bluuebunny/crossref_metadata_2025 huggingface dataset:
Antibodies Against Rat Brain Vesicle‐Associated Membrane Protein (Synaptobrevin) Prevent Inhibition of Acetylcholine Release by Tetanus Toxin or Botulinum Neurotoxin Type B
Authors: Poulain Bernard , Rossetto Ornella , Deloye Florence , Schiavo Giampietro , Tauc Ladislav , Montecucco Cesare
Published: October 2006

Abstract: Tetanus and botulinum B neurotoxins are zinc endopeptidases that cleave vesicle‐associated membrane protein (VAMP or synaptobrevin) at a single peptide bond. To test the possibility that in vivo also the toxin‐induced blockade of neurotransmission is due to cleavage of VAMP, rat brain VAMP‐specific antibodies were raised in rabbits. IgGs purified from one antiserum, which bind specifically to rat brain VAMP, also specifically recognize proteins from Aplysia californica in immunoblotting. When injected into neurons in the buccal ganglion of Aplysia, these IgGs did not affect the release of acetylcholine but effectively prevented the inhibitory action of both toxins on neurotrans‐ mitter release, thus indicating that the block of neurotransmission by these neurotoxins is consequent to the cleavage of VAMP or specific interaction with VAMP.
τ in Paired Helical Filaments Is Functionally Distinct from Fetal τ: Assembly Incompetence of Paired Helical Filament‐τ
Authors: Yoshida Hirotaka , Ihara Yasuo
Published: October 2006

Abstract: τ in paired helical filaments (PHF‐τ) and fetal τ share several phosphorylated epitopes, as revealed by several phosphorylation‐dependent PHF monoclonals. We have asked whether there is any difference between the two molecules in tubulin assembly‐promoting activity and found that PHF‐τ is almost assembly incompetent, whereas fetal τ is low in this activity but still assembly competent. This indicates that despite substantial similarities in immunoreactivities, PHF‐τ and fetal τ are quite distinct from each other in function.
Pre‐ and Posttranslational Regulation of β‐Endorphin Biosynthesis in the CNS: Effects of Chronic Naltrexone Treatment
Authors: Bronstein David M. , Day Nicola C. , Gutstein Howard B. , Trujillo Keith A. , Akil Huda
Published: October 2006

Abstract: There appear to be two anatomically distinct β‐endorphin (βE) pathways in the brain, the major one originating in the arcuate nucleus of the hypothalamus and a smaller one in the area of the nucleus tractus solitarius (NTS) of the caudal medulla. Previous studies have shown that these two proopiomelanocortin (POMC) systems may be differentially regulated by chronic morphine treatment, with arcuate cells down‐regulated and NTS cells unaffected. In the present experiments, we examined the effects of chronic opiate antagonist treatment on βE biosynthesis across different CNS regions to assess whether the arcuate POMC system would be regulated in the opposite direction to that seen after opiate agonist treatment and to determine whether different βE‐containing areas might be differentially regulated. Male adult rats were administered naltrexone (NTX) by various routes for 8 days (subcutaneous pellets, osmotic minipumps, or repeated intraperitoneal injections). Brain and spinal cord regions were assayed for total βE‐ir, different molecular weight immunoreactive β‐endorphin (βE‐ir) peptides, and POMC mRNA. Chronic NTX treatment, regardless of the route of administration, reduced total βE‐ir concentrations by 30–40% in diencephalic areas (the arcuate nucleus, the remaining hypothalamus, and the thalamus) and the midbrain, but had no effect on βE‐ir in the NTS or any region of the spinal cord. At the same time, NTX pelleting increased POMC mRNA levels in the arcuate to ∼ 140% of control values. These data suggest that arcuate POMC neurons are up‐regulated after chronic NTX treatment (whereas NTS and spinal cord systems remain unaffected) and that they appear to be under tonic inhibition by endogenous opioids. Chromatographic analyses demonstrated that, after chronic NTX pelleting, the ratio of full length βE1–31 to more processed βE‐ir peptides (i.e., βE1–27 and βE1–26) tended to increase in a dose‐dependent manner in diencephalic areas. Because βE1–31 is the only POMC product that possesses opioid agonist properties, and βE1–27 has been posited to function as an endogenous anatgonist of βE1–31, the NTX‐induced changes in the relative concentrations of βE1–31 and βE1–27/βE1–26 may represent a novel regulatory mechanism of POMC cells to alter the opioid signal in the synapse.
Expression of Neurotransmitter Receptors by mRNAs from Neurons Developing In Vitro: A Xenopus Oocyte Expression Study
Authors: Wahl Philip , Ragsdale David , Schousboe Arne , Miledi Ricardo
Published: October 2006

Abstract: Poly(A)+ mRNA was extracted from cultures of neurons isolated from mouse embryonic day 14 cerebral cortex and injected into Xenopus oocytes. This led to the expression of receptors for γ‐aminobutyric acid (GABA), glycine, acetylcholine, serotonin, glutamate, kainate, N‐methyl‐D‐aspartate, and quisqualate. Northern blot analysis of poly(A)+ mRNA from the cultured neurons with a GluRI cDNA probe revealed the presence of three hybridization bands with estimated mRNA sizes of 5.1, 4.0, and 3.1 kb, respectively. The development of mRNAs coding for neurotransmitter receptors was investigated by isolating mRNA from neurons cultured for 2, 8, and 14 days in vitro and injecting it into Xenopus oocytes. The amplitude of membrane currents elicited by the transmitters gave a measure of the relative amounts of the different mRNAs. The size of the responses to kainate, aspartate (together with glycine), glutamate, acetylcholine, GABA, serotonin, and glycine increased with the time of culture in vitro. However, in contrast to all other agonist‐induced currents, the current induced by glycine failed to increase further from 8 to 14 days in culture. It is concluded that the time course of receptor development in cortical neurons in vitro is similar to the development in vivo.
Ischemia‐Induced Accumulation of Extracellular Amino Acids in Cerebral Cortex, White Matter, and Cerebrospinal Fluid
Authors: Shimada Nobumitsu , Graf Rudolf , Rosner Gerd , Heiss Wolf‐Dieter
Published: October 2006

Abstract: In a global model of brain ischemia, accumulation of amino acids was studied in the extracellular space of the auditory cortex and the internal capsule using microdialysis, and in CSF of halothane anesthetized cats. In both brain regions, blood flow determined by hydrogen clearance decreased below 10 ml/100 g/min after extracranial multiple‐vessel occlusion, and extracellular potassium activity (Ke) measured in the dialysate increased significantly. A delayed rise in Ke was observed in CSF. In contrast, ischemic amino acid accumulation differed markedly between the two brain regions investigated. In cortex, transmitter amino acids glutamate, aspartate, and γ‐aminobutyric acid (GABA) rose almost immediately after onset of ischemia, and increased 30‐, 25‐, and 250‐fold, respectively, after 2 h of ischemia. The nontransmitter amino acids taurine, alanine, and serine increased 10‐, seven‐, and fourfold, respectively, whereas glutamine and essential amino acids (valine, phenylalanine, isoleucine, and leucine) increased only 1.5‐fold. In the internal capsule, increases in amino acids, if any, were delayed and much smaller than in cortex. The largest alteration was a fivefold elevation of GABA. In CSF, changes in amino acids were small and comparable to those in the internal capsule. Our results demonstrate that ischemia‐induced extracellular amino acid accumulation is a well localized phenomenon restricted to gray matter structures that possess release and reuptake systems for these substances. We assume that amino acids diffuse slowly into adjacent white matter structures, and into CSF.
Evidence for a GABAergic Projection from the Dorsal Nucleus of the Lateral Lemniscus to the Inferior Colliculus
Authors: Shneiderman A. , Chase M. B. , Rockwood J. M. , Benson C. G. , Potashner S. J.
Published: October 2006

Abstract: This study attempts to determine whether the pathways from the guinea pig dorsal nucleus of the lateral lemniscus (DNLL) to the inferior colliculus (IC) use γ‐aminobutyric acid (GABA) as a transmitter. Injections of kainic acid (KA) were used to destroy neurons in the left DNLL. Two to 4 days after the injection, Nissl‐stained sections through the lesion site showed destruction of the DNLL neurons. The lesions varied in size; 12–100% of the DNLL neurons were destroyed on the injected side without damage to the ipsilateral IC. Two to 4 days after the injection, the electrically evoked, Ca2+‐dependent release and high‐affinity uptake of [3H]GABA were measured in dissected pieces of the left and right IC. These activities were compared with those in the IC taken from unlesioned controls and from sham controls, which received injections of saline instead of KA. Each IC was divided into a dorsal piece, which contained the dorsal cortex and dorsomedial nucleus, and a ventral piece, which contained the central and lateral nuclei. Lesions of the left DNLL depressed the release and uptake of [3H]GABA in the ventral pieces of the IC, but there was a greater depression in the ventral IC contralateral to the lesioned DNLL. There were good correlations between the percentage of neuronal loss in the left DNLL and deficits in [3H]GABA release and uptake activities in the ipsi‐ and contralateral ventral IC. By contrast, there was no depression of [3H]GABA release and uptake in the dorsal pieces of the IC. The localization of the deficits in release and uptake appears to match the distribution of the synaptic endings of the DNLL pathways in the IC. This correspondence associates GABA release and uptake activities with the DNLL projections to the IC and, therefore, suggests that GABA may be a transmitter of these pathways. The release and uptake of [14C]glycine was also measured to determine whether glycine might be a transmitter of the DNLL pathways to the IC. Lesions of the left DNLL failed to alter the Ca2+‐dependent release or the uptake of [14C]glycine, suggesting that DNLL neurons are unlikely to use this compound as a transmitter.
Extracellular Dopamine and Serotonin in the Rat Striatum During Transient Ischaemia of Different Severities: A Microdialysis Study
Authors: Richards D. A. , Obrenovitch T. P. , Symon L. , Curzon G.
Published: October 2006

Abstract: Generalised neurotransmitter overflow into the extracellular space on cerebral ischaemia has been widely reported and implicated in events leading to subsequent neu‐ronal death. As little is known about the effect of depth of ischaemia on these changes, we have subjected anaesthetised rats to a sequence of four challenges [high K+ stimulus, moderate (penumbral) ischaemia, severe ischaemia, cardiac arrest] and have concurrently monitored both electrophysio‐logical parameters and changes in extracellular dopamine, serotonin, and their metabolites in the striatum. Oi'particu‐lar relevance to human stroke therapy was penumbral ischaemia, where ionic homeostasis was maintained even though electrical function was lost. All challenges increased extracellular monoamines, although levels were significantly greater when ischaemia was severe enough to produce sustained anoxic depolarisation. Baseline levels were rapidly restored during recovery phases. Acidic monoamine metabolites decreased significantly during each insult, returning to basal levels during reperfusion after moderate ischaemia, and to significantly higher levels after severe ischaemia. Results indicate that sustained anoxic depolarisation may be a critical factor in determining outcome after ischaemia, being associated with significantly greater release of monoamines, and impairment of electrical function recovery.
Kainate‐Induced Changes in Opioid Peptide Genes and AP‐1 Protein Expression in the Rat Hippocampus
Authors: Pennypacker K. R. , Walczak D. , Thai L. , Fannin R. , Mason E. , Douglass J. , Hong J. S.
Published: October 2006

Abstract: In the rat hippocampus, jun, c‐fos, and fos‐related antigen immunoreactivity, AP‐1 DNA binding, and opioid peptide gene expression were examined after kainate treatment to determine whether the induction and DNA binding of AP‐1 transcription factors are correlated with the expression of the opioid peptide genes. One and one‐half hours after kainate administration, fos‐related antigen and jun immunoreactivity and AP‐1 DNA binding were induced; maximal elevation was observed after 4.5 h. Transcription factor expression and DNA binding increased in a dose‐dependent manner. Preprodynorphin and preproenkephalin mRNA induction was also dose dependent. The anticonvulsants, pentobarbital and diazepam, effectively blocked electroencephalographic seizure activity caused by kainate treatment, whereas valproic acid was approximately 50% effective. Opioid peptide gene expression, fos‐related antigen and jun immunoreactivity, and AP‐1 DNA binding all reflected similar reductions after anticonvulsant treatment. Therefore, expression and DNA binding activity of the AP‐1 transcription factors are correlated with opioid peptide gene expression in the rat hippocampus.
Phospholipase A2 Modulates Different Subtypes of Excitatory Amino Acid Receptors: Autoradiographic Evidence
Authors: Catania M. V. , Hollingsworth Z. , Penney J. B. , Young A. B.
Published: October 2006

Abstract: Exogenous phospholipases have been used extensively as tools to study the role of membrane lipids in receptor mechanisms. We used in vitro quantitative autoradiography to evaluate the effect of phospholipase A2 (PLA2) on N‐methyl‐D‐aspartate (NMDA) and non‐NMDA glutamate receptors in rat brain. PLA2 pretreatment induced a significant increase in α‐[3H]amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate ([3H]AMPA) binding in the stratum radiatum of the CA1 region of the hippocampus and in the stratum moleculare of the cerebellum. No modification of [3H]AMPA binding was found in the stratum pyramidale of the hippocampus at different ligand concentrations. [3H]‐Glutamate binding to the metabotropic glutamate receptor and the non‐NMDA‐, non‐kainate‐, non‐quisqualate‐sensitive [3H]glutamate binding site were also increased by PLA2 pretreatment. [3H]Kainate binding and NMDA‐sensitive [3H]glutamate binding were minimally affected by the enzyme pretreatment. The PLA2 effect was reversed by EGTA, the PLA2 inhibitor p‐bromophenacyl bromide, and prolonged pretreatment with heat. Bovine serum albumin (1%) prevented the increase in metabotropic binding by PLA2. Arachidonic acid failed to mimic the PLA2 effect on metabotropic binding. These results indicate that PLA2 can selectively modulate certain subtypes of excitatory amino acid receptors. This effect is due to the enzymatic activity but is probably not correlated with the formation of arachidonic acid metabolites. Independent of their possible physiological implications, our results provide the first autoradiographic evidence that an enzymatic treatment can selectively affect the binding properties of excitatory amino acid receptors in different regions of the CNS.
An Improved Solid‐Phase Bioassay for Evaluation of Neurotrophic Factors
Authors: Nakajima Mitsunari , Furukawa Shoei , Hayashi Kyozo , Kawashima Takuji , Hayashi Yokichi
Published: October 2006

Abstract: A bioassay system originally described by other investigators was improved to evaluate the biological activities of neurotrophic factors (NTFs) and is referred to as a solid‐phase bioassay (SPB). The principle and an outline of the SPB are as follows: (a) Test samples containing NTFs are applied to polyacrylamide gel electrophoresis (PAGE) of the sodium dodecyl sulfate (SDS) or two‐dimensional type and then transferred onto nitrocellulose membranes, (b) neurons are cultured on the protein‐blotted membranes, and (c) the distributions of the surviving neurons are estimated following fixation and clarification of the nitrocellulose membranes. The rationale is that neuron survival will be restricted to the migration positions of the NTF(s). We used nerve growth factor (NGF) as a NTF and sympathetic neurons of 10‐ to 12‐day‐old chick embryos as NGF‐responsive neurons. Neuronal survival was observed in the areas coinciding with the positions of NGF migration on the membranes in both systems following SDS‐PAGE and two‐dimensional PAGE. These results indicate that the SPB can identify the positions of NGF‐like molecules on nitrocellulose membranes. Using this system, we found active entities, with molecular masses of ∼ 100–200 kDa, different from NGF in crude extracts of mouse submaxillary glands. The SPB developed is considered to be a useful tool for obtaining information on the physicochemical and/or biological properties of putative NTFs in crude samples.
Arachidonic Acid and Other Unsaturated Fatty Acids Alter Membrane Potential in PC12 and Bovine Adrenal Chromaffin Cells
Authors: Ehrengruber M. U. , Deranleau D. A. , Kempf C. , Zahler P. , Lanzrein M.
Published: October 2006

Abstract: The action of arachidonic acid and other fatty acids on membrane potential in PC 12 and bovine chromaffin cells was investigated using a membrane potential‐sensitive fluorescent dye. Arachidonic acid (1–40 μM) provoked dose‐dependent membrane hyperpolarization, thereby reducing hyperpolarization induced by the K+‐selective ionophore valinomycin. Other cis‐unsaturated fatty acids, but not lipoxygenase products or the saturated fatty acid palmitic acid, also affected membrane potential. Tetraethylammonium blocked the arachidonic acid‐induced hyperpolarization. These data suggest that cis‐unsaturated fatty acids alter membrane potential in PC 12 and bovine chromaffin cells by modulating K+ conductances. Valinomycin‐generated hyperpolarization had no effect on agonist‐induced Ca2+ influx into bovine chromaffin cells, whereas preincubation with arachidonic acid and other cis‐unsaturated fatty acids blocked Ca2+ influx and secretion. We propose a model where internally generated fatty acids act as a feed‐back to desensitize the stimulated cell via inhibition of receptor‐dependent Ca2+ influx and induction of membrane hyperpolarization.
Morphine Inhibits Calcium Influx and the Response to Acetylcholine in Xenopus Oocytes
Authors: Lipinsky D. , Oron Y. , Sarne Y.
Published: October 2006

Abstract: Incubation of intact Xenopus oocytes with the opioid radioligand [3H]diprenorphine (0.5 nM) resulted in specific binding of 1.7 ± 0.3 fmol per oocyte. Morphine (10 μM) inhibited the uptake of 45Ca2+ into the oocyte by 66 ± 9%. The opioid antagonist naltrexone partially blocked this effect of morphine. Preincubation of oocytes with morphine (10 μM, 2 min) partially inhibited the fast and slow responses of the oocyte to acetylcholine by 26 and 52%, respectively. We conclude that native Xenopus oocytes possess opioid receptors that may modulate the muscarinic response by limiting calcium influx into the cell.
Type I Calmodulin‐Sensitive Adenylyl Cyclase Is Neural Specific
Authors: Xia Zhengui , Choi Eui‐Ju , Wang Fan , Blazynski Christine , Storm Daniel R.
Published: October 2006

Abstract: The distribution of type I calmodulin‐sensitive adenylyl cyclase in bovine and rat tissues was examined by northern blot analysis and in situ hybridization. Northern blot analysis using poly(A)+‐selected RNA from various bovine tissues indicated that mRNA for type I adenylyl cyclase was found only in brain, retina, and adrenal medulla, suggesting that this enzyme is neural specific. In situ hybridization studies using bovine, rabbit, and rat retina indicated that mRNA for type I adenylyl cyclase is found in all three nuclear layers of the neural retina and is particularly abundant in the inner segment of the photoreceptor cells. The neural‐specific distribution of type I adenylyl cyclase mRNA and its restricted expression in areas of brain implicated in neuroplasticity are consistent with the proposal that this enzyme plays an important role in various neuronal functions including learning and memory.
N‐Methyl‐d‐Aspartate Receptor Excitotoxicity Involves Activation of Polyamine Synthesis: Protection by α‐Difluoromethylornithine
Authors: Trout Jerome J. , Koenig Harold , Goldstone Alfred D. , Iqbal Zafar , Lu Chung Y. , Siddiqui Farida
Published: October 2006

Abstract: We investigated the role of polyamines and their regulatory enzyme ornithine decarboxylase in N‐Methyl‐D‐aspartate‐induced excitotoxicity in embryonic chick retina. N‐Methyl‐D‐aspartate (200 μM) produced an early increase in ornithine decarboxylase activity, putrescine concentration, and Ca2+ entry, leading to selective neuronal death by 30 min. This response was attenuated by the ornithine decarboxylase inhibitor α‐difluoromethylornithine and the N‐methyl‐D‐aspartate receptor antagonist 5‐aminophosphonovaleric acid. Exogenous putrescine increased intracellular putrescine and spermine levels and reversed neuroprotection by α‐difluoromethylornithine, but not by 5‐aminophosphonovaleric acid. N‐Methyl‐D‐aspartate‐receptor stimulation of putrescine/polyamine synthesis mediates abnormal Ca2+ entry and acute excitotoxic neuronal death. Postreceptor inhibition of the ornithine decar‐boxylase/polyamine cascade by α‐difluoromethylornithine may provide neuroprotection against N‐methyl‐D‐aspartate‐induced excitotoxicity.
Brain α‐Ketoglutarate Dehydrotenase Complex Activity in Alzheimer's Disease
Authors: Mastrogiacomo Frank , Bergeron Catherine , Kish Stephen J.
Published: October 2006

We measured the activity of the a‐ketoglutarate dehydrogenase complex (α‐KGDHC), a rate‐limiting Krebs cycle enzyme, in postmortem brain samples from 38 controls and 30 neuropathologically confirmed Alzheimer's disease (AD) cases, in both the presence and absence of thiamine pyrophosphate (TPP), the enzyme's cofactor. Statistically significant correlations between brain pH and lactate levels and α‐KGDHC activity in the controls were observed, suggesting an influence of agonal status on the activity of α‐KGDHC. As compared with the controls, mean α‐KGDHC activity, with added TPP, was significantly (p < 0.005) reduced in AD brain in frontal (‐56%), temporal (‐60%), and parietal (‐68%) cortices, with the reductions (‐25 to ‐53%) in the occipital cortex, hippocampus, amygdala, and caudate failing to reach statistical significance. In the absence of exogenously administered TPP, mean a‐KGDHC activity was reduced to a slightly greater extent in all seven AD brain areas (‐39 to ‐83%), with the reductions now reaching statistical significance in the four cerebral cortical areas and hippocampus. A statistically significant negative correlation was observed between α‐KGDHC activity and neurofibrillary tangle count in AD parietal cortex, the brain area exhibiting the most marked reduction in enzyme activity; this suggests that the enzyme activity reduction in AD brain may be related to the disease process and severity. In each brain area examined, TPP produced a greater stimulatory effect on α‐KGDHC activity in the AD group (23–280% mean stimulation) as compared with the controls (‐4 to ±50%); this TPP effect could be explained by reduced endogenous TPP levels in AD brain. Reduced brain α‐KGDHC activity could be consequent to loss of neurons preferentially enriched in α‐KGDHC, a premortem reduction in TPP levels (which may have affected enzyme stability), elevated brain levels of the α‐KGDHC inhibitor ammonia, or an actual failure in the expression of the gene encoding the enzyme. We suggest that a defect in this key Krebs cycle enzyme could contribute to an impairment of cerebral energy metabolism and the brain dysfunction in AD.
Expression of Facilitative Glucose Transporter Isoforms in Human Brain Tumors
Authors: Nagamatsu Shinya , Sawa Hiroki , Wakizaka Akira , Hoshino Takao
Published: October 2006

The expression of facilitative glucose transporter (GLUT) isoforms in human astrocytic tumors was examined. Reverse transcriptase‐polymerase chain reaction of a surgically biopsied glioblastoma was carried out using the degenerative oligonucleotide primers corresponding to the sequences of the human facilitative glucose transporter family, and polymerase chain reaction products were hybridized with human GLUT1, GLUT2, GLUT3, GLUT4, and GLUT5 cDNA probes. The results showed that a biopsied glioblastoma expressed GLUT1, GLUT3, and GLUT4 glucose transporter genes. Northern blot analysis of total RNA (10 μg) from a biopsied glioblastoma showed the transcripts of only GLUT1 and GLUT3, suggesting that the expression of insulin‐responsive glucose transporter GLUT4 mRNA is relatively low. Immunoblot analysis of biopsied glioblastoma tissues by polyclonal antibodies against the C‐terminal synthetic peptides of GLUT1, GLUT3, and GLUT4 showed a single band of each polypeptide. However, elevated expression of GLUT1 and GLUT3 glucose transporters was not observed in the glioblastoma. Astrocytic tumor tissues (n = 14) were also examined immunohistochemically. Reactive products for GLUT1 were observed in the luminal surface of capillaries in all cases, whereas tumor cells were positive for GLUT1 in only two of 14 cases. GLUT3 was positive in astrocytic tumor cells in all cases. Three of 14 cases expressed the GLUT4 protein, which was localized in the cytoplasm of tumor cells. These results suggest that the facilitative glucose transport may be altered in astrocytic tumor cells and thus display a significant change in glucose metabolism.
Kynurenine Pathway Enzymes in Brain: Responses to Ischemic Brain Injury Versus Systemic Immune Activation
Authors: Saito Kuniaki , Nowak Thaddeus S. , Suyama Kazuhiko , Quearry Bonnie J. , Saito Misato , Crowley Jeffrey S. , Markey Sanford P. , Heyes Melvyn P.
Published: October 2006

Accumulation of l‐kynurenine and quinolinic acid (QUIN) in the brain occurs after either ischemic brain injury or after systemic administration of pokeweed mitogen. Although conversion of l‐[13C6]tryptophan to [13C6]‐QUIN has not been demonstrated in brain either from normal gerbils or from gerbils given pokeweed mitogen, direct conversion in brain tissue does occur 4 days after transient cerebral ischemia. Increased activities of enzymes distal to indoleamine‐2,3‐dioxygenase may determine whether l‐kynurenine is converted to QUIN. One day after 10 min of cerebral ischemia, the activities of kynureninase and 3‐hydroxy‐3,4‐dioxygenase were increased in the hippocampus, but local QUIN levels and the activities of the indoleamine‐2,3‐dioxygenase and kynurenine‐3‐hydroxylase were unchanged. By days 2 and 4 after ischemia, however, the activities of all of these enzymes in the hippocampus as well as QUIN levels were significantly increased. Kynurenine aminotransferase activity in the hippocampus was unchanged on days 1 and 2 after ischemia but was decreased on day 4, at a time when local kynurenic acid levels were unchanged. A putative precursor of QUIN, [13C6]anthranilic acid, was not converted to [13C6]‐QUIN in the hippocampus of either normal or 4‐day postischemic gerbils. Gerbil macrophages stimulated by endo‐toxin in vitro converted l‐[13C6]tryptophan to [13Ce]QUIN. Kinetic analysis of kynurenine‐3‐hydroxylase activity in the cerebral cortex of postischemic gerbils showed that Vmax increased, without changes in Km. Systemic administration of pokeweed mitogen increased indoleamine‐2,3‐dioxygenase and kynureninase activities in the brain without significant changes in kynurenine‐3‐hydroxylase or 3‐hydroxyanthranilate‐3,4‐dioxygenase activities. Increases in kynurenine‐3‐hydroxylase activity, in conjunction with induction of indoleamine‐2,3‐dioxygenase, kynureninase, and 3‐hydroxyanthranilate‐3,4‐dioxygenase in macro‐phage infiltrates at the site of brain injury, may explain the ability of postischemic hippocampus to convert l‐[13C6]tryptophan to [13C6]QUIN.
Interconvertible Kinetic States of t‐Butylbicycloorthobenzoate Binding Sites of the γ‐Aminobutyric AcidA Ionophores
Authors: Maksay Gabor , Van Rijn Clementina M.
Published: October 2006

The kinetics of t‐[3H]butylbicycloorthobenzoate (TBOB) binding to the convulsant sites of the γ‐aminobutyric acidA (GABAA) receptor‐ionophore complex were examined in synaptosomal membrane preparations of rat brain. On and off rates of TBOB binding were accelerated by 1 μM GABA and decelerated by 1 μM bicuculline methochloride, a GABAA antagonist. The presence of GABA and bicuculline methochloride created rapid and slow phases of dissociation, respectively. The three groups of rate constants distinguished for the dissociation of 4 nM and 30 nM [3H]TBOB represent multiaffinity states of the convulsant sites depending on the presence of GABA or bicuculline methochloride. Apparent association rate constants do not obey the equation kapp=koff±kon [TBOB] without assuming interconvertibility of the kinetic states during binding. Avermectin B1a (AVM B1a), a chloride channel opening agent, accelerated the association and dissociation of TBOB and resulted in a biphasic effect on TBOB binding, i.e., enhancement at low concentrations (EC50, 7.8 nM) followed by displacement at high concentrations (IC50 6.3 μM) of AVM B1a. AVM B1a resulted in similar biphasic effects on t‐ [35S]butylbicyclophosphorothionate binding. DIDS, an isothiocyanatostilbene derivative with irreversible anion channel blocking effect, selectively inhibited basal [3H]TBOB binding (IC50 125 μM DIDS) leaving the enhancement by AVM B1a unaffected.
Soluble and Particulate Organophosphorus Neuropathy Target Esterase in Brain and Sciatic Nerve of the Hen, Cat, Rat, and Chick
Authors: Tormo Nieves , Gimeno Juan Ramón , Sogorb Miguel Angel , Díaz‐Alejo Nuria , Vilanova Eugenic
Published: October 2006

Considerable evidence exists suggesting that the so‐called neuropathy target esterase (NTE) is involved in the mechanisms responsible for organophosphorus‐induced delayed polyneuropathy (OPIDP). Earlier studies in the adult hen, the habitually employed experimental model in OPIDP, have shown that most NTE activity in the brain is centered in paniculate fractions, whereas approximately 50% of this activity in the sciatic nerve is encountered in soluble form, with the rest being paniculate NTE. In the present work, we have studied the paniculate and soluble fractional distribution of paraoxon‐resistant phenylvalerate esterase activity (B activity), parabxon‐ and mipafox‐resistant phenylvalerate esterase activity (C activity), and NTE activity (B ‐ C) according to ultracentrifugation criteria (100,000 g for 1 h). To this effect, two sensitive (adult hen and cat) and two scarcely sensitive (rat and chick) models were used. In all four experimental models, the distribution pattern was qualitatively similar: B activity and total NTE were much greater in brain (900–2, 300 nmol/min/g of tissue) than in sciatic nerve (50–100 nmol/min/g of tissue). The proportion of soluble NTE in brain was very low (<2%), whereas its presence in sciatic nerve was substantial (30–50%). The NTE/B ratio in brain was high for the particulate fraction (>60%) and low in the soluble fraction (7–30%); in sciatic nerve the ratio was about 50% in both fractions. Slight quantitative differences were observed in terms of OPIDP sensitivity: the proportion of soluble NTE in sciatic nerve was slightly higher in the sensitive animals (hen and cat: 49 and 44%, respectively) than in the rat and chick (41 and 37%, respectively), although no differences were noted in terms of concentration (in nanomoles per minute per gram of tissue). It is concluded that the distribution pattern of the activities studied is similar in all four experimental models, with no important quantitative differences directly related to species sensitivity or age.
Decline in Response to Nicotine in Aged Rat Striatum: Correlation with a Decrease in a Subpopulation of Nicotinic Receptors
Authors: Schulz David W. , Kuchel George A. , Zigmond Richard E.
Published: October 2006

Specific and reproducible changes involving the cholinergic and dopaminergic systems have been described in both the aging rodent and the human nervous system. Nevertheless, relatively little information is available on changes in nicotinic cholinergic receptors occurring in normal aging, and there have been few attempts to correlate alterations in receptor densities with changes in nicotinic actions. We have utilized the nicotine‐mediated stimulation of endogenous dopamine efflux in a striatal slice preparation as a functional index of responsiveness to nicotine in aging. Following incubation with nicotine, this efflux was significantly lower in 25‐month‐old (aged) as opposed to 4‐month‐old (young) rats. In contrast, the release of striatal dopamine following a high‐potassium stimulus was similar at both ages. Binding studies in young and aged animals did not reveal any significant change with age in the total number of striatal nicotinic receptors recognized by either [3H]nicotine or the neuronal nicotinic antagonist 125l‐neuronal bungarotoxin. However, there was a nearly 80% decline in the subpopulation of striatal nicotinic receptors jointly recognized by both nicotine and neuronal bungarotoxin, but not by α‐bungarotoxin. Quantitative autoradiography demonstrated declines with age in this receptor subtype in several brain regions examined. Decrements in this specific subpopulation of nicotinic receptors or in the nerve cells expressing these receptors may contribute to the functional declines that take place in the aging motor and visual systems.
The Ontogeny of Apomorphine‐Induced Alterations of Neostriatal Dopamine Release: Effects on Spontaneous Release
Authors: Andersen Susan L. , Gazzara Russell A.
Published: October 2006

The effects of apomorphine (0.05, 0.1, and 1.0 mg/kg, s.c.) on the extracellular levels of dopamine and the dopamine metabolite 3, 4‐dihydroxyphenylacetic acid were studied through the use of in vivo microdialysis in the neostriatum of developing and adult rats. Fifteen‐minute samples were collected from urethane‐anesthetized rats 5, 10–11, 21–22, and 35–36 days old and adults and quantified by HPLC with electrochemical detection. Apomorphine attenuated extracellular levels of dopamine in all age groups, suggesting that the dopamine autoreceptor modulating release in the neostriatum is functional by 5 days of age. A dose‐response effect of apomorphine on extracellular dopamine was observed in all age groups except at 10–11 days of age. Extracellular levels of 3, 4‐dihydroxyphenylacetic acid were also significantly decreased in all age groups, consistent with the hypothesis that synthesis‐modulating dopamine autoreceptors in the neostriatum are functional by 5 days of age. Apomorphine had a significantly greater effect on extracellular 3, 4‐dihydrpxyphenylacetic acid levels at the 0.05 and 0.1 mg/kg doses in the 5‐ and 10–11‐day‐old age groups compared with the other ages. Absolute levels of extracellular dopamine were significantly attenuated at 5 days of age compared with the other ages, and absolute levels of extracellular 3, 4‐dihydroxyphenylacetic acid monotonically increased with age.
Sex Differences in Acute Swim Stress‐Induced Changes in the Binding of MK‐801 to the NMDA Subclass of Glutamate Receptors in Mouse Forebrain
Authors: Akinci Muallã K. , Johnston Graham A. R.
Published: October 2006

Acute swim stress (3 min at 32°C) in mice produces increases in the binding of MK‐801 to the NMDA subclass of glutamate receptors to forebrain membranes prepared from male mice. Scatchard analyses indicate that the observed increases in the binding of MK‐801 in membranes from male mice are the result of changes in the affinity and density of low‐affinity binding sites and in the density of high‐affinity binding sites. In female mice, any changes in the binding of MK‐801 appear to be much less pronounced and restricted to the low‐affinity binding sites. These results are in contrast to the situation with binding to GABA receptors where acute swim stress increases GABA binding in forebrain membranes much more in female than in male mice. This indicates significant sex differences in the responses of receptors for the major excitatory and inhibitory transmitters to acute swim stress. These rapid changes in MK‐801 binding may result from changes in endogenous modulators as appears to be the case in the acute swim stress‐induced changes in GABA binding. As with GABA binding, the endogenous modulators are likely to include steroids, the sex differences reflecting differences in modulation by gonadal steroids and the stress‐induced changes reflecting differences in modulation by adrenal steroids. Estradiol, progesterone, and corticosterone treatments have been reported by other workers to influence the properties of glutamate receptors.
Lithium Decreases Membrane‐Associated Protein Kinase C in Hippocampus: Selectivity for the α Isozyme
Authors: Manji Husseini K. , Etcheberrigaray René , Chen Guang , Olds James L.
Published: October 2006

We investigated the effects of lithium on alterations in the amount and distribution of protein kinase C (PKC) in discrete areas of rat brain by using [3H]phorbol 12, 13‐dibutyrate quantitative autoradiography as well as western blotting. Chronic administration of lithium resulted in a significant decrease in membrane‐associated PKC in several hippocampal structures, most notably the subiculum and the CA1 region. In contrast, only modest changes in [3H]phorbol 12, 13‐dibutyrate binding were observed in the various other cortical and subcortical structures examined. Immunoblotting using monoclonal anti‐PKC antibodies revealed an isozyme‐specific 30% decrease in hippocampal membrane‐associated PKC α, in the absence of any changes in the labeling of either the β(I/II) or γ isozymes. These changes were observed only after chronic (4 week) treatment with lithium, and not after acute (5 days) treatment, suggesting potential clinical relevance. Given the critical role of PKC in regulating neuronal signal transduction, lithium's effects on PKC in the limbic system represent an attractive molecular mechanism for its efficacy in treating both poles of manic‐depressive illness. In addition, the decreased hippocampal membrane‐associated PKC observed in the present study offers a possible explanation for lithium‐induced memory impairment.
Inhibition of β‐Amyloid Production by Activation of Protein Kinase C
Authors: Gabuzda Dana , Busciglio Jorge , Yankner Bruce A.
Published: October 2006

The cellular factors regulating the generation of β‐amyloid from the amyloid precursor protein (APR) are unknown. Activation of protein kinase C (PKC) by phorbol ester treatment inhibited the generation of the 4‐kDa β‐amyloid peptide in transfected COS cells, a human glioma cell line, and human cortical astrocytes. An analogue of diacylglycerol, the endogenous cellular activator of PKC, also inhibited the generation of β‐amyloid. Activation of PKC increased the level of secreted APP in transfected COS cells but did not significantly affect the level of secreted APP in primary human astrocytes or in the glioma cell line. Cell‐associated APP and the secreted APP derivative, but not β‐amyloid, were phosphorylated on serine residues. Activation of PKC did not increase the level of APP phosphorylation, suggesting that PKC modulates the proteolytic cleavage of APP indirectly by phosphorylation of other substrates. These results indicate that PKC activation inhibits β‐amyloid production by altering APP processing and suggest that β‐amyloid production can be regulated by the phospholipase C‐diacylglycerol signal transduction pathway.
GABA Alters GABAA Receptor mRNAs and Increases Ligand Binding
Authors: Kim Helen Y. , Sapp Douglas W. , Olsen Richard W. , Tobin Allan J.
Published: October 2006

In addition to its role as an inhibitory neurotransmitter, γ‐aminobutyric acid (GABA) influences the cytodifferentiation of developing neurons both in culture and in vivo. Here, we report some of the targets of GABA action and the mechanism through which GABA acts. In primary cultures of cerebellar granule cells, GABA specifically stimulates an increase in the levels of mRNAs for α 1 and β 2 GABAA receptor subunits. The GABAA agonist 4, 5, 6, 7‐tetrahydroisoxazolo [5, 4‐c]pyridin‐3‐ol (THIP) mimics this effect, and the GABAA antagonist bicuculline prevents it. In addition, GABA and THIP trigger an increase in the number of GABA binding sites. This increase parallels that seen in vivo, where the total number of GABAA receptor sites increases during postnatal cerebellar development. It is interesting that the period of the greatest increase in the number of receptor sites coincides with the development of the granule cells. Taken together, our data suggest that GABA may play an important role during maturation of cerebellar granule cells by influencing the number and composition of its own receptors.
Serotonin and Dopamine Sensitization in the Nucleus Accumbens, Ventral Tegmental Area, and Dorsal Raphe Nucleus Following Repeated Cocaine Administration
Authors: Parsons L. H. , Justice J. B.
Published: October 2006

Abstract— The present study was designed to examine the effects of chronic cocaine administration on the extracellular response of serotonin (5‐HT) and dopamine (DA) to a peripheral cocaine injection using in vivo brain microdialysis in awake rats. Two different dual probe preparations were used: One group of animals had guide cannulae aimed at the ventral tegmental area (VTA) and nucleus accumbens (N ACC) and a second group of animals had guide cannulae aimed at the dorsal raphe nucleus (DRN) and N ACC. Rats from both groups were given daily injections of either cocaine (20 mg/kg i.p.) or saline (0.9%; 0.05 ml/kg i.p.) for 10 consecutive days. On day 11, baseline dialysate levels of DA, 5‐HT, dihydroxyphenylacetic acid, and 5‐hydroxyindoleacetic acid were obtained from either the N ACC and VTA or the N ACC and DRN, followed by a 10 mg/kg i.p. cocaine injection and an additional 150 min of dialysate sampling. The percent baseline increases of both 5‐HT and DA were significantly higher in the N ACC, VTA, and DRN of animals that received daily injections of cocaine compared with saline controls (p < 0.05, in each region). Maximum dialysate 5‐HT concentrations after cocaine challenge were significantly higher in the N ACC and VTA (p < 0.05) and DRN (p < 0.01) of chronically treated animals compared with saline controls. Maximum dialysate DA concentrations were significantly higher in the N ACC and DRN (p < 0.05) of chronically treated animals compared with saline controls. There was no significant difference between acute and chronic animals in the maximum dialysate DA concentration from the VTA after cocaine challenge. 5‐HT was significantly more sensitized in the 5‐HT cell body region (DRN) than the N ACC terminal field (p < 0.05), whereas DA was significantly more sensitized in the N ACC terminal field than the DA cell bodies of the VTA (p < 0.05).
Messenger RNA Levels of Chromogranin B, Secretogranin II, and VGF in Rat Brain After AF64A‐Induced Septohippocampal Cholinergic Lesions
Authors: Mahata Manjula , Hörtnagl Heide , Mahata Sushil K. , Fischer‐Colbrie Reiner , Winkler Hans
Published: October 2006

Abstract— The mRNA levels of secretogranin II, chromo‐granin B, and VGF were compared in brains of control and AF64A‐treated rats. This toxin induces specific lesions of the septohippocampal cholinergic pathway. As a consequence of this treatment, the Chromogranin B message was elevated in the dentate gyrus granule cells of the hippocampus. In the paraventricular nucleus of the hypothalamus, a concomitant elevation of the messages of secretogranin II and corticotropin‐releasing factor occurred in the parvocellular neurons, and an increase of those of secretogranin II and VGF occurred in a subgroup of magnocellular neurons. Further increases for secretogranin II were seen in the amygdaloid nuclei and the reticular thalamic nuclei and increases for Chromogranin B in the temporal cortex, substantia nigra compacta, and ventral tegmental area. These results indicate that the toxin‐induced lesion of the cholinergic pathway innervating the hippocampus apparently leads to the stimulation of several defined groups of neurons that react with an increase in the mRNA levels of their secretory peptides. We suggest that changes in mRNA expression of these peptides are useful parameters for defining neurons under chronic stimulation. Key Words: Secretory peptides—Large dense core vesicles—Corticotropin releasing factor—Septohippocampal cholinergic system—Hippocampus—AF64A.
Coupling Among Energy Failure, Loss of Ion Homeostasis, and Phospholipase A2 and C Activation During Ischemia
Authors: Katsura Ken‐ichiro , Rodriguez de Turco Elena B. , Folbergrová Jaroslava , Bazan Nicolas G. , Siesjö Bo K.
Published: October 2006

Abstract— The objective of the present experiments was to correlate changes in cellular energy metabolism, dissipative ion fluxes, and lipolysis during the first 90 s of ischemia and, hence, to establish whether phospholipase A2or phospholipase C is responsible for the early accumulation of phospholipid hydrolysis products. Ischemia was induced for 15–90 s in rats, extracellular K+ (K+e) was recorded, and neocortex was frozen in situ for measurements of labile tissue metabolites, free fatty acids, and diacylglycerides. Ischemia of 15‐and 30‐s duration gave rise to a decrease in phosphocreatine concentration and a decline in the ATP/free ADP ratio. Although these changes were accompanied by an activation of K+ conductances, there were no changes in free fatty acids until after 60s, when free arachidonic acid accumulated. An increase in other free fatty acids and in total diacylglyceride content did not occur until after anoxic depolarization. The results demonstrate that the early functional changes, such as activation of K+ conductances, are unrelated to changes in lipids or lipid mediators. They furthermore suggest that the initial lipolysis occurs via both phospholipase A2 and phospholipase C, which are activated when membrane depolarization leads to influx of calcium into cells.
Effect of Systemic Administration of N‐Methyl‐d‐Aspartic Acid on Extracellular Taurine Level Measured by Microdialysis in the Hippocampal CA1 Field and Striatum of Rats
Authors: Shibanoki Shinji , Kogure Miki , Sugahara Megumi , Ishikawa Koichi
Published: October 2006

Abstract: The extracellular concentrations of amino acids in the hippocampal CA1 field and striatum of conscious freely moving rats were monitored simultaneously by in vivo brain microdialysis using HPLC with electrochemical detection. Under basal conditions, aspartate, glutamate, glutamine, glycine, taurine, and alanine were detected, but γ‐aminobutyric acid was undetectable in both regions. In‐traperitoneal injection of N‐methyl‐d‐aspartic acid (NMDA; 10 mg/kg) caused a significant increase (three‐to fivefold) in the taurine concentration in the dialysate obtained from both the hippocampal CA1 and striatum, whereas other amino acids (aspartate, glutamate, and alanine) did not show significant changes. Local application of NMDA (300 γ) to both regions via the dialysis probes also caused a similar increase (three‐to fivefold) in both regions. Under infusion of hypertonic Ringer's solution containing 150 mM sucrose, the effect of NMDA on the level of taurine in both the regional dialysates was not affected. The effect of NMDA was totally reduced by intraperitoneal administration of MK‐801 (0.3–1.0 mg/kg), a noncompetitive antagonist of NMDA receptors. Continuous infusion of dl‐2‐amino‐5‐phosphonovaleric acid (1.0 mM), a competitive antagonist of NMDA receptors, via the dialysis probes completely inhibited the effect of NMDA. These findings suggest that systemic administration of NMDA is effective as well as local administration into the brain and that NMDA receptors might be involved in the regulation of the extracellular taurine level in the brain without dependence on cell swelling.