Specific Regions Of Rat Brain Research Articles

High fat diet increases the incidence of orofacial dyskinesia and oxidative stress in specific brain regions of rats

Haloperidol-induced orofacial dyskinesia (OD) is a putative animal model of tardive dyskinesia (TD) whose pathophysiology has been related to free radical generation and oxidative stress. Schizophrenic patients have been reported to eat a diet higher in fat than the general population and dietary fat intake can lead to an increase in oxidative stress in animal models. The objective of this study was to determine whether association of ingestion of a high fat diet with prolonged haloperidol treatment could lead to OD and oxidative stress in the rat brain. Haloperidol decanoate administration (38 mg/kg, IM, which is equivalent to 1 mg/kg/day) monthly for a period of 6 months to rats fed previously with a high fat and normo fat diets (6 months) caused a increase in vacuous chewing (VCM) and duration of facial twitching (FT). Haloperidol caused a reduction in body weight gain and the loss of body weight occurred after 4 months of treatment with haloperidol. The effects on body weight were more accentuated in HF diet group. HF diet ingestion was associated with an increase in TBARS levels in cerebellum and cerebral cortex (regardless of haloperidol treatment). A significant diet × haloperidol treatment interaction in striatum, subcortical parts and the region containing the substantia nigra was observed for TBARS. In fact, haloperidol caused an increase in TBARS levels of these regions only in rats fed with the HF. These results indicate that a high fat diet caused a transitory increase in haloperidol-induced OD in rats and this in part can be related to the haloperidol-induced oxidative stress in brain structures involved with OD.

Expression and distribution of kinin B1 receptor in the rat brain and alterations induced by diabetes in the model of streptozotocin

A role for kinin B1 receptors was suggested in the spinal cord and peripheral organs of streptozotocin (STZ)-diabetic rats. The present study aims at determining whether B1 receptors are also induced and over-expressed in the brain of STZ-rats at 2, 7, and 21 days post-treatment. This was addressed by in situ hybridization using the [35S]-UTPalphaS-labeled riboprobe and by in vitro autoradiography with the radioligand [125I]-HPP-des-Arg10-Hoe 140. In control rats, B1 receptor mRNA was found widely distributed in many brain regions. Low mRNA levels were found in thalamus and hypothalamus (7-12 nCi/g) while high mRNA signals were detected in cortical regions and hippocampus (18-29 nCi/g). In diabetic rats, B1 receptor mRNA was markedly increased in hippocampus, temporal/parietal cortices and amygdala at 2 and 7 days (+88 to +150%). Low densities of B1 receptor binding sites were detected in all analyzed regions in control rats (0.18-0.37 fmol/mg tissue). In diabetic rats, B1 receptor binding sites were significantly increased in hippocampus, amygdala, temporal/parietal, and perhinal/piriform cortices (+ 55 to + 165 %) at 7 days only. Results highlight an early but transient and reversible up-regulation of B1 receptors in specific brain regions of STZ-diabetic rats. This may offer the advantage of reducing putative central side effects with B1 receptor antagonists if used for the treatment of diabetic complications in the periphery.

Metabolic characterization of distinct neuroanatomical regions in rats by magic angle spinning 1H nuclear magnetic resonance spectroscopy

High-resolution magic angle spinning (HRMAS) (1)H NMR spectroscopy has been applied to the biochemical characterization of specific brain regions in rats in order to establish baseline levels of tissue metabolite profiles with which to compare models of neuropathology or toxic lesion. Cores of tissue (20 mg) from the brain stem, cerebellum, frontal cortex, and hippocampus were obtained from histologically defined coronal slices of brain from 18 male Sprague-Dawley rats. HRMAS (1)H NMR spectra were acquired for each of the regions sampled and the degree of intersample variability, as assessed by principal components analysis and discriminant analysis by projection to latent structure was found to be low. Clear region-specific differences in the biochemical profiles were observed using both comparison of metabolite ratios and/or pattern recognition methods. Relatively low concentrations of GABA in the cerebellum, high concentrations of taurine and N-acetylaspartate in the cortex, and high levels of choline, glycerophosphocholine, and phosphocholine in the hippocampus predominantly influenced the classification of the different brain regions. Additionally, N-acetylaspartylglutamate was detected in the brain stem, but was largely absent from the other regions examined. Such analyses provide a baseline reference for further HRMAS NMR spectroscopic studies to monitor disease and pharmacological insults in specific regions of the brain.

The study on learning and memory ability and some amino acid content in specific brain regions of rats

目的 探讨不同学习记忆能力大鼠的不同脑区一些氨基酸含量的差异,以揭示学习记忆能力与特定脑区某些氨基酸含量的关系.方法 wistar大鼠在MG-2型三等分辐射式迷宫中进行学习记忆能力的检测后,根据学习记忆能力分为两组(学习记忆能力均好者为Ⅰ组;学习记忆能力均差者为Ⅱ组).将大鼠立即断头处死,分别测定大脑皮层、小脑、纹状体、海马、下丘脑五个脑区Asp、Glu、Ala、Gly、GABA、Tau的含量.结果学习记忆能力均差的大鼠与学习记忆能力均好的大鼠比较:海马、纹状体Asp含量显著降低(P＜0.05);皮层、海马、纹状体Glu含量显著降低(P＜0.05);皮层、小脑、海马、纹状体Tau含量显著降低(P＜0.05和P＜0,01);皮层、纹状体Gly含量显著降低(P＜0.05);而纹状体GABA含量则显著升高(P＜0.05);各脑区Ala含量在两组间无显著性差异(P﹥0.05).结论海马、皮层、小脑、纹状体内氨基酸水平和比例与学习记忆密切相关。

Modulation of dopamine transporter function by alpha-synuclein is altered by impairment of cell adhesion and by induction of oxidative stress.

Human alpha-synuclein accumulates in dopaminergic neurons as intraneuronal inclusions, Lewy bodies, which are characteristic of idiopathic Parkinson's disease (PD). Here, we suggest that modulation of the functional activity of the dopamine transporter (DAT) by alpha-synuclein may be a key factor in the preferential degeneration of mesencephalic dopamine (DA)-synthesizing neurons in PD. In cotransfected Ltk-, HEK 293, and SK-N-MC cells, alpha-synuclein induced a 35% decrease in [3H]DA uptake. Biotinylated DAT levels were decreased by 40% in cotransfected cells relative to cells expressing only DAT. DAT was colocalized with alpha-synuclein in mesencephalic neurons and cotransfected Ltk- cells. Coimmunoprecipitation studies showed the existence of a complex between alpha-synuclein and DAT, in specific rat brain regions and cotransfected cells, through specific amino acid motifs of both proteins. The attenuation of DAT function by alpha-synuclein was cytoprotective, because DA-mediated oxidative stress and cell death were reduced in cotransfected cells. The neurotoxin MPP+ (1-methyl-4-phenylpyridinium), oxidative stress, or impairment of cell adhesion ablated the alpha-synuclein-mediated inhibition of DAT activity, which caused increased uptake of DA and increased biotinylated DAT levels, in both mesencephalic neurons and cotransfected cells. These studies suggest a novel normative role for alpha-synuclein in regulating DA synaptic availability and homeostasis, which is relevant to the pathophysiology of PD.

Effect of Ethanol Self‐Administration on μ‐ and δ‐Opioid Receptor‐Mediated G‐Protein Activity

This study examined the effects of ethanol self-administration on mu- and delta-opioid receptor-mediated G-protein activity in specific brain regions of male Long Evans rats. Rats were trained to self-administer ethanol by using a home-cage modification of the sucrose substitution paradigm. After 30 to 40 days of sucrose or sucrose/15% ethanol self-administration (20 min sessions, Monday-Friday), rats were killed for autoradiographic assays. Coronal sections of brains from sucrose and ethanol self-administering rats were collected and processed for basal and mu- and delta-stimulated [35S]guanosine-5'-O-(gamma-thio)-triphosphate (GTPgammaS) binding. Sections were exposed to film and then analyzed by using computer-assisted densitometry to determine levels of basal and agonist-stimulated [35S]GTPgammaS binding. Mu-opioid-stimulated [35S]GTPgammaS binding was decreased in the prefrontal cortex of brains from ethanol compared with sucrose self-administering rats. Mu-opioid-stimulated [35S]GTPgammaS binding was unchanged in the cingulate cortex, caudate-putamen, nucleus accumbens, amygdala, hypothalamus, thalamus, and locus ceruleus of ethanol compared with sucrose self-administering rats. Basal and delta-opioid-stimulated [35S]GTPgammaS binding did not differ between the two groups in the prefrontal cortex or any other region analyzed. These data demonstrate decreased mu-opioid-mediated G-protein activity in the prefrontal cortex of ethanol self-administering rats and suggest an interaction between ethanol and mu-opioid receptors in this region.

Hydroethidine detection of superoxide production during the lithium–pilocarpine model of status epilepticus

Hydroethidine is reported to be selectively oxidized to ethidium by superoxide. Using digital imaging and fluorescence microscopy it is possible to evaluate neuronal ethidium accumulation in specific brain regions of rats damaged in the lithium–pilocarpine model of status epilepticus. Intravenous or intraperitoneal administration of hydroethidine prior to 1 h of status epilepticus produced diffuse cytosolic distribution of ethidium fluorescence suggesting an increased neuronal production of superoxide that was not observed in control animals. A significantly increased number of neurons with the enhanced ethidium fluorescence was observed in parietal cortex, piriform cortex, perirhinal cortex, lateral amygdala, mediodorsal thalamus and laterodorsal thalamus, suggesting superoxide as a mechanism of neuronal injury in those regions. Other regions injured by lithium-pilocarpine seizures, such as the basolateral amygdala and hippocampus, did not demonstrate the enhanced neuronal ethidium fluorescence. In such regions it is possible that superoxide is not a mechanism of injury or that 1 h of status epilepticus is not sufficient to produce superoxide or other reactive oxygen species.

A neuroproteomic approach to targeting neuropeptides in the brain.

A novel universal neuropeptide display approach in the mass range of 300-5000 Da was developed to complement two-dimensional gel electrophoresis in the analysis of peptides and small proteins from brain tissue samples. For the analysis of neuropeptides we utilized on-line nanoscale capillary reversed phase liquid chromatography and electrospray ionization quadrupole-time of flight mass spectrometry. The method was employed for the analysis of a large number of peptides from three specific rat brain regions. Approximately 1500 peptides from each brain region were detected in the same analysis. Several of these peptides were sequenced using collision-induced dissociation and identified by database search tools. In addition, a method for comparing peptide elution profiles between samples was developed, to provide two- and three-dimensional computer graphics of the profiles and to pinpoint differences for statistical measurements. Among the characterized peptides were fragments from proteins such as hemoglobin, alpha-synuclein, stathmin, cyclophilin, actin, NADH dehydrogenase, cytochrome c oxidase and prosomatostatin, as well as the bioactive neuropeptides W-hemorphin-4, and LW-hemorphin-7. The present study showed that the combination of nanoscale reversed phase liquid chromatography and high-resolution tandem mass spectrometry provides a novel and powerful approach to investigate a large number of peptides and protein fragments in the brain.

Endogenous morphine and codeine: Possible role as endogenous anticonvulsants

Exogenously administered morphine can have both convulsive or anticonvulsive effects, depending on the dose and species. The levels of the endogenous opiate alkaloids morphine and codeine were significantly elevated in specific rat brain regions by the convulsive drug, pentylenetetrazole, as well as by the anticonvulsant drugs, carbamazepine and phenytoin. Morphine and codeine levels in peripheral tissues (heart, lung, spleen and adrenal) were unaffected by these drugs. Maximal increases in morphine levels were seen in the hypothalamus and striatum (2–10-fold), while lesser increases occurred in the midbrain and brain stem (2–4-fold). Codeine levels were also markedly increased in hypothalamus (5–10 fold), In contrast to morphine, codeine levels were also increased in the hippocampus (2–10-fold), but were unchanged in the striatum. These studies suggest that the endogenous alkaloids morphine and codeine are involved in the modulation of convulsions and that morphine and/or codeine may act as an endogenous anticonvulsant.

Subcellular localization of a glutathione-dependent dehydroascorbate reductase within specific rat brain regions.

Recently, we described the occurrence of a dehydroascorbate reductase within the rat CNS. This enzyme regenerates ascorbate after it is oxidized during normal aerobic metabolism. In this work, we describe the neuronal compartmentalization of the enzyme, using transmission electron microscopy of those brain areas in which the enzyme was most densely present when observed under light microscopy. In parallel biochemical studies, we performed immunoblotting and measured the enzyme activity of the cytoplasm and different nuclear fractions. Given the abundance of ascorbate in the caudate-putamen, we focused mostly on the occurrence of dehydroascorbate reductase at the striatal subcellular level. We also studied cerebellar Purkinje cells, hippocampal CA3 pyramidal cells and giant neurons in the magnocellular part of the red nucleus. In addition to neurons, immunolabeling was found in striatal endothelial cells, in the basal membrane of blood vessels and in perivascular astrocytes. In neuronal cytosol, the enzyme was observed in a peri-nuclear position and on the nuclear membrane. In addition, in both the striatum and the cerebellum, we found the enzyme within myelin sheets. Dehydroascorbate reductase was also present in the nucleus of neurons, as further indicated by measuring enzyme activity and by immunoblotting selected nuclear fractions. Immunocytochemical labeling confirmed that the protein was present in isolated pure nuclear fractions. Given the great amount of free radicals which are constantly generated in the CNS, the discovery of a new enzyme with antioxidant properties which translocates into neuronal nuclei appears to be a potential starting point to develop alternative strategies in neuroprotection.

Chronic valproate treatment increases expression of endoplasmic reticulum stress proteins in the rat cerebral cortex and hippocampus

Background: Sodium valproate is a highly effective treatment for bipolar disorder, but its mechanism of action remains poorly understood. We recently found with differential display polymerase chain reaction that valproate regulates the expression of the endoplasmic reticulum stress protein GRP78 in the rat cerebral cortex. In our study, we investigated the effect of this drug on the other members of the endoplasmic reticulum stress protein family, GRP94 and calreticulin, and we studied the brain regional distribution of GRP78, GRP94, and calreticulin. Methods: Immunohistochemistry was used to measure protein levels of GRP78, GRP94, and calreticulin after treatment with sodium valproate (300 mg/kg, intraperitoneal) in specific rat brain regions. Results: We report here that chronic treatment with valproate also increased expression of other members of the endoplasmic reticulum stress protein family, such as GRP94 and calreticulin. The brain regional distribution of these changes was similar for all three proteins, with marked increase detected in the frontal cortex, parietal cortex, and CA1 region of the hippocampus. Conclusions: Because GRP78, GRP94, and calreticulin possess molecular chaperone activity and bind Ca 2+ in the endoplasmic reticulum, the pharmacologic action of valproate may involve one or more of these processes.

Chronic Heroin Self-Administration Desensitizes μ Opioid Receptor-Activated G-Proteins in Specific Regions of Rat Brain

In previous studies from our laboratory, chronic noncontingent morphine administration decreased mu opioid receptor-activated G-proteins in specific brainstem nuclei. In the present study, mu opioid receptor binding and receptor-activated G-proteins were examined after chronic heroin self-administration. Rats were trained to self-administer intravenous heroin for up to 39 d, achieving heroin intake up to 366 mg. kg(-1). d(-1). mu opioid-stimulated [(35)S]GTPgammaS and [(3)H]naloxone autoradiography were performed in adjacent brain sections. Agonist-stimulated [(35)S]GTPgammaS autoradiography also examined other G-protein-coupled receptors, including delta opioid, ORL-1, GABA(B), adenosine A(1), cannabinoid, and 5-HT(1A). In brains from heroin self-administering rats, decreased mu opioid-stimulated [(35)S]GTPgammaS binding was observed in periaqueductal gray, locus coeruleus, lateral parabrachial nucleus, and commissural nucleus tractus solitarius, as previously observed in chronic morphine-treated animals. In addition, decreased mu opioid-stimulated [(35)S]GTPgammaS binding was found in thalamus and amygdala after heroin self-administration. Despite this decrease in mu-activated G-proteins, [(3)H]naloxone binding demonstrated increased mu opioid receptor binding in several brain regions after heroin self-administration, and there was a significant decrease in mu receptor G-protein efficiency as expressed as a ratio between agonist-activated G-proteins and mu receptor binding. No effects on agonist-stimulated [(35)S]GTPgammaS binding were found for any other receptor examined. The effect of chronic heroin self-administration to decrease mu-stimulated [(35)S]GTPgammaS binding varied between regions and was highest in brainstem and lowest in the cortex and striatum. These results not only provide potential neuronal mechanisms that may contribute to opioid tolerance and dependence, but also may explain why various chronic effects of opioids develop to different degrees.

Relationship of mu opioid receptor binding to activation of G-proteins in specific rat brain regions

This study investigated the relationship between mu receptor binding and mu agonist activation of G-proteins in the rat brain. To directly compare agonist potencies in receptor binding ( K i values) and G-protein activation ( K s values), both agonist-stimulated [ 35S]guanosine-5′- O-(γ-thio)-triphosphate ([ 35S]GTPγS) and [ 3H]naloxone binding assays were conducted under identical conditions, using the full mu agonist [d-Ala 2, N-Me 4, Gly 5-ol]-enkephalin (DAMGO). DAMGO exhibited biphasic competition of [ 3H]naloxone binding and stimulation of [ 35S]GTPγS binding in most regions. Whereas the high-affinity component represented a large percentage (50–80%) of total receptor sites, the high-affinity component of DAMGO-stimulated [ 35S]GTPγS binding was much lower, <30% of the total, and in most regions significant stimulation of [ 35S]GTPγS binding did not occur until the high-affinity binding sites were completely occupied. Moreover, the low-affinity potencies for DAMGO in receptor binding and G-protein activation were the same across different regions. Receptor–transducer amplification factors were calculated by the ratio of the apparent B max of net agonist-stimulated [ 35S]GTPγS binding to the B max of receptor binding. Amplification factors for the nine regions examined were relatively high and varied significantly across regions, from a ratio of 8 in the thalamus to 38 in the cortex, suggesting that the efficiency of mu opioid receptor coupling to G-proteins varies across brain regions.

Effects of chronic electroconvulsive shock on the expression of beta-adrenergic receptors in rat brain: immunological study.

The purpose of the present study is to determine the effect of chronic electroconvulsive shock (ECS) on the expression of beta-adrenergic receptors in rat brain by Western blot using mAb beta CO2, a monoclonal antibody against beta-adrenergic receptors. Rats in ECS treated groups received maximal ECS (70 mA, 0.5 second, 60 Hz) through ear-clip electrodes for 12 consecutive days. The experiment was carried out in 14 discrete regions of brain. Chronic ECS reduced the expression of beta-adrenergic receptors in frontal cortex, temporal cortex, parietooccipital cortex, hippocampus and limbic forebrain, but not in other areas of brain. The regional specificity and the magnitude of the reduction of receptor expression are well correlated with those of the reduction of receptor ligand binding, which was determined using [3H]dihydroalprenolol. To the best of our knowledge, this is the first report to demonstrate that chronic ECS decreases the expression of receptor protein in specific regions of rat brain.

Brain cytokine mRNAs in anorectic rats bearing prostate adenocarcinoma tumor cells.

Cancer is consistently associated with anorexia. The Lobund-Wistar rat model of prostate cancer exhibits clinical manifestations (including anorexia) that resemble many aspects of the human disease. Cytokines are proposed to be involved in cancer-associated anorexia. Here we investigated mRNA profiles of feeding-modulatory cytokines and neuropeptides in specific brain regions of anorectic Lobund-Wistar rats bearing prostate adenocarcinoma tumor cells. Interleukin (IL)-1beta system components (ligand, signaling receptor, receptor accessory proteins, receptor antagonist), tumor necrosis factor-alpha, transforming growth factor-beta1, glycoprotein 130 (IL-6 receptor signal transducer), proopiomelanocortin (POMC, opioid peptide precursor), and neuropeptide Y (NPY) mRNAs were analyzed with sensitive and specific RNase protection assays. The same brain region sample was assayed for all components. The data show that early anorexia in tumor-bearing rats was associated with an upregulation of IL-1beta mRNA in the brain regions examined (cerebellum, cortex, and hypothalamus). IL-1 receptor antagonist (IL-1Ra) mRNA and IL-1 receptor type I mRNA levels were also significantly increased in the cortex and hypothalamus. All other cytokine components, POMC, or NPY mRNA levels were not significantly different between tumor-bearing and pair-fed (control) rats. IL-1beta mRNA and IL-1Ra mRNA were also significantly upregulated in the spleen of tumor-bearing rats. These data suggest that 1) IL-1beta mRNA upregulation in the brain may be relevant to the anorexia exhibited by the tumor-bearing Lobund-Wistar rat and 2) in vivo characterization of cytokine components in discrete brain regions during cancer is necessary to understand underlying molecular mechanisms responsible for cancer-associated neurological manifestations.

Effect of nicotine use and withdrawal on brain preproenkephalin A mRNA

Although the effect of nicotine on brain neurotransmitters and behavior has been studied, the mechanism(s) by which nicotine contributes to tobacco use remains unclear. One transmitter that may relate to long-term nicotine use and its withdrawal is enkephalin, a five-amino acid opioid peptide derived from the proenkephalin A family. In the present study we determined the effect of acute and chronic nicotine treatment and its withdrawal on preproenkephalin A mRNA levels (PPE mRNA) in specific rat brain regions using Northern blot analysis. Acute treatment with nicotine produced a significant increase in PPE mRNA in striatum and hippocampus. Chronic treatment with nicotine caused a significant decrease in PPE mRNA in these brain regions. In both striatum and hippocampus there was a rebound increase in PPE mRNA 24 h after nicotine cessation which approached the saline level 7 days later. Nicotine withdrawal 24 h following nicotine cessation, caused a significant increase in PPE mRNA in both brain regions. These effects of nicotine were blocked by pretreating rats with the nicotinic antagonist, mecamylamine. These data strongly suggest that brain opioid system(s) are involved in mediating nicotinic responses and its withdrawal and may have clinical implications in treating nicotine addiction.

Interleukin-1β System (Ligand, Receptor Type I, Receptor Accessory Protein and Receptor Antagonist), TNF-α, TGF-β1 and Neuropeptide Y mRNAs in Specific Brain Regions During Bacterial LPS-Induced Anorexia

Bacterial lipopolysaccharide (LPS) or endotoxin induces neurological manifestations including anorexia. It is proposed that LPS-induced cytokine production is involved in the generation of neurological manifestations and in neuroinflammatory/immunological responses during Gram-negative infections. For example, LPS-induced effects can be blocked or ameliorated by the interleukin-1 receptor antagonist (IL-1Ra). Here, sensitive and specific RNase protection assays were used to investigate the effects of the intracerebroventricular (i.c.v.) administration of LPS on mRNA levels of interleukin-1β (IL-1β) system components, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, and neuropeptide Y (NPY) in the cerebellum, hippocampus, and hypothalamus. The same brain region sample was analyzed with all of the antisense probes. The data show simultaneous local induction of multiple cytokine components messenger ribonucleic acids (mRNAs) within specific brain regions in anorectic rats responding to i.c.v. administered LPS (500 ng/rat). Interleukin-1β and IL-1Ra had a similar mRNA induction profile (hypothalamus > cerebellum > hippocampus). Interleukin-1 receptor type I (IL-1RI) mRNA also increased in all three brain regions examined, and the soluble form of IL-1 receptor accessory protein (IL-1R AcP II) mRNA was induced in the hypothalamus. Tumor necrosis factor-α mRNA levels increased in the hypothalamus > hippocampus > cerebellum. Levels of membrane bound IL-1R AcP, TGF-β1, and NPY mRNAs did not change significantly in any brain region. The results suggest that: (1) endogenous up-regulation of IL-1β and TNF-α in the hypothalamus contribute to LPS-induced anorexia; and (2) the ratio IL-1Ra/IL-1β, and IL-1β ↔ TNF-α interactions may have implications for Gram-negative infections associated with high levels of LPS in the brain-cerebrospinal fluid.

Diazepam alters caffeine-induced effects on β-endorphin levels in specific rat brain regions

The present study was conducted to evaluate the effects of caffeine and the benzodiazepine agonist diazepam, and a combination of both on β-endorphin (BEN) levels in specific rat brain regions. Male Sprague-Dawley rats (150–200 g) adapted to a 12-hour light: 12-hour dark illumination cycle were used in this study. Caffeine (10 mg/kg), diazepam (2 mg/kg) or a combination of caffeine (10 mg/kg) and diazepam (2 mg/kg) were administered intraperitoneally to rats at 11:00 hr. Control animals were injected with saline. Animals were sacrificed by decapitation 1 h after injection, the brains were immediately removed; the cortex, hippocampus, hypothalamus and midbrain were dissected and their B-EN levels measured by radioimmunoassay. Caffeine administration significantly increased B-EN levels in the cortex. Similarly, administration of diazepam alone resulted in a significant increase of B-EN levels in cortex. However, concurrent administration of diazepam and caffeine resulted in higher increase of B-EN levels in cortex. No significant changes in B-EN levels were detected in hippocampus and midbrain after administration of either caffeine or diazepam alone. On the other hand, when diazepam and caffeine were concurrently administered a significant increase of B-EN levels were observed in the midbrain. Moreover, administration of diazepam alone resulted in a significant increase of B-EN levels in hypothalamus. This increase was still observed following concurrent administration of diazepam and caffeine. These results clearly indicate that diazepam alters caffeine-induced effects on B-EN in specific rat brain regions.

Effects of pentylenetetrazol-induced status epilepticus on c-Fos and HSP72 immunoreactivity in the immature rat brain

Pentylenetetrazol (PTZ)-induced status epilepticus (SE) leads to acute and long-term metabolic decreases in specific brain regions of rats at 10 (P10) or 21 days after birth (P21). These decreases are not related to apparent neuronal damage. Therefore, to better understand the neuronal activation and stress response to PTZ in immature rats, we mapped the expression of c-Fos and of the 72 kDa heat-shock protein (HSP72) in the same model of severe SE induced by the repetitive i.p. injections of subconvulsive doses of PTZ. Rats were sacrificed either at 2 or 24 h after the onset of SE in order to reveal c-Fos immunoreactivity, and at 24 and 72 h for HSP72 expression. Hematoxylin-eosin staining was performed at 24, 72 and 144 h after SE. The expression of c-Fos at 2 h after SE was more marked at P21 than at P10 and was prominent at both ages in the hippocampal dentate gyrus, cerebral cortex and amygdala. Some immunoreactivity was also present in the hypothalamus, thalamus and a few brainstem and cerebellar regions at both ages. There was a good relation between the regions expressing c-Fos and those exhibiting acute metabolic decreases at P21. Conversely, PTZ seizures did not lead to any expression of c-Fos at 24 h after SE or of HSP72 at 24 or 72 h at any age. Cell density was not affected by PTZ-induced SE at any age and at any time. These results suggest that c-Fos is a useful marker of neuronal activation induced by severe and prolonged seizures in the immature brain. The lack of HSP72 and of late c-Fos expression likely reflect the absence of neuronal damage in this model of PTZ-induced SE in the immature rat.

Persistent Alterations in Regional Brain Glial Fibrillary Acidic Protein and Synaptophysin Levels Following Pre- and Postnatal Polychlorinated Biphenyl Exposure

Pregnant Wistar WU rats were exposed to 0, 5, and 25 mg of the commercial polychlorinated biphenyl (PCB) mixture Aroclor 1254 per kilogram of body weight on Days 10 to 16 of gestation. Pregnant rats were sacrificed on Gestation Day 20 to observe effects on fetal body and brain weights. Male and female offspring were sacrificed on Postnatal Days 21 and 90 (PND21 and PND90, respectively) and examined for treatment-related effects on neurochemical parameters. The concentrations of the neuronal and glial cell markers, synaptophysin and glial fibrillary acidic protein (GFAP), were measured in diverse brain regions from the offspring using immunochemical techniques. The level of calcineurin (a calmodulin-regulated protein phosphatase) activity was measured in cerebellar homogenates. In addition, ethoxyresorufinO-deethylase (EROD) activity was determined in hepatic microsomes as a measure of a well-characterized response to PCB exposure in experimental animals. The major alterations of GFAP levels following maternal PCB treatment were significant increases in the lateral olfactory tract (LOT) and the cerebellum (CB) and significant decreases in the brain stem (BS) of the offspring on PND21 and 90. Synaptophysin levels were significantly decreased relative to controls in the LOT, prefrontal cortex, and striatum of the offspring on PND90. In the BS, synaptophysin levels were significantly decreased relative to controls in male and female weanlings on PND21 and males on PND90; however, significant increases were observed in the BS of females on PND90. No effect of maternal PCB treatment was observed on levels of GFAP and synaptophysin in the dorsal hippocampus on PND21 and 90. Due to analytical restrictions statistical comparisons of GFAP levels were limited to examining the effect of maternal PCB treatment per brain region per sex per time point. Calcineurin activity was decreased in the female CB on PND21, but a significant increase in activity was observed in the female CB on PND90. No effect of maternal PCB treatment was observed on the cerebellar calcineurin activity in male offspring on PND21 and 90. EROD activity was highly induced in maternal microsomes from both PCB treatment groups, but only slightly induced in fetal hepatic microsomes. On PND21 weanling hepatic microsomal EROD activity was highly induced following gestational and lactational PCB exposure; however, on PND90 EROD activity was unaffected by maternal PCB treatment in male offspring and significantly decreased in female offspring. The results of the present study indicate that gestational and lactational exposure to the commercial PCB mixture results in long-term alterations in a neuronal and glial cell markers in specific brain regions of rats. These marker proteins may be useful for determining the strucure–activity relationships in PCB-induced developmental neurotoxicity.