Effect of Exogenous Gangliosides on Amino Acid Uptake and Na+,K+-ATPase Activity in Superior Cervical and Nodose Ganglia of Rats

The effects of some gangliosides on active uptake of nonmetabolizable alpha-aminoisobutyric acid (AIB) and Na+, K+-ATPase and Ca2+, Mg2+-ATPase activities in superior cervical ganglia (SCG) and nodose ganglia (NG) excised from adult rats were examined during aerobic incubation at 37 degrees C for 2 h. In NG, amino acid uptake was greatly accelerated with the addition of galactosyl-N-acetylgalactosaminyl-[N-acetylneuraminyl]-galactosylgluc osyl ceramide (GM1) (85%) and also with N-acetylgalactosaminyl-[N-acetylneuraminyl]-galactosylglucosyl ceramide (GM2) or [N-acetylneuraminyl]-galactosyl-N-acetylgalactosaminyl-[N-acetyl- neuraminyl]-galactosylglucosyl ceramide (GD1a) (43% each) compared with a nonaddition control at a 5 nM concentration. Under identical conditions, Na+, K+-ATPase activity was strongly stimulated with GM1 (180%) and GD1a (93%), whereas Ca2+, Mg2+-ATPase activity showed no change. In SCG, on the other hand, AIB uptake was apparently inhibited (-27%) by addition of GM1, with a slight decrease in Na+, K+-ATPase but no change in Ca2+, Mg2+-ATPase activity in the tissue. Both asialo-GM1, in which N-acetylneuraminic acid is deficient, and Forssman glycolipid, which is not present in nervous tissue, failed to produce any significant increase in both SCG and NG not only in amino acid uptake, but also in Na+, K+-ATPase activity. A kinetic study of active AIB uptake showed that GM1 ganglioside produced an increase in Km with no change in Vmax in SCG, whereas it caused a decrease in Km with a slight increase in Vmax in NG. Treatment of NG and SCG with neuraminidase from Vibrio cholerae, an enzyme that split off sialic acid from polysialoganglioside, leaving GM1 intact, caused little inhibition of the amino acid uptake.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of Quinolinic Acid in the Nucleus Basalis Magnocellularis on Cortical High-Affinity Choline Uptake

A transient 45% increase in cortical high-affinity choline uptake (HACU) was observed after an injection of quinolinic acid (QUIN) into the nucleus basalis magnocellularis (nbM) of the rat. This was followed by a steady decline in choline uptake, which resulted in a 46% decrease by day 7. Specific [3H]hemicholinium-3 binding to coronal brain sections showed a similar pattern following injections of QUIN into the nbM. The increase in cortical HACU elicited by QUIN appeared to be dose dependent.     

γ-Hydroxybutyric Acid in Subcellular Fractions of Rat Brain

The presence of gamma-hydroxybutyric acid (GHB) in synaptosome-enriched fractions of rat brain was ascertained using a GLC technique. The stability of GHB in synaptosomes was evaluated by addition of various gamma-aminobutyric acid (GABA) transaminase (GABA-T) inhibitors, GHB, or ethosuximide to the homogenizing medium. Furthermore, changes in whole brain GHB levels were compared with those in the synaptosomal fraction in animals treated with GABA-T inhibitors, GABA, or ethosuximide. GHB was present in synaptosome-enriched fractions in concentrations ranging from 40 to 70 pmol/mg of protein. There was no evidence for redistribution, leakage, or metabolism of GHB during the preparation of synaptosomes. The elevations of whole brain GHB level associated with GABA-T or ethosuximide treatment were reflected by a parallel increase in synaptosomal GHB content. These data add to the growing evidence that GHB may have neurotransmitter or neuromodulator function.     

Time Course of Adaptations in Dopamine Biosynthesis, Metabolism, and Release Following Nigrostriatal Lesions: Implications for Behavioral Recovery from Brain Injury

Alterations in neostriatal dopamine metabolism, release, and biosynthesis were determined 3, 5, or 18 days following partial, unilateral destruction of the rat nigrostriatal dopamine projection. Concentrations of dopamine and each of its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 3-methoxytyramine (3-MT) were markedly decreased in the lesioned striata at 3, 5, or 18 days postoperation. The decline in striatal high-affinity [3H]dopamine uptake closely matched the depletion of dopamine at 3 and 18 days postoperation. However, neither DOPAC, HVA, nor 3-MT concentrations were decreased to as great an extent as dopamine at any time following lesions that depleted the dopamine innervation of the striatum by greater than 80%. In these more severely lesioned animals, dopamine metabolism, estimated from the ratio of DOPAC or HVA to dopamine, was increased two- to four-fold in the injured hemisphere compared with the intact hemisphere. Dopamine release, estimated by the ratio of 3-MT to dopamine, was more increased, by five- to sixfold. Importantly, the HVA/dopamine, DOPAC/dopamine, and 3-MT/dopamine ratios did not differ between 3 and 18 days postlesioning. The rate of in vivo dopamine biosynthesis, as estimated by striatal DOPA accumulation following 3,4-dihydroxyphenylalanine (DOPA) decarboxylase inhibition with NSD 1015, was increased by 2.6- to 2.7-fold in the surviving dopamine terminals but again equally at 3 and 18 days postoperation. Thus, maximal increases in dopamine metabolism, release, and biosynthesis occur rapidly within neostriatal terminals that survive a lesion. This mobilization of dopaminergic function could contribute to the recovery from the behavioral deficits of partial denervation by increasing the availability of dopamine to neostriatal dopamine receptors. However, these presynaptic compensations are not sufficient to account for the protracted (at least 3-week) time course of sensorimotor recovery that has been observed following partial nigrostriatal lesion.     

N1-Methyl-2-125I-Lysergic Acid Diethylamide, a Preferred Ligand for In Vitro and In Vivo Characterization of Serotonin Receptors

Methylation of 2-125I-lysergic acid diethylamide (125I-LSD) at the N1 position produces a new derivative, N1-methyl-2-125I-lysergic acid diethylamide (125I-MIL), with improved selectivity and higher affinity for serotonin 5-HT2 receptors. In rat frontal cortex homogenates, specific binding of 125I-MIL represents 80-90% of total binding, and the apparent dissociation constant (KD) for serotonin 5-HT2 receptors is 0.14 nM (using 2 mg of tissue/ml). 125I-MIL also displays a high affinity for serotonin 5-HT1C receptors, with an apparent dissociation constant of 0.41 nM at this site. 125I-MIL exhibits at least 60-fold higher affinity for serotonin 5-HT2 receptors than for other classes of neurotransmitter receptors, with the dopamine D2 receptor as its most potent secondary binding site. Studies of the association and dissociation kinetics of 125I-MIL reveal a strong temperature dependence, with very slow association and dissociation rates at 0 degree C. Autoradiographic experiments confirm the improved specificity of 125I-MIL. Selective labeling of serotonin receptors was observed in all brain areas examined. In vivo binding studies in mice indicate that 125I-MIL is the best serotonin receptor label yet described, with the highest frontal cortex to cerebellum ratio of any serotonergic radioligand. 125I-MIL is a promising ligand for both in vitro and in vivo labeling of serotonin receptors in the mammalian brain.     

Calcium-Dependent and-Independent Release of Glutamate from Synaptosomes Monitored by Continuous Fluorometry

An enzyme-linked fluorometric assay is described for the continuous monitoring of the unidirectional efflux of glutamate from guinea-pig synaptosomes. Glutamate efflux from freshly suspended, polarized synaptosomes occurs at 0.35-0.39 nmol min-1 mg of protein-1 and is not significantly affected by external Ca2+. KCl depolarization (30 mMKCl) in the absence of Ca2+ doubles this rate, whereas in the presence of Ca2+, the initial kinetics of the assay are consistent with the release in the first 5 s of 0.6 nmol mg of protein-1. The final extent of Ca2+-dependent release amounts to 1.9 nmol mg of protein-1, or 8.5% of the total intrasynaptosomal glutamate content. Preincubation of synaptosomes at 30 degrees C for 2 h before depolarization leads to a decrease in Ca2+-independent release and an increase in Ca2+-dependent release, consistent with an intrasynaptosomal relocation of the amino acid.     

Postmortem Changes in Rat Brain: Studies on Membrane-Bound Enzymes and Receptors

The relationship between the stability of potential neurochemical markers and autolysis time was studied at 4 degrees C and 25 degrees C using postmortem brain samples from two rat strains. In general, qualitatively similar results were obtained with either N/Nih or Sprague-Dawley rats; however, quantitative differences were often observed, particularly in regard to benzodiazepine receptor changes. For every enzyme activity or binding property examined, no significant change was found when brains were kept at 4 degrees C for up to 72 h prior to freezing at -70 degrees C. Na,K-ATPase and low-affinity Ca-ATPase activities were also stable in brains kept at 25 degrees C for up to 72 h. Mg-ATPase activity was reduced in brains kept at 25 degrees C for 24 and 48 h. [3H]Guanidinoethylmercaptosuccinic acid [( 3H]GEMSA) binding to enkephalin convertase in the cytosol was not significantly changed in brains kept at 25 degrees C; however, a small increase was seen for [3H]GEMSA binding to the membrane fraction at 24, but not 48 and 72 h postmortem. [3H]Quinuclidinyl benzilate [( 3H]QNB) binding to muscarinic cholinergic receptors decreased in brains kept at 25 degrees C for 72 h. Opioid receptor binding also decreased in brains kept at 25 degrees C. Using [3H]2-D-alanine-5-D-leucine enkephalin to label delta opioid receptors, a statistically significant decrease in binding was observed as early as 6 h postmortem, and was completely abolished after 72 h at 25 degrees C. In contrast, [3H]naloxone binding was unchanged after 24 h at 25 degrees C, but was decreased after 48 and 72 h.(ABSTRACT TRUNCATED AT 250 WORDS)     

2-Amino-7-Phosphonoheptanoic Acid Inhibits Insulin-Induced Convulsions and Striatal Aspartate Accumulation in Rats with Frontal Cortical Ablation

Pretreatment of rats with the excitatory amino acid antagonist 2-amino-7-phosphonoheptanoic acid (2-APH; 0.5 mmol/kg, i.p.) protected against insulin-induced clonic seizures. Complete protection was observed in 38% of the rats and partial protection in an additional 50%. Lesioning of the corticostriatal pathway by frontal cortical ablation caused decreases in the striatal levels of aspartate (-28%) and glutamate (-18%), an increase in striatal glutamine level (45%), and decreased high-affinity uptake of D-[3H]aspartate (-27%) in the lesioned dorsal neostriatum. Insulin-induced hypoglycemia caused a predicted sharp increase in aspartate level (165%) and decreased glutamate (-20%) and glutamine (-38%) levels in the intact striatum. Pretreatment of rats with 2-APH significantly reversed the insulin-induced changes in striatal aspartate, glutamate, and glutamine levels, especially in the intact hemisphere. In normoglycemic control rats, the "metabolic," i.e., concentration in the lesioned hemisphere, aspartate pool constituted 72% and the "synaptic," i.e., the concentration difference between the intact and lesioned hemispheres, 28% of the total striatal aspartate pool. 2-APH had no effect on the level of "metabolic" aspartate in the striata of normoglycemic rats but caused an almost complete suppression of "synaptic" aspartate. Following insulin-induced hypoglycemia, the "metabolic" aspartate pool doubled, whereas the "synaptic" aspartate pool increased 3.5-fold in the absence of 2-APH. The insulin-induced rise in "synaptic" aspartate level was almost completely blocked by 2-APH (a 5% rise instead of a 3.5-fold rise).(ABSTRACT TRUNCATED AT 250 WORDS)     

A systematic study of the amino acid compositions and D/L ratios in fossil bones from two french neanderthalian sites

The reaction of racemization in which the L amino acids are reversibly converted into the corresponding D amino acids, proceeds in geological environment at such a slow rate that it may be used as a geochronometer. However, in fossils several parameters may affect the rate of racemization, i.e. moisture, surface, pH buffer and metal cations. This work consists of a systematic study of total amino acid content in fossil bones from two neanderthalian sites. The amino acid distributions of all specimens were determined and compared to that of fresh bone. The D/L amino acid were quantified and expressed in terms of age as a function of the temperature. The results led us to consider the «La Roquette» site older than «Les Canalettes» site.     

用薄层色谱分离提纯单半乳糖和双半乳糖双甘油酯

本文选择两种溶剂体系,用两次单向薄层层析,从小麦抗寒与不抗寒品系天然橡胶胶乳分离提纯出6种单半乳糖和双半乳糖双甘油酯。并比较了它们的疏水侧链的脂肪酸组成。小麦糖脂疏水侧链脂肪酸的不饱和指数远大于天然胶乳糖脂。抗寒品系胶乳糖脂疏水侧链脂肪酸不饱和指数大于不抗寒品系。双半乳糖双甘油酯疏水侧链脂肪酸不饱和指数均大于其单半乳糖糖脂。