bluuebunny/crossref_metadata_2025 huggingface dataset:
Purification and Characterization of Clathrin from Bovine Brain
Authors: Garbern James Y. , Wu Jang‐Yen
Published: October 2006

Abstract: Clathrin has been purified to electrophoretic homogeneity by initial extraction of clathrin from purified coated vesicle fraction, followed by column chromatographies with gel filtration. DEAE‐cellulose, and hydroxylapatite and finally by preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE). Antibody specific to clathrin has also been obtained. Two forms of native clathrin, fast and slow components, have been prepared to about 95% purity by hydroxylapatite column chromatography. Both fast and slow components are believed to represent two different aggregates of clathrin subunit because they comigrate in agarose electrophoresis. pH 7.4, and also migrate as clathrin subunit on SDS‐PAGE with a molecular weight of 175,000. Furthermore, both components cross‐react with antibody against purified clathrin and compete for antibody binding site with labeled fast component. The fast component can also be converted to the slow component. In addition to clathrin, two proteins of about 38,000 and 35,000 M.W. that consistently co‐purified with native clathrin are probably also intrinsic to coated vesicle.
Metabolism of Acetate to Amino Acids in Brains of Rats After Complete Hepatectomy
Authors: Tyce Gertrude M. , Ogg Jauneita , Owen Charles A.
Published: October 2006

Abstract: The concentration of glutamine increases in the brain after hepatectomy. In the present studies the conversion of intravenously given [14C]acetate to [14C]glutamate and [14C]glutamine was studied in control rats and in rats at 6 h after complete hepatectomy. The incorporation of label into glutamate was only slightly inhibited, but the further incorporation into glutamine was greatly inhibited, after hepatectomy. These data, and previous data using [14C]glucose as precursor, indicate that synthesis of glutamine in brain is inhibited after hepatectomy, and suggest that its concentration must increase because degradation is inhibited to an even greater extent.
A Gas Chromatographic Method for the Determination of N‐Acetyl‐l‐Aspartic Acid, N‐Acetyl‐α‐ Aspartylglutamic Acid and β‐Citryl‐l‐Glutamic Acid and Their Distributions in the Brain and Other Organs of Various Species of Animals
Authors: Miyake Masaham , Kakimoto Yasuo , Sorimachi Masaru
Published: October 2006

Abstract: A simple and sensitive gas‐chromatographic method for the determination of N‐acetyl‐l‐aspartic acid (NA‐Asp), N‐acetyl‐α‐aspartylglutamic acid (NA‐Asp‐Glu) and β‐citryl‐l‐glutamic acid (β‐CG) was developed. The organ, regional and phylogenetic distributions of these compounds were studied. NA‐Asp and NA‐Asp‐Glu were highly concentrated in nervous tissue, and less than 1% of the amounts in the nervous tissues were found in nonnervous organs. These two compounds showed a reciprocal relationship in their regional distribution in mature brains, but such a relationship was not evident or was even reversed in immature brains. The two compounds also showed different developmental changes in different regions of the brain. Fish brain contained a relatively high concentration of NA‐Asp, but only a trace amount of NA‐Asp‐Glu. By contrast, a 10 times higher concentration of NA‐Asp‐Glu than NA‐Asp was found in frog brain. Reptilian brain contained similar amounts of each compound. Avian and mammalian brain had NA‐Asp at a roughly 10 times higher concentration than NA‐Asp‐Glu. β‐CG occurred at the highest concentration in the immature brain of rat and guinea pig, but disappeared in the mature brains. The adult frog brain, however, contained a large amount of β‐CG. In the adult rat, testis contained the highest concentration of β‐CG.
Uptake and Release of N‐Methyl‐d‐Aspartate by Rat Brain Slices
Authors: Skerritt J. H. , Johnston G. A. R.
Published: October 2006

Abstract: The excitant amino acid, N‐methyl‐d‐aspartate, was actively taken up by slices of rat cerebral cortex. This uptake was Na+ ‐ and temperature‐dependent, but was relatively inefficient (Km 3 MM, Vmax 0.07 μmol/g/min) compared with that of other acidic amino acids. The uptake of N‐methyl‐d‐aspartate does not appear to have a rate‐limiting influence on the time course of N‐methyl‐d‐aspartate‐induced excitation since potent uptake inhibitors, such as threo‐3‐hydroxy‐l‐aspartate, do not influence the excitant action of N‐methyl‐d‐aspartate. The relatively prolonged excitant action of this acidic amino acid may be the result of relatively slow dissociation of the activated receptor complex. Reloaded N‐methyl‐d‐aspartate can be released from rat brain slices by stimulation with K+ ions. Such K+‐stimulated release appeared to be Ca2+‐independent, unlike the K+‐stimulated release of preloaded d‐aspartate. These findings suggest that N‐methyl‐d‐aspartate may be a weak but selective substrate for a glial acidic amino acid uptake system.
Characterization of an Initiating Cell‐Free Protein Synthesis System Derived from Rabbit Brain
Authors: Cosgrove James W. , Brown Ian R.
Published: October 2006

Abstract: Protein synthesis in the brain is known to be affected by a wide range of treatments. The detailed analysis of the mechanisms that are involved would be facilitated by the development of cell‐free translation systems derived from brain tissue. To date, brain cell‐free systems have not been fully characterized to demonstrate a capacity for initiation of translation. The following criteria were utilized to demonstrate that a cell‐free protein synthesis system derived from rabbit brain was capable of initiation in vitro: (a) sensitivity of cell‐free translation to the initiation inhibitor aurintricarboxylic acid (ATA); (b) binding of [35S]Met‐tRNAf to 40S and 80S initiation complexes; (c) incorporation of labeled initiation methionine into high‐molecular‐weight proteins; and (d) the association of labeled exogenous mRNA with polysomes. The optimum conditions for amino acid incorporation in this system were 4 mM‐Mg2+, 140 mM‐K+, and pH 7.55. Incorporation was dependent on the addition of ATP, GTP, and an energy‐generating system. Cell‐free protein synthesis reflected the normal process, since a similar spectrum of proteins was synthesized in vitro and in vivo. This initiating cell‐free translation system should have wide application in the analysis of the mechanisms whereby various treatments affect protein synthesis in the brain.
Abnormality of Long‐Chain Fatty Acids in Erythrocyte Membrane Sphingomyelin from Patients with Adrenoleukodystrophy
Authors: Tsuji Shoji , Suzuki Minoru , Ariga Toshio , Sekine Michiko , Kuriyama Masaru , Miyatake Tadashi
Published: October 2006

Abstract: We have devised an analytical method for the determination of fatty acid composition of erythrocyte membrane sphingomyelin by chemical ionization mass spectrometry combined with capillary column gas‐liquid chromatography. Fatty acid composition of erythrocyte membrane sphingomyelin from 8 patients with adrenoleukodystrophy (ALD) and 16 healthy controls were examined by this method. The ratio of hexacosanoic acid (C26.0) to docosanoic acid (C22:0) in erythrocyte membrane sphingomyelin from ALD patients was 2.6‐fold higher than that of the controls. This result suggests that biochemical diagnosis of ALD is possible by the analysis of fatty acid composition of erythrocyte membrane sphingomyelin. Furthermore, it demonstrates that biochemical abnormality in ALD is the generalized abnormal metabolism of very long‐chain saturated fatty acids.
Biosynthesis of Prostacyclin in Rat Cerebral Microvessels and the Choroid Plexus
Authors: Goehlert Uwe G. , Kin N. M. K. Ng Ying , Wolfe Leonhard S.
Published: October 2006

Abstract: Microvessels, predominantly capillaries, were isolated from rat cerebrum by a modification of published procedures. The morphology and purity of the preparations were monitored by light and electron microscopy and by enrichment in alkaline phosphatase, γ‐glutamyl transpeptidase, and prostacyclin synthetase. A reversed‐phase high‐pressure liquid chromatographic method was used in the purification of prostaglandins after extraction from aqueous incubation solutions. Prostacyclin synthesis in brain is localized in cerebral blood vessels and capillaries. The endogenous biosynthetic capacity of the isolated cerebral capillary fractions for prostacyclin, measured as its chemically stable breakdown product, 6‐keto‐prostaglandin F1α, was 11 ng/mg protein/10 min. Choroid plexus and intact surface vessels synthesized 6‐keto‐prostaglandin F1α at 37 and 35 ng/mg protein/10 min, respectively. The prostacyclin‐synthesizing enzyme of the cerebral capillaries also converted the exogenously added prostaglandin endoperoxides to 6‐keto‐prostaglandin F1α. Comparison of the synthesis of prostaglandins 6‐keto‐F1α, E2, and F2α showed that 6‐keto‐prostaglandin F1α was the major prostaglandin formed in the microvessels, in the larger surface vessels, and in the choroid plexus. Prostaglandin D2 was not detected. Prostacyclin synthesis by the cerebral vasculature is similar to that in other blood vessels and cultured human endothelial cells. Possible physiological roles of prostacyclin in the cerebral microvasculature are discussed with special regard to the autoregulation of cerebral blood flow.
Radioimmunoassay for Central Nervous System Myelin‐Specific Proteolipid Protein
Authors: Trotter John L. , Lieberman Lawrence , Margolis Frank L. , Agrawal Harish C.
Published: October 2006

Abstract: A double‐antibody radioimmunoassay (RIA) has been developed with antisera to purified rat brain myelin proteolipid protein (PLP). The addition of Triton X‐100 allowed antibody‐antigen interaction and immune precipitation in the presence of sodium dodecyl sulfate (SDS). The RIA will accurately measure 8‐80 ng of PLP in buffer or human serum. The RIA is highly specific for myelin PLP and does not cross‐react with material in tissues (heart, kidney, muscle, testicle, and intestine) other than the central nervous system. The antibodies to rat myelin PLP cross‐react with PLP from bovine brain homogenate or myelin. Myelin PLP was found to account for 55 and 52% of total myelin protein from bovine and rat brain, respectively. Furthermore, there is a higher concentration of PLP in white than in gray matter corresponding to the degree of myelination. Unlike myelin basic protein, myelin PLP was undetectable in both bovine and rat peripheral nervous system.
Quantitative Changes in Myelin Proteins in a Peripheral Neuropathy Caused by Tullidora (Karwinskia humboldtiana)
Authors: Aoki Kazuko , Muñoz‐Martinez E. J.
Published: October 2006

Peripheral nerve demyelination was induced in cats by oral administration of ether extracts of Tullidora (Karwinskia humboldtiana). Proteins from several hindlimb nerves, spinal roots, and dorsal columns of the spinal cord were subjected to slab gel electrophoresis and quantified by densitometry. In Tullidora‐treated cats with severe motor disturbances, specific myelin proteins were reduced by at least 50% in motor nerves and less than 25% in cutaneous axons. There was a greater decrease of these proteins in the distal than in the cephalad segments of the sciatic nerve; no changes were detected either in the spinal roots or in the white matter of the spinal cord. Electron microscopy revealed intense demyelination in the motor nerves only. Both the density of the 100 Å‐thick neurofilaments and the relative proportion of a polypeptide with a molecular weight of 68,000 were considerably increased in the affected nerves. It is tentatively concluded that the active principles of Tullidora may enter the axons through the motor nerve terminals. The distal segments of the motor nerves would then be preferentially affected and demyelination could result from axonal damage.
Noradrenaline, a Transmitter Candidate in the Retina
Authors: Osborne N. N.
Published: October 2006

The occurrence, metabolism, uptake, and release of noradrenaline were studied in the bovine retina with the following results. (1) Small amounts of noradrenaline occur in the retina and are restricted to the area corresponding to the inner nuclear and plexiform layers. (2) Retinal tissue can metabolise [14C]dopamine to form quantities of [14C]noradrenaline. (3) [14C]Noradrenaline can also be partly metabolised to form [14C]normetanephrine. (4) When bovine retinas were incubated with 5 × 10‐7 M‐[3H]noradrenaline for 20 min and processed for autoradiography, most of the label was associated with apparent nerve processes in the inner plexiform layer. Biochemical analysis showed that more than 95% of the label was noradrenaline. (5) [14C]Noradrenaline uptake saturated with increasing noradrenaline concentrations and followed Michaelis‐Menten kinetics. This uptake could be accounted for by two processes, a high‐affinity system with a Km1 of 5 × 10‐8 M and a Vmax1 of 0.193 pmol/mg/10 min and a low‐affinity system with a Km2 of 6.3 × 10‐5 M and a Vmax2 of 0.109 nmol/mg/10 min. (6) Noradrenaline uptake was strongly dependent on temperature and sodium, less dependent on potassium, and independent of calcium and magnesium ions. (7) Centrally acting drugs, such as desipramine, imipramine, desmethylimipramine, and amitriptyline, inhibited noradrenaline uptake by more than 55% at the concentration of 5 × 10‐5 M. These drugs at the same concentration diminished dopamine uptake by less than 30%. (8) Noradrenaline uptake is stereospecific, the (‐) isomer having a greater affinity for the uptake sites than the (+) isomer. (9) [14C]Noradrenaline in the retina could be released by increasing the external potassium concentration. This release was calcium‐dependent and was blocked by 20 mM‐cobalt chloride. The present studies could be interpreted as supporting the idea that noradrenaline acts as a transmitter in the retina.
The Metabolism of Purine and Pyrimidine Nucleotides in Rat Cortex During Insulin‐Induced Hypoglycemia and Recovery
Authors: Chapman Astrid G. , Westerberg Eva , Siesjö Bo K.
Published: October 2006

Brains of paralysed rats with insulin‐induced hypoglycemia were frozen in situ after spontaneous EEG activity had been absent for 5 or 15 min (“coma”). Recovery (30 min) was achieved in a different group of rats by administering glucose after a 30‐min coma period. Purine and pyrimidine nucleotides, nucleosides and free bases were determined in the cortical extracts by high pressure liquid chromatography (HPLC). The ATP values obtained with the HPLC method were in excellent agreement with those obtained using standard enzymatic/fluorometric techniques, while values for ADP and AMP obtained with the HPLC method were significantly lower. Comatose animals showed a severe (40‐80%) reduction in the concentrations of all nucleoside triphosphates (ATP. GTP, UTP and CTP) and a simultaneous increase in the concentrations of all nucleoside di‐ and monophosphates, including that of IMP. The adenine nucleotide pool size decreased to 50% of control level. The concentrations of the nucleosides adenosine, inosine, and uridine increased 50‐ to 250‐fold, while the concentrations of the purine bases, xanthine and hypoxanthine, rose 2‐ and 30‐fold, respectively. There were no increases in the concentrations of adenine, guanine, or xanthosine. Following glucose administration there was a partial (ATP, UTP and CTP) or almost complete (GTP) recovery of the nucleoside triphosphate levels. During recovery, the levels of nucleosidc di‐ and monophosphates and of adenosine decreased to values close to control; the rise in the inosine level was only partially reversed, and the concentrations of hypoxanthine and xanthine rose further. The adenine nucleotide pool size was only partially restored (to 67% of control value). The adenine nucleotide pool size was not increased by i.p. injection of adenosine or adenine under control condition, or during the posthypoglycemic recovery period.
Regional CNS Levels of Acetylcholine and Choline During Hypoglycemic Stupor and Recovery
Authors: Gorell Jay M. , Navarro Carolyn P. , Schwendner Susan P. W.
Published: October 2006

During insulin stupor in mice, acetylcholine levels in cerebral cortex, cerebellum. brainstem, striatum, and hippocampus were unchanged from control values despite brain glucose concentrations 3‐10% of normal, whereas choline levels rose 2.4‐3.6‐fold in all five CNS regions. Brain acetylcholine and choline levels did not change during recovery following glucose injection. The data suggest that. in hypoglycemic stupor, (1) overall rates of acetylcholine synthesis and degradation remain balanced within each of the CNS regions studied: (2) the biochemical mechanism that elevates brain choline levels is unlikely to be related only to cholinergic synaptic processes: and (3) brain choline levels need not rise for stupor to occur.
Cerebral Protective Effect and Radical Scavenging Action
Authors: Yasuda Hiroshi , Shimada Osafumi , Nakajima Akira , Asano Takao
Published: October 2006

Abstract The role of radical scavenging action in cerebral protective effect of drugs was investigated in vitro. Incubation of rat brain mitochondrial suspension with ascorbic acid and Fe2+ resulted in the formation of malondialdehyde and a decrease in the turbidity of the suspension, indicating that the mitochondria were peroxidatively disintegrated. Nizofenone at 10 μm or more inhibited the peroxidative disintegration of mitochondria, and complete inhibition was observed at 100–200 μm. The action of nizofenone was also ascertained by experiments with rat liver mitochondria. The anti‐peroxidative activity of nizofenone was estimated to be approximately equivalent to that of α‐tocopherol, and this property was unique. Among the cerebral protective drugs tested, thiopental was only slightly efficient, and pentobarbital, phenobarbital, and dimethyl sulfoxide had no effect. In addition, nizofenone was found to scavenge a stable free radical, diphenyl‐p‐picrylhydrazyl, but the barbiturates did not. These findings suggest that there is no intimate relationship between cerebral protective effect and free radical scavenging action.
Platelet Glutamate and Aspartate Uptake in Huntington's Disease
Authors: Mangano Richard M. , Schwarcz Robert
Published: October 2006

Abstract A possible involvement of amino acid uptake mechanisms in the etiology of the human neurodegenerative disease, Huntington's disease (HD), was investigated. Measurement of glutamate (Glu) and aspartate (Asp) uptake was performed in blood platelets, which have previously been shown to constitute a peripheral model system for central amino acid uptake processes. Analyses of Glu and Asp accumulation at 10−7 M and kinetic examination of the high affinity site for Glu indicate no significant differences between control and HD platelets. A genetically determined defect in amino acid uptake therefore does not seem to underlie the nerve cell loss observed in HD patients.
Comparison of the Binding of [3H]Spiperone and [3H]Domperidone in Homogenates of Mammalian Retina and Caudate Nucleus
Authors: Watling Keith J. , Iversen Leslie L.
Published: October 2006

Abstract— The specific binding of [3H]spiperone and [3H]domperidone, as defined by 1 μm‐(+)butaclamol, was compared in homogenates of bovine retina and caudate nucleus. Scatchard analyses of saturation data for [3H]spiperone binding yielded dissociation constants (Kd) of 0.35 nm in the retina and 0.64 nm in the caudate nucleus. Comparison of the maximum number of binding sites (Bmax) present in each tissue indicated that the density of sites in bovine caudate nucleus (270 fmol/mg protein) was approximately three times higher than in bovine retina (92 fmol/mg protein). This difference was even more marked in guinea pig tissues, with a ratio of 7:1 between corpus striatum and retina. The pharmacological analysis of [3H]spiperone binding in both the bovine retina and caudate nucleus indicated an interaction with dopaminergic rather than serotonergic sites. However, inhibition curves obtained to dopaminergic agonists in the bovine retina were significantly steeper than those observed in the bovine caudate nucleus, as reflected in the greater Hill coefficients obtained for these agents in the retina. Furthermore, only a small amount of specific [3H]domperidone binding was observed in either the bovine caudate nucleus or the guinea pig striatum, whilst no specific [3H]domperidone binding was detectable in homogenates of either bovine or guinea pig retina. These data suggest that the retina possesses only a small population of dopaminergic D2 sites and that these binding sites may differ from those present in the caudate nucleus.
Changes in Chromatin Structure Associated with Alzheimer's Disease
Authors: Lewis P. N. , Lukiw W. J. , De Boni U. , Crapper McLachlan D. R.
Published: October 2006

Abstract— The enzyme micrococcal nuclease was used to examine the accessibility of chromatin extracted from brains of 13 patients with senile and presenile dementia of the Alzheimer type. Compared with chromatin extracted from brains of 8 patients without neurological signs or brain pathology and brains of 7 patients with nonAlzheimer dementia, Alzheimer chromatin was less accessible to this enzyme‐. Reduced accessibility was reflected by a reduced yield of mononucleosomes in comparison with dinucleosomes and larger oligomers. Both neuronal and glial chromatin were found to be similarly affected. The reduced yield of mononucleosomes from Alzheimer chromatin is not due to their increased breakdown, but is probably related to protein associated with the internucleosomal linker region that retards nuclease action. Dinucleosomes isolated from control and Alzheimer nuclease digests were examined for their protein complement. Three perchloric acid‐soluble proteins situated in the histone HI region of sodium dodecyl sulfate (SDS) gels were present in elevated levels in Alzheimer dinucleosomes. These results represent the first example of altered chromosomal proteins associated with a diseased state of the brain.
Metabolic Changes in Cerebral Cortex, Hippocampus, and Cerebellum During Sustained Bicuculline‐Induced Seizures
Authors: Folbergrová Jaroslava , Ingvar Martin , Siesjö Bo K.
Published: October 2006

Abstract— The objective of the present experiments was to study metabolic correlates to the localization of neuronal lesions during sustained seizures. To that end, status epilepticus was induced by i.v. administration of bicuculline in immobilized and artificially ventilated rats, since this model is known to cause neuronal cell damage in cerebral cortex and hippocampus but not in the cerebellum. After 20 or 120 min of continuous seizure activity, brain tissue was frozen in situ through the skull bone, and samples of cerebral cortex, hippocampus, and cerebellum were collected for analysis of glycolytic metabolites, phosphocreatine (PCr), ATP, ADP, AMP, and cyclic nucleotides. After 20 min of seizure activity, the two “vulnerable” structures (cerebral cortex and hippocampus) and the “resistant” one (cerebellum) showed similar changes in cerebral metabolic state, characterized by decreased tissue concentrations of PCr, ATP, and glycogen, and increased lactate concentrations and lactate/ pyruvate ratios. In all structures, though, the adenylate energy charge remained close to control. At the end of a 2‐h period of status epilepticus, a clear deterioration of the energy state was observed in the cerebral cortex and the hippocampus, but not in the cerebellum. The reduction in adenylate energy charge in the cortex and hippocampus was associated with a seemingly paradoxical decrease in tissue lactate levels and with failure of glycogen resynthesis (cerebral cortex). Experiments with infusion of glucose during the second hour of a 2‐h period of status epilepticus verified that the deterioration of tissue energy state was partly due to reduced substrate supply; however, even in animals with adequate tissue glucose concentrations, the energy charge of the two structures was significantly lowered. The cyclic nucleotides (cAMP and cGMP) behaved differently. Thus, whereas cAMP concentrations were either close to control (hippocampus and cerebellum) or moderately increased (cerebral cortex), the cGMP concentrations remained markedly elevated throughout the seizure period, the largest change being observed in the cerebellum. It is concluded that although the localization of neuronal damage and perturbation of cerebral energy state seem to correlate, the results cannot be taken as. evidence that cellular energy failure is the cause of the damage. Thus, it appears equally probable that the pathologically enhanced neuronal activity (and metabolic rate) underlies both the cell damage and the perturbed metabolic state. The observed changes in cyclic nucleotides do not appear to bear a causal relationship to the mechanisms of damage.
Barbiturate Attenuation of Brain Free Fatty Acid Liberation During Global Ischemia
Authors: Shiu Gerald K. , Nemoto Edwin M.
Published: October 2006

Abstract: To find a biochemical basis for the increased tolerance of the brain to anoxia during barbiturate anesthesia, we studied whole‐brain free fatty acids (FFA) at various times after decapitation of awake and pentobarbital‐anesthetized rats. Post‐decapitation, the brains were kept at 37°C for 1 to 60 min before freezing in liquid N2. Nonischemic brains were frozen in liquid N2, using a rapid sampling technique. Whole‐brain arachidonic, stearic, oleic, linoleic, and palmitic acids were quantitated by gas‐liquid chromatography. In unanesthetized, nonischemic brain, total FFA was 1226 ± 121 nmol/g brain (n= 12) and was unaffected by pentobarbital anesthesia (1126 ± 86 nmol/g brain, n= 11), except for a reduction in arachidonic acid. Total FFA in unanesthetized and pentobarbital‐anesthetized rats transiently declined between 0 and 1 min of ischemia, and then rose linearly for up to 60 min, with consistently lower values in pentobarbital‐treated rats, the greatest attenuation being that of arachidonic and stearic acid liberation. Brain FFA liberation during global ischemia is the first known biochemical variable directly correlated with the duration (i.e., severity) of global ischemia. The attenuation of brain FFA liberation and especially of arachidonic and stearic acids may be the biochemical basis of barbiturate attenuation of ischemic brain injury.
Phosphorylation‐Mediated Changes in Pyruvate Dehydrogenase Activity Influence Pyruvate‐Supported Calcium Accumulation by Brain Mitochondria
Authors: Browning M. , Baudry M. , Bennett W. F. , Lynch G.
Published: October 2006

Abstract: Changes in the activity of pyruvate dehydrogenase [pyruvate: lipoamide oxidoreductase (decarboxylating and acceptor‐acetylating), EC 1.2.4.1, PDH], elicited by inhibition of the phosphorylation of its 40,000 Mrα‐subunit, were compared with changes in pyruvate‐supported calcium accumulation by rat brain mitochondria. Dichloroacetate (DCA) produces concentration‐dependent inhibition of the phosphorylation of intramitochondrial PDH a‐subunit, which is accompanied by stimulation of PDH activity and calcium accumulation. DCA did not affect succinate‐ or ATP‐supported mitochondrial calcium accumulation. The concentration of DCA giving half‐maximal inhibition of the phosphorylation was almost identical to that giving half‐maximal stimulation of PDH activity and calcium accumulation. PDH activity and pyruvate‐supported calcium accumulation showed similar dependence on pyruvate concentration with respective apparent affinities for pyruvate of 40 μm and 30 μm, and both activities exhibited positive cooperativity. DCA modified only the maximal activity of PDH or the maximal calcium accumulation without changing either the apparent affinities for pyruvate or calcium or the Hill coefficients. These data provide evidence that calcium accumulation by mitochondria is tightly linked to PDH activity and that changes in the phosphorylation of the PDH α‐subunit can be reflected in changes in the calcium‐buffering ability of mitochondria. This suggests a possible mechanism by which a variety of manipulations, such as repetitive synaptic stimulation, can alter the regulation of internal calcium levels.
Glucocorticoids Increase Tyrosine Hydroxylase Activity in Cultured Murine Neuroblastoma
Authors: Williams Lawrence R. , Sandquist Dean , Black Asa C. , Williams Terence H.
Published: October 2006

Abstract: Cyclic AMP and glucocorticoids appear to have a role in regulating the activity of tyrosine hydroxylase (TH), as well as the expression of “morphological differentiation” in murine neuroblastoma. Monolayer cultures of C‐1300 murine neuroblastoma (clone NBP2) were treated with the following compounds in ethanol: dexamethasone, triamcinolone acetonide, hydrocortisone, cortexolone, androstenedione, testosterone, estradiol‐17β; or with the phosphodiesterase inhibitor Ro20–1724 [4‐(3‐butoxy‐4‐methoxybenzyl)‐2‐imidazolidinone]. Treatment with either 200 μg/ml Ro20–1724 or 50 μg/ml dexamethasone produced significant increases in TH activity compared to alcohol controls (1.44 vs. 0.82 nmol 14CO2/mg protein/hr compared to 0.095). Triamcinolone acetonide or hydrocortisone also produced smaller, but significant, increases in TH activity compared to dexamethasone. When steroid activities were compared at 25 μm concentration and after 60 min of incubation (to maximize TH activity), triamcinolone acetonide was not as effective (62%) as dexamethasone. The relatively inactive glucocorticoid cortexolone produced a slight but significant increase, while the androgens androstenedione and testosterone and the estrogen estradiol‐17β were without effect.
Circadian Rhythm in Pineal Methionine S‐Adenosyltransferase
Authors: Guchhait Ras B. , Monjan Andrew A.
Published: October 2006

Abstract: The circadian rhythm of methionine S‐adenosyltransferase, which catalyzes the formation of S‐adenosylmethionine, a cosubstrate for melatonin in the pineal gland, follows the pattern of hydroxyindole‐O‐methyltransferase. Around the middle of the dark period, methionine S‐adenosyltransferase and hydroxyindole‐O‐methyltransferase appear to be elevated by 2.5‐ and 1.5‐fold, respectively, and tend to fall back during the light period.
Transsynaptic Regulation of Olfactory Bulb Catecholamines in Mice and Rats
Authors: Kawano T. , Margolis F. L.
Published: October 2006

Abstract: Norepinephrine (NE), dopamine (DA), 3,4‐dihydroxyphenylalanine (DOPA), and 3,4‐dihydroxyphenylacetic acid (DOPAC) were measured simultaneously by high performance liquid chromatography with electrochemical detection in extracts of olfactory bulbs at various intervals after chemical or surgical deafferentation. Chemical deafferentation of mice by intranasal irrigation with Triton X‐100 or of rats by olfactory axotomy resulted in a rapid progressive decline of DA and DOPAC and an associated rise in NE in the olfactory bulb. However, after several weeks, these values returned to pre‐lesion levels concomitant with reinnervation of the bulb by the afferent neurons. In contrast, deafferentation by procedures known to prevent reinnervation of the bulb by the afferent chemoreceptor neurons (i.e., a ZnSO4 solution in mice or a surgical procedure in rats) completely blocked the return to pre‐lesion values of DA, DOPAC, and NE. The specificity of these effects was demonstrated by the inability of intranasal administration of the neurotoxin 6‐hydroxydopamine to alter DA levels, resulting instead in a significant decline in olfactory bulb NE content. These data demonstrate that the DA content of the olfactory bulb can be influenced by either chemical or surgical modulation of the afferent pathway in two different species. This offers additional support for our hypothesis of transsynaptic regulation of intrinsic DA neurons of the bulb by the afferent olfactory chemoreceptor neurons.
Cerebral Metabolic Responses to Electroconvulsive Shock and Their Modification by Hypercapnia
Authors: Miller Alexander L. , Shamban Ava T. , Corddry David H. , Kiney Colleen A.
Published: October 2006

Abstract: Brain glucose metabolism was studied in paralyzed, ventilated rats given electroconvulsive shock (ECS) under normocapnic and hypercapnic conditions. Brains were obtained with a freeze‐blowing apparatus. Rates of glucose utilization were determined with [2‐14C]glucose and[3H]deoxyglucose as tracers. In normocapnic rats, ECS caused a large increase in the rate of glycolysis to 5–6 μmol/g/min. Brain lactate levels increased three‐ to fourfold. The stimulation of glucose metabolism was reflected in decreased brain glucose 6‐phosphate concentration as early as 2–3 s after ECS. There were significant decreases in brain glucose and glycogen levels at 20 and 30 s after ECS. The decreases in endogenous brain glucose accounted for most of the increases in glucose utilization measured isotopically, implying that influx of glucose from blood into brain did not increase greatly over these time periods. Animals made hypercapnic by respiration with 10% CO2 for 2 min prior to ECS were different in their metabolic responses to ECS in several ways. The increases in glycolyt‐ic rate and lactate content of brain were half of those found in normocapnic rats. Brain glycogen and glucose concentrations did not change significantly in the hypercapnic rats during seizure activity. Thus, hypercapnia lessened the stimulation of glycolysis caused by ECS, but increased net influx of glucose from blood to brain. The mechanisms of these effects of hypercapnia are uncertain, but it is postulated that the effect on glycolytic activity is due to the acidosis and that the effect on glucose transport is due to an increase in capillary surface area.
Evidence for Two Distinct Forms of Fatty Acid Cyclooxygenase in Brain
Authors: Lysz Thomas W. , Needleman Philip
Published: October 2006

Abstract: The enzymatic metabolism of [14C]arachidonic acid (AA) was studied with microsomes prepared from rabbit medulla. Prostaglandin E2 (PGE2) levels, measured either by radiochemistry or radioimmunoassay, rose rapidly and abruptly plateaued within 5 min, while prostaglandin F2a (PGF2a) levels continued to rise for 30 min. The rapid termination of PGE2 biosynthesis was not the result of limited cofactor, substrate, or product feedback inhibition, nor was it due to PGE2‐9‐ketoreductase activity. Inhibition of the PGH2→ PGE2 isomerase by arachidonic acid or its metabolites could not explain the abrupt halt in PGE2 biosynthesis. Proof for two separate cyclooxygenases comes from our observation that a preincubation of the brain microsomes with unlabeled AA eliminated PGE2 biosynthesis while PGF2o production continued. Further evidence to suggest two cyclooxygenases in brain is derived from the observation that indomethacin inhibited PGE2 production at concentrations that did not affect PGF2a biosynthesis. These results suggest that one fatty acid cyclooxygenase is closely associated with PGH2→ PGE2 isomerase and readily undergoes autodestruction and the second cyclooxygenase is associated with a PGH2→ PGF2a reductase and is somewhat resistant to arachidonate‐induced destruction and to nonsteroidal antiinflammatory agents.
No Effect of Hyperketonemia on Local Cerebral Glucose Utilization in Conscious Rats
Authors: Corddry David H. , Rapoport Stanley I. , London Edythe D.
Published: October 2006

Abstract: Local cerebral glucose utilization was measured by the [14C]2‐deoxy‐d‐glucose method in conscious control and hyperketonemic rats. Hyperketonemia was induced by 3 days of starvation or by infusion of 3‐ hydroxybutyrate in fed rats. These treatments produced combined blood ketone body concentrations (acetoacetate + 3‐hydroxybutyrate) of from 1.2 to 2.4 mM. Neither treatment significantly affected glucose utilization in any of the 15 brain regions studied. These observations indicate that hyperketonemia in resting, conscious rats does not interfere with brain uptake and phosphorylation of glucose.
Regional Prostaglandin Levels in Cerebral Ischaemia
Authors: Crockard H. Alan , Bhakoo Kishore K. , Lascelles Peter T.
Published: October 2006

Abstract: Regional quantitative studies of brain prostaglandins (PG) in gerbils with bilateral carotid ligation have shown increases in PG2α in all areas of ischaemia but with no increases in PGE2. Pretreatment of animals with indomethacin (3 mg/kg) reduced this response, whereas dexamethasone (2.5 mg/kg) had no such effect. PGE1 and thromboxane‐B2 were found not to be present in our preparation. The significance of these findings lies in the fact that both drugs, which are known to inhibit the development of ischaemic oedema, have now been shown to produce markedly different effects on prostaglandin levels. This may indicate either, that prostaglandins are not involved in is chaemic oedema formation, or, that the drugs have different mechanisms of action.
High‐Affinity Calcium Binding in Rat Synaptosomal Plasma Membranes: Decrease in Young, Genetically Hypertensive Rats
Authors: Nunez Anne‐Marie , Devynck Marie‐Aude , Meyer Philippe
Published: October 2006

Abstract: A large number (about 4‐5 nmoVmg of protein) of high‐affinity (apparent dissociation constant at 37°C: KD37°C= 5 × 10−8M) calcium binding sites was characterized in synaptosomal membrane fractions enriched in plasma membranes that were isolated from rat brain. These sites were studied simultaneously in membranes from spontaneously hypertensive young rats (SHR) and their normotensive controls. No difference was observed between whole synaptosomes from these two substrains. However, plasma membraneenriched fractions from SHR exhibited a reduced calcium binding capacity without a significant change in affinity. This decrease which averaged 15‐20% was not due to any variation in the accessibility of calcium to its binding sites, as similar results were obtained in the presence of the calcium ionophore A23,R7. The reduction found in calcium binding is very similar to that previously described in erythrocyte membranes. It is envisaged that such an abnormality at nerve endings might play a role in the pathogenesis of genetic hypertension.
Human 14‐3‐3 Protein: Radioimmunoassay, Tissue Distribution, and Cerebrospinal Fluid Levels in Patients with Neurological Disorders
Authors: Boston Paul F. , Jackson Peter , Thompson R. J.
Published: October 2006

Abstract: An antiserum to human 14‐3‐3 protein has been produced in rabbits. The protein was a poor antigen and attempts to improve immunogenicity were unsuccessful. A radioimmunoassay was developed using the antiserum, 125I‐ 14‐3‐3‐2, and unlabelled 14‐3‐3‐2 as standards. The assay had a sensitivity limit of 2.5 ng.m1−1. The minor component of human 14‐3‐3 protein (14‐3‐3‐1 protein) cross‐reacted to approximately 10% in the assay. Human tissues were surveyed for 14‐3‐3 protein by two‐dimensional electrophoresis and by radioimmunoassay. Two‐dimensional electrophoresis showed a 14‐3‐3 protein complex in brain, intestine, and testis, but not in other tissues. Radioimmunoassay showed that although brain had the highest concentration of 14‐3‐3 (13.3 μg. mg−1 soluble protein), immunoreactivity was present in all tissues, with the concentration in intestine and testis approaching 50% of the brain level. Lower levels (less than 1.0 μg. mg−1 soluble protein) were seen in liver, kidney, skeletal muscle, and erythrocytes. The immunoreactivity present in tissues other than brain showed the same molecular weight and charge characteristics as authentic 14‐3‐3 protein. The radioimmunoassay also detected 14‐3‐3 protein in serum (50 ng.m1−1) and in CSF (5‐130 ng.ml−1). The immunoreactivity present in CSF appeared to be intact 14‐3‐3 protein. CSF 14‐3‐3 levels were measured in 82 patients with various neurological disorders. Measurements of this protein did not appear sufficiently discriminating to be o f diagnostic value.
Potassium‐Stimulated Release of Radiolabelled Taurine and Glycine from the Isolated Rat Retina
Authors: Smith L. F. P. , Pycock C. J.
Published: October 2006

Abstract: The release of preloaded [3H]glycine and [3H]taurine in response to a depolarising stimulus (12.5‐50 mM KCl) has been studied in the superfused rat retina. High external potassium concentration immediately increased the spontaneous efflux of [3H]glycine, the effect of 50 mM K+ apparently being abolished by omitting calcium from the superfusing medium. In contrast, although high potassium concentrations increased the spontaneous emux of [3H]taurine from the superfused rat retina, this release was not evident until the depolarising stimulus was removed from the superfusing medium. The magnitude of this “late” release of [3H]taurine was dependent on external K+ concentrations, and appeared immediately after cessation of the stimulus irrespective of whether it was applied for 4, 8, or 12 min. Potassium (50 mM)‐induced release of taurine appeared partially calcium‐dependent, being significantly reduced (p < 0.01) but not abolished by replacing calcium with 1 mM EDTA in the superfusate. High‐affinity uptake systems for both [3H]glycine and [3H]taurine were demonstrated in the rat retina in vitro (Km values, 1.67 μM and 2.97 μM; Vmax values, 19.3 and 23.1 nmol/g wet weight tissue/h, respectively). The results are discussed with respect to the possible neuro‐transmitter roles of both amino acids in the rat retina.
Rapid Cellular Regulation of D‐Glucose Transport in Cultured Neural Cells
Authors: Lange Klaus , Keller Konrad , Ludwig Wolf‐Dieter , Monden Ingrid , Reinsch Ingeborg , Brandt Ursula
Published: October 2006

Abstract: Previous studies have revealed two different kinds of regulation of glucose utilization in cell lines derived from the nervous system (Keller et al., 1981). We found glucose metabolism of C‐6 glioma cells to be limited and regulated by membrane transport. In contrast, glucose utilization of C‐1300 neuroblastoma (N2A) cells was limited by the known regulatory enzymes of the Embden‐Meyerhof pathway. Under the given experimental conditions the “membrane‐limited” C‐6 glioma cells were characterized by periodically changing glucose transport rates and very low intracellular glucose concentrations, which remained constant in spite of widely differing transport rates. These findings suggest the close functional coupling between transport and phosphorylation required for the regulation of glucose transport by cellular metabolic needs.
Exogenous Choline Enhances the Synthesis of Acetylcholine Only Under Conditions of Increased Cholinergic Neuronal Activity
Authors: Trommer Brad A. , Schmidt Dennis E. , Wecker Lynn
Published: October 2006

Abstract: The effect of choline (60 mg/kg, i.p.) on fluphenazine‐ and pentylenetetrazol‐induced alterations in the concentration of acetylcholine (ACh) and/or the rate of sodium‐dependent high‐affinity choline uptake (HACU) in rat striatum and hippocampus was studied. Systemic administration of the dopamine receptor blocking agent fluphenazine hydrochloride (0.5 mg/kg, i.p.) decreased the concentration of ACh in the striatum; this effect was prevented by the prior administration of choline. The central nervous system stimulant pentylenetetrazol (30 mg/kg, i.p.) reduced the concentration of ACh in both striatum and hippocampus and increased the velocity of HACU in the hippocampus. Pretreatment with choline totally prevented the depletion of ACh induced by pentylenetetrazol in the striatum. In the hippocampus, prior administration of choline prevented the pentylenetetrazol‐induced increase in the rate of HACU and attenuated the effect of pentylenetetrazol on the levels of ACh. Results indicate that the acute administration of choline antagonizes pharmacologically induced alterations in cholinergic activity as assessed by the rate of HACU and the steady‐state concentration of ACh. Furthermore, data support the hypothesis that the administration of choline increases the ability of central cholinergic neurons to synthesize ACh under conditions of increased neuronal activity.
A Search for Discrete Cholinergic Nuclei in the Human Ventral Forebrain
Authors: Davies Peter , Feisullin Sophia
Published: October 2006

Abstract: Slices cut from five frozen human brains were dissected into 2‐mm cubes and assayed for choline acetyltransferase (ChAT) activity and protein content. A pattern of enrichment of ChAT activity was found ventral to the anterior commissure; this finding is consistent with the location of the enzyme in the cells of the nucleus basalis of Meynert. The region beneath the anterior commissure was the only place a discrete enrichment of activity could be found, and the precise topography of the enrichment was somewhat variable from brain to brain. The results are discussed in the light of recent knowledge concerning the source of the cortical cholinergic innervation.
Different Forms of Adrenal Phenylethanolamine N‐Methyltransferase: Species‐Specific Posttranslational Modification
Authors: Park Dong H. , Baetge E. Edward , Kaplan Barry B. , Albert Vivian R. , Reis Donald J. , Joh Tong H.
Published: October 2006

Abstract: Phenylethanolamine N‐methyltransferase was purified from rat and cow adrenal glands. The enzymes from the two species have the same molecular weight of 31,000, but differ in electrophoretic mobility. During polyacrylamide gel electrophoresis, the rat form migrates faster than the bovine form. Antibodies to bovine enzyme precipitated equally well the rat and cow form of the enzyme, but antibodies against rat enzyme precipitated poorly the bovine form. In contrast, both antibodies recognized a similar protein in the in vitro translation products of poly(A+)mRNA isolated from cow adrenal glands. The results suggest that the primary protein structure of rat and bovine enzyme is similar and that differences in electrophoretic mobility are due to posttranslational modification of the enzyme molecule.
Calcium Uptake and Catecholamine Secretion by Cultured Bovine Adrenal Medulla Cells
Authors: Kilpatrick Daniel L. , Slepetis Ruta J. , Corcoran James J. , Kirshner Norman
Published: October 2006

Abstract: The uptake of 45Ca2+ and secretion of catecholamines by primary cultures of adrenal medulla cells were studied. Nicotine, veratridine, potassium, and Ionomycin stimulate both the accumulation of 45Ca2+ and the secretion of catecholamines. Nicotinic antagonists block 45Ca2+ uptake induced by nicotine, tetrodotoxin blocks 45Ca2+ uptake induced by veratridine, and D600 blocks uptake induced by K+, nicotine, and veratridine, but not 45Ca2+ uptake or secretion induced by Ionomycin. The EC50 for nicotine is 3 μm for catecholamine secretion and 10 μm for 45Ca2+ uptake, while the EC50S for veratridinestimulated uptake and secretion are approximately the same (75 μm). Kinetic studies show that the uptake of Ca2+ is rapid and appears to precede the secretion of catecholamines, and that the rate of uptake declines rapidly. The uptake of 45Ca2+ and secretion of catecholamines stimulated by veratridine and 50 mm‐K+ show saturation kinetics with respect to external calcium concentrations at about 2 mm. On the other hand, the uptake of 45 Ca2+ stimulated by nicotine does not become saturated at external calcium concentrations of 10 mm although the secretion of catecholamines reaches a maximum at external calcium concentrations of 2 mm. The data suggest that depolarizing agents such as veratridine and 50 mm‐K+ stimulate 45Ca2+ entry through voltage‐sensitive calcium channels, while nicotinic agonists stimulate calcium entry through the acetylcholine receptor ion channels as well as through voltage‐sensitive calcium channels.