SAD-B modulates epileptic seizure by regulating AMPA receptors in patients with temporal lobe epilepsy and in the PTZ-induced epileptic model

α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors are the predominant mediators of glutamate-induced excitatory neurotransmission. It is widely accepted that AMPA receptors are critical for the generation and spread of epileptic seizure activity. Dysfunction of AMPA receptors as a causal factor in patients with intractable epilepsy results in neurotransmission failure. Brain-specific serine/threonine-protein kinase 1 (SAD-B), a serine-threonine kinase specifically expressed in the brain, has been shown to regulate AMPA receptor-mediated neurotransmission through a presynaptic mechanism. In cultured rat hippocampal neurons, the overexpression of SAD-B significantly increases the frequency of miniature excitatory postsynaptic currents (mEPSCs). Here, we showed that SAD-B downregulation exerted antiepileptic activity by regulating AMPA receptors in patients with temporal lobe epilepsy (TLE) and in the pentylenetetrazol (PTZ)-induced epileptic model. We first used immunoblotting and immunohistochemistry analysis to demonstrate that SAD-B expression was increased in the epileptic rat brain. Subsequently, to explore the function of SAD-B in epilepsy, we used siRNA to knock down SAD-B protein and observed behavior after PTZ-induced seizures. We found that SAD-B downregulation attenuated seizure severity and susceptibility in the PTZ-induced epileptic model. Furthermore, we showed that the antiepileptic effect of SAD-B downregulation on PTZ-induced seizure was abolished by CNQX (an AMPA receptor inhibitor), suggesting that SAD-B modulated epileptic seizure by regulating AMPA receptors in the brain. Taken together, these findings suggest that SAD-B may be a potential and novel therapeutic target to limit epileptic seizures.

Neuroprotective properties of RT10, a fraction isolated from Parawixia bistriata spider venom, against excitotoxicity injury in neuron-glia cultures

L-Glutamate (L-Glu), the major excitatory neurotransmitter in the mammalian Central Nervous System (CNS), is essential to cognitive functions. However, when L-Glu is accumulated in large concentrations at the synaptic cleft, it can induce excitotoxicity that results in secondary damage implicated in many neurological disorders. Current therapies for the treatment of neurological disorders are ineffective and have side effects associated with their use; therefore, there is a need to develop novel treatments. In this regard, previous studies have shown that neuroactive compounds obtained from the venom of the spider Parawixia bistriata have neuroprotective effects in vitro and in vivo. In this sense, this work aimed to evaluate potential neuroprotective effects of fraction RT10, obtained from this spider venom, on primary cultures of neuron and glial cells subjected to glutamate excitotoxicity insults. Methods: Primary cultures of neurons and glia were obtained from the cerebral tissue of 1-day-old postnatal Wistar rats. After 7 days in vitro (DIV), the cultures were incubated with fraction RT10 (0.002; 0.02; 0.2 and 2 µg/µL) or riluzole (100 µM) for 3-hours before application of 5 mM L-Glu. After 12 hours, the resazurin sodium salt (RSS) test was applied to measure metabolic activity and proliferation of living cells, whereas immunocytochemistry for MAP2 was performed to measure neuronal survival. In addition, the cells were immunolabeled with NeuN and GFAP in baseline conditions. Results: In the RSS tests, we observed that pre-incubation with RT10 before the excitotoxic insults from L-Glu resulted in neuroprotection, shown by a 10% reduction in the cell death level. RT10 was more effective than riluzole, which resulted in a cell-death reduction of 5%. Moreover, qualitative analysis of neuronal morphology (by MAP2 staining, expressed as fluorescence intensity (FI), an indirect measure of neuronal survival) indicate that RT10 reduced the toxic effects of L-Glu, as shown by a 38 % increase in MAP2 fluorescence when compared to L-Glu insult. On the other hand, the riluzole treatment resulted in 17% increase of MAP2 fluorescence; therefore, the neuroprotection from RT10 was more efficacious. Conclusion: RT10 fraction exhibits neuroprotective effects against L-Glu excitotoxicity in neuron-glia cultured in vitro.(AU)

Spinal CCL2 Promotes Central Sensitization, Long-Term Potentiation, and Inflammatory Pain via CCR2: Further Insights into Molecular, Synaptic, and Cellular Mechanisms / 神经科学通报·英文版

Mounting evidence supports an important role of chemokines, produced by spinal cord astrocytes, in promoting central sensitization and chronic pain. In particular, CCL2 (C-C motif chemokine ligand 2) has been shown to enhance N-methyl-D-aspartate (NMDA)-induced currents in spinal outer lamina II (IIo) neurons. However, the exact molecular, synaptic, and cellular mechanisms by which CCL2 modulates central sensitization are still unclear. We found that spinal injection of the CCR2 antagonist RS504393 attenuated CCL2- and inflammation-induced hyperalgesia. Single-cell RT-PCR revealed CCR2 expression in excitatory vesicular glutamate transporter subtype 2-positive (VGLUT2) neurons. CCL2 increased NMDA-induced currents in CCR2/VGLUT2 neurons in lamina IIo; it also enhanced the synaptic NMDA currents evoked by dorsal root stimulation; and furthermore, it increased the total and synaptic NMDA currents in somatostatin-expressing excitatory neurons. Finally, intrathecal RS504393 reversed the long-term potentiation evoked in the spinal cord by C-fiber stimulation. Our findings suggest that CCL2 directly modulates synaptic plasticity in CCR2-expressing excitatory neurons in spinal lamina IIo, and this underlies the generation of central sensitization in pathological pain.

Increasing Burden of Alzheimer's Disease by Aging

No abstract available.

Microinjection of NMDA-type glutamate receptor agonist NMDA and antagonist D-AP-5 into the central nucleus of the amygdale alters water intake rather than food intake / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the role of N-Methyl-D-aspartic acid (NMDA)-type glutamate receptors in the central nucleus of the amygdale (CeA) in food and water intake.</p><p><b>METHODS</b>Male Sprague-Dawley rats with stainless steel cannulae implanted unilaterally into the CeA were used. The prototypic NMDA receptor agonist NMDA, or the selective NMDA receptor antagonist D(-)-2-amino-5-phosphonopentanoic acid (D-AP-5) was microinjected into the CeA of satiated and euhydrated rats.</p><p><b>RESULTS</b>Intra-CeA injection of 8.50, 17.00, or 34.00 nmol NMDA did not alter food intake but significantly increased water intake 0-1 h after the injection (F(3,32)=3.191, P=0.037) independent of food intake. Without affecting the food intake, injection of 6.34, 12.70, or 25.40 nmol D-AP-5 into the CeA significantly decreased water intake 0-1 h after the injection (F(3,28)=3.118, P=0.042) independent of food intake.</p><p><b>CONCLUSION</b>NMDA receptors in the CeA may participate in the control of water intake rather than food intake.</p>

Neurotrasmisores en niños con meningitis bacteriana / Neurotransmitters in children with bacterial meningitis

La meningitis bacteriana continúa siendo una enfermedad potencialmente fatal, especialmente en países en vías de desarrollo. Los aminoácidos excitatorios están fuertemente implicados en la patogénesis del daño neuronal en meningitis bacteriana. El objetivo fue medir niveles de glutamato, GABA, glicina y taurina en liquido cefalorraquídeo y correlacionarlos con el grado de severidad, complicaciones y secuelas. Estudio prospectivo en 31 pacientes con meningitis bacteriana y 10 pacientes con líquido cefalorraquídeo normal (control), con edades de 1 mes - 13 años de edad. El análisis de aminoácidos se realizó al ingreso y al tercer día mediante cromatografía líquida de alta presión. De los 31 pacientes que ingresaron al estudio 64,5 % fueron de género femenino, 13 lactantes, 8 preescolares y 10 escolares. El promedio de aminoácidos en los niños con meningitis fue más alto que en el grupo control (P<0,01). El glutamato disminuyó significativamente en pacientes con hidrocefalia. El GABA está disminuido en pacientes con parálisis cerebral y la taurina está disminuida en higroma y aumentada en lesión de pares craneales, trastornos de la conducta e hipoacusia. Los cambios en los niveles de aminoácidos en líquido cefalorraquídeo refleja el estado patológico y severidad del daño cerebral. Este estudio provee información del eventual papel de la inmunomodulación y posible uso de antagonistas de aminoácidos excitatorios, con efecto neuroprotector, en el tratamiento de meninigitis bacteriana e indica que esta clase de molécula neurotóxica puede represetar un importante blanco en la terapia adyuvante para meningitis bacteriana.

Bacterial meningitis rmains a potentially fatal disease, especially in developing countries. Exitatory amino acids are strongly implicated in the pathogenesis of neuronal damage in bacterial meningitis. To measure levels of glutamate, GABA, glycine and taurine in cerebroespinal fluid and correlate with the degree of severity, complications and sequelae. Prospective study in 31 patients with bacterial meningitis and 10 patients with normal cerebrospinal fluid (control), aged 1 month - 13 years old. Amino acid analysis was performed on admission and on the third day using high pressure liquid chromatography. Of the 31 patients entering the study 64.5 % were females, 13 infants, 8 preschoolers and 10 elementary school students. The average number of amino acids in children with meningitis was higher than in the control group (P<0.01). Glutamate levels significantly decreased in patients with hydrocephalus. GABA levels decreased in patients with cerebral pasly, and taurine diminished in hygroma, and increased in cranial nerve injury, eating disorders and hearing loss. Changes in amino acid levels in cerebrospinal fluid reflect pathological state and severity of brain damage. This study provides information on the possible role of immunomudulation and possible use of excitatory amino acid antogonists with neuroprotective effects in the treatment of bacterial meningitis, indicating that this class of neurotoxic molecules may represent important target in adjuvant therapy for bacterial meningitis.

Sex-dependent behavioral effects and morphological changes in the hippocampus after prenatal invasive interventions in rats: implications for animal models of schizophrenia

OBJECTIVES: Although schizophrenia affects both human genders, there are gender-dependent differences with respect to age of onset, clinical characteristics, course and prognosis of the disease. METHODS: To investigate sex-dependent differences in motor coordination and activity as well as in cognitive and social behavior, we repeatedly tested female (n = 14) and male (n = 12) Fisher rats (postnatal days, PD 56-174) that had received intracerebroventricular injections of kainic acid as well as female (n = 15) and male (n = 16) control animals. The hippocampus was examined histologically. RESULTS: Compared to male controls, in the alcove test both female controls and female animals with prenatal intervention spent less time in a dark box before entering an unknown illuminated area. Again, animals that received prenatal injection (particularly females) made more perseveration errors in the T-maze alternation task compared to controls. Female rats exhibited a higher degree of activity than males, suggesting these effects to be sex-dependent. Finally, animals that received prenatal intervention maintained longer lasting social contacts. Histological analyses showed pyramidal cells in the hippocampal area CA3 (in both hemispheres) of control animals to be longer than those found in treated animals. Sex-dependent differences were found in the left hippocampi of control animals and animals after prenatal intervention. CONCLUSION: These results demonstrate important differences between males and females in terms of weight gain, response to fear, working memory and social behavior. We also found sex-dependent differences in the lengths of hippocampal neurons. Further studies on larger sample sets with more detailed analyses of morphological changes are required to confirm our data.

Neurotoxicity Screening in a Multipotent Neural Stem Cell Line Established from the Mouse Brain

Neural stem cells (NSCs) have mainly been applied to neurodegeneration in some medically intractable neurologic diseases. In this study, we established a novel NSC line and investigated the cytotoxic responses of NSCs to exogenous neurotoxicants, glutamates and reactive oxygen species (ROS). A multipotent NSC line, B2A1 cells, was established from long-term primary cultures of oligodendrocyte-enriched cells from an adult BALB/c mouse brain. B2A1 cells could be differentiated into neuronal, astrocytic and oligodendroglial lineages. The cells also expressed genotypic mRNA messages for both neural progenitor cells and differentiated neuronoglial cells. B2A1 cells treated with hydrogen peroxide and L-buthionine-(S,R)-sulfoximine underwent 30-40% cell death, while B2A1 cells treated with glutamate and kainate showed 25-35% cell death. Cytopathologic changes consisting of swollen cell bodies, loss of cytoplasmic processes, and nuclear chromatin disintegration, developed after exposure to both ROS and excitotoxic chemicals. These results suggest that B2A1 cells may be useful in the study of NSC biology and may constitute an effective neurotoxicity screening system for ROS and excitotoxic chemicals.

Effects of triiodothyronine on the learning and memory behaviors in neonatal mice following excitotoxic brain damage / 中国当代儿科杂志

<p><b>OBJECTIVE</b>Some research has shown that learning and memory function impairments in rats with hypothyroidism are associated with triiodothyronine (T3) deficiency in neurons. This study aimed to investigate the effects of L-T3 administration on learning and memory behaviors in neonatal mice with excitotoxic brain damage.</p><p><b>METHODS</b>Seventy-one 5-day-old ICR neonatal mice were randomly assigned to five groups: controls that received intracerebral and intraperitoneal injections of phosphate buffered saline (PBS) (n=14); a group that received intracerebral injections of ibotenic acid (IA) and intraperitoneal injection of PBS (n=14); 3 groups that received intracerebral injections of IA and intraperitoneal injection of L-T3 at 0.2, 0.5, and 1 microg/kg, respectively (n=14-15). Intraperitoneal injections were done 1, 24, 48, 72 and 96 hrs after intracerebral injections. Learning and memory functions were evaluated by the Y-maze discrimination learning test on postnatal days 33-34.</p><p><b>RESULTS</b>The learning and memory functions in the highest L-T3 dose group were significantly better than those in the IA, and the lower L-T3 dose groups, presenting with decreased number of trials to criterion[15.8 + or - 4.5 vs 21.3 + or - 6.3 (IA group), 20.5 + or - 6.0 (0.2 microg/kg L-T3 group) or 21.0 + or - 6.5 (0.5 microg/kg L-T3 group); P<0.05], and achieving a higher correct percentage [91.4+ or - 9.5% vs 79.3 + or - 10.0% (IA group), 77.9 + or - 14.2% (0.2 microg/kg L-T3 group) or 80.7 + or - 12.2% (0.5 microg/kg L-T3 group); P<0.05].</p><p><b>CONCLUSIONS</b>High-dose L-T3 (1 microg/kg) may improve learning and memory functions in mice following excitotoxic brain damage.</p>

Neurobehavioral function of neonatal mice following excitotoxic brain damage / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To assess the changes of neurobehavioral function in a neonatal mouse model of excitotoxic brain damage.</p><p><b>METHODS</b>Fifty-five 5-day-old ICR neonatal mice were randomly assigned to three groups: blank (no intravenous) control (n=20), saline control (n=20) and excitotoxic brain damage model (ibotenic acid treatment, n=15). Behavioral function was evaluated by the surface righting reflex test (postnatal days 6-10), the swimming test (postnatal days 8-12) and the Y-maze discrimination learning test (postnatal days 33-34).</p><p><b>RESULTS</b>Righting time in the surface righting reflex test in the ibotenic acid treatment group on postnatal days 6-10 was more prolonged than that in the two control groups (p<0.05). Swimming test scores in the ibotenic acid treatment group were significantly lower than those in the two control groups (p<0.05). In the Y-maze discrimination learning test, the mice from the ibotenic acid treatment group performed significantly worse than two control groups, presenting with increased learning times (19.79+/-2.42 vs 16.29+/-2.48 or 16.30+/-2.37; p<0.05) and achieving a lower correct percentage (86.7% vs 96.5% or 95.0%) (p<0.05).</p><p><b>CONCLUSIONS</b>The developmental reflexes and learning and memory functions were impaired in neonatal mice following excitotoxic brain damage. Behavioral testing is useful in the evaluation of early developmental reflexes and long-term neurobehavioral outcome in neonatal mice with excitotoxic brain damage.</p>

The anteromedial extrastriate complex is critical for the use of allocentric visual cues and in the retention of the Lashley III maze task in rats

The anteromedial extrastriate complex has been proposed to play an essential role in a spatial orientation system in rats. To gain more information about that possible role, in the present work, two questions were addressed: 1. Are allocentric visual cues relevant for acquisition of the orientation task in the Lashley III maze? 2. Is this integration of allocentric inputs in the anteromedial visual complex relevant in the retention of this test? While a control group of rats was trained keeping the maze in the same position, the experimental group was trained with the maze rotated counterclockwise by 144 degrees from session to session. Control rats reached learning criterion significantly earlier and with less errors than the experimental ones (p<.05). After 11 sessions, rats of both groups received stereotaxic injections of ibotenic acid in the anteromedial complex. In the retention test one week after surgery, the control group, which had been able to learn using egocentric and allocentric visual cues, showed a greater deficit than the experimental animals (p<.05). These results confirm the role of the anteromedial complex in the processing of visuospatial orientation tasks and demonstrate the integration of allocentric visual cues in the solution of those tasks.

Protein tyrosine kinase inhibitors modify kainic acid-induced epileptiform activity and mossy fiber sprouting but do not protect against limbic cell death

Intrahippocampal administration of kainic acid (KA) induces synaptic release of neurotrophins, mainly brain-derived neurotrophic factor, which contributes to the acute neuronal excitation produced by the toxin. Two protein tyrosine kinase inhibitors, herbimycin A and K252a, were administered intracerebroventricularly, in a single dose, to attenuate neurotrophin signaling during the acute effects of KA, and their role in epileptogenesis was evaluated in adult, male Wistar rats weighing 250-300 g. The latency for the first Racine stage V seizure was 90 ± 8 min in saline controls (N = 4) which increased to 369 ± 71 and 322 ± 63 min in animals receiving herbimycin A (1.74 nmol, N = 4) and K252a (10 pmol, N = 4), respectively. Behavioral alterations were accompanied by diminished duration of EEG paroxysms in herbimycin A- and K252a-treated animals. Notwithstanding the reduction in seizure severity, cell death (60-90 percent of cell loss in KA-treated animals) in limbic regions was unchanged by herbimycin A and K252a. However, aberrant mossy fiber sprouting was significantly reduced in the ipsilateral dorsal hippocampus of K252a-treated animals. In this model of temporal lobe epilepsy, both protein kinase inhibitors diminished the acute epileptic activity triggered by KA and the ensuing morphological alterations in the dentate gyrus without diminishing cell loss. Our current data indicating that K252a, but not herbimycin, has an influence over KA-induced mossy fiber sprouting further suggest that protein tyrosine kinase receptors are not the only factors which control this plasticity. Further experiments are necessary to elucidate the exact signaling systems associated with this K252a effect.

Anti-seizure efficacy of nimodipine in pentylenetetrazole and kainic acid combined seizure models in mice.

The present study aimed at establishing two models of experimental seizures by combination treatment with subconvulsive doses of PTZ and kainic acid in adult male mice and evaluating the modulatory role of cerebroselective dihydropyridine calcium channel blocker, nimodipine. The CD50 +/- SEM value for PTZ was found to be 20.00 +/- 0.92 mg/kg, ip in kainic acid (administered at per se subconvulsive dose of 1.00 mg/kg, ip) pretreated mice while CD50 +/- SEM value for kainic acid was found to be 0.30 +/- 0.08 mg/kg, ip in PTZ (administered at per se subconvulsive dose of 30.00 mg/kg, ip) pretreated mice. Nimodipine (5.00 mg/kg, ip) significantly protected the mice from seizure in both of the combination in vivo seizure models. The results suggested synergistic interaction between PTZ and kainic acid at subconvulsive dose combination while the protective efficacy of nimodipine suggested the role of calcium ion as an important mediator for the genesis of seizures.

Protective effect of maternal immunoglobulin G against NMDA-induced neurotoxicity on hippocampus neurons / 中华儿科杂志

<p><b>OBJECTIVE</b>To investigate possible protective effect of maternal immunoglobulin G (IgG) against N-methyl-D-aspartate-mediated neurotoxicity on primary-cultured rat hippocampal neurons and the mechanism of the effect.</p><p><b>METHODS</b>An in vitro system had been developed for the study of hippocampal neurons. Intracellular lactic dehydrogenase (LDH) release was used as a marker to measure the rates of neuronal damage. The cells were stained with Trypan blue to measure the rate of neuronal death.</p><p><b>RESULTS</b>N-methyl-D-aspartate (NMDA) at a concentration of 50 micromol/L resulted in increased release of LDH and the cell mortality (P < 0.01, respectively). Maternal IgG of different concentration (10 mg/L, 100 mg/L) inhibited NMDA-induced intracellular LDH release (P < 0.01, respectively) and cell mortality (P < 0.05, 0.01, respectively), and larger dose had stronger effect (P < 0.05).</p><p><b>CONCLUSIONS</b>Maternal IgG had protective effect on primary-cultured rat hippocampal neurons injured by NMDA and the effect was dose-dependent.</p>

Effect of ketogenic diet on hippocampus synaptic reorganization and GluR5 expression in kainic acid induced rat model of epilepsy / 中华儿科杂志

<p><b>OBJECTIVE</b>Ketogenic diet (KD) is a high fat, low protein, low carbohydrate diet. Its antiepileptic effect is certain but the underlying mechanism is unknown. The aim of the study was to reveal the possible mechanism from the view points of synaptic reorganization and GluR(5) expression in hippocampus.</p><p><b>METHODS</b>Epilepsy was induced in Sprague-Dawley rats by kainic acid at postnatal day 28, all control animals were fed with normal rodent chow, whereas experimental rats were fed with ketogenic feed for 8 weeks. Spontaneous recurrent seizures were recorded. Mossy fiber sprouting and neuron damage in hippocampus were investigated by Timm staining and Nissl staining. Western blot and RT-PCR methods were applied to detect the expression of GluR(5) and GluR(5) mRNA in hippocampus.</p><p><b>RESULTS</b>KD-fed rats (1.40 +/- 1.03) had significantly fewer spontaneous recurrent seizures than control diet-fed rats (7.36 +/- 3.75). The mean A of mossy fiber sprouting in the inner molecular layer of dentate gyrus was markedly higher in KA induced animals than that in saline control animals but it was similar in different diet fed groups. No significant differences were found in the mean A of Timm staining in CA(3) area and Nissl staining of neuron in hilus, CA(3) and CA(1) area. After KA kindling, KD-fed animals [(189.38 +/- 40.03)/mg pro] had significantly higher GluR(5) expression in hippocampus than control diet-fed animals [(128.79 +/- 46.51)/mg pro] although their GluR(5) mRNA was the same.</p><p><b>CONCLUSION</b>Mossy fiber sprouting may be responsible for epileptogenesis in KA induced model and KD can suppress seizures in these animals. KD may upregulate young rat GluR(5) in inhibitory interneurons of CA(1) thus lead to an increased inhibition to prevent the propagation of seizure.</p>

Chronic excitotoxic lesion of the dorsal raphe nucleus induces sodium appetite

We determined if the dorsal raphe nucleus (DRN) exerts tonic control of basal and stimulated sodium and water intake. Male Wistar rats weighing 300-350 g were microinjected with phosphate buffer (PB-DRN, N = 11) or 1 æg/0.2 æl, in a single dose, ibotenic acid (IBO-DRN, N = 9 to 10) through a guide cannula into the DRN and were observed for 21 days in order to measure basal sodium appetite and water intake and in the following situations: furosemide-induced sodium depletion (20 mg/kg, sc, 24 h before the experiment) and a low dose of dietary captopril (1 mg/g chow). From the 6th day after ibotenic acid injection IBO-DRN rats showed an increase in sodium appetite (12.0 ± 2.3 to 22.3 ± 4.6 ml 0.3 M NaCl intake) whereas PB-DRN did not exceed 2 ml (P < 0.001). Water intake was comparable in both groups. In addition to a higher dipsogenic response, sodium-depleted IBO-DRN animals displayed an increase of 0.3 M NaCl intake compared to PB-DRN (37.4 ± 3.8 vs 21.6 ± 3.9 ml 300 min after fluid offer, P < 0.001). Captopril added to chow caused an increase of 0.3 M NaCl intake during the first 2 days (IBO-DRN, 33.8 ± 4.3 and 32.5 ± 3.4 ml on day 1 and day 2, respectively, vs 20.2 ± 2.8 ml on day 0, P < 0.001). These data support the view that DRN, probably via ascending serotonergic system, tonically modulates sodium appetite under basal and sodium depletion conditions and/or after an increase in peripheral or brain angiotensin II.

Sleep changes during chronic cold exposure showed that the homeostatic requirement of sleep is reduced in the medial preoptic area lesioned rats.

The effects of chronic exposure to a mildly cold ambient temperature (T(a)) of 18 degrees C on sleep wakefulness (S-W) and brain temperature (T(br)) were studied in the medial preoptic area (mPOA) lesioned male Wistar rats. Electroencephalogram (EEG), electrooculogram (EOG) and electromyogram (EMG) electrodes were chronically implanted to assess S-W, and a thermocouple above the dura to record the T(br). After three recordings (24 h each) of S-W and T(br) at 24 degrees C, N-methyl D-aspartic acid (NMDA) was intracerebrally injected to produce bilateral destruction of neurons in the mPOA. There was decreased sleep and increased T(br) even four weeks after the mPOA lesion. T(a) of the environmental chamber was then reduced to 18 degrees C, and the S-W and T(br) were again recorded for 24 h each on the 1st, 7th, 14th, 21st, and on 28th days of continuous exposure to the mild cold T(a). Exposure to the cold produced further decrease in sleep and increase in the T(br). However, sleep came back to the pre-exposure level by the 14th day. An increase in the duration of sleep episodes was responsible for the restoration of sleep during chronic cold exposure. The study showed that the requirement of sleep was reset at a lower level in the mPOA lesioned rats. The mPOA lesion affected the sleep maintenance and sleep initiation, though the latter became evident only during chronic cold exposure. The magnitude of the acute changes in T(br) and S-W were less in the lesioned rats, as compared to those observed in the normal rats exposed to similar cold T(a). On the basis of these observations, it could be proposed that the mPOA plays some role in cold induced changes in thermoregulation and sleep regulation. The T(br) remained elevated throughout the period of cold exposure. Resetting of the T(br), at a higher level may be part of the homeostatic readjustment to restore sleep.

Neurogenesis of dentate granule cells following kainic acid induced seizures in immature rats / 中华儿科杂志

<p><b>OBJECTIVE</b>Data accumulated over the past years have led to widespread recognition that neurogenesis, the emergence of new neurons, persists in the hippocampal dentate gyrus of the adult mammalian brain, and can be increased by seizures in multiple models. Also, aberrant reorganization of dentate granule cell axons, the mossy fiber sprouting, occurs in human temporal lobe epilepsy and rodent epilepsy models. However a number of studies suggest that the immature brain is less vulnerable to the morphologic alteration of hippocampus after seizures. The goal of this study was to determine whether the seizures can induce dentate granule cell neurogenesis and mossy fiber sprouting in the immature rat.</p><p><b>METHODS</b>Seizures was elicited by unilateral microinfusion of kainic acid (KA, 1 micro g) into the amygdula at postnatal day 15 (P15). Rat pups were given bromodeoxyuridine (BrdU) intraperitoneally on day 5 after KA administration and killed 7 d or 21 d later. The brains were processed for BrdU mitotic labeling combined with double-label immunohistochemistry using neuron-specific, early differentiation marker TuJ1 (betaIII tubulin) or granule-specific marker CaBP (calcium-binding protein calbindin D28k) as well as glia-specific marker GFAP (glial fibrillary acidic protein). Mossy fiber sprouting in intermolecular layer and CA3 subfield was assessed in Timm-stained sections both 1 month and 3 months after KA administration by using a rating scale and density measurement.</p><p><b>RESULTS</b>The dentate BrdU-immunoreactive cells of the KA-treated rats increased significantly compared with those of control rats on day 7 and 21 after BrdU administration (7 d: 244 +/- 15 vs. 190 +/- 10; 21 d: 218 +/- 19 vs. 133 +/- 12, P < 0.05). Approximately 80.2% and 78.7% of BrdU-labeled cells coexpressed TuJ1 in KA-treated rats and control rats on day 7 after BrdU respectively (P > 0.05). On 21 d after BrdU, 60.2% and 58.2% of dentate BrdU-labeled cells coexpressed GaBP in KA-treated rats and control rats respectively (P > 0.05). GFAP colocalized with 3%-5% dentate BrdU-labeled cells in the rats of both groups on day 7 and 21 after BrdU. It was also demonstrated that status epilepticus at P15 did not result in any detectable mossy fiber sprouting within the hippocampus both 1 month and 3 months after KA administration.</p><p><b>CONCLUSIONS</b>KA induced seizures can increase granule cell neurogenesis in the immature rat. Most of newly appeared cells migrate from subgranular proliferation zone (SGZ) into granule cell layer, the hilus as well as the molecular layer, and there they can differentiate into granule neurons. These observations also indicate that there is an early developmental resistance to seizure-induced mossy fiber sprouting in the immature brain.</p>

AMPA, not NMDA, activates RhoA GTPases and subsequetly phosphorylates moesin

Glutamate induced rapid phosphorylation of moesin, one of ERM family proteins involved in the ligation of membrane to actin cytoskeleton, in rat hippocampal cells (JBC, 277:16576-16584, 2002). However, the identity of glutamate receptor has not been explored. Here we show that a-amino- 3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor is responsible for glutamate-induced RhoA activation and phosphorylation of moesin. Glutamate induced phosphorylation at Thr-558 of moesin was still detectible upon chelation of Ca(2+), suggesting involvement of AMPA receptor instead of N-methyl D-Aspartate (NMDA) receptor in this phosphorylation of moesin. AMPA but not NMDA- induced moesin phosphorylation was independent of Ca(2+). Both AMPA and NMDA but not Kainate induced moesin phosphorylation at similar levels. However, the kinetics of phosphorylation varied greatly between AMPA and NMDA where AMPA treatment rapidly increased phosphomoesin, which reached a maximum at 10 min after treatment and returned to a basal level at 30 min. In contrast, NMDA-induced phosphorylation of moesin reached a maximum at 30 min after treatment and was remained at higher levels at 60 min. A possible involvement of RhoA and its downstream effector, Rho kinase in the AMPA receptor-triggered phosphorylation of moesin was also explored. The kinetics for the glutamate- induced membrane translocation of RhoA was similar to that of moesin phosphorylation induced by AMPA. Moreover, Y-27632, a specific Rho kinase inhibitor, completely blocked AMPA-induced moesin phosphorylation but had no effect on NMDA-induced moesin phosphorylation. These results suggest that glutamate-induced phosphorylation of moesin may be mediated through the AMPA receptor/RhoA/Rho kinase pathway.

Changes in metabotropic glutamate receptor 4 expression and the effects of L-2-amino-4-phosphonobutyrate in a rodent model of diffuse brain injury / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To examine the changes in the expression of mGluR4 after diffuse brain injury (DBI) and to determine the role of its specific agonist L-2-amino-4-phosphonobutyrate (L-AP4) in vivo.</p><p><b>METHODS</b>A total of 161 male SD rats were randomized into the following groups. Group A included normal control, sham-operated control and DBI group. DBI was produced according to Marmarou's diffuse head injury model. mRNA expression of mGluR4 was detected by hybridization in situ. Group B included DBI alone, DBI treated with normal saline and DBI treated with L-AP4. All DBI rats were trained in a series of performance tests, following which they were subjected to DBI. At 1 and 12 hours, animals were injected intraventricularly with L-AP4 (100 mmol/L, 10 microl) or normal saline. Motor and cognitive performances were tested at 1, 3, 7, 14 days after injury and the damaged neurons were also detected.</p><p><b>RESULTS</b>There was no significant difference between normal control group and sham-operated group in the expression of mGluR4 (P>0.05). The animals exposed to DBI showed significantly increased expression of mRNA of mGluR4 compared with the sham-operated animals 1 h after injury (P<0.05). At 6 hours, the evolution of neuronal expression of mGluR4 in the trauma alone group was relatively static. Compared with saline-treated control animals, rats treated with L-AP4 showed an effective result of decreased number of damaged neurons and better motor and cognitive performances.</p><p><b>CONCLUSIONS</b>Increased expression of mGluR4 is important in the pathophysiological process of DBI and its specific agonist L-AP4 can provide remarkable neuroprotection against DBI not only at the histopathological level but also in the motor and cognitive performance.</p>