bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantitation of Rat Dopamine Transporter mRNA: Effects of Cocaine Treatment and Withdrawal
Authors: Xia Yue , Goebel Dennis J. , Kapatos Gregory , Bannon Michael J.
Published: October 2006

Abstract: Dopamine transporter mRNA levels in the rat substantia nigra were quantified using a sensitive nuclease protection assay with a highly homologous human dopamine transporter cDNA clone. The same probe was also used to visualize dopamine transporter mRNA in the substantia nigra by in situ hybridization. Repeated cocaine administration (15 mg/kg, twice a day for 6.5 days) resulted in a >40% decrease in nigral dopamine transporter mRNA levels. In contrast, dopamine transporter mRNA levels were unchanged after either acute treatment (4 h before death) or repeated cocaine treatment followed by a 72‐h withdrawal period. Thus, blockade of the dopamine transporter by repeated cocaine administration may result in the down‐regulation of dopamine transporter gene expression in dopamine neurons.
Correlation Between Electroencephalogram Isoelectric Time and Hippocampal Norepinephrine Levels, Measured by Microdialysis, During Ischemia in Rats
Authors: Perego C. , Gatti S. , Vetrugno G. C. , Marzatico F. , Algeri S.
Published: October 2006

Abstract: It is suggested that norepinephrine (NE) plays a role during transient forebrain ischemia. NE may have a protective action against neuronal cell death in the hippocampus, or it may be one of the causes of injurious ischemic effects. We used the microdialysis technique to study extracellular NE levels in the rat hippocampus before, during, and after 30 min of transient incomplete forebrain ischemia (induced by four‐vessel occlusion) to describe the time course of NE in this condition. There was a maximal increase (fivefold) in extracellular NE after 10 min of reflow only when the electroencephalogram was isoelectric. NE levels returned to baseline 40 min after release of the carotid clamps and remained constant for the next 80 min. Thus there appears to be a transient NE overflow in the hippocampus during ischemia, closely related to the complete loss of brain electrical activity.
Chromatographic Resolution and Characterization of a Nerve Growth Factor‐Dependent Kinase That Phosphorylates Microtubule‐Associated Proteins 1 and 2 in PC12 Cells
Authors: Sano Mamoru
Published: October 2006

Abstract: When the supernatant fractions from extracts of control and nerve growth factor (NGF)‐ or dibutyryl cyclic AMP‐treated PC12D cells were applied to DEAE‐Sepharose columns and proteins were eluted with a gradient of NaCl, three separate peaks of kinase activity that phosphorylated microtubule‐associated proteins (MAPs) were recovered. Enhancement of the kinase activity in peak 1 was noted in the case of dibutyryl cyclic AMP‐treated cells. In contrast, the kinase activity in the third peak was markedly elevated, in terms of the ability to phosphorylate MAP1 and MAP2, in the case of the extract from NGF‐treated cells. This activity was designated previously as NGF‐dependent MAP kinase. The apparent molecular mass of the active kinase was 45–50 kDa. The apparent Km value was 35 μM for ATP with either MAP1 or MAP2 as substrate. When the kinase activity in the fractions from the DEAE‐Sepharose column was assayed in the presence of Mn2+ instead of Mg2+, another NGF‐stimulated kinase activity was detected in the fractions eluted by a lower concentration of NaCl than that which eluted the Mg2+‐activated kinase. Other growth factors, namely, epidermal growth factor and basic fibroblast growth factor, also stimulated the activity of NGF‐dependent MAP kinase. Possible involvement of the kinase in the outgrowth of neurites has been suggested. The NGF‐induced activation of NGF‐dependent MAP kinase was blocked by the presence of K‐252a. In contrast, the activation of NGF‐dependent MAP kinase by basic fibroblast growth factor and by epidermal growth factor was not blocked, but actually stimulated by K‐252a, a result that correlates well with the analogous actions of the drug on the outgrowth of neurites that is induced by these growth factors. The latter observation strengthens the possibility of a close relationship between the outgrowth of neurites and the activation of NGF‐dependent MAP kinase.
Characterization of the Irreversible Inhibition of High‐Affinity Choline Transport Produced by Hemicholinium Mustard
Authors: Gylys Karen Hoppens , Mellin Charlotte , Amstutz René , Jenden Donald J.
Published: October 2006

Abstract: The inhibition of high‐affinity choline transport by hemicholinium mustard (HCM), an alkylating analogue of hemicholinium‐3, was examined in rat brain synaptosomes and guinea pig myenteric plexus. In synaptosomes, 50% high‐affinity choline transport inhibition occurs with an HCM concentration of 104 nM (4‐min incubation). A 10‐min preincubation with 10 nM HCM results in essentially complete (>95%) inactivation that persists after washing. Low‐affinity choline transport in synaptosomes is unaffected by HCM inhibition at all concentrations examined (1–50 μM). Time course experiments indicate that the maximum irreversible inhibition (58%) seen after a 1‐min preincubation with 500 nM HCM decreases to 46% inhibition after a 15‐min preincubation; however, analysis of variance reveals that this difference is not significant. HCM inhibition of acetylcholine release from myenteric plexus‐longitudinal muscle preparations persists for at least 2 h after removal of drug from the incubation bath; this inactivation can be prevented by coincubation with a high choline concentration during treatment with the mustard. In contrast, inhibition produced by the parent compound hemicholinium‐3 is largely reversed by washing in both preparations: examined. The observed potency and selectivity of HCM suggest its usefulness as a covalent probe for high‐affinity choline transport.
Expression of Nerve Growth Factor and Nerve Growth Factor Receptor Genes in Human Tissues and in Prostatic Adenocarcinoma Cell Lines
Authors: MacGrogan Donal , Saint‐André Jean‐Paul , Dicou Eleni
Published: October 2006

Abstract: Nerve growth factor (NGF) mRNAs were detected and quantified in a variety of normal and neoplastic human tissues by northern blot hybridization. Human heart contained the highest NGF mRNA levels, whereas lower but comparable levels were found in the placenta, prostate, and kidney. All tissues examined coexpressed the low‐affinity NGF receptor (LNGFR), whereas none of these tissues expressed the high‐affinity NGF receptor encoded by the trk protooncogene. The widespread distribution of the LNGFR suggests that it plays a role in the regulation of normal cell growth. No overexpression of NGF or LNGFR mRNA was detected in neoplastic tissues, whereas LNGFR‐like immunoreactivity was localized outside of tumor cells. Transforming growth factor‐α and protooncogene c‐fos expression in these tissues did not show a systematic correlation with NGF/LNGFR expression. Furthermore, regulation of the human NGF gene was studied in DU145 cells, a prostatic adenocarcinoma cell line that synthesizes significant NGF mRNA levels. Serum induced, whereas dexamethasone inhibited, NGF mRNA synthesis in these cells. Serum induction was preceded by a rapid and transient activation of the c‐fos protooncogene.
5,5′‐Dihydroxy‐4,4′‐Bitryptamine: A Potentially Aberrant, Neurotoxic Metabolite of Serotonin
Authors: Wrona Monika Z. , Goyal R. N. , Turk D. J. , Blank C. LeRoy , Dryhurst Glenn
Published: October 2006

Abstract: Previous investigators have detected unknown oxidized forms of 5‐hydroxytryptamine (5‐HT) in the CSF of Alzheimer's disease (AD) patients. Furthermore, an unidentified autoxidation product of this neurotransmitter is an inhibitor of acetylcholinesterase (AChE), an enzyme compromised in the Alzheimer brain. In this study it is demonstrated that the major product of autoxidation of 5‐HT is 5,5′‐dihydroxy‐4,4′‐bitryptamine (DHBT). Central administration of DHBT to mice at a dose of 40 μg (free base) evokes profound behavioral responses, which persist until the animals die (∼24 h). One hour after central administration of DHBT, the levels of norepinephrine, dopamine, 5‐HT, and acetylcholine and their metabolites in whole brain are greatly elevated. Disturbances to the catecholaminergic and serotonergic systems were still evident shortly before the death of animals. DHBT is also shown to be a noncompetitive inhibitor of AChE in vitro. These observations suggest that if DHBT is formed as an aberrant metabolite of 5‐HT in the human brain, it could potentially be neurotoxic and contribute to the neuronal degeneration and other neurochemical and neurobiochemical changes associated with AD or perhaps other neurodegenerative diseases.
An Antibody Specific for Component 8 of Myelin Basic Protein from Normal Brain Reacts Strongly with Component 8 from Multiple Sclerosis Brain
Authors: McLaurin J. , Hashim G. , Moscarello M. A.
Published: October 2006

Abstract: Myelin basic protein (MBP) consists of several components or charge isomers (C‐1 through C‐8) generated by one or a combination of posttranslational modifications. One of these, C‐8, has been shown to contain citrulline (Cit) at defined sites formed by deimination of six arginyl residues. This unusual modification has allowed us to raise antibodies specific for this charge isomer only. To do this, a synthetic peptide, Gly‐Cit‐Cit‐Cit‐Cit, was coupled to keyhole limpet hemocyanin and injected into rabbits. The antibodies so generated reacted only with C‐8 and not with any of the other charge isomers. A second antibody fraction was raised against the synthetic peptide ACitHGFLPCitHR naturally occurring between residues 24 and 33 of C‐8 (all other charge isomers contain R instead of Cit at positions 25 and 31). These antibodies preferred C‐8 but reacted with the other charge isomers, to the extent of ∼25–30% of the reactivity shown with C‐8. In studies with C‐8 from multiple sclerosis (MS) MBP, much greater reactivity was obtained with these antibodies when compared with their reactivity with C‐8 from normal MBP. Because the total number of Cit residues in C‐8 from MS and normal MBP is the same, the difference in reactivity may be related to structural factors. The antibodies raised with the tetra‐Cit peptide were reacted with three pairs of synthetic peptides: 24ARHGFLPRHR33 and ACitHGFLPCitHR; 120GQRPGFGYGGRAS132 and GQCitPGFGYGGCitAS; and 157GGRDSRSGSPMARR170 and GGCitDSRSGSPMACitR. They reacted only with the Cit‐containing peptides in the order 157–170 > 120–130 > 24–33. Because all three peptides contained two Cit residues, the greater reactivity of 157–170 suggested structural factors may modulate the activity. With S49, an antibody raised against peptide 69GSLPQKSQ‐RSQDEN84 (rat sequence), a region in which Cit was not found and therefore is identical for both C‐1 and C‐8, showed greater reaction with C‐1 than C‐8, further supporting a role for conformational factors. Because the major difference between C‐1 and C‐8 is the presence of Cit in the later, conformational changes in this region of MBP must have been induced by Cit located even at some distance in the linear sequence of amino acids.
Effects of Vasopressin on Blood‐Brain Transfer of Methionine in Dogs
Authors: Brust Peter , Shaya Elias K. , Jeffries Keith J. , Dannals Robert F. , Ravert Hayden T. , Wilson Alan A. , Conti Peter S. , Wagner Henry N. , Gjedde Albert , Ermisch Armin , Wong Dean F.
Published: October 2006

Abstract: We used a simplified probe detection system for positron‐emitting radionuclides in order to measure blood‐brain barrier transport of amino acids in anesthetized dogs. Plasma and brain time‐activity curves were recorded after intravenous bolus injection of L‐[11C]methionine before and after administration of 1 μg of vasopressin. Three‐compartment models with three or four transfer coefficients were used to derive the kinetics of L‐[11C]methionine uptake in brain. The blood‐brain clearance of the tracer (K1) was 0.075 ml ml‐1 min‐1 before and 0.041 ml ml‐1 mir‐1 after injection of vasopressin. The partition volume and the initial distribution (plasma) volume of methionine were unchanged and within the expected limits. The net accumulation rate of methionine (K), estimated by both the four‐parameter (kinetic) and three‐parameter (graphic) approaches, decreased after vasopressin injection in all six studies.
Accumulation of Lysosphingolipids in Tissues from Patients with GM1 and GM2 Gangliosidoses
Authors: Kobayashi Takuro , Goto Ikuo , Okada Shintaro , Orii Tadao , Ohno Kousaku , Nakano Takeshi
Published: October 2006

Abstract: By using a sensitive method, we assayed lysocom‐pounds of gangliosides and asialogangliosides in tissues from four patients with GM2 gangliosidosis (one with Sand‐hoff disease and three with Tay‐Sachs disease) and from three patients with GM1 gangliosidosis [one with infantile type (fetus), one with late‐infantile, and one with adult type]. In the brain and spinal cord of all the patients except for an adult GM 1 gangliosidosis patient, abnormal accumulation of the lipids was observed, though the concentration in the fetal tissue was low. In GM2 gangliosidosis, the amounts of lyso GM2 ganglioside accumulated in the brain were similar among the patient with Sandhoff disease and the patients with Tay‐Sachs disease, whereas the concentration of asialo lyso GM2 ganglioside in the brain was higher in the former patient than in the latter patients. By comparing the sphingoid bases of neutral sphingolipids, gangliosides, and lysosphingolipids, it was suggested that lysosphingolipids in the diseased tissue are synthesized by sequential glycosylation from free sphingoid bases, but not by deacyla‐tion of the sphingolipids. Because lysosphingolipids are known to be cytotoxic, the abnormally accumulated lysosphingolipids may well be the pathogenetic agent for the neuronal degeneration in gangliosidoses.
Phorbol Ester‐Mediated Stimulation of Phospholipase D Activity in Sciatic Nerve from Normal and Diabetic Rats
Authors: Eggen Bart J. L. , Eichberg Joseph
Published: October 2006

Abstract: Evidence for the presence of phospholipase D activity in sciatic nerve was obtained by incubation of 32P‐prelabeled nerve segments in the presence of ethanol and measurement of [32P]phosphatidylethanol (PEth) formation expressed as a fraction of total phospholipid radioactivity. PEth synthesis was enhanced with increasing concentrations of ethanol (100 mM‐2 M). 4‐β‐Phorbol dibutyrate (100 nM‐1μM) stimulated PEth formation up to twofold in a time‐ and dose‐dependent manner. The stimulatory effect evoked by 100 nM phorbol ester was completely abolished by Ro 31–8220 (compound 3), a selective protein kinase C inhibitor. Efforts to identify the phospholipid precursor of PEth were unsuccessful, suggesting this product arises from a small discrete precursor pool. On subcellular fractionation of nerve, the ratio of basal and 4‐β‐phorbol dibutyrate‐stimulated phospholipase D activity recovered in a myelin‐enriched fraction, compared with a nonmyelin fraction, was 0.5 when results are expressed as a percentage of total phospholipid radioactivity. This ratio rises to 1.2 if the results are calculated assuming only phosphatidylcholine and phosphatidylethanolamine are potential precursors. The results suggest that myelin is a major locus of phospholipase D activity. Nerve from streptozotocin‐induced diabetic and control animals displayed the same basal phospholipase D activity, but the enzyme in diabetic nerve was stimulated to a greater extent by a suboptimal concentration of 4‐β‐phorbol dibutyrate. These results support the conclusion that protein kinase C modulates phospholipase D activity in nerve and suggest that in diabetic nerve the enzyme activation mechanism may possess increased sensitivity.
Increased Neopterin Levels in Brains of Patients with Human Immunodeficiency Virus Type 1 Infection
Authors: Shaskan Edward G. , Brew Bruce J. , Rosenblum Marc , Thompson Randall M. , Price Richard W.
Published: October 2006

Abstract: Postmortem levels of native neopterin (D‐erythroneopterin) were measured in cerebral cortical samples from 44 human immunodeficiency virus type 1‐infected and eight uninfected, nonneurological control patients. Cerebral cortical gray and white matter neopterin levels for the controls ranged from 0.5 to 7.2 pmol/mg of protein in contrast to neopterin levels in brains of the virus‐infected patients, which frequently were more than threefold and occasionally more than 30‐fold higher than mean control levels. Cortical neopterin levels did not correlate with severity of the acquired immunodeficiency syndrome dementia complex, but subcortical levels correlated with the presence of active human immunodeficiency virus type 1 infection, as reflected by pathological evidence of multinucleated giant cell encephalitis. Evidence of opportunistic cytomegalovirus infections in ∼25% of the human immunodeficiency virus type 1‐infected patients was associated with enhanced levels of neopterin in frontal cortex.
Pathological Phosphorylation Causes Neuronal Death: Effect of Okadaic Acid in Primary Culture of Cerebellar Granule Cells
Authors: Candeo Paola , Favaron Marco , Lengyel Imre , Manev Radmila M. , Rimland Joseph M. , Manev Hari
Published: October 2006

Abstract: We have investigated the role of protracted phosphatase inhibition and the consecutive protracted protein phosphorylation on neuronal viability. We found that in primary cultures of cerebellar granule neurons, the protracted (24‐h) inhibition of the serine/threonine protein phosphatases 1 and 2A (EC 3.1.3.16) by treatment of the cultures with okadaic acid (OKA; 5–20 nM) caused neurotoxicity that could be inhibited by the protein kinase inhibitor l‐(5‐isoquinolinylsulfonyl)‐2‐methylpiperazine (H7) or by the previous down‐regulation of the neuronal protein kinase C (PKC; ATP:protein phosphotransferase; EC 2.7.1.37). PKC was down‐regulated by exposure of the cultures for 24 h to 100 nM phorbol 12‐myristate 13‐acetate (TPA). The effect of the drugs used in the viability studies on the pattern of protein phosphorylation was measured by quantitative autoradiography. In particular, the 50‐ and 80‐kDa protein bands showed dramatic changes in the degree of phosphorylation: increase by OKA and brief TPA treatment; decrease by H7 or 24 h of TPA treatment; and inhibition of the OKA‐induced increase by H7 or 24 h of TPA treatment. The results suggest that the protracted phosphorylation, in particular that mediated by PKC, may lead to neuronal death and are in line with our previous suggestion that prolonged PKC translocation is operative in glutamate neurotoxicity.
Expression of Plasma Membrane Calcium‐Pumping ATPase mRNAs in Developing Rat Brain and Adult Brain Subregions: Evidence for Stage‐Specific Expression
Authors: Brandt Paul , Neve Rachael L.
Published: October 2006

Abstract: The plasma membrane calcium‐pumping ATPases (Ca2+‐ATPases) maintain resting free cytosolic calcium concentrations in cells at the submicromolar level. These Ca2+‐ATPases are encoded by four genes that can be alternately spliced to produce nine different mRNAs, each of which has a unique tissue‐specific distribution. Examination of the expression of these mRNAs in rat brain during development revealed that transcripts from three of the four known genes are expressed by the end of gestation. However, the stage of transcription induction varies among the isoforms. The mRNA encoding plasma membrane Ca2+‐ATPase (PMCA) lb, the isoform thought to maintain a housekeeping function, was present from embryonic day 10. The other alternately spliced PMCA1 mRNAs, PMCAla and c, which are preferentially expressed in the brain, did not appear until embryonic day 14. PMCA2a mRNA and the alternatively spliced PMCA2b and c transcripts were coordinately induced on embryonic day 18. The PMCA3a transcript first appeared on embryonic day 18 but did not reach steady‐state levels until postnatal day 3, whereas production of PMCA3b mRNA first occurred on embryonic day 10 and reached steady‐state expression by embryonic day 18. Several PMCA mRNAs tested varied in expression in specific regions of the brain that were examined at three postnatal time points.
Involvement of Growth‐Associated Protein‐43 with Irreversible Neurite Outgrowth by Dibutyryl Cyclic AMP and Phorbol Ester in NG108–15 Cells
Authors: Kumagai Chihiro , Tohda Michihisa , Isobe Masaharu , Nomura Yasuyuki
Published: October 2006

Simultaneous treatment with 12‐O‐tetradecanoylphorbol 13‐acetate (TPA) and dibutyryl cyclic AMP (diBu‐cAMP) for 72 h induced neurites in NG108–15 cells significantly longer than treatment with each alone. Treatment for 72 h with both drugs induced irreversible neurite extension and a decline in protein kinase C activity, although neurites extended by diBu‐cAMP alone disappeared after the withdrawal of the drug. The expression of growth‐associated protein‐43 (GAP‐43) mRNA was also observed by a combined application of TPA and diBu‐cAMP. The increased level of GAP‐43 mRNA induced by treatment with both drugs for 72 h was maintained at least 24 h after withdrawal of the drugs. In cells transfected with GAP‐43 cDNA, neurites induced by treatment with diBu‐cAMP alone for 72 h were maintained at least 48 h after removal of the drugs. These results suggest that GAP‐43 could be involved in the maintenance of elongated neurites and that a decline in protein kinase C activity may be involved in the accumulation of GAP‐43.
Mapping of Histamine H1 Receptors in the Human Brain Using [11C]Pyrilamine and Positron Emission Tomography
Authors: Yanai Kazuhiko , Watanabe Takehiko , Yokoyama Hiroyuki , Hatazawa Jun , Iwata Ren , Ishiwata Kiichi , Meguro Kenichi , Itoh Masatoshi , Takahashi Toshihiro , Ido Tatsuo , Matsuzawa Taiju
Published: October 2006

We have studied the characteristics of carbon‐11 labeled pyrilamine as a radioligand for investigating histamine H1 receptors in human brain with positron emission tomography (PET). [11QPyrilamine is distributed evenly in proportion to cerebral blood flow at initial PET images. Later (after 45–60 min), 11C radioactivity was observed at high concentrations in the frontal and temporal cortex, hippocampus, and thalamus, and at low concentrations in the cerebellum and pons. The regional distribution of the carbon‐11 labeled compound in the brain corresponded well with that of the histamine H1 receptors determined in vitro in autopsied materials. In six controls, the frontal and temporal cortices/cerebellum ratio increased during the first 60 min to reach a value of 1.22 ± 0.071. Intravenous administration of d‐chlorpheniramine (5 mg) completely abolished the specific binding in vivo in the frontal cortex and temporal cortex (cortex/cerebellum ratio, 0.955 ±0.015). The availability of this method for measuring histamine H1 receptors in vivo in humans will facilitate studies on neurological and psychiatric disorders in which histamine H1 receptors are thought to be abnormal.
Temporal Profiles of Nerve Growth Factor β‐Subunit Level in Rat Brain Regions After Transient Ischemia
Authors: Shozuhara Hidetaka , Onodera Hiroshi , Katoh‐Semba Rituko , Kato Kanefusa , Yamasaki Yasundo , Kogure Kyuya
Published: October 2006

To determine the role of nerve growth factor (NGF) in ischemic brain damage, we measured the temporal and regional changes in the level of NGF in the hippo‐campal subfields, the cerebral cortex, the striatum, and the septum at 1, 2, 7, and 30 days after transient forebrain ischemia using a highly sensitive sandwich‐type enzyme im‐munoassay system for the β‐subunit of mouse 7S NGF (β‐NGF). We also analyzed glial fibrillary acidic protein immunoreactivity in the hippocampus to ascertain the contribution of reactive astrocytes to NGF production after an ischemic insult. In the CA1 subfield of the hippocampus, the level of β‐NGF decreased slightly 2 days after ischemia (not significant), at which time CA1 pyramidal cell loss began to occur, and increased by 40% 30 days after ischemia (p < 0.05). A marked increase in glial fibrillary acidic pro‐tein‐positive astrocytes in the CA1 subfield 2–30 days after ischemia suggests that the reactive astrocytes participated in a gradual increase in the level of β‐NGF after recirculation. The level of β‐NGF in the dentate gyrus decreased transiently 2 days (p < 0.05) and 7 days (p < 0.01) after ischemia, followed by recovery to the level of control animals 30 days after ischemia. The level of β‐NGF in the septum gradually decreased 7 days (−27%, p < 0.05) and 30 days (−43%, p < 0.01) after ischemia. The levels of β‐NGF in the cerebral cortex and striatum remained unaltered throughout the observation period. These results suggest that the alteration of NGF content occurs not only in the CA1 subfield, where pyramidal cells are depleted, but also in areas with minimal histological damage. Septal decrease in NGF may reflect the reduction of retrograde NGF transport from hippocampus to septal cholinergic neurons.
A Cyclic AMP Mechanism Mediates the Serotonin‐Induced Increase in Glutamic Acid Decarboxylase Activity in the Preoptic Area and Hypothalamus
Authors: Munaro N. I. , Taleisnik S.
Published: October 2006

Previous studies have shown that the injection of 5‐hydroxytryptamine (5‐HT) into the third ventricle of rats on the afternoon of proestrus increases glutamic acid decarboxylase (GAD) activity in the preoptic area and the hypothalamus. In the present report we examine the adenylate cyclase‐cyclic AMP (cAMP) system as mediator of that effect. The increase in GAD activity induced by intraventricu‐lar injection of 5‐HT was completely blocked by injecting an antiserum against cAMP into the third ventricle 30 min earlier, whereas an injection of serum from normal rabbits was ineffective. On the contrary, activation of adenylate cy‐clase activity by intraventricular injection of forskolin increased GAD activity, an effect that was also blocked by anti‐cAMP serum. Anti‐cAMP serum also lowered GAD activity in the preoptic area and hypothalamus when injected on the morning of proestrus but not when injected in the afternoon, when the values of GAD activity were already low. The results suggest that a cAMP mechanism may be involved in the changes in preoptic‐area and hypotha‐lamic GAD activity such as the rise in enzyme activity induced by intraventricular injection of 5‐HT.
Phosphoethanolamine Enhances High‐Affinity Choline Uptake and Acetylcholine Synthesis in Dissociated Cell Cultures of the Rat Septal Nucleus
Authors: Bostwick J. Robert , Abbe Russell , Appel Stanley H.
Published: October 2006

Dissociated rat septal nucleus cells cultured in defined medium exhibited twofold increases in the maximal rates of sodium‐dependent, high‐affinity choline uptake and acetylcholine formation when grown in the presence of phosphoethanolamine. The effect was concentration‐dependent (EC50= 15 μM) and appeared to be associated with in vitro maturation of cholinergic neurons rather than with enhanced survival. Choline acetyltransferase, acetylcholin‐esterase, and choline kinase activities were unaffected by this treatment. The effect of phosphoethanolamine was specific for cholinergic neurons, because treatment with this compound did not alter the kinetic constants for high‐affinity neuronal uptake of γ‐aminobutyric acid or dopamine. The action appeared to be mediated primarily through activation of the sodium‐dependent, high‐affinity transport mechanism for choline as opposed to alterations in the storage and release of acetylcholine.
Metabotropic Glutamate Receptor Activation Produces Extrapyramidal Motor System Activation That Is Mediated by Striatal Dopamine
Authors: Sacaan Aida I. , Bymaster Frank P. , Schoepp Darryle D.
Published: October 2006

Little is known about the in vivo function of the GTP‐binding protein‐coupled “metabotropic” excitatory amino acid (EAA) receptor. In vitro studies on agonist‐induced brain phosphoinositide hydrolysis have shown that (15,3i?)‐1‐aminocyclopentane‐l,3‐dicarboxylic acid is a highly selective and efficacious metabotropic EAA agonist. We have recently reported that in vivo unilateral intrastria‐tal injection of (15′,3.K)‐l‐aminocyclopentane‐l,3‐dicar‐boxylic acid induces transient extrapyramidal motor activation that manifests itself as contralateral turning. In this study, we fully characterized the onset of turning behavior following intrastriatal (1S,3.R)‐1‐aminocyclopentane‐l,3‐dicarboxylic acid injection and the possible involvement of striatal dopamine neurons in the mediation of this effect. Rats were anesthetized with the short‐acting agent halo‐thane to allow for rapid surgical recovery and thus early behavioral measurements. Intrastriatal (1S,3R)‐1‐aminocy‐clopentane‐l,3‐dicarboxylic acid (1 μmol/2 μl) produced an incremental increase in contralateral turning starting at 1 h and plateauing 3–6 h after injection (peak effect, 39.1 ± 6.7 rotations per 5 min). Dopamine depletion with α‐methyl‐DL‐p‐tyrosine (250 mg/kg i.p., 80% depletion) resulted in >85% inhibition of (1S,3R)‐1‐aminocyclopentane‐l,3‐di‐carboxylic acid‐induced contralateral turning. The dopamine antagonist haloperidol (0.3 mg/kg i.p.) produced 48% inhibition of the (1S,3R)‐1‐aminocyclopentane‐1,3‐dicar‐boxylic acid response. In time course studies, turning behavior correlated with increases in levels of the dopamine metabolites 3,4‐dihydroxyphenylacetic acid and homovanillic acid. These results suggest a functional interaction between the metabotropic EAA receptor and the dopami‐nergic system in the striatum.
Differences in Dopamine Clearance and Diffusion in Rat Striatum and Nucleus Accumbens Following Systemic Cocaine Administration
Authors: Cass Wayne A. , Gerhardt Greg A. , Mayfield R. Dayne , Curella Pamela , Zahniser Nancy R.
Published: October 2006

Acute cocaine administration preferentially increases extracellular dopamine levels in nucleus accumbens as compared with striatum. To investigate whether a differential effect of cocaine on dopamine uptake could explain this observation, we used in vivo electrochemical recordings in anesthetized rats in conjunction with a paradigm that measures dopamine clearance and diffusion without the confounding effects of release. When a finite amount of dopamine was pressure‐ejected at 5‐min intervals from a micropipette adjacent to the electrode, transient and reproducible increases in dopamine levels were detected. In response to 15 mg/kg of cocaine‐HCl (i.p.), these signals increased in nucleus accumbens, indicating significant inhibition of the dopamine transporter. The time course of the dopamine signal increase paralleled that of behavioral changes in unanesthetized rats receiving the same dose of cocaine. In contrast, no change in the dopamine signal was detected in dorsal striatum; however, when the dose of cocaine was increased to 20 mg/kg, enhancement of the dopamine signal occurred in both brain areas. Quantitative auto‐radiography with [3H]mazindol revealed that the affinity of the dopamine transporter for cocaine was similar in both brain areas but that the density of [3H]mazindol binding sites in nucleus accumbens was 60% lower than in dorsal striatum. Tissue dopamine levels in nucleus accumbens were 44% lower. Our results suggest that a difference in dopamine uptake may explain the greater sensitivity of nucleus accumbens to cocaine as compared with dorsal striatum. Furthermore, this difference may be due to fewer dopamine transporter molecules in nucleus accumbens for cocaine to inhibit, rather than to a higher affinity of the transporter for cocaine.
Somatostatin Content Increases Following Norepinephrine Depletion in Frontal Cortex of l‐Methyl‐4‐Phenyl‐1,2,3,6‐Tetrahydropyridine‐Treated Mice
Authors: Mitsuo Kunihiko , Schwartz Joan P.
Published: October 2006

The effects of l‐methyl‐4‐phenyl‐l,2,3,6‐tetrahy‐dropyridine (MPTP) on somatostatin (SS)‐containing neurons were examined by measuring dopamine, norepinephrine (NE), SS, and SS mRNA in striatum and frontal cortex of C57/B16 mice at various times following treatment with MPTP‐HC1 (96 mg/kg i.p.). MPTP caused a 70% depletion of dopamine in striatum by 1 day and a 40% depletion of NE in frontal cortex within 3 days. SS content was increased in frontal cortex 4 days later, but not in striatum; there were no changes in SS mRNA. Maprotiline, a specific NE‐uptake blocker, prevented both the depletion of NE and the increase of SS in frontal cortex due to MPTP administration. These results support the possibility that NE can regulate SS in frontal cortex and are discussed in terms of the decrease of SS seen in parkinsonian patients with dementia.
Pharmacological Characterization and Autoradiographic Localization of Histamine H2 Receptors in Human Brain Identified with [125I]Iodoaminopotentidine
Authors: Traiffort E. , Pollard H. , Moreau J. , Ruat M. , Schwartz J. C. , Martinez‐Mir M. I. , Palacios J. M.
Published: October 2006

125I‐Aminopotentidine (125I‐APT), a reversible probe of high specific radioactivity and high affinity and selectivity for the H2 receptor, was used to characterize and localize this histamine receptor subtype in human brain samples obtained at autopsy. On membranes of human caudate nucleus, specific 125I‐APT binding at equilibrium revealed a single component, with a dissociation constant of 0.3 nM and maximal capacity of about 100 fmol/mg of protein. At 0.2 nM, 125I‐APT specific binding, as defined with tiotidine, an H2‐receptor antagonist chemically unrelated to iodoaminopotentidine, represented 40–50% of the total. Specific 125I‐APT binding was inhibited by a series of typical H2‐receptor antagonists that displayed apparent dissociation constants closely similar to corresponding values at the reference biological system, i.e., guinea pig atrium. This indicates that the pharmacology of the H2 receptor is the same in the human brain as on this reference system. However, histamine was about 10‐fold more potent in inhibiting 125I‐APT binding to membranes of human brain than of guinea pig brain. 125I‐APT binding was also inhibited by amitriptyline and mianserin, two antidepressant drugs, in micromolar concentrations corresponding to effective plasma concentrations of treated patients. The distribution of H2 receptors was established autoradiographically with 125I‐APT on a series of coronal sections of human brain after assessing the pharmacological specificity of the labeling. The highest density of 125I‐APT sites was found in the basal ganglia, various parts of the limbic system, e.g., hippocampus or amygdaloid complex, and the cerebral cortex. H2 receptors displayed a laminar distribution in cerebral cortex and hippocampal formation. A low density of sites was found in cerebellum as well as in hypothalamus, the brain area where all the perikarya and the largest number of axons of histaminergic neurons are found. The widespread distribution of H2 receptors in the human brain is consistent with the alleged modulatory role of histamine mediated by this subtype of receptor.
Cell Swelling, Blebbing, and Death Are Dependent on ATP Depletion and Independent of Calcium During Chemical Hypoxia in a Glial Cell Line (ROC‐1)
Authors: Jurkowitz‐Alexander Marianne S. , Altschuld Ruth A. , Hohl Charlene M. , Johnson J. David , McDonald John S. , Simmons Todd D. , Horrocks Lloyd A.
Published: October 2006

The morphological and biochemical changes that occur during chemical hypoxic injury in a neural cell line were studied in the presence and absence of calcium. Oligo‐dendroglial‐glioma hybrid cells (ROC‐1) were subjected to inhibitors of glycolytic and oxidative ATP synthesis (chemical hypoxia). Complete respiratory inhibition depleted [ATP] to <5% of control by 4 min. Blebs appeared on the cell surfaces and cells began to swell within a few minutes of ATP depletion. A 200% increase in cell volume and bleb coalescence preceded irreversible cell injury (lactate dehy‐drogenase release) which began at ∼20 min with 50% cell death by 40 min. In energized cells an equivalent degree of osmotic swelling induced by ouabain inhibition of the Na+, K+‐ATPase pump did not produce blebbing or cell death. Partial inhibition of respiration decreased [ATP] to ∼10% of control by 40 min. Blebbing and swelling began at 40 min and bleb coalescence preceded plasma membrane disruption which began at ∼55 min. ATP depletion, blebbing, swelling, and death followed similar time courses in the presence or absence of extracellular calcium ([Ca2+]e). Intracel‐lular calcium ([Ca2+]i) was measured using fura‐2. In calcium‐containing medium metabolic inhibition caused a transient increase in resting [Ca2+]i (100 ± 17 nM) followed by a low steady‐state level preceding plasma membrane disruption. Following deenergization in calcium‐free medium, [Ca2+]i remained below 60 nM throughout injury and death. These data suggest that decreased ATP initiates a sequence of events including bleb formation and cell swelling that lead to irreversible cell injury in the absence of large increases in [Ca2+]i.
Effect of Experimental Diabetes on the Catecholamine Metabolism in Rat Brain
Authors: Gupta G. , Azam M. , Baquer N. Z.
Published: October 2006

Abstract: The levels of epinephrine, norepinephrine, and dopamine and the activities of tyrosine hydroxylase and monoamine oxidase were estimated in four regions of rat brain during alloxan‐induced hyperglycemia and insulin‐induced hypoglycemia. Catecholamine levels were estimated by HPLC, and the insulin levels were quantified by radioimmunoassay. The results demonstrated significant increases in the activities of the metabolizing enzymes and levels of catecholamines during experimental conditions. The levels of catecholamines were highest in the cerebral hemispheres, the region associated with high activities of the metabolizing enzymes. Insulin‐induced hypoglycemia caused a decrease in the activities of the metabolizing enzymes followed by their recovery within 2 h.
Decarboxylation of Exogenous l‐3,4‐Dihydroxyphenylalanine in Rat Striatum as Studied by In Vivo Voltammetry
Authors: Nakazato Taizo , Akiyama Akitane
Published: October 2006

Abstract: An in vivo voltammetric technique was used to determine whether striatal nondopaminergic neurons take up and decarboxylate exogenous l‐3,4‐dihydroxyphenylalanine (L‐DOPA) and release it as dopamine. After the striatal serotonergic neurons of the rat had been destroyed by intraventricular injection of 5,7‐dihydroxytryptamine, l‐DOPA was administered intraperitoneally. It was found that changes in the dopamine concentration in the striatal extracellular fluid of the rat were the same as those in the nonlesioned rat. l‐DOPA was also administered to the rat after the striatal perikarya had been destroyed by the intrastriatal injection of kainate. The striatal dopamine concentrations of the lesioned rat changed in parallel with 5,7‐dihydroxytryptamine‐lesioned rats, as well as the nonlesioned rats. Moreover, when normal rats were administered l‐DOPA, the dopamine concentration was not increased in the cerebellum, where dopamine neurons do not exist. From these observations, it is concluded that exogenous l‐DOPA is taken up, decarboxylated to dopamine, and released only in the striatal dopamine neurons.
Increased Intracellular γ‐Aminobutyric Acid Selectively Lowers the Level of the Larger of Two Glutamate Decarboxylase Proteins in Cultured GABAergic Neurons from Rat Cerebral Cortex
Authors: Rimvall Karin , Martin David L.
Published: October 2006

Abstract: The regulation of glutamate decarboxylase (GAD; EC 4.1.1.15) was studied by using cultures of cerebral cortical neurons from rat brain grown in serum‐free medium. About 50% of the neurons in the cultures were γ‐aminobutyric acid (GABA)ergic as determined by two double‐staining procedures. Immunoblotting experiments with four anti‐GAD sera that recognize the two forms to varying degrees, demonstrated that the cultures contained the two forms of GAD that are present in rat brain (apparent molecular masses = 63 and 66 kDa). GAD activity was reduced by 60–70% when intracellular GABA levels were increased by incubating the cultures with the GABA‐transaminase inhibitor γ‐vinyl‐GABA for >5–10 h or with 1 mM GABA itself. Neither baclofen nor muscimol (100 μM) affected GAD activity. Immunoblotting experiments showed that only the larger of the two forms of GAD (66 kDa) was decreased by elevated GABA levels. These results, together with previous results indicating that the smaller form of GAD is more strongly regulated by pyridoxal 5′‐phosphate (the cofactor for GAD), suggest that the two forms of GAD are regulated by different mechanisms.
Evidence for a Hyperexcitability State of Staggerer Mutant Mice Macrophages
Authors: Kopmels B. , Mariani J. , Delhaye‐Bouchaud N. , Audibert F. , Fradelizi D. , Wollman E. E.
Published: October 2006

Abstract: We recently reported an abnormal production of interleukin‐1 (IL‐1) in peripheral macrophages of several neurological mutant mice that exhibit patterns of neuronal degeneration, especially in the cerebellum. After in vitro activation by lipopolysaccharide acid (LPS), these macrophages hyperexpress IL‐1β mRNA and hyperproduce IL‐1 protein in comparison with +/+ controls. In the present study, focused on the staggerer mutant mice, we investigate if this genetic dysregulation is specific for IL‐1β or if it reflects a generalized hyperexcitability of these macrophages. The hyperexpression of IL‐1β mRNA in sg/sg macrophages is present whatever the duration of LPS stimulation, even for periods as short as 15 min, although it reaches a maximum after 4 h of stimulation. The hyperinducibility of sg/sg macrophages is observed even when very low doses of LPS are used (0.01 μg/ml) and reaches its maximum for 5 μg/ml LPS. Synthetic molecules (muramyl dipeptides), such as N‐acetylmuramyl‐L‐alanyl‐D‐isoglutamine or murabutide, known as macrophage activators, are also efficient in revealing the cytokine hyperexpression in sg/sg macrophages. In addition, hyperexpression of two other cytokines, i.e., tumor necrosis factor‐α and IL‐1α mRNAs, is also detected in LPS‐stimulated macrophages of mutant mice. Finally, the effect of an inhibitor of protein synthesis, cycloheximide, is similar in +/+ and sg/sg macrophages. As a whole, these data lead us to conclude that the sg/sg macrophages are in a state of general hyperexcitability when compared with +/+ ones.
Human Brain Lectin: A Soluble Lectin That Binds Actin
Authors: Joubert Raymonde , Caron Michel , Avellana‐Adalid Virginia , Mornet Dominique , Bladier Dominique
Published: October 2006

Abstract: A biotinylated probe was used for detection of endogenous ligands of a human brain lectin on blotted human brain soluble proteins. Of the various proteins from brain extract resolved by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, five reacted with the biotinylated probe. After elimination of saccharidic moieties by periodic treatment of the same extract, a single band with Mr∼ 43,000 was recognized by the lectin. This band was identified as actin using an anti‐actin antibody. These results were confirmed by binding of biotinylated lectin to purified actin.
Different Natures of Supersensitivity of Adenylate Cyclase Stimulated by Calcitonin Gene‐Related Peptide and Isoproterenol in Rat Diaphragm After Denervation and Reserpine Treatment
Authors: Matsumoto Naoko , Wang Xiao‐Bing , Uchida Shuji
Published: October 2006

Abstract: In skeletal muscles, calcitonin gene‐related peptide (CGRP) released from motor nerve terminals and humoral catecholamines stimulate adenylate cyclase (AC) and enhance muscle contraction. The effects of denervation and treatment with reserpine on twitch contraction and the AC system in rat diaphragm were investigated. The basal levels of twitch contraction and AC activity of the diaphragm of rats were both increased 2 weeks after phrenic nerve denervation but were not altered by treatment with reserpine. Reserpine treatment provoked supersensitivity of AC to isoproterenol, without affecting the response to CGRP. On the other hand, denervation decreased the activation of AC and enhancement of twitch contraction by CGRP, without affecting the responses to isoproterenol. These data suggest that denervation causes up‐regulation of AC as a result of loss of CGRP release from nerve terminal and that depletion of catecholamines by reserpine treatment supersensitizes the responses at the β‐adrenoceptor level. Thus, nervous and humoral factors regulate the AC system in striated muscle by different mechanisms.
Induction of Interleukin‐1β mRNA in Rat Brain After Transient Forebrain Ischemia
Authors: Minami Masabumi , Kuraishi Yasushi , Yabuuchi Kazuki , Yamazaki Akira , Satoh Masamichi
Published: October 2006

Abstract: The expression of interleukin‐1β (IL‐1β) mRNA in the cerebral cortex, hippocampus, striatum, and thalamus of rats was studied after transient forebrain ischemia. IL‐1β mRNA was not detected in all these regions of sham‐operated control rats. IL‐1β mRNA was induced after transient forebrain ischemia and reached a detectable level in all regions examined 15 min after the start of recirculation. The induction of IL‐1β mRNA had a few peaks, that is, peaks were observed at 30 and 240 min in the four regions examined, and another peak was observed at 90 min in the striatum. One day after the start of recirculation, IL‐1β mRNA levels were markedly decreased, but even 7 days after that, IL‐1β mRNA was found at very low levels in all regions examined. The amounts of c‐fos and β‐actin mRNAs on the same blots were also examined. The induction of c‐fos mRNA was transient and had only one peak in all regions examined, whereas the levels of β‐actin mRNA in these regions were fairly constant throughout the recirculation period. Thus, we provide the first evidence for a characteristic expression of IL‐1β mRNA in several brain regions after transient forebrain ischemia.
Uptake and Metabolism of Choline by Rat Brain After Acute Choline Administration
Authors: Klein Jochen , Köppen Andrea , Löffelholz Konrad , Schmitthenner Jorg
Published: October 2006

Abstract: The present study is concerned with the uptake and metabolism of choline by the rat brain. Intraperitoneal administration of choline chloride (4‐60 mg/kg) caused a dose‐dependent elevation of the plasma choline concentration from 11.8 to up to 165.2 μM within 10 min and the reversal of the negative arteriovenous difference (AVD) of choline across the brain to positive values at plasma choline levels of >23 μM. Net choline release and uptake were linearly dependent on the plasma choline level in the physiological range of 10‐50 μM, whereas the CSF choline level was significantly increased only at plasma choline levels of >50 μM. The bolus injection of 60 mg/kg of [3H]choline chloride caused the net uptake of > 500 μMol/g of choline by the brain as calculated from the AVD, which was reflected in a minor increase of free choline level and a long‐lasting increase of brain phosphorylcholine content, which paralleled the uptake curve. Loss of label from phosphorylcholine 30 min to 24 h after choline administration was accompanied by an increase of label in phosphatidylcholine, an indication of a delayed transfer of newly taken‐up choline into membrane choline pools. In conclusion, homeostasis of brain choline is maintained by a complex system that interrelates choline net movements into and out of the brain and choline incorporation into and release from phospholipids.