Rat Brain mRNA Research Articles

Taohong Siwu Decoction Regulates Cell Necrosis and Neuroinflammation in the Rat Middle Cerebral Artery Occlusion Model.

Cell necrosis and neuroinflammation play an important role in brain injury induced by ischemic stroke. Previous studies reported that Taohong Siwu decoction (THSWD)can reduce heart muscle cell necrosis and has anti-inflammatory properties. In this study, we investigated the effects of THSWD on cell necrosis and neuroinflammation in a rat model of middle cerebral artery occlusion (MCAO). Thirty-six male Sprague-Dawley (SD) rats were randomly divided into three groups with 12 rats in each group. They were the sham operation group, MCAO model group, and MCAO + THSWD group. We used ELISA to determine the levels of TNF-α, Mcp-1, and IL-1β inflammatory factors in rat serum, qRT‐PCR to detect the expression of TNF‐α, Mcp‐1 and IL‐1β mRNA in rat brain, and immunohistochemistry to detect the number of microglia and neutrophils in rat brain. qRT-PCR and Western blot were used to detect the mRNA and protein expression levels of IBA-1 and MPO inflammatory factors and the TNF-α/RIP1/RIP3/MLKL pathway in the rat brain and protein expression levels. Compared with the sham operation group, the expression of MCP-1, IL-1β, IBA-1, and MPO inflammatory factors and the TNF-α/RIP1/RIP3/MLKL pathway were significantly upregulated in the MCAO group. Compared with the MCAO group, the expressions of MCP-1, IL-1β, IBA-1, and MPO inflammatory factors and the TNF-α/RIP1/RIP3/MLKL pathway were significantly downregulated in the MCAO + THSWD group. THSWD can reduce the expression levels of MCP-1, IL-1β, IBA-1, and MPO inflammatory factors as well as the TNF-α/RIP1/RIP3/MLKL pathway. Meanwhile, it can reduce the necrosis and inflammation of brain cells after cerebral ischemia, so as to protect the brain tissue of rats.

Biosynthesis and Extracellular Concentrations of N,N-dimethyltryptamine (DMT) in Mammalian Brain

N,N-dimethyltryptamine (DMT), a psychedelic compound identified endogenously in mammals, is biosynthesized by aromatic-L-amino acid decarboxylase (AADC) and indolethylamine-N-methyltransferase (INMT). Whether DMT is biosynthesized in the mammalian brain is unknown. We investigated brain expression of INMT transcript in rats and humans, co-expression of INMT and AADC mRNA in rat brain and periphery, and brain concentrations of DMT in rats. INMT transcripts were identified in the cerebral cortex, pineal gland, and choroid plexus of both rats and humans via in situ hybridization. Notably, INMT mRNA was colocalized with AADC transcript in rat brain tissues, in contrast to rat peripheral tissues where there existed little overlapping expression of INMT with AADC transcripts. Additionally, extracellular concentrations of DMT in the cerebral cortex of normal behaving rats, with or without the pineal gland, were similar to those of canonical monoamine neurotransmitters including serotonin. A significant increase of DMT levels in the rat visual cortex was observed following induction of experimental cardiac arrest, a finding independent of an intact pineal gland. These results show for the first time that the rat brain is capable of synthesizing and releasing DMT at concentrations comparable to known monoamine neurotransmitters and raise the possibility that this phenomenon may occur similarly in human brains.

Aerobic exercise-stimulated Klotho upregulation extends life span by attenuating the excess production of reactive oxygen species in the brain and kidney.

Aerobic exercise induces many adaptive changes in the whole body and improves metabolic characteristics. Klotho, an anti-aging gene, is mainly expressed in the brain and kidney. The roles of Klotho in the brain and kidney during aerobic exercise remain largely unknown. The present study aimed to determine whether aerobic exercise could influence the expression of Klotho, decrease reactive oxygen species (ROS) and prolong life span. Sprague Dawley rats were exercised on a motor treadmill. Klotho mRNA and protein expression levels in rat brain and kidney tissues were examined using reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. ROS production was detected following intermittent aerobic exercise (IAE) or continuous aerobic exercise (CAE). Kaplan-Meier curve analysis demonstrated that aerobic exercise significantly improved rat survival (P<0.001). The ROS levels in rat brain and kidney tissues were decreased in the aerobic exercise groups compared with the control group (P<0.05). In addition, Klotho mRNA and protein expression levels were increased significantly following aerobic exercise compared with controls (P<0.05). There was no significant difference between the IAE and CAE groups in any experiments (P>0.05). These results suggest that aerobic exercise-stimulated Klotho upregulation extends the life span by attenuating the excess production of ROS in the brain and kidney. As Klotho exhibits a potential anti-aging effect, promoting Klotho expression through aerobic exercise may be a novel approach for the prevention and treatment of aging and aging-related diseases.

Glial- and Neuronal-Specific Expression of CCL5 mRNA in the Rat Brain.

Chemokine (C-C motif) ligand 5 (CCL5) belongs to a group of chemokines that play a role in the peripheral immune system, mostly as chemoattractant molecules, and mediate tactile allodynia. In the central nervous system (CNS), CCL5 and its receptors have multiple functions, including promoting neuroinflammation, insulin signaling, neuromodulator of synaptic activity and neuroprotection against a variety of neurotoxins. Evidence has also suggested that this chemokine may regulate opioid response. The multifunctional profile of CCL5 might correlate with its ability to bind different chemokine receptors, as well as with its unique cellular expression. In this work, we have used fluorescence in situ hybridization combined with immunohistochemistry to examine the expression profile of CCL5 mRNA in the adult rat brain and provide evidence of its cellular localization. We have observed that the highest expression of CCL5 mRNA occurs in all major fiber tracts, including the corpus callosum, anterior commissure, and cerebral peduncle. In these tracts, CCL5 mRNA was localized in oligodendrocytes, astrocytes and microglia. Astrocytic and microglial expression was also evident in several brain areas including the cerebral cortex, caudate/putamen, hippocampus, and thalamus. Furthermore, using a specific neuronal marker, we observed CCL5 mRNA expression in discrete layers of the cortex and hippocampus. Interestingly, in the midbrain, CCL5 mRNA co-localized with tyrosine hydroxylase (TH) positive cells of the ventral tegmental area, suggesting that CCL5 might be expressed by a subset of dopaminergic neurons of the mesolimbic system. The expression of CCL5 mRNA and protein, together with its receptors, in selected brain cell populations proposes that this chemokine could be involved in neuronal/glial communication.

NAC1 Regulates Somatic Cell Reprogramming by Controlling Zeb1 and E-cadherin Expression.

SummaryReprogramming somatic cells to induced pluripotent stem cells (iPSCs) is a long and inefficient process. A thorough understanding of the molecular mechanisms underlying reprogramming is paramount for efficient generation and safe application of iPSCs in medicine. While intensive efforts have been devoted to identifying reprogramming facilitators and barriers, a full repertoire of such factors, as well as their mechanistic actions, is poorly defined. Here, we report that NAC1, a pluripotency-associated factor and NANOG partner, is required for establishment of pluripotency during reprogramming. Mechanistically, NAC1 is essential for proper expression of E-cadherin by a dual regulatory mechanism: it facilitates NANOG binding to the E-cadherin promoter and fine-tunes its expression; most importantly, it downregulates the E-cadherin repressor ZEB1 directly via transcriptional repression and indirectly via post-transcriptional activation of the miR-200 miRNAs. Our study thus uncovers a previously unappreciated role for the pluripotency regulator NAC1 in promoting efficient somatic cell reprogramming.

Characterization of ectonucleotidase expression in the rat carotid body: regulation by chronic hypoxia.

The carotid body (CB) chemoreflex maintains blood Po2 and Pco2/H+ homeostasis and displays sensory plasticity during exposure to chronic hypoxia. Purinergic signaling via P1 and P2 receptors plays a pivotal role in shaping the afferent discharge at the sensory synapse containing catecholaminergic chemoreceptor (type I) cells, glial-like type II cells, and sensory (petrosal) nerve endings. However, little is known about the family of ectonucleotidases that control synaptic nucleotide levels. Using quantitative PCR (qPCR), we first compared expression levels of ectonucleoside triphosphate diphosphohydrolases (NTPDases1,2,3,5,6) and ecto-5'-nucleotidase (E5'Nt/CD73) mRNAs in juvenile rat CB vs. brain, petrosal ganglia, sympathetic (superior cervical) ganglia, and a sympathoadrenal chromaffin (MAH) cell line. In whole CB extracts, qPCR revealed a high relative expression of surface-located members NTPDase1,2 and E5'Nt/CD73, compared with low NTPDase3 expression. Immunofluorescence staining of CB sections or dissociated CB cultures localized NTPDase2,3 and E5'Nt/CD73 protein to the periphery of type I clusters, and in association with sensory nerve fibers and/or isolated type II cells. Interestingly, in CBs obtained from rats reared under chronic hypobaric hypoxia (~60 kPa, equivalent to 4,300 m) for 5-7 days, in addition to the expected upregulation of tyrosine hydroxylase and VEGF mRNAs, there was a significant upregulation of NTPDase3 and E5'Nt/CD73 mRNA, but a downregulation of NTPDase1 and NTPDase2 relative to normoxic controls. We conclude that NTPDase1,2,3 and E5'Nt/CD73 are the predominant surface-located ectonucleotidases in the rat CB and suggest that their differential regulation during chronic hypoxia may contribute to CB plasticity via control of synaptic ATP, ADP, and adenosine pools.

MiRNA and mRNA expression profiling in rat brain following alcohol dependence and withdrawal.

Long- lasting alterations in brain gene expression in alcohol addiction have been determined although no clear mechanism has yet been elucidated. There exist many factors regulating the mechanism of gene expression. We aimed in this study to detect miRNA (microRNA) and mRNA expression profile at the specific brain regions regarding ethanol exposure and withdrawal. Rats were exposed to liquid alcohol consumption for 21 days. Oligonucleotides microarrays and bioinformatics analyses were used to identify gene expression, miRNA expression and their functions in the Prefrontal cortex, Hippocampus and Corpus striatum of wistar rats. A bioinformatics strategy with microarray analysis, quantitative real time PCR, bioinformatics and mRNA (messenger RNA) miRNA- miRNA integrative analyses revealed that expression models interact with neuroplasticity and synaptic processes. Those significantly changed after ethanol exposure and withdrawal processes included 160 mRNAs and 29 rat-miRNAs at prefrontal cortex, 142 mRNAs and 26 rat-miRNAs at hippocampus, and 143 mRNAs and 30 rat-miRNAs at corpus striatum. Gene ontology and ingenuity pathway analyses revealed that most of the altered genes were responsible for synaptic plasticity, neuron differentiation, chromatin organization and some certain important signaling pathways. In conclusion, consistent and integrated variations in miRNA expression and in their focus mRNAs in rat brain were noted after alcohol exposure and withdrawal. Besides, understanding the molecular mechanisms of alcohol abuse will no doubt guide to development of significant cure methods for addiction. We are of the opinion that our findings may shed light on classification of novel biomarkers.

Expression and cellular localization of hepcidin mRNA and protein in normal rat brain

BackgroundHepcidin is a peptide hormone belonging to the defensin family of cationic antimicrobial molecules that has an essential role in systemic iron homeostasis. The peptide is synthesised by hepatocytes and transported in the circulation to target tissues where it regulates the iron export function of the ferrous iron permease, ferroportin. In the brain hepcidin protein has been identified using immuno-histochemistry and mRNA by real-time PCR but not by in situ hybridisation raising the question of whether there is measurable transcription of the hepcidin gene in the central nervous system. Alternatively hepcidin could be transported as a hormone to the brain via the circulation.ResultsBy RT-PCR hepcidin mRNA was present at low level throughout normal rat brain while in situ hybridisation to detect low-abundant mRNA revealed that transcripts were restricted to endothelium of blood vessels and choroid plexus. In contrast, hepcidin protein analysed by immuno-histochemistry was highly expressed in blood vessels, in endothelium and in pericytes. Hepcidin was also present in glial cells and in the olfactory bulb, sub-ventricular zone and dentate gyrus, areas where neurogenesis and synaptic plasticity are maintained throughout adult life. The hepcidin species identified by Western blotting in sub-ventricular zone, cortex and hippocampus migrated as a ~2.8 kDa band, identical in size to hepcidin present in normal rat serum suggesting that hepcidin in brain was the full-length biologically active 25 amino acid peptide. Hepcidin co-localised with ferroportin in ependymal cells of the sub-ventricular zone and in the corpus callosum consistent with a regulatory role in iron metabolism at these sites.ConclusionsHepcidin protein was widely expressed in brain parenchyma while levels of hepcidin gene transcription appeared to be below the limits of detection of the in situ hybridisation probes. This disparity suggests that not all hepcidin in the brain is transcribed in situ and may originate in part outside the brain. The properties of hepcidin as a cationic peptide hormone are reflected in the finding of hepcidin in the walls of blood vessels and in pericytes and glia, cells that may be involved in transporting the peptide into brain interstitium.

Relaxin-3 receptor (RXFP3) signalling mediates stress-related alcohol preference in mice.

Stressful life events are causally linked with alcohol use disorders (AUDs), providing support for a hypothesis that alcohol consumption is aimed at stress reduction. We have previously shown that expression of relaxin-3 mRNA in rat brain correlates with alcohol intake and that central antagonism of relaxin-3 receptors (RXFP3) prevents stress-induced reinstatement of alcohol-seeking. Therefore the objectives of these studies were to investigate the impact of Rxfp3 gene deletion in C57BL/6J mice on baseline and stress-related alcohol consumption. Male wild-type (WT) and Rxfp3 knockout (KO) (C57/B6JRXFP3TM1/DGen) littermate mice were tested for baseline saccharin and alcohol consumption and preference over water in a continuous access two-bottle free-choice paradigm. Another cohort of mice was subjected to repeated restraint followed by swim stress to examine stress-related alcohol preference. Hepatic alcohol and aldehyde dehydrogenase activity was assessed in mice following chronic alcohol intake and in naive controls. WT and Rxfp3 KO mice had similar baseline saccharin and alcohol preference, and hepatic alcohol processing. However, Rxfp3 KO mice displayed a stress-induced reduction in alcohol preference that was not observed in WT littermates. Notably, this phenotype, once established, persisted for at least six weeks after cessation of stress exposure. These findings suggest that in mice, relaxin-3/RXFP3 signalling is involved in maintaining high alcohol preference during and after stress, but does not appear to strongly regulate the primary reinforcing effects of alcohol.

The effects of dental pulp cell and dental pulp cell-derived neurosphere on the expression of Brain-derived neurotrophic factor mRNA in rat brain after global ischemia

The effects of dental pulp cell and dental pulp cell-derived neurosphere on the expression of Brain-derived neurotrophic factor mRNA in rat brain after global ischemia

The effects of dietary restriction and aging on amyloid precursor protein and presenilin-1 mRNA and protein expression in rat brain

The objective of this study was to examine the effects of aging and long-term dietary restriction (DR) on the level of amyloid precursor protein (APP) and presenilin-1 (PS-1), proteins that are critically involved in Alzheimer's disease. Changes in mRNA and protein expression were assessed by real-time PCR and western blot analysis during aging and long-term DR in the cortex and hippocampus of 6-, 12-, 18-, and 24-month-old rats. Prominent regional changes in expression were observed in response to aging and DR. Although the hippocampus displayed significant alterations in APP mRNA and protein expression and no significant changes in PS-1 expression, an opposite pattern was observed in the cortex. DR counteracted the age-related changes in APP mRNA expression in both structures of old animals. The observed DR-induced increase in mRNA levels in the hippocampus was accompanied by an increase in the level of full-length protein APP. These results indicate that although both structures are very sensitive to aging, a specific spatial pattern of changes in APP and PS-1 occurs during aging. Furthermore, these findings provide evidence that DR can affect APP and PS-1 expression in a manner consistent with its proposed 'antiaging' effect.

New insights concerning insulin synthesis and its secretion in rat hippocampus and cerebral cortex: Amyloid-β1–42-induced reduction of proinsulin level via glycogen synthase kinase-3β

The reduction of insulin levels in hippocampal areas is associated with Alzheimer's disease. The present study using rat brain explores the mechanisms of insulin synthesis and secretion, as well as amyloid-β1–42 (Aβ1–42)-induced reduction of proinsulin expression. After confirming the expression of insulin mRNA and proinsulin in rat brain, we visualized and analyzed the motion of insulin secretion in rat hippocampal neurons using pH-sensitive green fluorescent protein (pHluorin) fused to the insulin. In the rat hippocampal neurons expressing insulin–pHluorin, time-lapse confocal laser scanning microscopy revealed the appearance of fluorescent spots induced by depolarization after stimulation with 50mM KCl. In these fluorescent spots, Ca2+-dependent activator protein for secretion 2 (CAPS2), which is the regulator of the dense-core vesicle involving neuronal peptides, was co-localized with insulin–pHluorin. However, Aβ1–42-induced reduction of proinsulin in rat hippocampal neurons was inhibited by treatment with lithium and transfection with glycogen synthase kinase-3β (GSK-3β) siRNA. These results demonstrate that synthesized insulin is secreted from rat hippocampal and cortical neuron's dense-core vesicles, and that activation of GSK-3β in Aβ1–42-induced Alzheimer's model hippocampal neurons decreases the insulin synthesis.

Arcuate NPY Controls Sympathetic Output and BAT Function via a Relay of Tyrosine Hydroxylase Neurons in the PVN

Neuropepetide Y (NPY) is best known for its powerful stimulation of food intake and its effects on reducing energy expenditure. However, the pathways involved and the regulatory mechanisms behind this are not well understood. Here we demonstrate that NPY derived from the arcuate nucleus (Arc) is critical forthe control of sympathetic outflow and brown adipose tissue (BAT) function. Mechanistically,a key change induced by Arc NPY signaling is a marked Y1 receptor-mediated reduction in tyrosine hydroxylase (TH) expression in the hypothalamic paraventricular nucleus (PVN), which is also associated with a reduction in TH expression in the locus coeruleus (LC) and other regions in the brainstem. Consistent with this, Arc NPY signaling decreased sympathetically innervated BAT thermogenesis, involving the downregulation of uncoupling protein 1 (UCP1) expression in BAT. Taken together, these data reveal a powerful Arc-NPY-regulated neuronal circuit that controls BAT thermogenesis and sympathetic output via TH neurons.

Controlled cortical impact injury and craniotomy result in divergent alterations of pyruvate metabolizing enzymes in rat brain

Dysregulated glucose metabolism and energy deficit is a characteristic of severe traumatic brain injury (TBI) but its mechanism remains to be fully elucidated. Phosphorylation of pyruvate dehydrogenase (PDH) is the rate-limiting mitochondria enzyme reaction coupling glycolysis to the tricarboxylic acid cycle. Phosphorylation of PDH E1α1 subunit catalyzed by PDH kinase (PDK) inhibits PDH activity, effectively decoupling aerobic glycolysis whereas dephosphorylation of phosphorylated PDHE1α1 by PDH phosphatase (PDP) restores PDH activity. We recently reported altered expression and phosphorylation of pyruvate dehydrogenase (PDH) following TBI. However, little is known about PDK and PDP involvement. We determined PDK (PDK1–4) and PDP isoenzyme (PDP1–2) mRNA and protein expression in rat brain using immunohistochemistry and in situ hybridization techniques. We also quantified PDK and PDP mRNA and protein levels in rat brain following TBI using quantitative real-time PCR and Western blot, respectively. Controlled cortical impact-induced TBI (CCI-TBI) and craniotomy significantly enhanced PDK1-2 isoenzyme mRNA expression level but significantly suppressed PDP1 and PDK4 mRNA expression after the injury (4h to 7days). CCI-TBI and craniotomy also significantly increased PDK1–4 isoenzyme protein expression but suppressed PDP1–2 protein expression in rat brain. In summary, the divergent changes between PDK and PDP expression indicate imbalance between PDK and PDP activities that would favor increased PDHE1α1 phosphorylation and enzyme inhibition contributing to impaired oxidative glucose metabolism in TBI as well as craniotomy.

Fish Oil Feeding Up-Regulates the Expression of 5-Aminolevulinate Synthase 2 mRNA in Rat Brain

In this study, we investigated the effect of fish oil on gene expression in the cerebral cortex, and found that 5-aminolevulinate synthase 2 (ALAS2) mRNA expression was up-regulated by fish oil feeding. ALAS2 promoter activity was found to be regulated by retinoic acid. Our results suggest that fish oil modulates neuronal functions via heme synthesis.

Chronic phencyclidine (PCP)-induced modulation of muscarinic receptor mRNAs in rat brain : Impact of antipsychotic drug treatment

Many antipsychotics (APDs) have a high affinity for muscarinic receptors, which is thought to contribute to their therapeutic efficacy, or side effect profile. In order to define how muscarinic receptor gene expression is affected by atypical or typical APDs, rats were treated with chronic (2.58 mg/kg) PCP (a psychotomimetic) or vehicle, plus clozapine (20 mg/kg/day) or haloperidol (1 mg/kg/day), and M1, M2 and M3 receptor mRNA levels were determined in brain sections. Negligible changes in M2 or M3 muscarinic mRNA were detected in any region after clozapine or haloperidol. Chronic PCP administration increased M1 mRNA expression in the prefrontal cortex, which was not reversed by either chronic clozapine or haloperidol treatment. Chronic clozapine treatment in combination with PCP treatment decreased M1 receptor mRNA levels in the nucleus accumbens core, whereas chronic haloperidol in combination with PCP treatment increased M1 receptor mRNA levels in the ventromedial hypothalamus and medial amygdala. Thus M1 receptor gene expression is targeted by APDs, although the regions affected differ according to the APD treatment and whether PCP has been administered. The different brain circuitry modulated, may reflect the differing modes of action of typical and atypical APDs. These data provide support for the dysregulation of M1 receptors in schizophrenia, and furthermore, modulation by antipsychotic agents in the treatment of schizophrenia.This article is part of a Special Issue entitled ’Schizophrenia’.

EKSPRESI RELATIF mRNA HIF-1α PADA JANTUNG, OTAK DAN DARAH TIKUS SELAMA INDUKSI HIPOKSIA SISTEMIK

Normal 0 false false false IN X-NONE X-NONE MicrosoftInternetExplorer4 Relative Expression of HIF-1 &alpha; mRNA in Rat Heart, Brain and Blood During Induced Systemic Hypoxia. Hypoxia is a pathological condition in which the body as a whole or region of the body (tissue or cell) deprived of adequate oxygen supply. The transcriptional regulator hypoxia inducible factor-1 (HIF-1) is an essential mediator of O 2 homeostasis. Unlike the &beta; sub unit (HIF-1 &beta; ), the activity of HIF-1 &alpha; is controlled in an oxygen-dependent manner. It has been reported that the stability and expression of HIF-1 &alpha; during hypoxia is remarkably higher than those under normoxic conditions.The aim of this study was to analyze the adaptive tissue responses during induced systemic hypoxia by comparation of relative expression of mRNA HIF-1 &alpha; in rat heart, brain and blood. Twenty-five male Sprague Dawley rats were subjected to systemic hypoxia by placing them in the hypoxic chamber supplied by 8-10% of O 2 for 0, 1, 7, 14 and 21 days, respectively. The relative expression level of HIF-1 &alpha; mRNA in brain, heart and leucocyte cells were analyzed using quantitative RT-PCR assay (Real Time PCR) based on Pfaff&rsquo;s formula. This study demonstrates that the increased of relative expression of HIF-1 &alpha; mRNA during induced systemic hypoxia reached its maximum level at day 7 (in heart) or at day 14 (in brain), whereas in leucocyte cells the stimulation of HIF-1 &alpha; expression was intensively maintained up to 21 days although the expression has reached the remarkably high level. We could conclude that HIF-1 &alpha; as an oxygen sensing during systemic hypoxia has different capacity and sensitivity in brain, heart and blood tissues, due to the importance of oxygen homeostasis in each tissue. Keywords: induced systemic hypoxia, quantitative RT-PCR, relative expression of HIF-1 &alpha; mRNA

Electrophysiological characterization of a novel small peptide from the venom of Conus californicus that targets voltage-gated neuronal Ca2+ channels

Conus californicus belongs to a genus of marine gastropods with more than 700 extant species. C. californicus has been shown to be distantly related to all Conus species, but showing unusual biological features. We report a novel peptide isolated from C. californicus with a significant inhibitory action over neuronal voltage-gated calcium channels. The new toxin is formed by 13-amino acid residues with two disulfide bonds, whose sequence (NCPAGCRSQGCCM) is strikingly different from regular ω-conotoxins. In the HPLC purification procedure, the venom fraction eluted in the first 10–15 min produced a significant decrease (54% ± 3%) of the Ca2+ current in Xenopus laevis oocytes transfected with purified rat-brain mRNA. A specific peptide obtained from the elution at 13 min decreased the Ca2+ current in the adult rat dorsal-root ganglion neurons in a primary culture by 34% ± 2%. The cysteine pattern of this peptide corresponds to the framework XVI described for the M-superfamily of conopeptides and is unprecedented among Conus peptides acting on Ca2+ channels.

Kaitocephalin Antagonism of Glutamate Receptors Expressed in Xenopus Oocytes

Kaitocephalin is the first discovered natural toxin with protective properties against excitotoxic-death of cultured neurons induced by N-methyl-d-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/kainic acid (kainate, KA) receptors. Nevertheless, the effects of kaitocephalin on the function of these receptors were unknown. In this work we report some pharmacological properties of synthetic (-)-kaitocephalin on rat brain glutamate receptors expressed in Xenopus laevis oocytes and, on the homomeric AMPA-type GluR3 and KA-type GluR6 receptors. Kaitocephalin was found to be a more potent antagonist of NMDA receptors (IC(50) = 75 +/- 9 nM) than of AMPA receptors from cerebral cortex (IC(50) = 242 +/- 37 nM) and from homomeric GluR3 subunits (IC(50) = 502 +/- 55 nM). Moreover, kaitocephalin is a weak antagonist of the KA-type receptor GluR6 (IC(50) ~ 100 muM) and of metabotropic (IC(50) > 100 muM) glutamate receptors expressed by rat brain mRNA.

Localization of zip1 and zip4 mRNA in the adult rat brain

The localization of two members of the Slc39a (zip1 and zip4) family of zinc transporters was examined in the brains of adult mice. Zip1 was highly enriched in brain regions with high densities of neuronal cell bodies, including the hippocampus, thalamus, and perifontal cortex. Zip1 was also expressed in commissural fiber tracts such as the corpus callosum and anterior commissure, but little was found in the internal and external capsules. Also, very low amounts of zip1 mRNA were detected in resting astrocytes and reactive astrocytes that were examined at 14 days after inflicting a stab wound. Zip1 mRNA was detected in ependymal cells lining the third and lateral ventricles and epithelium cells in the choroid plexus. Interestingly, zip4 mRNA was detected in the choroid plexus but not in the ependymal cells or other neural elements. Zip4 mRNA was also detected in brain capillaries, but zip1 mRNA was not. In zip4 knockout heterozygotes that express green fluorescent protein regulated by the zip4 promoter, green fluorescent protein was detected in brain capillaries. Because zip4 levels are regulated by dietary Zn, our studies suggest that the brain has the potential of adapting to changes in Zn status.