bluuebunny/crossref_metadata_2025_split huggingface dataset:
D1 dopamine receptor stimulation increases GluR1 phosphorylation in postnatal nucleus accumbens cultures
Authors: Chao Steven Z. , Lu Wenxiao , Lee Hey‐Kyoung , Huganir Richard L. , Wolf Marina E. 
Published: March 2003

AbstractPostsynaptic interactions between dopamine and glutamate receptors in the nucleus accumbens are critical for acute responses to drugs of abuse and for neuroadaptations resulting from their chronic administration. We tested the hypothesis that D1 dopamine receptor stimulation increases phosphorylation of the AMPA receptor subunit GluR1 at the protein kinase A phosphorylation site (Ser845). Nucleus accumbens cell cultures were prepared from postnatal day 1 rats. After 14 days in culture, GluR1 phosphorylation was measured by western blotting using phosphorylation site‐specific antibodies. The D1 receptor agonist SKF 81297 increased Ser845 phosphorylation in a concentration‐	dependent manner, with marked increases occurring within 5 min. This was prevented by the D1 receptor antagonist SCH 23390 and the protein kinase A inhibitor H89, and reproduced by forskolin. The D2 receptor agonist quinpirole attenuated the response to D1 receptor stimulation. Neither D1 nor D2 receptor agonists altered GluR1 phosphorylation at Ser831, the site phosphorylated by protein kinase C and calcium/calmodulin‐dependent protein kinase II. In other systems, phosphorylation of GluR1 at Ser845 is associated with enhancement of AMPA receptor currents. Thus, the present results suggest that AMPA receptor transmission in the nucleus accumbens may be augmented by concurrent D1 receptor stimulation.
PKC and Raf‐1 inhibition‐related apoptotic signalling in N2a cells
Authors: Bronisz Agnieszka , Gajkowska Barbara , Doma Krystyna , Ska‐Janik ń 
Published: March 2003

AbstractIn this study, a neuroblastoma N2a cell line was applied to investigate mechanisms of apoptosis induced either by selective inhibition of protein kinase C (PKC) by low amounts of staurosporine (STS10) or by inhibition PI3‐K after wortmannin (WM) treatment. We present evidence that, in the absence of serum in the medium, decreased phosphorylation of Raf‐1 and BAD112, as well as Akt and BAD136, proteins and their translocation to mitochondria coincided with STS10‐ or WM‐induced apoptosis, respectively. Concomitantly, release of cytochrome c into the cytosol indicated a BCL‐2‐dependent mode of cell death after both treatments. Furthermore, in typical ‘gain of function’ experiments, cells with overexpression of permanently active Raf‐1 or Akt transgenes displayed a significantly higher and independent resistance to either STS10 or WM. Thus, our results indicate that PKC/Raf‐1/BAD112, as well as PI3‐K/Akt/BAD136 signalling pathways, are both necessary for N2a cell survival and thus are unable to functionally substitute for each other as long as the cells do not receive additional signal(s) derived from serum. However, in the presence of serum, undefined trophic signal(s) can stimulate cross‐talk between these two pathways at a level upstream from Raf‐1 and Akt phosphorylation. In this case, only simultaneous inhibition of PKC and PI3‐K is able to induce apoptosis.
Involvement of P2X7 receptors in the regulation of neurotransmitter release in the rat hippocampus
Authors: Sperlágh Beáta , Köfalvi Attila , Deuchars Jim , Atkinson Lucy , J. Milligan Carol , Buckley Noel J. , Vizi E. Sylvester 
Published: March 2003

AbstractAlthough originally cloned from rat brain, the P2X7 receptor has only recently been localized in neurones, and functional responses mediated by these neuronal P2X7 receptors (P2X7R) are largely unknown. Here we studied the effect of P2X7R activation on the release of neurotransmitters from superfused rat hippocampal slices. ATP (1–30 mm) and other ATP analogues elicited concentration‐dependent [3H]GABA outflow, with the following rank order of potency: benzoylbenzoylATP (BzATP) > ATP > ADP. PPADS, the non‐selective P2‐receptor antagonist (3–30 µm), Brilliant blue G (1–100 nm) the P2X7‐selective antagonist and Zn2+ (0.1–30 µm) inhibited, whereas lack of Mg2+ potentiated the response by ATP. In situ hybridization revealed that P2X7R mRNA is expressed in the neurones of the cell body layers in the hippocampus. P2X7R immunoreactivity was found in excitatory synaptic terminals in CA1 and CA3 region targeting the dendrites of pyramidal cells and parvalbumin labelled structures. ATP (3–30 µm) and BzATP (0.6–6 µm) elicited concentration‐dependent [14C]glutamate efflux, and blockade of the kainate receptor‐mediated transmission by CNQX (10–100 µm) and gadolinium (100 µm), decreased ATP evoked [3H]GABA efflux. The Na+ channel blocker TTX (1 µm), low temperature (12°C), and the GABA uptake blocker nipecotic acid (1 mm) prevented ATP‐induced [3H]GABA efflux. Brilliant blue G and PPADS also reduced electrical field stimulation‐induced [3H]GABA efflux. In conclusion, P2X7Rs are localized to the excitatory terminals in the hippocampus, and their activation regulates the release of glutamate and GABA from themselves and from their target cells.
Glucose transport and utilization are altered in the brain of rats deficient in n‐3 polyunsaturated fatty acids
Authors: Ximenes da Silva Adriana , Lavialle Françoise , Gendrot Ghislaine , Guesnet Philippe , Alessandri Jean‐Marc , Lavialle Monique 
Published: March 2003

AbstractLong‐chain polyunsaturated (n‐3) fatty acids have been reported to influence the efficiency of membrane receptors, transporters and enzymes. Because the brain is particularly rich in docosahexaenoic acid (DHA, 22:6 n‐3), the present study addresses the question of whether the 22:6 n‐3 fatty acid deficiency induces disorder in regulation of energy metabolism in the CNS. Three brain regions that share a high rate of energy metabolism were studied: fronto‐parietal cortex, hippocampus and suprachiasmatic nucleus. The effect of the diet deficient in n‐3 fatty acids resulted in a 30–50% decrease in DHA in membrane phospholipids. Moreover, a 30% decrease in glucose uptake and a 20–40% decrease in cytochrome oxidase activity were observed in the three brain regions. The n‐3 deficient diet also altered the immunoreactivity of glucose transporters, namely GLUT1 in endothelial cells and GLUT3 in neurones. In n‐3 fatty acid deficient rats, GLUT1‐immunoreactivity readily detectable in microvessels became sparse, whereas the number of GLUT3 immunoreactive neurones was increased. However, western blot analysis showed no significant difference in GLUT1 and GLUT3 protein levels between rats deficient in n‐3 fatty acids and control rats. The present results suggest that changes in energy metabolism induced by n‐3 deficiency could result from functional alteration in glucose transporters.
Temporal patterns of cytokine and apoptosis‐related gene expression in spinal cords of the G93A‐SOD1 mouse model of amyotrophic lateral sclerosis
Authors: Hensley Kenneth , Floyd Robert A. , Gordon Brian , Mou Shenyun , Pye Quentin N. , Stewart Charles , West Melinda , Williamson Kelly 
Published: March 2003

AbstractFamilial amyotrophic lateral sclerosis (FALS) is often caused by gain‐of‐function mutations in Cu,Zn‐superoxide dismutase (SOD1). Multiprobe ribonuclease protection assays (RPAs) were used to investigate expression of 36 different cytokines and apoptosis‐related genes in spinal cords of mice that ubiquitously express human SOD1 bearing a glycine (r) alanine substitution at residue 93 (G93A‐SOD1). Mice were studied at late presymptomatic stage (80 days), and at 120 days when the animals experience severe hindlimb paralysis and accumulation of oxidatively modified proteins. Spinal cord tissue from G93A‐SOD1 mice expressed a selective subset of macrophage‐typical cytokines (monokines) including interleukin (IL)1α, IL1β and IL1RA at 80 days increasing by 120 days. Contrastingly, T‐cell derived cytokines (lymphokines) including IL2, IL3 and IL4 were detected at low levels in non‐transgenic mice but these were not elevated in G93A‐SOD1 mice even at 120 days. Apoptosis‐related genes were generally unaffected at 80 days but multiple caspases and death receptor components were up‐regulated at 120 days; the only exceptions being FADD and the tumor necrosis factor (TNF)α receptor p55 which was up‐regulated at 80 days and increased further at 120 days. These data indicate that in the G93A‐SOD1 mouse: (i) cytokine expression changes precede bulk protein oxidation and apoptosis gene expression; (ii) lymphocyte contributions to cytokine expression in FALS are likely minor; and (iii) TNFα and its receptors may link inflammation to apoptosis in ALS.
Oxidative damage increases with age in a canine model of human brain aging
Authors: Head E. , Liu J. , Hagen T. M. , Muggenburg B. A. , Milgram N. W. , Ames B. N. , Cotman C. W. 
Published: March 2003

AbstractWe assayed levels of lipid peroxidation, protein carbonyl formation, glutamine synthetase (GS) activity and both oxidized and reduced glutathione to study the link between oxidative damage, aging and β‐amyloid (Aβ) in the canine brain. The aged canine brain, a model of human brain aging, naturally develops extensive diffuse deposits of human‐type Aβ. Aβ was measured in immunostained prefrontal cortex from 19 beagle dogs (4–15 years). Increased malondialdehyde (MDA), which indicates increased lipid peroxidation, was observed in the prefrontal cortex and serum but not in cerebrospinal fluid (CSF). Oxidative damage to proteins (carbonyl formation) also increased in brain. An age‐dependent decline in GS activity, an enzyme vulnerable to oxidative damage, and in the level of glutathione (GSH) was observed in the prefrontal cortex. MDA level in serum correlated with MDA accumulation in the prefrontal cortex. Although 11/19 animals exhibited Aβ, the extent of deposition did not correlate with any of the oxidative damage measures, suggesting that each form of neuropathology accumulates in parallel with age. This evidence of widespread oxidative damage and Aβ deposition is further justification for using the canine model for studying human brain aging and neurodegenerative diseases.
α‐Synuclein exhibits competitive interaction between calmodulin and synthetic membranes
Authors: Lee Daekyun , Lee Sun‐Young , Lee Eui‐Nam , Chang Chung‐Soon , Paik Seung R. 
Published: March 2003

Abstractα‐Synuclein, a pathological component of Parkinson's disease by constituting the Lewy bodies, has been suggested to be involved in membrane biogenesis via induction of amphipathic α‐helices. Since the amphipathic α‐helix is also known as a recognition signal of calmodulin for its target proteins, molecular interaction between α‐synuclein and calmodulin has been investigated. By employing a chemical coupling reagent ofN‐(ethoxycarbonyl)‐2‐ethoxy‐1,2‐dihydroquinoline, α‐synuclein has been shown to yield a heterodimeric 1 : 1 complex with calmodulin on sodium dodecyl sulfate–polyacrylamide gel electrophoresis in the presence and even absence of calcium, whereas β‐synuclein was more dependent upon calcium for its calmodulin interaction. The selective calmodulin interaction of α‐synuclein in the absence of calcium was also demonstrated with the aggregation kinetics of the synucleins in which only the α‐synuclein aggregation was affected by calmodulin. A reversible binding assay confirmed that α‐synuclein interacted with the Ca2+‐free as well as the Ca2+‐bound calmodulins with almost identicalKds of 0.35 µmand 0.31 µm, respectively, while β‐synuclein preferentially recognized the Ca2+‐bound form with aKdof 0.68 µm. By using a C‐terminally truncated α‐synuclein of α‐syn97, the calmodulin binding site(s) on α‐synuclein was(were) shown to be located on the N‐terminal region where the amphipathic α‐helices have been suggested to be induced upon membrane interaction. By employing liposome and calmodulin in a state of being either soluble or immobilized on agarose, actual competition of α‐synuclein between membranes and calmodulin was demonstrated with the observation that α‐synuclein previously bound to the liposome was released upon specific interaction with the calmodulins. Taken together, these data may suggest that α‐synuclein could act not only as a negative regulator for calmodulin in the presence and even absence of calcium, but it could also exert its activity at the interface between calmodulin and membranes.
Differential effects of hypoxia–ischemia on subunit expression and tyrosine phosphorylation of the NMDA receptor in 7‐ and 21‐day‐old rats
Authors: Gurd James W. , Bissoon N. , Beesley P. W. , Nakazawa Takanobu , Yamamoto Tadashi , Vannucci S. J. 
Published: March 2003

AbstractThe effect of cerebral hypoxia–ischemia (HI) on levels and tyrosine phosphorylation of the NMDA receptor was examined in 7‐ (P7) and 21 (P21)‐day‐old rats. Unilateral HI was administered by ligation of the right common carotid artery and exposure to an atmosphere of 8% O2/92% N2for 2 (P7) or 1.5 (P21) h. This duration of HI produces significant infarction in nearly all of the survivors with damage being largely restricted to the cortex, striatum, and hippocampus of the hemisphere ipsilateral to the carotid artery ligation. NR2A levels in the right hemisphere of P7 pups were markedly reduced after 24 h of recovery, while NR1 and NR2B remained unchanged. In contrast, NR2B, but not NR2A, was reduced after HI at P21. At both ages, HI resulted in a transient increase in tyrosine phosphorylation of a number of forebrain proteins that peaked between 1 and 6 h of recovery. At both P7 and P21, tyrosine phosphorylation of NR2B was enhanced 1 h after HI and had returned to basal levels by 24 h. HI induced an increase in tyrosine phosphorylation of NR2A in 21 day, but not in 7‐day‐old animals. The differential effects of HI on the NMDA receptor at different post‐natal ages may contribute to changing sensitivity to hypoxia–ischemia.
A new brain‐derived neurotrophic factor transcript and decrease inbrain‐derived neurotrophic factor transcripts 1, 2 and 3 in Alzheimer's disease parietal cortex
Authors: Garzon Diego , Yu Guanhua , Fahnestock Margaret 
Published: March 2003

AbstractBrain‐derived neurotrophic factor (BDNF) supports hippocampal, cortical and basal forebrain cholinergic neurons, which lose function in Alzheimer's disease. In Alzheimer's tissues such as hippocampus and parietal cortex, brain‐ derived neurotrophic factor mRNA is decreased three‐ to four‐fold compared with controls. However, the molecular mechanism of the down‐regulation of BDNF in Alzheimer's disease is unknown. The human brain‐derived neurotrophic factor gene has multiple promoters governing six non‐coding upstream exons that are spliced to one downstream coding exon, leading to six different transcripts. Here we report an alternate human splice variant within exon 4I for a total of seven transcripts. Previous brain‐derived neurotrophic factor mRNA measurements in Alzheimer's disease tissue were done using the downstream coding exon present in all transcripts. Using RT‐PCR primers specific for each upstream exon, we observe a significant decrease in three human brain‐derived neurotrophic factor mRNA transcripts in Alzheimer's disease samples compared with controls. Transcripts 1 and 3 each exhibit a two‐fold decrease, and transcript 2 shows a five‐fold decrease. There are no significant differences between control and Alzheimer's disease samples for the other transcripts, including the new splice variant. In rat, both transcripts 1 and 3 are regulated through the transcription factor cAMP response element binding protein, whose phosphorylation is decreased in the Alzheimer's disease brain. This could lead to specific down‐regulation of the brain‐derivedneurotrophic factor transcripts shown here.
Restricted, but abundant, expression of the novel rat gene‐3 (R3) relaxin in the dorsal tegmental region of brain
Authors: Burazin Tanya C. D. , Bathgate Ross A. D. , Macris Mary , Layfield Sharon , Gundlach Andrew L. , Tregear Geoffrey W. 
Published: March 2003

AbstractRelaxin is a peptide hormone with known actions associated with female reproductive physiology, but it has also been identified in the brain. Only one relaxin gene had been characterized in rodents until recently when a novel human relaxin gene, human gene‐3 (H3) and its mouse equivalent (M3) were identified. The current study reports the identification of a rat homologue, rat gene‐3 (R3) relaxin that is highly expressed in a discrete region of the adult brain. The full R3 relaxin cDNA was generated using RT‐PCR and 3′ and 5′ RACE protocols. The derived amino acid sequence of R3 relaxin retains all the characteristic features of a relaxin peptide and has a high degree of homology with H3 and M3 relaxin. The distribution of R3 relaxin mRNA in adult rat brain was determined and highly abundant expression was only detected in neurons of the ventromedial dorsal tegmental nucleus (vmDTg) in the pons, whereas all other brain areas were unlabelled or contained much lower mRNA levels. Relaxin binding sites and relaxin immunoreactivity were also detected in the vmDTg. These together with earlier findings provide strong evidence for a role(s) for multiple relaxin peptides as neurotransmitters and/or modulators in the rat CNS.
Selective expression of Narp, a secreted neuronal pentraxin, in orexin neurons
Authors: Reti Irving M. , Reddy Radhika , Worley Paul F. , Baraban Jay M. 
Published: March 2003

AbstractRecent studies have provided compelling evidence demonstrating that orexin (also known as hypocretin) neurons play a central role in the pathophysiology of narcolepsy. However, targeted deletion of orexin does not fully mimic the functional deficits induced by selective ablation of these neurons; implying that other secreted signaling molecules expressed in these neurons mediate key aspects of their function. In this study, we demonstrate that orexin neurons display robust expression of neuronal activity‐regulated pentraxin (Narp), a secreted neuronal pentraxin, implicated in regulating clustering of α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate (AMPA) receptors. Furthermore, we have found that hypothalamic melanin‐concentrating hormone (MCH) neurons, which form a peptidergic pathway thought to oppose the effects of the orexin system, express another neuronal pentraxin, NP1. Thus, these findings suggest that these pathways utilize neuronal pentraxins, in addition to neuropeptides, as synaptic signaling molecules.
Regulation of hydrogen peroxide production by brain mitochondria by calcium and Bax
Authors: Starkov Anatoly A. , Polster Brian M. , Fiskum Gary 
Published: March 2003

AbstractAbnormal accumulation of Ca2+ and exposure to pro‐apoptotic proteins, such as Bax, is believed to stimulate mitochondrial generation of reactive oxygen species (ROS) and contribute to neural cell death during acute ischemic and traumatic brain injury, and in neurodegenerative diseases, e.g. Parkinson's disease. However, the mechanism by which Ca2+ or apoptotic proteins stimulate mitochondrial ROS production is unclear. We used a sensitive fluorescent probe to compare the effects of Ca2+ on H2O2 emission by isolated rat brain mitochondria in the presence of physiological concentrations of ATP and Mg2+ and different respiratory substrates. In the absence of respiratory chain inhibitors, Ca2+ suppressed H2O2 generation and reduced the membrane potential of mitochondria oxidizing succinate, or glutamate plus malate. In the presence of the respiratory chain Complex I inhibitor rotenone, accumulation of Ca2+ stimulated H2O2 production by mitochondria oxidizing succinate, and this stimulation was associated with release of mitochondrial cytochrome c. In the presence of glutamate plus malate, or succinate, cytochrome c release and H2O2 formation were stimulated by human recombinant full‐length Bax in the presence of a BH3 cell death domain peptide. These results indicate that in the presence of ATP and Mg2+, Ca2+ accumulation either inhibits or stimulates mitochondrial H2O2 production, depending on the respiratory substrate and the effect of Ca2+ on the mitochondrial membrane potential. Bax plus a BH3 domain peptide stimulate H2O2 production by brain mitochondria due to release of cytochrome c and this stimulation is insensitive to changes in membrane potential.
Ethanol impairs monosaccharide uptake and glycosylation in cultured rat astrocytes
Authors: Tomás M. , Fornas E. , Megías L. , Durán J.‐M. , Portolés M. , Guerri C. , Egea G. , Renau‐Piqueras J. 
Published: March 2003

AbstractAstrocyte and glial–neuron interactions have a critical role in brain development, which is partially mediated by glycoproteins, including adhesion molecules and growth factors. Ethanol affects the synthesis, intracellular transport, subcellular distribution and secretion of these glycoproteins, suggesting alterations in glycosylation. We analyzed the effect of long‐term exposure to low doses of ethanol (30 mm) on glycosylation process in growing cultured astrocytes in vitro. Cells were incubated for short (5 min) and long (90 min) periods with several radioactively labeled carbohydrate precursors. The uptake, kinetics and metabolism of these precursors, as well as the radioactivity distribution in protein gels were analyzed. The levels of GLUT1 and mannosidase II were also determined. Ethanol increased the uptake of monosaccharides and the protein levels of GLUT1 but decreased those of mannosidase II. It altered the carbohydrate moiety of proteins and increased cell surface glycoproteins containing terminal non‐reduced mannose. These results indicate that ethanol impairs glycosylation in rat astrocytes, thus disrupting brain development.
Interaction between GABAA receptor subunit intracellular loops: implications for higher order complex formation
Authors: Nymann‐Andersen Jesper , Wang Hongbing , Sawyer Gregory W. , Olsen Richard W. 
Published: March 2003

AbstractThe majority of fast inhibitory neurotransmission in the CNS is mediated by the GABA type‐A (GABAA) receptor, a ligand‐gated chloride channel. Of the approximately 20 different subunits composing the hetero‐pentameric GABAA receptor, the γ2 subunit in particular seems to be important in several aspects of GABAA receptor function, including clustering of the receptor at synapses. In this study, we report that the intracellular loop of the γ2 subunit interacts with itself as well as with γ1, γ3 and β1–3 subunits, but not with the α subunits. We further show that γ2 subunits interact with photolabeled pentameric GABAA receptors composed of α1, β2/3 and γ2 subunits, and calculate the dissociation constant to be in the micromolar range. By using deletion constructs of the γ2 subunit in a yeast two‐hybrid assay, we identified a 23‐amino acid motif that mediates self‐association, residues 389–411. We confirmed this interaction motif by inhibiting the interaction in a glutathione‐S‐transferase pull‐down assay by adding a corresponding γ2‐derived peptide. Using similar approaches, we identified the interaction motif in the γ2 subunit mediating interaction with the β2 subunit as a 47‐amino acid motif that includes the γ2 self‐interacting motif. The identified γ2 self‐association motif is identical to the interaction motif reported between GABAA receptor and GABAA receptor‐associated protein (GABARAP). We propose a model for GABAA receptor clustering based on GABARAP and GABAA receptor subunit–subunit interaction.