetized animals that were immersed in liquid nitrogen the changes were less marked and mainly affected the concentrations of phosphocreatine, ADP and lactate. When paralysed animals that were anaesthetized with N2O were immersed in liquid nitrogen the only significant change from the control was a decrease in phosphocreatine content. The virtual absence of autolytic changes in this group of animals was not related to the anaesthesia since more pronounced changes were observed in phenobarbitone‐anaesthetized rats immersed in the coolant. These differences could be explained by the fact that spontaneously breathing animals immersed in liquid nitrogen developed arterial hypoxia much faster than paralysed animals. It is concluded that an optimal metabolite pattern can only be obtained in anaesthetized animals, frozen with a method that was described by Kerr almost 40 years ago (Kerr, 1935). If unanaesthetized animals must be used, greater attention should be paid to the oxygenation of the blood during the freezing than to such factors as speed of freezing or depth of anaesthesia.
THE BIOSYNTHESIS AND CONCENTRATION OF GALACTOSYL DIGLYCERIDE IN GLIAL AND NEURONAL ENRICHED FRACTIONS OF ACTIVELY MYELINATING RAT BRAIN
Authors: Deshmukh D. S. , Flynn T. J. , Pieringer R. A.
Published: October 2006

—In continuation of our studies on the association of the galactosyl diglycerides of brain with myelination, we have measured the biosynthesis and concentration of these glyceride glycolipids, in oligodendroglial, astroglial, neuronal, and myelin enriched fractions from brains of rats of postnatal age 16, 19 and 29 days. The relative purity of cell fractions and myelin derived from 50 to 60 brains of each age‐group was checked by phase contrast microscopy and 2′,3′‐cyclic nucleotide‐3′‐phosphohydrolase activity. The relative purity was comparable to that reported by other investigators for cell fractions from bovine brain. Of the three cell types, the oligodendroglia had the highest and the neurons had the lowest capacity to enzymatically synthesize and to accumulate monogalactosyl diglyceride. The amount of monogalactosyl diglyceride found in myelin compared to that found in oligodendroglial fraction greatly increased during development between 16 and 29 days of age. The biosynthesis of galactosyl ceramide but not glucosyl ceramide was highest in oligodendroglial enriched cell fraction. However, ceramide glucosyl‐transferase activity, which was greatly affected by the method used for cellular separation, was highest in a microsomal fraction derived from grey matter. Our results support the contention that the oligodendroglial cells are the site of synthesis of myelin constituents of the central nervous system, and that there is a temporal relationship between this site of synthesis and the site of deposition (myelin).
ACTIVATION OF HEXOSE MONOPHOSPHATE PATHWAY IN BRAIN BY ELECTRICAL STIMULATION IN VITRO
Authors: Kimura H. , Naito K. , Nakagawa K. , Kuriyama K.
Published: October 2006

Abstract— The incubation of cerebral cortical slices for 15 min in Krebs‐Ringer‐tris (pH 7.6) solution at 37°C with [1‐14C]glucose or [6‐14C]glucose as substrates yielded a C‐1:C‐6 14CO2 ratio of 1.21, whereas this ratio increased to 3.01 after the application of electrical stimulation (ES). Tissue levels of 6‐phosphoglu‐conate (6PG) and glucose 6‐phosphate (G6P), intermediary metabolites of hexose monophosphate (HMP) pathway, were 7 and 180 nmol/g tissue following 15 min incubation, and increased by 33 and 45 per cent respectively following the application of ES. Activities of 6‐phosphogluconate dehydrogenase (6PGDH, 6‐phospho‐d‐gluconate: NADP+ 2‐oxidoreductase, EC 1.1.1.44) and glucose‐6‐phosphate dehydrogenase (G6PDH, d‐glucose‐6‐phosphate: NADP+ 1‐oxidoreductase, EC 1.1.1.49), important enzymes in regulating the activity of HMP pathway, in cerebral cortical slices were 689 and 907 pmol/mg protein/min and were increased by 66 and 25 per cent respectively by the application of ES. Synaptosomal G6PDH and 6PGDH activities were maximally activated by the addition of 40 mm‐Na+ to the reaction mixture, whereas no activation by Na+ was observed in microsomal G6PDH and 6PGDH. Amobarbital inhibited more strongly the Embden–Meyerhof (EM) pathway than the HMP pathway, while imipramine had a stronger inhibitory effect on HMP pathway than on EM pathway in the electrically stimulated cerebral tissues.

The present results indicate that the HMP shunt pathway in the cerebral cortex is activated by the application of ES in vitro, possibly at synaptic regions and may play an important metabolic and functional role in the brain.
OESTROGEN EFFECTS ON BRAIN AND PITUITARY ENZYME ACTIVITIES
Authors: Luine Victoria N. , Khylchevskaya R. I. , McEwen B. S.
Published: October 2006

Abstract— Ovariectomized female rats were treated daily with oestradiol‐17β benzoate for intervals up to one week and enzyme activities were measured in the pituitary and various brain regions. Brain regions were selected for study on the basis of their previously demonstrated content of putative oestradiol receptor sites. (1) Pituitary showed oestrogen‐dependent increases in glucose‐6‐phosphate dehydrogenase (G6PDH), 6‐phosphogluconate dehydrogenase (6PGDH) and lactic dehydrogenase (LDH), and no change in NADP+‐dependent isocitric dehydrogenase (ICDH), NADP+‐dependent malic dehydrogenase (MDH) or hexokinase (HK). MDH and ICDH were elevated in whole hypothalamus. Enzyme activities did not change significantly in whole amygdala, cerebral cortex, or hippocampus. (2) Sub‐regions of the preoptic area, hypothalamus and amygdala were dissected to obtain more highly concentrated populations of cells containing putative oestrogen receptor sites. In the basomedial sub‐region of hypothalamus, activities of MDH, ICDH and G6PDH were elevated by oestrogen treatment. In the corticomedial sub‐region of amygdala, MDH and ICDH were elevated by oestrogen treatment. No change was observed in any of the six enzymes in medial preoptic area. (3) Increases in enzyme activities were related to the total in vivo dose of oestradiol benzoate given. (4) Hypophysectomy or adrenalectomy did not prevent the enzymatic responses to oestrogen. (S) Oestrogen added directly to the enzyme incubation medium did not change enzyme activities. (6) Weight loss in ovariectomized rats due to reduced food intake did not increase enzyme activities. (7) In the pituitary, good correlation was obtained between the known receptor binding properties of various oestrogenic and non‐oestrogenic steroids and the elevation in G6PDH activity. The results indicate that oestradiol acts directly to cause changes in activities of some brain and pituitary enzymes. The possibility is discussed that these changes may result from oestrogen interaction with putative receptor sites found in pituitary and certain brain regions.
ISOLATION AND PARTIAL CHARACTERIZATION OF RAT BRAIN SYNAPTIC PLASMA MEMBRANES1
Authors: Gurd J. W. , Jones L. R. , Mahler H. R. , Moore W. J.
Published: October 2006

Abstract— Synaptic plasma membranes from the cortices of adult rat brain were isolated from synaptosomes prepared by flotation of a washed mitochondrial pellet (P2) in a discontinuous Ficoll‐sucrose gradient. Contamination of the synaptosome fraction by microsomes was estimated by enzymic and chemical analysis to be less than 15 per cent. (2) The purified synaptosome fraction was subjected to osmotic shock, subfractionated on a discontinuous sucrose gradient and the distribution of enzymic and chemical markers for synaptic plasma membranes, microsomal membranes and mitochondria was determined. (3) Comparison of synaptosome subfractions prepared in the presence and absence of 1 mM NaH2 PO4/0.1 mM EDTA buffer pH 7.5, indicated that the ionic composition of the isolation medium markedly affected the distribution and enzymic composition of the subfractions. (4) Synaptic plasma membranes prepared in the presence of PO4/EDTA exhibited a 10‐fold enrichment in [Na++ K+] ATPase and were characterized by less than 15 and 10 per cent contamination by microsomes and mitochondria respectively. (5) The polypeptide composition of the purified synaptic plasma membranes was compared with the microsomes and mitochondria by polyacrylamide gel electrophoresis in sodium dodecyl sulphate. No differences between the protein and glycoprotein composition of the synaptic plasma membranes and microsomes were detected. The mitochondria, in contrast, possessed a unique protein composition.
THE RELATIONSHIP BETWEEN NORADRENALINE TURNOVER IN CEREBRAL CORTEX AND ELECTRICAL SELF‐STIMULATION THROUGH ELECTRODES IN THE REGION OF LOCUS COERULEUS
Authors: Anlezark G. M. , Walter D. S. , Arbuthnott G. W. , Crow T. J. , Eccleston D.
Published: October 2006

—Rats allowed to stimulate themselves through electrodes implanted in the region of the noradrenaline‐containing cells of the locus coeruleus had increased concentrations in the cerebral cortex of 4‐hydroxy‐3‐methoxy‐phenylglycol (a noradrenaline metabolite). In a group of animals stimulated under anaesthesia a rise was present if, and only if, the electrodes had previously supported self‐stimulation behaviour. It is concluded that the increase in cortical 4‐hydroxy‐3‐methoxy‐phenylglycol reflects an increase in noradrenaline turnover which results from self‐stimulation through electrodes in the region of the locus coeruleus. The results constitute further evidence for the involvement of a specific catecholamine‐containing pathway, the coerulo‐cortical noradrenergic system, in central reward mechanisms.
d‐ASPARTATE OXIDASE ACTIVITY IN EXTRACTS OF MAMMALIAN CENTRAL NERVOUS TISSUE
Authors: Daves L. P. , Johnston G. A. R.
Published: October 2006

Abstract— d‐Aspartate oxidase activity has been measured in water extracts of acetone powders prepared from cat forebrain, cerebellum and spinal cord, rat brain, hog brain and sheep brain stem, and compared with that found in rabbit and cat kidney. The results suggest that the brain enzyme has very similar properties to the n‐aspartate oxidase (d‐aspartate: oxygen oxidorcductase (deaminating), EC 1.4.3.1) of kidney. Crude extracts (ammonium sulphate fractions of water extracts of acetone powders) displayed little activity without added FAD. FMN could not replace FAD. With oxygen as electron acceptor, the enzyme oxidized d‐aspartate much more rapidly than d‐glutamate, and displayed quite high activities with N‐substituted derivatives of d‐aspartate as substrates. Those amino acids susceptible to oxidation by d‐amino acid oxidase were not oxidized by the d‐aspartate oxidase. The regional distribution of the d‐aspartate oxidase activity within the CNS differed from that of d‐amino acid oxidase. As has been previously observed for kidney d‐aspartate oxidase activity, dicarboxylic acids competitively inhibited this enzymic activity in brain extracts, while sodium benzoate and sodium barbitone, inhibitors of d‐amino acid oxidase, were without effect.
MULTIPLE BINDING SITES OF HUMAN BRAIN MONOAMINE OXIDASE AS INDICATED BY SUBSTRATE COMPETITION
Authors: White Helen L. , Wu Joyce Chen
Published: October 2006

Abstract—Six endogenous substrates of monoamine oxidase (EC 1.4.3.4) (serotonin,l‐norepinephrine, dopamine, tyramine, tryptamine andβ‐phenethylamine) were used separately and in pairs with human brain mitochondrial extracts. ApparentK1values were obtained from experiments in which only 1 of 2 substrates was isotopically labelled, and these values were compared with experimentalKmvalues.β‐Phenethylamine appears to be metabolized at enzyme active sites independent from those which bind serotonin. The substratel‐norepinephrine competes with serotonin for an enzyme site, but also may be catalysed at an additional site which is independent of serotonin binding. Experiments in which [14C]tryptamine was combined with [3E]serotonin indicated that tryptamine is a much more potent inhibitor of serotonin oxidation than was predicted fromKmvalues. It is suggested that the competition among substrates of MA0 which is observedin uitromay have relevance toin uiuomechanisms for control of biogenic amine concentrations.
2′,3′‐CYCLIC NADP AS A SUBSTRATE FOR 2′,3′‐CYCLIC NUCLEOTIDE 3′‐PHOSPHOHYDROLASE
Authors: Sogin D. C.
Published: October 2006

Abstract– 2′,3′‐Cyclic NADP has been prepared by cyclization of NADP at pH 6 in the presence of l‐ethyl‐(3‐dimethylaminopropyl)‐carbodiimide. The NADP derivative is readily hydrolyzed to NADP by the enzyme in brain and nerve that hydrolyzes 2′,3′‐cyclic nucleotides to 2′‐phospho esters. The Km for this substrate is the same as that for 2′,3′‐cyclic AMP (0.22 mm) at pH 6 and 25°C. The two substrates are hydrolyzed by the phosphohydrolase at similar maximum velocities. The nicotinamide moiety in cyclic NADP thus has little effect on the enzyme‐substrate interaction. This synthetic substrate can be used in a rapid (2 min) and sensitive (10 ng of 31‐fold purified enzyme) spectrophotometric coupled enzyme assay for 2′,3′‐cyclic nucleotide 3′‐phosphohydrolase; in this assay the hydrolysis proceeds in the presence of glucose‐6‐phosphate dehydrogenase and its substrate and the NADPH formed is measured by the increase in absorbance at 340 nm. The assay is applicable to tissue extracts as well as to purified preparations of the enzyme. There is no interference from nucleases of the pancreatic RNase A type.
SELENIUM AND GLUTATHIONE PEROXIDASE IN DEVELOPING RAT BRAIN1
Authors: Prohaska J. R. , Ganther H. E.
Published: October 2006

Abstract– Week‐old rats were given a subcutaneous injection of carrier‐free Na275SeO3 and brain 75Se distribution was studied after 30 days, with special reference to the selenoprotein, glutathione peroxidase (GSH‐Px). Chemical fractionation studies showed the 75Se was associated mainly with protein and not extracted by hot trichloroacetic acid or chloroform‐methanol. Subcellular fractions also revealed a parallel distribution of 75Se and protein with the notable exception that 75Se was concentrated in the mitochondria and reduced in the cytosol. GSH‐Px activity was demonstrated in the isolated mitochondrial fraction. The estimated biological half‐life of brain 75Se was 45 days. Gel filtration (Sephadex G‐150) of brain cytosol resulted in four 75Se peaks: peak 1 was associated with the void volume, and had the greatest 75Se content; peak 2 (Ve/Vo= 1.4) contained nearly as much 75Se and had an apparent molecular weight of 94,000; peak 3 (Ve/V0= 2.4) had an apparent molecular weight of 13,500 and was markedly increased when brain was homogenized in the presence of Triton X‐100; peak 4 consisted of low molecular weight compounds. When fresh cytosol (with or without Triton X‐100) was chromatographed on Sephadex G‐150, GSH‐Px was detectable only in the void volume; however, storage of cytosol prepared in the presence of Triton X‐100 shifted most of the activity to peak 2 (94,000). The GSH‐Px activity in the void volume resembled the purified enzyme with regard to pH dependence, Km for cumene hydroperoxide at fixed [GSH], and first‐order kinetic behavior with respect to GSH. A minor peak of GSH‐Px activity showing zero‐order kinetics with respect to GSH concentration and an apparent molecular weight of 46,000 was detected when larger amounts of protein were chromatographed. The concentration of rat brain Se measured by chemical analysis reached adult levels by 2 weeks after birth, an age when the level of GSH‐Px had just begun to rise. It was estimated that only 1/5 of the total brain Se may be accounted for by its presence in GSH‐Px, suggesting that the function of the majority of brain Se remains to be determined.
ELEVATED EXTRACELLULAR POTASSIUM AS A STIMULUS FOR RELEASING [3H]NOREPINEPHRINE AND [14C]α‐AMINOISOBUTYRATE FROM NEOCORTICAL SLICES. SPECIFICITY AND CALCIUM DEPENDENCY OF THE PROCESS
Authors: Vargas Olga , Orrego F.
Published: October 2006

—The effect of short (4–6 min)‘pulses’ of elevated extracellular potassium ions K0, in the 10–50 mm range, on the efflux of [3H]norepinephrine [3H]NE) and [14C]α‐aminoisobutyrate (AIB) has been studied in a superfused neocortical thin slice system. At all the concentrations tested high K0 increases the efflux of both NE and AIB, although thc effects on the former are greater. In the absence of calcium ions, or in the presence of 8 mm‐MnCl2, the potassium‐stimulated release of both NE and AIB is severely depressed. However, potassium induced NE release is proportional to extracellular calcium ions in the 0–1.5 mm range, while that of AIB does not continue to increase above 0.2 mm‐calcium. This permissive role of calcium in amino acid efflux is interpreted as due to changes in the inactivation of membrane sodium conductance.
NEONATAL ASPHYXIA IN THE RAT: GREATER VULNERABILITY OF MALES AND PERSISTENT EFFECTS ON BRAIN MONOAMINE SYNTHESIS
Authors: Simon Nicole , Volicer L.
Published: October 2006

Abstract—

In the rat, neonatal asphyxia produced by suffocation did not leave permanent visible lesions in thc brain, nor did it result in permanent motor impairment, although a delay in the development of some reflexes was observed. A transient retardation of body and brain growth, which was more pronounced in males, was found. By 5‐6 weeks of age, body and brain weights of asphyxiated rats were no longer significantly different from control animals. However, an increase in brain norepinephrine synthesis was found to persist after maturation. An alteration of serotonin metabolism was found after maturation only in asphyxiated males. The possibility that neonatal asphyxia in the rat is a model for abnormal development of monoamine metabolism, relevant to early childhood behavior disorders such as infantile autism or the syndrome of minimal brain dysfunction, is discussed.
QUANTITATION OF ACTIN IN DEVELOPING BRAIN1
Authors: Pardee J. D. , Bamburg J. R.
Published: October 2006

Actin levels in the soluble fraction of chick embryo brain sonicates have been quantitated by polyacrylamide gel electrophoresis. Brain actin concentrations remained constant at 8.10 ± 0.9% in developing cnibryos but dropped to 6.0 ± 0.9% in adult brain. Over 80% of the total brain protein was solubilized by the sonication procedure and actin was not detected in the remaining insoluble fraction.
N‐ACETYLNEURAMINIC ACID MOLECULES AS POSSIBLE SEROTONIN BINDING SITES1
Authors: OCHOA E. L. M. , BANGHAM A. D.
Published: October 2006

5‐Hydroxytryptamine (5‐HT), but not acetylcholine, carbamylcholine or L‐D‐noradrenaline, binds to ox brain ganglioside micelles, to phosphatidylcholine smectic mesophases (liposomes) containing gangliosides and to the glycoprotein fetuin, through the negatively charged N‐acctylneuraminic acid (NeuNAc) residues. The 5‐HT binding to NeuNAc is reversible, saturable, prodeeds in a 1: 1 fashion and can be specifically blocked by 7‐methyltryptamine. Thc affinity constant at equilibrium for the reaction is of the order of 102 1. mol‐1. No special ganglioside was identified as specifically associating with the amine. A terminal NeuNAc in the gangliosides is not a prerequisite for binding, although it seems important for binding 5‐HT in entire mcmbrane preparations (MARCHBANKS, 1966) or for bringing about the 5‐HT induced contraction of smooth muscle cells (WOOLLCY & GOMML 1964a). It is proposed that in 5‐HT target cells, NeuNAc residues, probably attached to membrane surface glycoprotein(s) are involved in thc mechanism of action of the drug.
DECREASE OF HOMOVANILLIC, DIHYDROXYPHENYLACETIC ACID AND CYCLIC‐ADENOSINE‐3′,5′‐MONOPHOSPHATE CONTENT IN THE RAT CAUDATE NUCLEUS INDUCED BY THE ACUTE ADMINISTRATION OF AN AMINOACID MIXTURE LACKING TYROSINE AND PHENYLALANINE1
Authors: Biggio. G. , Porceddu M. L. , Gessa G. L.
Published: October 2006

The oral administration of an aminoacid mixture lacking tyrosine and phenylalanine induces, in rats, a profound depletion of tyrosine in serum and in brain. Brain tyrosine is maximally depleted by 73%, 2 h after treatment, when there is a concomitant decrease in the levels of HVA (by 50%), DOPAC (by 30%) and c‐AMP (by 28%) in the basal ganglia. However, 4 to 8 h after treatment, when brain tyrosine is still depleted by 47 and 28%, respectively, DA metabolites and c‐AMP levels have returned to normal. Our findings indicate that striatal tyrosine hydroxylase is fully saturated in v i m by the concentrations of tyrosine normally present in the basal ganglia. The results also suggest indirectly that decreased DA turnover results in decreased nerve activity, as judged by the decreased cyclic AMP levels.
CHARACTERIZATION OF AN α‐BUNGAROTOXIN BINDING COMPONENT FROM DROSOPHILA MELANOGASTER
Authors: Schmidt‐Nielsen B. K. , Gepner J. I. , Teng N. N. H. , Hall L. M.
Published: October 2006

Abstract— We describe an α‐bungarotoxin binding component from Dromphila melanoyaster that has the properties expected of an acetylcholine receptor. Toxin binding to a paniculate form of this component has been shown to be proportional to amount of extract, to be saturable and to be destroyed by heat. Localization studies using 125I‐α‐bungarotoxin binding to frozen sections has shown toxin binding to be restricted to synaptic areas of the Drosophila CNS. We have also shown that this toxinbinding component can be treated with Triton X‐100 without significantly altering its toxin‐binding and pharmacological specificity. The ability of preincubation with cholinergic ligands to block labeled α‐bungarotoxin binding to both particulate and detergent treated extracts has been studied. The nicotinic agents nicotine, d‐tubocurarine, and acetylcholine are the most effective blocking agents. All of the muscarinic agents tested and the nicotinic agent decamethonium were less effective than acetylcholine in preventing α‐bungarotoxin binding.
LOCALIZATION OF HEXOKINASE IN NEURAL TISSUE: LIGHT MICROSCOPIC STUDIES WITH IMMUNOFLUORESCENCE AND HISTOCHEMICAL PROCEDURES
Authors: Wilkin G. P. , Wilson J. E.
Published: October 2006

Abstract— The distribution of hexokinase (ATP: D‐hexose 6‐phosphotransferase, EC 2.7.1.1) in rat cerebellum, retina, hippocampus, choroid plexus and ependymal cells of the cerebral ventricles, and dorsal root ganglion has been determined, at the light microscopic level, by both immunofluorescence and a histochemical procedure using nitro blue tetrazolium. With the exception of an artifactual staining of the outer photoreceptor segments of retina when the histochemical procedure was used, both methods gave comparable results, from which the following conclusions are drawn:

The cytoplasm of neuronal cell bodies clearly contained hexokinase, although the relative levels varied markedly among different types of neurons; such variations have previously been detected by direct assay of hexokinase in dissected neuronal cell bodies (Kato & Lowry, 1973a).

Glial cells contained readily detectable levels of hexokinase: the immunofluorescence technique revealed spidery glial processes within the myelinated tracts; in other areas, glial cell cytoplasms were indistinguishable from surrounding neuropil, indicating comparable levels of hexokinase; the satellite glia of dorsal root ganglia actually contained higher levels than did adjacent large neurons. The present results, therefore, do not support previous suggestions that glia are characteristically low and neurons characteristically high in hexokinase content.

Hexokinase was distributed throughout neuropil areas, with a somewhat speckled appearance suggesting the existence of small localizations of relatively higher activity, the nature of which could not be determined at this level of resolution; the hexokinase level in neuropil was clearly higher than that of white fiber tracts, in agreement with previous direct biochemical measurements (Buellet al., 1958)

No detectable levels of hexokinase were found in cell nuclei.

Regions expected to be rich in nerve terminals (e.g. the cerebellar glomeruli, the plexiform layers of retina) showed relatively high hexokinase levels compared to the cytoplasm of adjacent neuronal perikarya, in agreement with previous subcellular fractionation experiments which indicated relatively high levels of hexokinase in nerve endings (Wilson, 1972). Considered along with the‘high affinity’glucose transport system in nerve endings (Diamond & Fishman, 1973), these results suggest nerve terminals are well adapted for the'efficient acquisition and introduction of glucose into metabolism.

In addition, high levels of hexokinase were observed in the inner photoreceptor segments of retina, and in the ependymal and choroid plexus cells of the ventricles.
THE BINDING OF 5‐HYDROXYTRYPTAMINE TO ACIDIC LIPIDS IN ISOBUTANOL1
Authors: Johnson David A. , Cho Tae Mook , Loh Horace H.
Published: October 2006

Abstract— In order to examine the possibility that acidic lipids can account for the binding of 5‐hydroxy [3H]tryptamine (5‐HT) to brain tissue, the binding to six acidic lipids was studied using an isobutanol‐water partition method. With the exception of the polyphosphoinositides, all the acidic lipids examined bind saturably and with high affinity. The apparent dissociation constants of 5‐HT to the acidic lipids were as follows: phosphatidylserine, 0.4 μM; phosphatidic acid, 0.6μM; diphosphoinositide, 0.8 μM; cerebroside sulfate, 1.4 μM; monophosphoinositide, 1.9 μM; and triphosphoinositide, 10 μM. The high affinity of these lipids to 5‐HT raises the possibility of some role for them in serotonergic activity.
THE DISTRIBUTION OF ACETYL‐CoA IN SPECIFIC AREAS OF THE CNS OF THE RAT AS MEASURED BY A MODIFICATION OF A RADIO‐ENZYMATIC ASSAY FOR ACETYLCHOLINE AND CHOLINE1
Authors: Shea P. A. , Aprison M. H.
Published: October 2006

Abstract—

A rapid and sensitive enzymatic assay for measuring picomole quantities of acetyl‐CoA, acetylcholine (ACh), and choline from the same tissue extract has been developed. After ACh and choline were extracted into 15% 1 N formic acid/85% acetone, the pellet was further extracted with 5% trichloroacetic acid (TCA) to remove the remaining acetyl‐CoA. The two extraction solvents were pooled and lipids, organic solvents, and TCA were removed first by a heptane‐chloroform wash followed by an ether extraction. In the acetyl‐CoA assay, endogenous ACh and choline were removed by extractions with sodium tetraphenylboron in butenenitrile prior to the enzymatic reactions. The acetyl‐CoA remaining in the aqueous phase was then converted enzymatically to labelled ACh in the presence of [Me‐14C]choline using choline acetyltransferase. The unreacted labelled precursor was converted to choline phosphate by the enzyme choline kinase. The [14C]ACh formed from acetyl‐CoA was extracted into sodium tetraphenylboron in butenenitrile and a portion of the organic phase containing the [14C]ACh was counted in a scintillation counter. Acetylcholine and choline were assayed from the same tissue extracts by a modification of the procedure by SHEA & APRISON (1973). Acetyl‐CoA levels in rat whole brain when killed by the near‐freezing procedure were found to be 5.50 ± 0.2 nmol/g. The content of acetyl‐CoA was the same whether the rats were killed by the near‐freezing method or by total freezing in liquid nitrogen. The levels of acetyl‐CoA did not change with time after death when the tissue was maintained at a temperature of −10°C. In the same tissue extracts from rat whole brain killed by the near‐freezing method, the content of ACh was 20.6 ± 0.7 nmol/g and choline 58.2 ± 1.2 nmol/g. Although reproducible, the level reported for choline is high when assayed under this condition. The content of choline however after total freezing was found to be 25.2 ± 2.0 nmol/g. The sensitivity (d. p. m. of sample twice blank) is 10 pmol for the acetyl‐CoA assay and 25 pmol for the ACh and choline assays. The regional distribution of these three compounds in the brain of rats as well as the content of acetyl‐CoA in heart, liver and kidney are presented.
CEREBRAL ENERGY METABOLISM DURING TRANSIENT ISCHEMIA AND RECOVERY IN THE GERBIL1
Authors: Levy D. E. , Duffy T. E.
Published: October 2006

Abstract—

Energy metabolism was studied in the cerebral cortex of gerbils during and following ischemia induced by 1 h of unilateral carotid artery occlusion. An aneurysm clip was applied to the right common carotid artery of 50‐70 g gerbils under brief halothane anesthesia, and the clip was removed 1 h later. Clinical state (gait, responsiveness, seizures) was evaluated during carotid occlusion, and 40% of the animals showed clinical evidence of stroke. Cortical energy stores (2 ATP + ADP + P‐creatine) were more than half depleted in the ipsilateral cortex of clinically‐affected gerbils, and glucose fell by 75%; lactate rose over 7‐fold in the same specimens. After release of the carotid clip, clinical state improved, and biochemical abnormalities partially resolved. However, even after 24 h, the concentration of ATP and the total pool of adenine nucleotides remained subnormal. Metabolic activity in the ischemic cortex, assessed as the utilization of high‐energy phosphates following decapitation, was normal after 1 h of recovery and decreased (‐50%) after 24 h but was increased by more than 50% after 4 h. Cerebral glucose utilization, evaluated from autoradiographs prepared after intravenous administration of 2‐[1‐14C]deoxyglucose, was also increased in the cortex, hippocampus, and thalamus after 4 h of recovery. This post‐ischemic hypermetabolism in tissue damaged by ischemia may identify a critical period for cell repair, when therapy could be decisive.
FORMATION OF GAMMA‐HYDROXYBUTYRATE IN BRAIN1
Authors: Anderson R. A. , Ritzmann R. F. , Tabakoff B.
Published: October 2006

Abstract— Gamma‐hydroxybutyric acid is a neuroactive compound which has been found to be a normal constituent of mammalian brain. The present report characterized enzymatic activity in brain forming gamma‐hydroxybutyrate (GHB) from succinic semialdehyde (SSA). When NADPH served as cofactor, whole brain homogenate was capable of forming nearly 300 nmol GHB/min/g brain when enzyme activity was measured at 37°C. GHB production was significantly less (50%) when NADH was the cofactor. A regional localization of these activities indicated that the cerebellum and septal area contained the highest capacity to form GHB in the presence of NADPH; intermediate to high activity was found in the cortex, medulla, superior colliculus and corpus striatum; low activity was present in the inferior colliculus, thalamus, pons, hippocampus, substantia nigra and hypothalamus. Activity in the presence of NADH was rather evenly distributed with the exceptions of the cerebellum and inferior colliculus, which contained high and low activity respectively. Both NADPH‐ and NADH‐dependent activities were found primarily in the cytosol. Pentobarbital inhibited enzyme activity and enzyme activity was differentiated from lactic dehydrogenase and alcohol dehydrogenase by use of specific inhibitors. In addition, mixed substrate experiments and kinetic analysis provided evidence for the presence of two reversible NADPH‐dependent enzymes capable of producing GHB from SSA.
THE [14C]DEOXYGLUCOSE METHOD FOR THE MEASUREMENT OF LOCAL CEREBRAL GLUCOSE UTILIZATION: THEORY, PROCEDURE, AND NORMAL VALUES IN THE CONSCIOUS AND ANESTHETIZED ALBINO RAT1
Authors: Sokoloff L. , Reivich M. , Kennedy C. , Rosiers M. H. Des , Patlak C. S. , Pettigrew K. D. , Sakurada O. , Shinohara M.
Published: October 2006

Abstract— A method has been developed for the simultaneous measurement of the rates of glucose consumption in the various structural and functional components of the brain in vivo. The method can be applied to most laboratory animals in the conscious state. It is based on the use of 2‐deoxy‐D‐[14C]glucose ([14C]DG) as a tracer for the exchange of glucose between plasma and brain and its phosphorylation by hexokinase in the tissues. [14C]DG is used because the label in its product, [14C]deoxyglucose‐6‐phosphate, is essentially trapped in the tissue over the time course of the measurement. A model has been designed based on the assumptions of a steady state for glucose consumption, a first order equilibration of the free [14C]DG pool in the tissue with the plasma level, and relative rates of phosphorylation of [14C]DG and glucose determined by their relative concentrations in the precursor pools and their respective kinetic constants for the hexokinase reaction. An operational equation based on this model has been derived in terms of determinable variables. A pulse of [14C]DG is administered intravenously and the arterial plasma [14C]DG and glucose concentrations monitored for a preset time between 30 and 45min. At the prescribed time, the head is removed and frozen in liquid N2‐chilled Freon XII, and the brain sectioned for autoradiography. Local tissue concentrations of [14C]DG are determined by quantitative autoradiography. Local cerebral glucose consumption is calculated by the equation on the basis of these measured values.

The method has been applied to normal albino rats in the conscious state and under thiopental anesthesia. The results demonstrate that the local rates of glucose consumption in the brain fall into two distinct distributions, one for gray matter and the other for white matter. In the conscious rat the values in the gray matter vary widely from structure to structure (54‐197 μmol/100 g/min) with the highest values in structures related to auditory function, e.g. medial geniculate body, superior olive, inferior colliculus, and auditory cortex. The values in white matter are more uniform (i.e. 33–40 μmo1/100 g/min) at levels approximately one‐fourth to one‐half those of gray matter. Heterogeneous rates of glucose consumption are frequently seen within specific structures, often revealing a pattern of cytoarchitecture. Thiopental anesthesia markedly depresses the rates of glucose utilization throughout the brain, particularly in gray matter, and metabolic rate throughout gray matter becomes more uniform at a lower level.