bluuebunny/crossref_metadata_2025 huggingface dataset:
CORRELATION OF DNA ACCUMULATION RATE WITH THYMIDYLATE SYNTHETASE AND THYMIDINE KINASE ACTIVITIES IN DEVELOPING RAT CEREBELLUM: EFFECT OF THYROXINE1
Authors: Clark B. R. , Weichsel M. E.
Published: October 2006

Abstract— Using the method of least squares, a logistic curve was fitted to the data points for DNA content in neonatal rat cerebellum versus postnatal age (day 0 is the day of birth). The resultant equation was differentiated to give an expression for the rate of cerebellar DNA accumulation in units of ng/h per mg wet cerebellum. The DNA accumulation rate in control rats increased from 77.0 at 2 days of age to a maximum of 108 at 7 days of age and declined thereafter to a minimum of 16.3 on day 15. Thyroxine treatment significantly (P < 0.05) increased the rate to 89.8 (117% of control) at 2 days of age, and a significant elevation was maintained to 6 days of age at which time a maximum rate of 115 (114% of control) was attained. The rate was significantly decreased below control at 9 and 12 days of age, and reached a minimum of 9.22 on day 15.

The developmental pattern for the activity of cerebellar thymidylate synthetase (EC 2.1.1.6), in units of pmol/h per mg wet cerebellum, closely paralleled the pattern for DNA accumulation rate in both control and thyroxine‐treated animals. In controls, thymidylate synthetase activity increased from 98.6 at 2 days of age to a maximum of 125 at 7 days of age and declined thereafter to a minimum of 30.0 at 15 days of age. In thyroxine‐treated animals, the activity was significantly increased to 118 (122% of control) at 4 days of age and remained significantly elevated through 6 days of age at which time a maximum activity of 154 (115% of control) was attained; thereafter, the activity was significantly decreased below control and reached a minimum of 16.9 (56.3% of control) on day 15. The developmental pattern for the activity of cerebellar thymidine kinase (EC 2.7.1.21) did not parallel the DNA accumulation rate quite so closely, in neither treated nor control animals, as did the pattern for thymidylate synthetase activity.

These data suggest that thymidylate synthetase activity in the developing rat cerebellum may be more important for maintenance of replicative DNA synthesis than is thymidine kinase activity. In addition, the thyroxine‐induced acceleration of the increase and subsequent decline in rate of DNA accumulation and in the activities of thymidylate synthetase and thymidine kinase in developing rat cerebella is probably the result of alterations in the number of external granular cells undergoing replicative DNA synthesis.
INTRACELLULAR AMINO ACID CONTENT OF NEURONAL, GLIAL, AND NON‐NEURAL CELL CULTURES: THE RELATIONSHIP TO GLUTAMIC ACID COMPARTMENTATION
Authors: Drummond R. J. , Phillips A. T.
Published: October 2006

Abstract— A protocol for the accurate determination of intracellular levels of amino acids in tissue cultured cells has been developed and used in the measurement of intracellular amino acids levels in neuronal, glial, and non‐neural cell lines, with the objective of establishing morphological correlates for large and small glutamic acid compartments and of examining hypotheses for the morphological basis of glutamic acid compartmentation. This survey of intracellular amino acid levels has revealed striking differences among the cell lines tested, but these differences did not correlate with cell type, i.e. neuronal vs glial, in contrast to earlier results (Rose, 1968) based on bulk separated neuronal and pial fractions from rat brain. Amino acid levels were found to be dependent upon tissue culture conditions, yet reproducible differences could be observed when growth and experimental conditions were carefully controlled. Glutamic acid levels for various cell lines ranged from 50.8 ± 14.3 to 158 ± 8.5 nmol/mg protein. Intracellular glutamine levels demonstrated even greater difference, with values ranging from 0.8 ± 0.2 to 107 ± 42.4 nmol/mg protein. Statistically significant differences in intracellular amino acid levels between cell lines were also observed for aspartic acid, praline, glycine, alanine, valine, cystathionine, isoleucine, and leucine. A number of cell lines demonstrating highly elevated elevated levels of γ‐aminobutyrate and β‐alanine were identified. The significance of neuronal and glial levels of glutamic acid, glutamine and γ‐aminobutyrate to models for glutamic acid compartmentation is discussed.
VITAMIN A RECEPTORS: RETINOL BINDING IN NEURAL RETINA AND PIGMENT EPITHELIUM
Authors: Wiggert B. , Bergsma D. R. , Lewis M. , Chader G. J.
Published: October 2006

Abstract— With sucrose density gradient analysis, bovine retinal cytosol demonstrates 2S and 7S retinol‐binding species; binding in pigment epithelial cytosol is predominantly to a 2S species. Binding of retinol to the 2S component in retina is unaffected by retinoic acid or retinyl palmitate whereas the ester effectively competes for 2S binding in pigment epithelium. Specific retinol binding can also be demonstrated by gel filtration on Sepharose 4B; no high molecular weight (> 100–200,000) retinol binding species are observed by this technique. Both the 2S and 7S binding species in retinal cytosol are protein in nature and differentially susceptible to proteolysis. The 2S and 7S species appear to be separate and distinct since chaotropic agents such as sodium thiocyanate or KCl and CaCl2 do not seem to convert the 7S species into 2S subunits. Scatchard plot analysis indicates high affinity retinol binding to the 2S receptor. Computer analysis of the binding data yields Ka=3 × 108, n = 1, a molecular size of 16,200 and ΔG0=−9.5 kcal/mol.
SIALYLTRANSFERASES IN RAT BRAIN: INTRACELLULAR LOCALIZATION AND SOME MEMBRANE PROPERTIES
Authors: Ng S.‐S. , Dain J. A.
Published: October 2006

Abstract— Total rat cerebral homogenate, with nuclei removed, yielded sialyltransferase activity peaks that were distinct from the protein distribution profile in a continuous sucrose density gradient. Marker enzyme studies and electron microscopic examinations on the gradient fractions suggested that most of the sialyltransferase activities were not associated with the synaptosomes.

The sialyltransferases appeared to be localized in the smooth microsomal membranes and the Golgi complex derivatives. The sialyltransferase activities were stimulated by non‐ionic detergent mixture, Triton CF‐54/Tween 80 (2/1, w/w), the effect being much more pronounced with exogenous substrates. The stimulatory effect was dependent on detergent concentration. With 1 mg detergent mixture per mg enzyme protein, the percent increases in enzyme activities with the different substrates were: endogenous glycolipids, 100; endogenous glycoproteins, 50; exogenous GM1a, 700; exogenous DS‐fetuin, 230. The action of the nonionic detergents appears to be on a hydrophobic segment of the enzyme molecule, bearing the active site, which is buried in the membrane lipid bilayer. This was substantiated by the partial trypsin resistance of the sialyltransferase activities and the abolition of that resistance when trypsiniza‐tion was performed in the presence of nonionic detergents. Furthermore, the sialyltransferase activities were markedly inhibited by organic solvents; and these inhibitory effects were inversely proportional to the solvent dielectric constants.
STUDIES ON THE TISSUE AND SUBCELLULAR DISTRIBUTION OF β‐N‐OXALYL‐l‐α, β‐DIAMINOPROPIONIC ACID, THE LATHYRUS SATIVUS NEUROTOXIN
Authors: Lakshmanan J. , Padmanaban G.
Published: October 2006

Abstract— β‐N‐Oxalyl‐l‐α, β‐diaminopropionic acid (ODAP), the Lathyrus sativus neurotoxin can be detected in significant concentrations in the synaptosomal fractions isolated from young rat brain and adult monkey spinal cord, when these animals manifest neurological symptoms after ODAP administration. However, isolated synaptosomes fail to exhibit any transport system for ODAP uptake. ODAP administered in vivo appears to get localized in a population of synaptosomes which exhibit a high affinity uptake system for glutamate.
ONTOGENESIS AND REGIONAL DISTRIBUTION OF HISTAMINE AND HISTAMINE‐N‐METHYLTRANSFERASE IN THE GUINEA PIG BRAIN1
Authors: TUOMISTO LEENA
Published: October 2006

Abstract—

The ontogenetic development and the regional distribution of histamine (HA) and histamine‐N‐methyltransferase (HMT, EC 2.1.1.8) in guinea pig brain and pituitary gland were studied. The samples were taken every fourth day beginning on the 28th foetal day. The HA concentration in the brains of the youngest foetuses was almost undetectable. A significant increase in HA concentrations occurred between days 40 and 44, which coincides with the period of rapid growth of nerve cell processes in this species. After this, a steep increase continued to the end of gestation in the hypothalamus, and to a lesser degree in the medulla‐midbrain and in the forebrain. In all parts of the brain the adult HA levels were reached by the time of birth. The HMT activity increased 15‐fold from the 28th foetal day to the adult and reached ca. 80% of the adult activity by the time of birth. The HMT activity developed earlier in the midbrain than in the forebrain or in the cerebellum, but after the birth the regional distribution of HMT was fairly even. In the pituitary gland the HA concentration and HMT activity increased hundredfold and tenfold, respectively.

The developmental patterns of HA and HMT in the guinea pig brain give support to the concept that HA might be a neurotransmitter in the brain.
5‐HYDROXYTRYPTAMINE CATABOLISM IN THE RAT BRAIN DURING ONTOGENESIS
Authors: BOURGOIN S. , ARTAUD F. , ADRIEN J. , HERY F. , GLOWINSKI J. , HAMON M.
Published: October 2006

Abstract—

Although the serotoninergic innervation is immature in the brains of young rats, the 5‐HIAA content is similar to that found in adults. As indicated by the ratio of 5‐HIAA to 5‐HT levels in the brain stem and the forebrain, the catabolism of the indolamine was more rapid during the first 3 postnatal weeks than in adults. This was contirmed by measuring the total formation of [3H]5‐HIAA from [3H]5‐HT newly synthesized from L‐[3H]tryptophan in brain stem slices of young and adult rats.

Electrolytic lesions of midbrain raphe nuclei (B7 and B8) performed on the 5th postnatal day resulted in parallel decreases in brain 5‐HT and 5‐HIAA levels; this ruled out the possibility that 5‐HIAA might be formed from 5‐HT synthesized outside serotoninergic neurons, using peripheral 5‐hydroxytryptophan. Inhibition of 5‐HT storage by reserpine pretreatment did not alter the higher capacity of newborn tissues to catabolize exogenous [3H]5‐HT. Therefore, possible differences in 5‐HT binding in serotoninergic neurons between newborn and adult rats were not likely to account for the differences in 5‐HT catabolism. Estimation of the rate of 5‐HIAA efflux from the brain after MAO inhibition did not reveal marked changes with age.

The activity of MAO type A, the enzyme involved in 5‐HT catabolism, was higher during early life than later on. This could be shown by using 5‐HT as substrate and clorgyline as a selective inhibitor. An opposite pattern of development was seen for MAO B, measured with benzylamine as substrate and deprenyl as selective inhibitor.

These results suggest that the high 5‐HIAA levels found in the brains of young rats can be attributed mainly to the presence of high MAO A activity during early life.
RAPID CONCURRENT AUTOMATED FLUORIMETRIC ASSAY OF NORADRENALINE, DOPAMINE, 3, 4‐DIHYDROXYPHENYLACETIC ACID, HOMOVANILLIC ACID AND 3‐METHOXYTYRAMINE IN MILLIGRAM AMOUNTS OF NERVOUS TISSUE AFTER ISOLATION ON SEPHADEX G10
Authors: WESTERINK B. H. C. , KORF J.
Published: October 2006

Abstract—

A semi‐automated fluorimetric assay technique for the concurrent estimations of noradrenaline, dopamine, 3, 4‐dihydroxyphenylacetic acid, homovanillic acid and 3‐methoxytyramine is described. The method is based on a rapid manually performed isolation of the catecholamines and metabolites on small columns of Sephadex G10, followed by automated fluorimetric dection in continuous flow systems. Samples containing no more that 2‐5 ng (NA, DA, DOPAC or HVA) or 10 ng 3‐MT could be reproducibly measured. The small Sephadex G10 columns has proven to be an excellent alternative to the currently used columns of alumina or ion‐exchange resins.
SYNTHESIS OF ETHANOLAMINE PHOSPHOGLYCERIDES BY MICROSOMES FROM THE BRAINS OF JIMPY AND QUAKING MICE
Authors: Dorman R. V. , Freysz L. , Horrocks L. A.
Published: October 2006

Abstract— —Brains of jimpy and quaking mice are known to be deficient in myelin and alkenylacyl‐glycero‐phosphorylethanolamines (alkenylacyl‐GPE, ethanolamine plasmalogens). Ethanolamine plasmalogen synthetic activity appeared to be normal and ethanolamine phosphotransferase (EC 2.7.8.1) activities are higher in the brain microsomes from jimpy and quaking mice than in their littermate controls when the activities are assayed with alkylacylglycerols and CDP[14C]ethanolamine. When endogenous diradylglycerols were the substrate, the rate of synthesis of diacyl‐GPE was normal but the rate of synthesis of the ether lipids, alkenylacyl‐GPE and alkylacyl‐GPE, was 33% and 8% below control levels for jimpy brain microsomes and quaking brain microsomes respectively. This difference is probably due to a normal content of diacylglycerols and a deficient content of alkylacylglycerols in the mutant brain microsomes. The apparent alkylacylglycerol deficiencies in the microsomes correspond with the ethanolamine plasmalogen deficiencies in the brains of these mutant mice.
EFFECTS OF POTASSIUM AND LITHIUM ON SODIUM TRANSPORT FROM BLOOD TO CEREBROSPINAL FLUID
Authors: Hesketh J. E.
Published: October 2006

Abstract— The relationship between isotopic sodium entry into cerebrospinal fluid (CSF) from blood and cisternal potassium concentration was studied using ventriculo‐cisternal perfusion in the rabbit. The entry of sodium into CSF was separated into 2 components. The fast component was significantly correlated with cisternal potassium concentration during perfusions with a potassium‐free artificial CSF. ATPase activity in the homogenised choroid plexus was shown to be sensitive to potassium over a range of concentrations similar to that in the perfusion studies. The results are interpreted as showing a potassium‐sensitive entry of 24Na across the choroid plexus due to a sodium‐pump situated in the apical membrane of the choroid plexus. The effects of low concentrations of lithium (0.6–1.2 mm) on 24Na entry into CSF and brain and on CSF secretion were studied. When applied via the ventricles lithium caused a 30–39% stimulation of the fast component of sodium entry and a 28% stimulation of CSF secretion. When given via the blood lithium inhibited the fast component of sodium entry and CSF secretion by 43% and 40% respectively. No effects of lithium were found on the slow component of sodium entry into CSF or sodium entry into brain. The results suggest that lithium at low (0.6–1.2 mM) concentrations can stimulate the choroid plexus sodium‐pump at the potassium‐sensitive side and inhibit it at the sodium‐sensitive side.
A SENSITIVE ENZYMATIC‐RADIOISOTOPIC ASSAY FOR 3,4‐DIHYDROXYPHENYLACETIC ACID
Authors: Kebabian J. W. , Saavedra J. M. , Axelrod J.
Published: October 2006

Abstract— An assay for 3,4‐dihydroxyphenylacetic acid (DOPAC) capable of detecting as little as 1 pmole of DOPAC is described. The basis of the assay is the O‐methylation of DOPAC utilizing S‐[methyl‐3H]adenosyl‐l‐methionine and a partially purified catechol‐O‐methyl transferase to form [methyl‐3H]homovanillic acid. The [methyl‐3H]homovanillic acid is purified with ion exchange resin and either solvent partitions or TLC. Utilizing this assay, the effect of either chlorpromazine or reserpine upon the content of DOPAC both in the caudate nucleus and in the substantia nigra was determined. Both of these drugs increase the level of DOPAC in the caudate nucleus but have minimal effects upon the level of DOPAC in the substantia nigra.
ACTIVATION OF GLYCOGEN PHOSPHORYLASE IN RAT CAUDATE NUCLEUS SLICES BY l‐ISOPROPYLNOREPINEPHERINE AND DIBUTYRYL CYCLIC AMP
Authors: Wilkening Doug , Makman Maynard H.
Published: October 2006

Abstract— l‐Isopropylnorepinepherine (IPNE), 3‐isobutyl‐1‐methylxanthine (IBMX) and N6,O2′‐dibutyryl cyclic AMP have been found to stimulate the conversion of glycogen phosphorylase (GPase) from b to a forms in rat caudate nucleus slices. The average percentage of total GPase in the a form in control incubations was 32%. The percentage of total GPase in the a form was increased to 1.5 times the control value in the presence of 1 mM‐IBMX, to twice the control value in the presence of 0.05 mM‐IPINE and to 2.5 times the control in the presence of 0.05 mM‐IPNE and 1 mM‐IBMX in combination. The increase in GPase activation correlated well with the elevation of cyclic AMP levels by these agents in caudate slices. The percentage of total GPase in the a form was also increased to 2.5 times the control by 1 mM dibutyryl cyclic AMP. Dopamine (DA) and 2‐chloroadenosine (2‐CI‐Ado), which also elevate cyclic AMP levels in rat caudate slices, were without significant effect on GPase. The results indicate that the β‐adrenergc adenylate cyclase in the rat caudate nucleus plays a role in the regulation of glycogen metabolism, while the DA‐stimulated adenylate cyclase is not significantly involved. 2‐CI‐Ado does have effects on glycogen metabolism in the caudate, but these effects do not appear to be mediated by GPase activation.
CHANGES IN PHOSPHOGLYCERIDE FATTY ACIDS OF RAT BRAIN INDUCED BY LINOLEIC AND LINOLENIC ACIDS AFTER PRE‐ AND POST‐NATAL FAT DEPRIVATION
Authors: Trapp B. D. , Bernsohn J.
Published: October 2006

Abstract— Pregnant rats were maintained on a fat‐free diet, starting at 10–12 days after impregnation and the offspring continued on the diet during a developmental period of 120 days. Brain fatty acids showed decreases in the level of (n‐3) and (n‐6) fatty acids of brain phophoglycerides, except for 22:5 (n‐6) which increased. These changes preceded an increase in the (n‐9) fatty acids (20:3 and 22:3). Supplementation with either linoleic or linolenic acid for 10 or 30 days after induction of the deficiency state caused an increase in the (n‐6) and (n‐3) fatty acids respectively, to control levels. The level of 22:5 (n‐6) was increased additionally by linoleic supplementation while linolenic refeeding to deficient animals decreased 22:5 (n‐6) to near control levels. The anomalous results obtained on 22:5 (n‐6) with 18:3 (n‐3) supplementation is attributed to competitive inhibition of linoleate desaturation by linolenate. Linoleic and linolenic acid were equally effective in reducing the elevated levels of the (n‐9) fatty acids toward control levels, although control levels with either fatty acid was not reached after 30 days supplementation. The increase of the (n‐6) and (n‐3) fatty acids to normal values precedes the decrease of (n‐9) fatty acids following supplementation of linoleic or linolenic acid to fat‐deficient rats. No change in fatty acid composition in control animals between 30 and 120 days was observed. In fat deficient as well as supplemented animals the total saturated, monounsaturated and polyunsatur‐ated fatty acid composition was constant as was the unsaturation index.
PHOSPHOPROTEIN PHOSPHATASE OF PINEAL GLAND: SOME PROPERTIES OF THE ENZYME AND THE IDENTIFICATION OF AN ENDOGENOUS ACTIVATOR
Authors: Yang H.‐Y. T. , Costa E. , Majane E. A. , Hong J.
Published: October 2006

Abstract— Both soluble and insoluble fractions of rat pineal glands catalyze the dephosphorylation of phosphohistone. The phosphoprotein phosphatase in cytosol as well as in insoluble fraction is inhibited by ZnCl2 and NaF. Guanosine triphosphate, ATP and MnCl2 activate the soluble enzyme but not the enzyme in the insoluble fraction suggesting that with solubilization from membranes some unfunctional changes of the enzyme may occur. Fractionation of the soluble enzyme preparation revealed the existence of two forms of enzyme differing in molecular weight. These two forms can be further differentiated by their sensitivities to MnCl2 and deoxycholate. A thermostable factor which activates the soluble but not the insoluble enzyme was demonstrated in both beef and rat pineal glands. The thermostable factor is protein in nature because it is nondialyzable and trypsin labile. Whether in vivo the endogenous activator mediates the regulation of the phosphoprotein phosphatase in pineal remains to be investigated.
AROMATIC ACID METABOLITES OF PHENYLALANINE IN THE BRAIN OF THE HYPERPHENYLALANINEMIC RAT: EFFECT OF PYRIDOXAMINE
Authors: Loo Yen Hoong , Scotto Lois , Horning Marjorie G.
Published: October 2006

Abstract— Phenylalanine levels approaching those found in clinical phenylketonuria were produced in the brain of suckling rats by injections of p‐chlorophenylalanine and L‐phenylalanine. The predominant aromatic acid metabolite found in the brain of these animals was phenylacetic acid with decreasing amounts of phenylpyruvic, phenyllactic, and mandelic acids.

The disposition of [3H]pyridoxamine in the brain of normal and hyperphenylalaninemic animals was found to be similar. Pyridoxamine was rapidly phosphorylated in the brain, and excess vitamer was converted mainly to pyridoxal. Pyridoxamine, when injected repeatedly, was effective in significantly reducing the amount of phenylacetate that accumulated in the brain over a period of 6 h. The significance of these findings is discussed.
DISAGGREGATION OF BRAIN POLYSOMES AFTER d‐LYSERGIC ACID DIETHYLAMIDE ADMINISTRATION IN VIVO: MECHANISM AND EFFECT OF AGE AND ENVIRONMENT1
Authors: Holbrook L. , Brown I. R.
Published: October 2006

Abstract— The cell‐free protein synthesis activity and tRNA content of the increased pool of brain monosomes produced after d‐lysergic acid diethylamide (LSD) administration were analyzed. Decreased reinitiation of protein synthesis rather than RNase activation or premature termination was shown to be the mechanism which results in brain polysome disaggregation after administration of the drug in vivo. At a constant dosage of 50 μg/kg the degree of polysome shift increases with age from 3‐week‐old rabbits to adults. There is also an extensive disaggregation of fetal brain polysomes when LSD is administered maternally. The LSD‐induced polysome shift was shown to be altered by holding cage environment, pre‐LSD sedation and post‐LSD handling with brief restraint. It was apparent that elements of environment and physiological arousal were involved in the macromolecular effect of the drug on the protein synthesis apparatus of the brain.
CHARACTERIZATION OF MEMBRANES OBTAINED FROM ELECTRIC ORGAN OF THE ELECTRIC EEL BY SUCROSE GRADIENT FRACTIONATION AND BY MICRODISSECTION
Authors: Rosenberg P. , Silman I. , Ben‐David E. , de Vries A. , Condrea E.
Published: October 2006

Abstract— Two membrane fractions were obtained from electric organ tissue of the electric eel by sucrose gradient centrifugation of tissue homogenates. Electron microscopic examination showed that both fractions contained mainly vesicular structures (microsacs). Both the light and heavy fractions had a‐bungarotoxin‐binding capacity and Na+‐K+ ATPase activity, while only the light fraction had AChE activity. The polypeptide patterns of vesicles derived from both the light and heavy fractions were examined by SDS‐polyacrylamide gel electrophoresis and found to be very similar. The ratio of protein to phospholipid in the light vesicles was much lower than in the heavy vesicles, but the relative amounts of individual phospholipids in the two fractions were similar.

A marked difference in the permeability of the light and heavy vesicles was observed by measuring efflux of both [14C]sucrose and 22Na+, and also by monitoring volume changes induced by changing the osmotic strength of the medium. All three methods showed the heavy vesicles to be much more permeable than the light ones. Only the light vesicles displayed increased sodium efflux in the presence of carbamylcholine.

The AChE in the light fraction does not appear to be membrane‐bound, but is rather a soluble enzyme, detached from the membrane during homogenization, which migrates on the gradient similarly to that of the light vesicles. This is supported by the fact that the bulk of the AChE is readily removed by washing the vesicles. Moreover, under the conditions employed in our sucrose gradient separations,‘native’14 S + 18 S AChE exists in the form of aggregates which migrate very similarly to the major peak of AChE activity of tissue homogenates.

Separated innervated and non‐innervated surfaces of isolated electroplax were obtained by microdissection. α‐Bungarotoxin‐binding capacity was observed only in the innervated membrane. About 80% of the AChE was in the innervated membrane, and about 70% of the Na+‐K+ ATPase in the non‐innervated membrane.

The data presented indicate that the light and heavy vesicle fractions separated by sucrose gradient centrifugation are not derived exclusively from the innervated and non‐innervated membranes respectively, as previously suggested by others, but contain membrane fragments from both sides of the electroplax. The separation of two populations on sucrose gradients may be explained both by the differences in permeability and in protein to phospholipid ratios.
PEROXIDASES IN THE NEURAL RETINA AND PIGMENT EPITHELIUM1
Authors: Armstrong D. , Connole E. , Feeney L. , Berman E. R.
Published: October 2006

Peroxidase activity, assayed with 2 mM‐H2O2 and suitable hydrogen donors (either p‐phenyl‐enediamine or diaminobenzidine), was demonstrated in homogenates of neural retina and pigment epithelium of both the dog and the cow. The enzyme is particle‐associated in the native state, but is readily extractable by brief sonication or freeze‐thawing. At optimum pH, which is between 4.0 and 4.5 for both sources, the specific activity is up to 40 times greater in pigment epithelial cells than in neural retina. Some catalase activity was detected in extracts from both bovine and canine neural retina, but catalase was essentially absent in pigment epithelium.

Fractionation of bovine pigment epithelial cells showed that peroxidase activity is associated mainly with heavy organelles sedimenting at low centrifugal forces. Melanosomes, nuclei, melanolysosomes and plasma membranes were the principal organelles identified in these low speed sediments. It was not possible to separate them either by differential centrifugation or on discontinuous sucrose gradients. However, melanosomes were excluded as the only source of peroxidase activity by isolating separately the melanotic and amelanotic cell populations; equal peroxidase was found in both cell types. Since nuclei are not a likely source of this enzyme, it is suggested that most of the peroxidase activity in bovine pigment epithelial cells is localized in either the melanolysosomes, plasma membranes, or both.
TURNOVER OF PHOSPHATIDYLCHOLINE IN MICROSOMES AND MYELIN IN BRAINS OF YOUNG AND ADULT RATS
Authors: Miller Sheldon L. , Morell Pierre
Published: October 2006

We have tested the hypothesis that the turnover of phosphatidylcholine in subcellular fractions of rat brain is a function of the age at which this lipid is deposited. Rats, 60 days of age, were injected intracranially with [2‐3H]glycerol and either [methyl‐14C]choline (to label the base moiety) or [U‐14C]glucose (to label acyl moieties). Littermates were killed up to 90 days after injection and brain microsomes and myelin isolated. Lipids were extracted and the phosphatidylcholine was isolated by 2‐dimensional TLC and hydrolyzed to its constituent moieties. The 3H in the glycerol backbone and 14C in the choline or acyl residues was quantitated. The microsomal and myelin 3H/14C ratios decreased with time with either set of precursors, indicating that labeled choline and acyl moieties were reutilized more efficiently than the glycerol backbone. The various precursors exhibited first order decay curves with half‐lives for the glycerol backbone of 6 and 11 days for the microsomal and myelin fractions respectively.

These results contrast with those previously obtained with identical experimental procedures when 17‐day‐old animals were injected. In that study, although much of the phosphatidylcholine turned over rapidly as for the older animals, by 2 weeks after injection most of the remaining phosphatidylcholine was turning over more slowly with a half‐life of 13 and 25 days for microsomes and myelin respectively (Miller et al., 1977). The base and acyl moieties also had a corresponding shorter half‐life in older animals relative to the slow turnover phase in younger rats.
A CHOLESTEROL‐ESTERIFYING ENZYME IN RAT CENTRAL NERVOUS SYSTEM MYELIN1
Authors: Choi Myung‐Un , Suzuki Kunihiko
Published: October 2006

Aontrary to our earlier finding (Eto & Suzuki, 1971), the myelin fraction purified from young adult rat brain consistently showed cholesterol‐esterifying activity. The specific activity in myelin was the highest among subcellular fractions. Extensive washing wiih various aqueous salt solutions failed to remove the activity from myelin. The enzyme was evenly distributed among the arbitrarily defined light, medium and heavy myelin subfractions. The myelin‐localized activity showed the pH optimum and heat stability identical to the microsome‐bound activity. Although there were minor differences in the effect of detergents or exogenous lipids added to the reaction mixture, no firm evidence was obtained to indicate that the myelin‐bound cholesterol‐esterifying enzyme is different from that in other subcellular fractions. On the other hand, the distribution among the myelin subfractions and heat stability of the myelin‐bound cholesterol‐esterifying activity were different from those of the myelin‐specific cholesterol ester hydrolase. Therefore, the esterification does not appear to be a mere reverse reaction catalyzed by the previously known myelin‐specific hydrolase. The rat brain myelin, therefore, is capable of both synthesizing and hydrolyzing cholesterol esters.
ACYL GROUP COMPOSITION OF METABOLICALLY ACTIVE LIPIDS IN BRAIN: VARIANCES AMONG SUBCELLULAR FRACTIONS AND DURING POSTNATAL DEVELOPMENT
Authors: Su Kwei Lee , Sun Grace Y.
Published: October 2006

Using a combination of preparative TLC and GLC technique, the content and acyl group composition of diacyl‐glycerophosphoinositols, diacyl‐glycerophosphates, diacylglycerols and triacyl‐glycerols in brain tissue were determined. The level of diacyl‐glycerophosphoinositols in 40 day‐old mouse brain was 2.7 μmol/g tissue as compared to 40–170 nmol/g for other minor lipids. The acyl groups of diacyl‐glycerophosphoinositols were enriched in 18:0 and 20:4 (n‐6). This characteristic acyl group profile was found in microsomes, synaptosomes, and in myelin. The acyl groups of diacyl‐glycerophosphates and diacylglycerols were comprised mainly of 16:0, 18:0, 18:1 and 20:4 (n‐6). In rat brain subcellular fractions, the acyl groups of diacylglycerols and diacyl‐glycerophosphates in the microsomal fraction had a higher proportion of 22:6 (n‐3) than those in the myelin and synaptosomal fractions. The acyl groups of the myelin lipids were higher in 18:l and lower in 20:4 (n‐6) as compared to those in the microsomal and synaptosomal fractions. The triacylglycerols in brain exhibited an unusual acyl group profile which included small proportions of 14:0, 16:1, 20:4 (n‐6), 22:4 (n‐6) and 22:6 (n‐3). Except for an increase in 18:1 and a corresponding decrease in 16:0 which was found in diacyl‐glycerophosphoinositols, no apparent acyl group change was observed in other metabolically active lipids during postnatal brain development.
THE TOXIC EFFECT OF SODIUM GLUTAMATE AND DL‐α‐AMINOADIPIC ACID ON RAT RETINA: CHANGES IN HIGH AFFINITY UPTAKE OF PUTATIVE TRANSMITTERS
Authors: Karlsen R. Lund
Published: October 2006

Subcutaneous administration of high doses of sodium glutamate to new born rats was used to destroy retinal interneurons and ganglion cells. Such treatment was accompanied by 90% reduction in the high affinity uptake of choline, 60–70% reductions in the uptakes of GABA, diamino‐n‐butyric acid and glycine and 30–40% reductions in the uptakes of asparatate and glutamate measured on retinal homogenates from 30‐day‐old rats. The high affinity uptakes of β‐alanine and taurine were unchanged. Preincubation of retinal homogenates with 1 mM β‐alanine or 100 μM diamino‐n‐butyric acid severely reduced the high affinity GABA uptake in control and experimental animals.

In intact retinae, however, the glutamate treatment increased the high affinity uptake of β‐alanine by 70%, whereas that of diamino‐n‐butyric acid was reduced by 40% and the high affinity uptakes of GABA and glutamate were unchanged.

Four hours after injection of the gliotoxic compound DL‐α‐aminoadipic acid into the vitreous body of 30‐day‐old rats, the Müller cells could no longer be identified. This lesion was accompanied by 55% reduction in the high affinity uptake of β‐alanine and 25% reduction in the uptakes of GABA and glutamate on intact retinae. The high affinity uptakes of diamino‐n‐butyric acid, choline and the enzyme activities of choline acetyltransferase and glutamate decarboxylase were unchanged under these conditions. After 24 h, however, the Müller cells could be recognized again, and the β‐alanine uptake had normalized.
CYCLIC NUCLEOTIDES and PROTEIN KINASE ACTIVITY IN THE RAT BRAIN POSTMORTEM
Authors: Schmidt Michael J. , Truex Lewis L. , Thornberry John F.
Published: October 2006

Abstract— The components of the cyclic nucleotide system were studied in rat brain at various times after death to determine the stability of the system postmortem. The concentration of cyclic AMP in 4 brain regions was highest 10 min after death and declined to stable levels within 6 h after death. At this time concentrations approximated those normally seen in vitro. The concentration of cyclic GMP in brain regions fell markedly postmortem and was undetectable 6 or 16 h after death. Nor‐epinephrine‐stimulated cyclic AMP accumulation measured in vitro in slices of the cerebral cortex was the same 10 min and 16 h postmortem. In the cerebellum, however, accumulated levels of cyclic AMP were lower 16 h postmortem, although the degree of stimulation elicited by norepinephrine was the same 10 min and 16 h after death. Cyclic AMP‐dependent and independent protein kinase activities were detected in the rat brain at all times after death. There was no change in the cyclic AMP‐dependent enzyme activity postmortem, but activity in the absence of cyclic AMP was significantly higher immediately after death compared to 6 or 16 h postmortem. These studies indicate that it should be possible to study the cyclic AMP system in human brain tissue obtained at autopsy.
STUDIES ON THE TRANSPORT OF GLUTAMINE IN VIVO BETWEEN THE BRAIN AND BLOOD IN THE RESTING STATE AND DURING AFFERENT ELECTRICAL STIMULATION
Authors: Abdul‐Ghani A.‐S. , Marton M. , Dobkin J.
Published: October 2006

Abstract— In this work we have studied the effect of afferent electrical stimulation (AES) of the contralateral brachial plexus on the release of glutamine and glutamate from the cat's brain into the cerebral venous blood, at rest and during continuous infusion of L‐glutamine and sucrose solutions.

(1) In the resting state, before stimulation, there was a net outflow of glutamine from the brain into the cerebral venous blood, but no release of glutamic acid. (2) AES caused release of glutamate and increased 3.5‐fold the release of glutamine. The increase in release of glutamine and glutamate was found to be reversed very shortly after stimulation. (3) Steady intravenous infusion of a 0.3 M‐gluta‐mine solution for 10 min changed the negative arterio‐venous difference in glutamine to a positive one and increased the content in brain by 15×20%. In this case AES caused a singificant drop, to zero of the glutamine arterio‐venous difference. (4) At the onset of pentamethylenetetrazole (PTZ) seizures, like AES, there was a significant reduction of the level of glutamine in the cats’cerebral cortex. This reduction vanished when the animals were infused with L‐glutamine solution but not with 0.3 M‐sucrose solution that was used as an inert electrolyte. (5) The kinetic behaviour of the glutamine transport is compatible with a carrier‐mediated process, but not with passive diffusion.
ENZYMATIC BIOSYNTHESIS OF ETHANOLAMINE‐ AND CHOLINE‐PHOSPHOGLYCERIDES IN RETINA DURING DEVELOPMENT. EFFECT OF LIGHT STIMULATION
Authors: Dreyfus H. , Harth S. , Urban P. F. , Mandel P. , Freysz L.
Published: October 2006

Choline‐ and ethanolamine‐phosphoglycerides (CPG and EPG) are the most abundant phospholipids of retinal membranes. We have investigated some regulatory mechanisms involved in the final steps of their biosynthesis, namely those catalysed by CDP‐choline 1,2 diradyl‐sn‐glycerol choline phosphotransferase (CPT) and CDP‐ethanolamine 1,2 diradyl‐sn‐glycerol ethanolamine phosphotransferase (EPT). We have studied both enzymes in the retina which offers an excellent model for the investigation of the molecular basis of the effect of its physiological stimulus, the light. In chick retina. the specific activity (SA) of EPT reached a maximum at the 18th day of embryonic life and decreased thereafter. In the case of CPT, a similar peak of SA was observed at hatching. The time of maximum SA of EPT and CPT corresponded to the period during which retinal rod outer segments are formed. The apparent Km values of EPT and CPT determined with whole retinal homogenates for CDP‐bases showed different profiles. The apparent Km of EPT decreased during embryonic life and increased thereafter whereas the apparent Km of CPT did not change during ontogenesis. Light stimulation of calf retinal homogenates had different effects on phosphotransferase activities. In the presence of only endogenous diacylglycerol (DAG) the SA of CPT was 2‐fold higher for dark‐adapted retinas, whereas no differences in EPT activities were observed. After addition of exogenous DAG (4mM) to the incubation medium, light stimulation of the retina led to a 50% increase of EPT activity whereas no effect was observed for CPT. These different effects could be related to the cyclic nucleotides present in retina before and after light stimulation. In addition all the data presented in this study indicate that, as in brain, CPT and EPT in retina are two different enzymes.
UPTAKE, RELEASE AND HOMO‐ AND HETERO‐EXCHANGE DIFFUSIONS OF INHIBITORY AMINO ACIDS IN GUINEA‐PIG CEREBELLAR SLICES
Authors: Okamoto Koichi , Namima Misako
Published: October 2006

Abstract— GABA, taurine and β‐alanine are taken up by guinea‐pig cerebellar slices by both the high‐and low‐affinity uptake processes, whereas glycine is taken up only by the low‐affinity process. A considerable amount of labelled GABA loaded in the slice is released by unlabelled external GABA and a minute amount is released by external β‐alanine, glycine and taurine. External glycine and β‐alanine releases labelled glycine loaded in the slice. Labelled taurine loaded is effectively released by external taurine and β‐alanine, while labelled β‐alanine loaded is released only by external β‐alanine.

It is suggested that hetero‐exchanges which are one‐directional in some cases also take place between the amino acids in addition to homo‐exchanges. Therefore, high‐affinity uptake processes observed with GABA and taurine could be the result of the homo‐exchange diffusions, while that of β‐alanine could be due to either the homo‐exchange or the hetero‐exchange diffusions or both.

K+′‐evoked releases of GABA and to a lesser extent, taurine are partially dependent upon the presence of Ca+ in the superfusion media, whereas that of glycine and probably that of β‐alanine, are not, K+ ‐evoked releases of labelled GABA and taurine are larger when loaded by their high‐affinity uptake systems than by their low‐affinity uptake processes. The reverse is the case with labelled glycine and β‐alanine. These results do not rule out the possibility that taurine might act as a neurotransmitter in the cerebellum.
CHARACTERIZATION OF THE ENZYMIC TRANSFER OF ARACHIDONOYL GROUPS TO 1‐ACYL‐PHOSPHOGLYCERIDES IN MOUSE SYNAPTOSOME FRACTION
Authors: Corbin D. R. , Sun G. Y.
Published: October 2006

Abstract—

In the presence of ATP, Mg2+ and CoA, mouse brain synaptosomes actively catalyze the transfer of arachidonate to 1‐acyl‐phosphoglycerides. The enzymic acylation process was inhibited greatly by 0.1 mm‐PCMB (parachloromercuric benzoate), and only partially by 0.1 mm‐DNP (2,4‐dini‐trophenol) and 0.001% (w/v) oligomycin. Arachidonoyl transfer to 1‐acyl‐glycerophosphorylcholines and 1‐acyl‐glycerophosphorylinositols was negligibly affected by 1 mm‐NaF and 10 mm‐glucose. Among the different 1‐acyl‐phosphoglycerides used as acceptor molecules, the relative order for arachidonoyl transfer activity was: 1‐acyl‐glycerophosphorylinositols > 1‐acyl‐glycerophosphorylcholines > l‐acyl‐glycerophosphorylethanolamines. The ester linkage at the C‐1 position of the 1‐acyl‐phosphoglycerides also appears necessary for activity for no arachidonoyl transfer activity was observed when 1‐alkenyl‐glycerophosphorylethanolamines were used as the acceptor molecules. Arachidonate transfer to 1‐acyl‐glycerophosphorylinositols was optimal at low acceptor concentrations (10–30 μm) whereas the transfer to 1‐acyl‐glycerophosphorylcholines occurred over a much broader range of acceptor concentrations (30–80 μm). At optimal acceptor concentration, maximal transfer activity was observed at arachidonate concentration of 50–80 μm. The newly synthesized phosphoglycerides were found distributed in the subsynaptic components with specific activity highest in the fraction containing synaptic vesicles.
INCORPORATION OF LIPIDS INTO SUBCELLULAR FRACTIONS OF BRAIN OF QUAKING MUTANT
Authors: Albers‐Jackson B. , Greenfield S. , Brostoffand S. W. , Hogan E. L.
Published: October 2006

Abstract—

The synthesis of lipids and their assembly into subcellular membrane fractions of the myelin deficient Quaking mutant and control brains was studied in 18‐, 24‐ and 41‐day‐old animals using a double label methodology with14C and 3H acetate as precursors. As a general procedure, Quaking mutants were injected intracranially with 50 μCi [14C]acetate and their littermate controls with 300 μCi [3H]acetate. The animals were killed 3 h post‐injection, their brains were pooled and subcellular fractions prepared from the common homogenate. An 80‐90% decrease in the incorporation of acetate into eleven lipids of myelin in the Quaking mutant was found. This occurred in the face of apparent normal incorporation (relative to microsomes) into lipids of the other main subcellular fractions (nuclear. mitochondrial and synaptosomal) with the exception of decreased incorporation into the myelin‐like fraction at 18 and 24 days. Cholesterol and cerebroside were less readily incorporated into Quaking myelin than the other lipids. Although the microsomal synthesis of cholesterol and cerebroside was depressed by about 30% in the Quaking mutant, the incorporation of cholesterol into nuclear, synaptosomal and mitochondrial fractions was unaffected in the mutant. This indicates that sufficient cholesterol is synthesized for the normal assembly of these organelles. In contrast the incorporation of acetate into cholesterol and cerebroside of Quaking myelin was decreased much more than microsomal synthesis. This latter result is consistent with a defect in the process of myclin membrane assembly
THYMIDINE PHOSPHORYLASE, THYMIDINE KINASE AND THYMIDYLATE SYNTHETASE ACTIVITIES IN CEREBRAL HEMISPHERES OF DEVELOPING CHICK EMBRYOS1
Authors: Hyndman A. G. , Zamenhof S.
Published: October 2006

Abstract—

The changes in activities of thymidine phosphorylase (EC 2.4.2.4), thymidine kinase (EC 2.7.1.75) and thymidylate synthetase (methylenetetrahydrofolate:dUrd‐5′‐P C‐methyltransferase, EC 2.1.1.‐) in the cerebral hemispheres of developing chick embryos were determined and compared with the course of DNA synthesis and of natural cell death in this organ.

Thymidine phosphorylase activity reaches a broad maximum at the 12th to 14th day of incubation, followed by a rapid decrease. Thymidine kinase and thymidylate synthetase activities are highest at the earliest time studied (day 10) and decrease until day 14, followed by an increase from day 14 to 16 and a further decrease from day 16 through 1 day post‐hatching. The rate of DNA synthesis essentially follows these activities, but the increase at day 16 is not discernible.

Our previous study revealed high DNA synthesis at day 10, with natural cell death concurring on days 12‐14, followed by another peak after day 16 (glial proliferation) and a decrease after day 16. It appears that thymidine phosphorylase activity reaches a maximum (days 12‐14) at the time of maximum cell death, which may be correlated with the degradative function of this enzyme. This was also the time for minimum activities of thymidine kinase and thymidylate synthetase; on the other hand, these activities reach a first (day 10) and second (day 16) maximum at the time of maximum DNA synthesis; this may be correlated with the synthetic functions of these enzymes.
RNA POLYMERASE ACTIVITY IN THE SUPERIOR CERVICAL GANGLION OF THE NEONATAL RAT: THE EFFECT OF NERVE GROWTH FACTOR
Authors: Huff Kenneth , Lakshmanan J. , Guroff Gordon
Published: October 2006

Abstract—

The activity of RNA polymerase in the superior cervical ganglia of the neonatal rat has been studied. The characteristics of the activity with respect to enzyme concentration, temperature, time, and ion concentrations are presented. Ionic conditions of the assay favoring polymerase I and polymerase II were each studied in the presence of specific inhibitors of DNA‐dependent RNA synthesis. The Km for GTP under either polymerase I or polymerase II conditions was found to be 10−5M, and the relative amounts of poly(A)‐containing RNA synthesized under both conditions was 4‐5%. A decrease in the activity of RNA polymerase with age has been observed.

The time course of increased activity following treatment of the animals in vivo or treatment of the isolated ganglia in organ culture with nerve growth factor is presented. The increase in activity observed after the administration of nerve growth factor is discussed in terms of possible mechanisms of action of nerve growth factor in this tissue.
EFFECT OF THE ADMINISTRATION OF L‐5‐HYDROXYTRYPTOPHAN AND A MONOAMINE OXIDASE INHIBITOR ON GLUCOSE METABOLISM IN RAT BRAIN
Authors: Wong Kar‐Lit , Tyce Gertrude M.
Published: October 2006

Abstract—

The effects of the presence of large amounts of 5‐HT and of its precursor 5‐HTP in brain on cerebral utilization of glucose were studied. [U‐14C]Glucose was injected to fed rats that had previously been treated with L‐5‐HTP, L‐5‐HTP and an inhibitor—N‐[β‐(2‐chlorophenoxy)‐ethyl]‐cyclopropylamine hydrochloride (Lilly‐51641)‐of MAO, or Lilly‐51641 alone. Such treatment increased the concentrations of 5‐HTP and 5‐HT in the brain. After treatment with 5‐HTP and Lilly‐51641, and to a lesser extent with Lilly‐51641 alone, the concentration of glucose in plasma was increased. However, the uptake of glucose by the brain did not appear to be proportionately increased, and this suggested an impairment in this mechanism. After the administration of Lilly‐51641 alone and more especially of Lilly‐51641 plus 5‐HTP, the concentration of glucose in the brain was increased. This increase was thought to be due to an impairment of glucose utilization, because the flux of 14C from glucose to amino acids in the brain was reduced. The concentrations of most major amino acids in the brain were not greatly affected by these treatments. GABA and alanine concentrations in the brain were modestly increased after treatment with 5‐HTP alone or in combination with Lilly‐51641. The present results suggest that the metabolism of glucose to amino acids in the brain is altered when the concentration of 5‐HTP, or more especially that of 5‐HT, in the brain is increased.