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Hippocampal function is important for learning and memory, and dysfunction of the hippocampus has been linked to the pathophysiology of neuropsychiatric diseases such as schizophrenia. Neuregulin1 (NRG1) and ErbB4, two susceptibility genes for schizophrenia, reportedly modulate long-term potentiation (LTP) at hippocampal Schaffer collateral (SC)-CA1 synapses. However, little is known regarding the contribution of hippocampal NRG1/ErbB4 signaling to learning and memory function. Here, quantitative real-time PCR and Western blotting were used to assess the mRNA and protein levels of NRG1 and ErbB4. Pharmacological and genetic approaches were used to manipulate NRG1/ErbB4 signaling, following which learning and memory behaviors were evaluated using the Morris water maze, Y-maze test, and the novel object recognition test. Spatial learning was found to reduce hippocampal NRG1 and ErbB4 expression. The blockade of NRG1/ErbB4 signaling in hippocampal CA1, either by neutralizing endogenous NRG1 or inhibiting/ablating ErbB4 receptor activity, enhanced hippocampus-dependent spatial learning, spatial working memory, and novel object recognition memory. Accordingly, administration of exogenous NRG1 impaired those functions. More importantly, the specific ablation of ErbB4 in parvalbumin interneurons also improved learning and memory performance. The manipulation of NRG1/ErbB4 signaling in the present study revealed that NRG1/ErbB4 activity in the hippocampus is critical for learning and memory. These findings might provide novel insights on the pathophysiological mechanisms of schizophrenia and a new target for the treatment of Alzheimer's disease, which is characterized by a progressive decline in cognitive function.

Special AT-rich sequence-binding protein 2 (Satb2) is a protein binding to the matrix attachment regions of DNA and important for gene regulation. Patients with SATB2 mutation usually suffer moderate to severe mental retardation. However, the mechanisms for the defects of intellectual activities in patients with SATB2 mutation are largely unclear. Here we established the heterozygous Satb2 mutant mice and Satb2 conditional knockout mice to mimic the patients with SATB2 mutation and figured out the role of Satb2 in mental activities. We found that the spatial memory and working memory were significantly damaged in the heterozygous Satb2 mutant mice, early postnatal Satb2-deficient mice (CaMKIIα-Cre + Satb2 fl/fl mice), and adult Satb2 ablation mice (Satb2 fl/fl mice injected with CaMKIIα-Cre virus). Functionally, late phase long-term potentiation (L-LTP) in these Satb2 mutant mice was greatly impaired. Morphologically, in CA1 neurons of CaMKIIα-Cre + Satb2 fl/fl mice, we found decreased spine density of the basal dendrites and less branches of apical dendrites that extended into lacunar molecular layer. Mechanistically, expression levels of immediate early genes (IEGs) including Fos, FosB, and Egr1 were significantly decreased after Satb2 deletion. And, Satb2 could regulate expression of FosB by binding to the promoter of FosB directly. In general, our study uncovers that Satb2 plays an important role in spatial memory and working memory by regulating IEGs-mediated hippocampal synaptic plasticity.

Neurotransmission requires precise control of neurotransmitter release from axon terminals. This process is regulated by glial cells; however, the underlying mechanisms are not fully understood. We found that glutamate release in the brain was impaired in mice lacking low-density lipoprotein receptor-related protein 4 (Lrp4), a protein that is critical for neuromuscular junction formation. Electrophysiological studies revealed compromised release probability in astrocyte-specific Lrp4 knockout mice. Lrp4 mutant astrocytes suppressed glutamatergic transmission by enhancing the release of ATP, whose level was elevated in the hippocampus of Lrp4 mutant mice. Consequently, the mutant mice were impaired in locomotor activity and spatial memory and were resistant to seizure induction. These impairments could be ameliorated by blocking the adenosine A1 receptor. The results reveal a critical role for Lrp4, in response to agrin, in modulating astrocytic ATP release and synaptic transmission. Our findings provide insight into the interaction between neurons and astrocytes for synaptic homeostasis and/or plasticity.

Background: The role of ERBB4 in liver disease has seldom been reported. This study aims to find genetic markers at ERBB4 for chronic hepatitis B virus (HBV) infection and determine the role of ERBB4 in liver injury., Methods: We selected and genotyped three single nucleotide polymorphisms and one insertion/deletion (Ins/Del) at the 5' and 3' untranslated region (UTR) of ERBB4 in a case-control study including 1344 pairs of HBV carriers and HBV natural clearance subjects. The luciferase reporter system was applied to study the regulative role of Ins/Del on ERBB4. Further, ERBB4 knockout mice were used to study the role of ERBB4 in liver injury. Proteomic quantification was performed by HPLC-MS/MS analysis to identify liver protein profile change between liver-specific ERBB4 knockout and control mice., Results: rs6147150 Ins/Del and rs1836724 T>C at the 3' UTR of ERBB4 were associated with reduced risk of chronic HBV infection (P = 0.002 and 0.004, respectively). Besides, the 12bp deletion at the 3' UTR increased ERBB4 expression due to lacking let-7c binding site. In addition, loss of ERBB4 led to more severe acute or chronic inflammation in mouse liver injury models. Further, quantitative proteomic analysis and data from the cancer genome atlas revealed that ACLY, an enzyme key for de novo lipogenesis, was negatively correlated with ERBB4., Conclusions: ERBB4 plays protective role from liver injury and its 3'UTR genetic variants could be genetic markers for chronic HBV infection.

Social isolation during the vulnerable period of adolescence produces emotional dysregulation that often manifests as abnormal behavior in adulthood. The enduring consequence of isolation might be caused by a weakened ability to forget unpleasant memories. However, it remains unclear whether isolation affects unpleasant memories. To address this, we used a model of associative learning to induce the fear memories and evaluated the influence of isolation mice during adolescence on the subsequent retention of fear memories and its underlying cellular mechanisms. Following adolescent social isolation, we found that mice decreased their social interaction time and had an increase in anxiety-related behavior. Interestingly, when we assessed memory retention, we found that isolated mice were unable to forget aversive memories when tested 4 weeks after the original event. Consistent with this, we observed that a single train of high-frequency stimulation (HFS) enabled a late-phase long-term potentiation (L-LTP) in the hippocampal CA1 region of isolated mice, whereas only an early-phase LTP was observed with the same stimulation in the control mice. Social isolation during adolescence also increased brain-derived neurotrophic factor (BDNF) expression in the hippocampus, and application of a tropomyosin-related kinase B (TrkB) receptor inhibitor ameliorated the facilitated L-LTP seen after isolation. Together, our results suggest that adolescent isolation may result in mental disorders during adulthood and that this may stem from an inability to forget the unpleasant memories via BDNF-mediated synaptic plasticity. These findings may give us a new strategy to prevent mental disorders caused by persistent unpleasant memories.

Ischemic injury in rodent models reliably leads to the activation of microglia, which might play a detrimental role in neuronal survival. Our preliminary studies suggest that nicotine plays a potential role in decreasing the numbers of cultured microglia in vitro. In the present study, we found treatment with nicotine 2, 6, and 12 h after ischemia for 7 days significantly increased the survival of CA1 pyramidal neurons in ischemia/reperfusion rats. This effect was accompanied by a significant reduction in the increase of microglia rather than astrocytes, as well as a significant reduction of enhanced expression of tumor necrosis factor-alpha (TNF-α) and interleukin-1beta (IL-1β) in CA1 induced by ischemia/reperfusion. Nicotine inhibits microglial proliferation in primary cultures with and without the stimulation of granulocyte-macrophage colony-stimulating factor (GM-CSF). Pre-treatment with α-bungarotoxin, a selective α7 nicotinic acetylcholine receptor (α7 nAChR) antagonist, could prevent the inhibitory effects of nicotine on cultured microglial proliferation suggesting that nicotine inhibits the microglial proliferation in an α7 nAChR-dependent fashion. Our results suggest that nicotine inhibits the inflammation mediated by microglia via α7 nAChR and is neuroprotective against ischemic stroke, even when administered 12 h after the insult. α7 nAChR agonists may have uses as anti-ischemic compounds in humans.

The trophic factor neuregulin 1 (Nrg1) and its receptor ErbB4 are schizophrenia candidate genes. NRG1-ErbB4 signaling was thought to regulate spine formation and function in a cell-autonomous manner. Yet, recent studies indicate that ErbB4 expression is largely restricted to GABAergic interneurons and is very low or absent in pyramidal cells. Here, we generated and characterized cell type-specific ErbB4 mutant and transgenic mice. Spine density and the number of excitatory synapses were unaltered by neither deletion nor overexpression of ErbB4 in pyramidal neurons. However, spine density and excitatory synapse number were reduced in PV-ErbB4(-/-) mice where ErbB4 was selectively ablated in parvalbumin-positive GABAergic interneurons. Concurrently, basal glutamate transmission was impaired in PV-ErbB4(-/-) mice, but not in mice where ErbB4 was deleted or overexpressed in pyramidal neurons. Our results demonstrate a role of ErbB4 in PV-positive interneurons for spine formation in excitatory neurons.

Excessive K(+) efflux promotes central neuronal apoptosis; however, the type of potassium channel that mediates K(+) efflux in response to different apoptosis-inducing stimuli is still unknown. It is hypothesized that the activation of large-conductance Ca(2+)-activated K(+) channels (BKCa) mediates hypoxia/reoxygenation (H/R)- and ischemia/reperfusion (I/R)-induced neuronal apoptosis. Rat hippocampal neuronal cultures underwent apoptosis after reoxygenation, as assessed by morphologic observation, terminal deoxynucleotidyl transferase dUTP nick end labeling staining, and caspase-3 activation. Single-channel recordings revealed upregulation of BKCa channel activity 6 h after reoxygenation, which might be caused by elevated cytosolic Ca(2+). The K(+) ionophore valinomycin and the BKCa channel opener NS1619 induced neuronal apoptosis. Transfection of the BKCa channel α subunit into Chinese hamster ovary (CHO-K1) cells, which do not express endogenous K(+) channels, or into neurons will induce cell apoptosis, indicating that the opening of the BKCa channel serves as a pivotal event in mediating cell apoptosis. The specific BKCa channel blockers charybdotoxin and iberiotoxin and the nonselective K(+) channel blocker tetraethylammonium at concentrations more specific to the BKCa channel were neuroprotective. The A-type potassium channel blocker 4-aminopyridine and apamin, a small-conductance Ca(2+)-activated K(+) channel blocker, were not protective. This result suggests the involvement of the BKCa channel in H/R-induced apoptosis. Similarly, specific BKCa channel blockers also showed neuroprotection in neurons subjected to oxygen-glucose deprivation/reoxygenation or animals subjected to forebrain ischemia-reperfusion. These results demonstrate that the over-activity of BKCa channels mediates hippocampal neuronal damage induced by H/R in vitro and I/R in vivo.

Revealing the neurobiological mechanism of depression has always been a big challenge in the field of neuroscience. Not only are depressive syndromes heterogeneous and their aetiologies diverse, but also some symptoms are impossible to reproduce in animal models. Nevertheless, great progress has been made on the understanding and treatment of depression in recent years. In this review, we focus on key leading hypotheses in the neurobiological mechanism of depression, examine their strengths and weaknesses critically, and also highlight new insights that promise to extend the understanding of depression and its treatment.

Neuregulin 1 (NRG1) is a trophic factor that acts by stimulating ErbB receptor tyrosine kinases and has been implicated in neural development and synaptic plasticity. In this study, we investigated mechanisms of its suppression of long-term potentiation (LTP) in the hippocampus. We found that NRG1 did not alter glutamatergic transmission at SC-CA1 synapses but increased the GABA(A) receptor-mediated synaptic currents in CA1 pyramidal cells via a presynaptic mechanism. Inhibition of GABA(A) receptors blocked the suppressing effect of NRG1 on LTP and prevented ecto-ErbB4 from enhancing LTP, implicating a role of GABAergic transmission. To test this hypothesis further, we generated parvalbumin (PV)-Cre;ErbB4(-/-) mice in which ErbB4, an NRG1 receptor in the brain, is ablated specifically in PV-positive interneurons. NRG1 was no longer able to increase inhibitory postsynaptic currents and to suppress LTP in PV-Cre;ErbB4(-/-) hippocampus. Accordingly, contextual fear conditioning, a hippocampus-dependent test, was impaired in PV-Cre;ErbB4(-/-) mice. In contrast, ablation of ErbB4 in pyramidal neurons had no effect on NRG1 regulation of hippocampal LTP or contextual fear conditioning. These results demonstrate a critical role of ErbB4 in PV-positive interneurons but not in pyramidal neurons in synaptic plasticity and support a working model that NRG1 suppresses LTP by enhancing GABA release. Considering that NRG1 and ErbB4 are susceptibility genes of schizophrenia, these observations contribute to a better understanding of how abnormal NRG1/ErbB4 signaling may be involved in the pathogenesis of schizophrenia.

Objective: To construct a RNA interfering plasmid targeting rat Bcl-2/E1B-19K-interacting protein 3 (BNIP3) and assess its effect on exogenous BNIP3 expression in HEK293 cells., Methods: The miRNA sequences were designed using Invitrogen BLOCK-iT RNAi Designer and synthesized into double-strand oligonucleotides, which were cloned into the pcDNATM6.2-GW/EmGFP-miR vector, followed by transformation of the product into competent Top10 E. coli cells. After expansion of the transformed bacteria, the plasmid was extracted and sequenced before its co-transfection with pEGFP-C3- rBNIP3 plasmid into HEK293 cells. The interference effect of the constructed plasmid on BNIP3 mRNA and protein expression were detected by real-time PCR and Western blotting., Results: The sequencing result indicated that the interfering plasmid targeting rat BNIP3 was constructed correctly. After transfection into HEK293 cells, the interfering plasmid significantly inhibited exogenous BNIP3 mRNA and protein expressions., Conclusion: The RNA interfering plasmid targeting rat BNIP3 has been constructed successfully, which provides a useful tool for studying the function of BNIP3.

Increasing evidence indicates that stimulating hippocampal neurogenesis could provide novel avenues for the treatment of depression, and recent studies have shown that in vitro neurogenesis is enhanced by hypoxia. The aim of this study was to investigate the potential regulatory capacity of an intermittent hypobaric hypoxia (IH) regimen on hippocampal neurogenesis and its possible antidepressant-like effect. Here, we show that IH promotes the proliferation of endogenous neuroprogenitors leading to more newborn neurons in hippocampus in adult rats. Importantly, IH produces antidepressant-like effects in multiple animal models screening for antidepressant activity, including the forced swimming test, chronic mild stress paradigm, and novelty-suppressed feeding test. Hippocampal x-ray irradiation blocked both the neurogenic and behavioral effects of IH, indicating that IH likely produces antidepressant-like effects via promoting neurogenesis in adult hippocampus. Furthermore, IH stably enhanced the expression of BDNF in hippocampus; both the antidepressant-like effect and the enhancement of cell proliferation induced by IH were totally blocked by pharmacological and biological inhibition of BDNF-TrkB (tyrosine receptor kinase B) signaling, suggesting that the neurogenic and antidepressant-like effects of IH may involve BDNF signaling. These observations might contribute to both a better understanding of physiological responses to IH and to developing IH as a novel therapeutic approach for depression.

Current antidepressants are clinically effective only after several weeks of administration. We show that Fuzi polysaccharide-1 (FPS), a new water-soluble polysaccharide isolated from Fuzi, which has been used to treat mood disorders in traditional Chinese medicine for centuries, increases the number of newborn cells in the dentate gyrus in adult mice, and most of these cells subsequently differentiate into new neurons. We also found that FPS administration reduces immobility in the forced swim test, and latency in the novelty suppressed-feeding test. Moreover, a 14-d regimen with FPS reverses avoidance behaviour and inhibition of hippocampal neurogenesis induced by chronic defeat stress. In contrast, imipramine, a well known antidepressant, reverses this avoidance behaviour only after 4 wk of continuous administration. Finally, acute treatment with FPS had no effect on brain monoamine levels in frontal cortex but significantly increases BDNF in the hippocampus, while the antidepressant effect and enhancement of cell proliferation induced by FPS administration were totally blocked by K252a, an inhibitor of trkB in a chronic social defeat depression model, suggesting that the neurogenic and antidepressant effects of FPS may involve BDNF signalling. In conclusion, our findings suggest that FPS could be developed as a putative antidepressant with a rapid onset of action.

Ethnopharmacological Relevance: Xiao-Xu-Ming decoction (XXMD) has long been employed clinically to treat stroke in traditional Chinese Medicine., Aim of the Study: To investigate the neuroprotective effects of XXMD in vivo and in vitro stroke models and determine involved mechanisms., Materials and Methods: Two models (four-vessel occlusion in adult Wistar rats and oxygen-glucose deprivation primary cultured neurons) were employed to mimic ischemia-reperfusion damage, in vivo and in vitro, respectively. The effects of XXMD were investigated with respect to neuronal damage, activity of caspase-3 and expression of Bcl-2 in CA1 region of hippocampus after ischemia. The cognitive ability was measured 7 days after ischemia/reperfusion by using Morris water maze., Results: Oral administration of XXMD significantly increased the density of neurons that survived in the CA1 region of hippocampus on the 3rd and 7th day after transient global ischemia was induced in a dose-dependent manner. XXMD ameliorated severe deficiencies in spatial cognitive performance induced by transient global ischemia. Inhibition of caspase-3 activity and up-regulation of Bcl-2 expression were induced in the high dose of XXMD-treated rats after ischemia. In oxygen-glucose deprivation model, both XXMD extract and drug-containing serum prepared from blood of high dose of XXMD-treated rats inhibited apoptotic neuronal death at 24h after reoxygenation., Conclusions: Our results clearly demonstrated that XXMD is neuroprotective and appears to influence deleterious pathological processes that are activated after the onset of ischemia., (Crown Copyright 2009. Published by Elsevier Ireland Ltd. All rights reserved.)

Objective: To study the effects of strychnos alkaloids on the proliferation of adult rat neuroprogenitor cells., Methods: Strychnos alkaloids free of strychnine and brucine were extracted from Strychnos nux vomica, and the effects of Strychnos alkaloids on the survival of HEK293 and PC12 cells were evaluated using MTT assay. In vitro cultured adult rat neuroprogenitor cells isolated from the hippocampus were treated with different concentrations of Strychnos alkaloids for 2 days, and the cell proliferation was assessed using BrdU incorporation assay., Results: At the concentration above 0.5 mg/ml, Strychnos alkaloids produced toxic effect against HEK293 cells (P<0.0001), while for PC12 cells, Strychnos alkaloids inhibited the cell survival at the concentration as low as 5 microg/ml (P<0.0001). After 2 days of exposure to 50 microg/ml Strychnos alkaloids, the neuroprogenitor cells showed significantly decreased number of BrdU-positive cells (P<0.01), but the total cell number remained stable when compared with that of the control cells (P>0.05), whereas at the concentration of 100 microg/ml, Strychnos alkaloids produced obvious cytotoxicity against the neuroprogenitor cells., Conclusion: Strychnos alkaloids can significantly inhibit the proliferation of adult rat neuroprogenitor cells, and this effect is probably selective, suggesting the potential of Strychnos alkaloids as a new drug for treatment of neurocytoma.

Objective: To investigate the effect of neurogulin-1 (NRG1) on the transmission and plasticity of CA1 synapses in mouse hippocampal slices at different temperatures., Methods: Under room temperature (26-/+1 degrees C) or physiological temperature (32-/+1 degrees C), field excitatory postsynaptic potential (fEPSP) evoked by extracellular microelectrode recording technique was recorded in the CA1 region in adult mouse hippocampal slices, and the long-term potentiation (LTP) was induced by high frequency stimulation (HFS). The effects of NRG1 on the baseline fEPSP and induction of LTP in CA1 region were observed., Results: No significant variation in the slope of fEPSP relative to the baseline fEPSP was observed after perfusion with NRG1 at room temperature or physiological temperature (P > 0.05). After HFS at the room temperature, the mean slope of fEPSP in the slices perfused with NRG1 was similar to that of the control group (P > 0.05), but HFS at the physiological temperature resulted in significant reduction in the mean slope of fEPSP in the slices perfused with NRG1 (P < 0.01)., Conclusion: NRG1 may temperature-dependently inhibit the induction of LTP in the CA1 region in mouse hippocampal slices.

Paeoniflorin is one of the bioactive components of Paeonia lactiflora, a traditional Chinese herbal medicine. It is the main monoterpene glucoside isolated from the P. lactiflora in 1963. Since then, researchers have found that paeoniflorin has multifold pharmacological effects. In this review, based on the recent available papers published in PubMed and National Knowledge Infrastructure Data Base, we present the major current approaches in understanding the detection methodology, pharmacokinetics and pharmacology, and toxicology of paeoniflorin.

Synapse formation requires proper interaction between pre- and postsynaptic cells. In anterograde signaling, neurons release factors to guide postsynaptic differentiation. However, less is known about how postsynaptic targets retrogradely regulate presynaptic differentiation or function. We found that muscle-specific conditional knockout of beta-catenin (Ctnnb1, also known as beta-cat) in mice caused both morphologic and functional defects in motoneuron terminals of neuromuscular junctions (NMJs). In the absence of muscle beta-catenin, acetylcholine receptor clusters were increased in size and distributed throughout a wider region. Primary nerve branches were mislocated, whereas secondary or intramuscular nerve branches were elongated and reduced in number. Both spontaneous and evoked neurotransmitter release was reduced at the mutant NMJs. Furthermore, short-term plasticity and calcium sensitivity of neurotransmitter release were compromised in beta-catenin-deficient muscle. In contrast, the NMJ was normal in morphology and function in motoneuron-specific beta-catenin-deficient mice. Taken together, these observations indicate a role for muscle beta-catenin in presynaptic differentiation and function, identifying a previously unknown retrograde signaling in the synapse formation and synaptic plasticity.

Objective: To investigate prolonged use of gonadotropin (Gn) stimulation for the patients with follicular development retardation during controlled hyperstimulation (COH) for in vitro fertilization (IVF) cycles., Methods: The inclusion criteria to categorize the good responder were that duration of Gn stimulation was < or = 15 days, total Gn dose used was < or = 3300 IU and total oocytes retrieved were > or = 4. The inclusion criteria for prolonged duration of Gn stimulation were that duration of stimulation was > or = 16 days. There were 69 oocyte retrieval cycles and 66 transfer cycles in prolonged stimulation group (group 1) and 483 oocyte retrieval cycles and 464 transfer cycles in good ovarian response group (group 2). The clinical characteristics and outcomes of two groups were analyzed., Results: Clinical 5 pregnancy rate, implantation rate and delivery rate were 45.5% (30/66) versus 51.7% (240/464), 30.5% (385/1262) versus 28.0% (46/164) and 37.9% (25/66) versus 39.4% (183/464), respectively, in groups 1 and 2. Duration of Gn stimulation and dose were (20.8 +/- 4.2) and (10.3 +/- 1.8) days, (3090 +/- 1140) and (2302 +/- 862) IU in groups 1 and 2, respectively. There was no statistical difference in patient's age, basal follicular stimulating hormone (FSH), clinical pregnancy rate, implantation rate, early miscarriage rate, ovarian hyperstimulation syndrome (OHSS) rate, delivery rate between two groups (P > 0.05). There were more polycystic ovary (PCO) and (or) polycystic ovarian syndrome (PCOS) patients, more basal antral follicles, longer duration of Gn stimulation (range 16 - 33 days), higher Gn dose, lower serum peak estradiol (E(2)) level, fewer oocytes, fewer embryos transferred, in group 1 compared with group 2 (P < 0.01)., Conclusions: These results demonstrate that prolonged Gn stimulation more than 16 days is a valuable alternative before cancellation of the IVF cycles for follicular development retardation during COH. Good clinical outcome can be achieved including pregnancy rate, implantation rate and delivery rate.

Objective: To investigate changes in synaptic and extrasynaptic N-methyl-D-aspartate receptors (NMDAR) during the development of cultured rat hippocampal neurons., Methods: Synaptic and extrasynaptic NMDAR channel currents were recorded from 1-day-old rat hippocampal neurons cultured for 1 and 2 weeks with patch-clamp technique in whole-cell configuration and outside-out configuration, respectively., Results: The amplitude of NMDAR-mediated miniature excited postsynaptic current (Meps(CNMDA)) decreased in neurons cultured for 2 weeks as compared with that recorded in neurons cultured for 1 week, and the 2-week neurons showed also much lowered sensitivity to selective NR2B blocker ifenprodil. The amplitude and open probability of extrasynaptic NMDAR in the 2-week neurons were significantly higher than those in the 1-week neurons, but the neurons differred little in conduction and reverse potential. Ifenprodil decreased the high conductance and open probability in both neurons, but the effect was more potent in the 2-week ones., Conclusions: There can be developmental changes in synaptic and extrasynaptic NMDAR channel currents in cultured rat hippocampal neurons, indicating that different NMDAR subtypes are expressed in the synaptic and extrasynaptic regions during the development of the hippocampal neurons. In 1-week neurons, NR2B are predominant both in synaptic and extrasynaptic regions, and at 2 weeks, synaptic NR2B are replaced by NR2A but NR2B still remains the predominant subtypes outside the synapses.

Objective: To investigate the periodic quantity variation of proliferating neuronal progenitors after global brain ischemia and provide evidence for choosing the time-window of drug therapy to promote neuronal regeneration after ischemia., Methods: Adult male Wistar rats were subjected to 15-min global brain ischemia (four-vessel occlusion model) and randomized subsequently into 8 groups (n=3). The rats were given intraperitoneal injections of BrdU (75 mg/kg) for 4 times daily (at a 2-hour interval) since day 7 till day 11 after ischemia, and on day 29, the rats were perfused transcardially for fixation. Another 3 normal rats were given BrdU in the same manner and killed the next day. Coronal sections of the brain tissue (30 microm) were prepared for immunocytochemical detection of BrdU-labeled cells and immunofluorescent detection of BrdU/NeuN double-labeled cells. The density of BrdU-positive cells and BrdU/NeuN double-labeled cells in the hippocampal dentate gyrus (DG) and CA1 region were counted and the density of proliferating cells at different days after ischemia were compared using one-way ANOVA., Results: The proliferation of the neuronal progenitors increased after global brain ischemia. The number of BrdU-positive cells in the DG and CA1 region decreased gradually in 7-10 days after ischemia, and reached the normal level during 11-14 days. The differentiation of the progenitors did not vary after ischemia., Conclusion: Increased proliferation of the neuroprogenitors occurs mainly within the initial 10 days after global ischemia in rats.

Activation of N-methyl-d-aspartic acid (NMDA) receptor plays an important role in neuronal apoptosis induced by cerebral ischemia but the underlying mechanisms are still unclear. The present study examined the neuroprotection of three chloride blockers in an in vitro cell model of cerebral ischemia established by treatment of cultured rat hippocampal neurons with NMDA. Hoechst 33258 staining and MTT assay were used to detect neuronal apoptosis and cell viability, respectively. The neuroprotective effects of chloride channel blockers on the cell viability and neuronal apoptosis were only observed when the blockers were applied before NMDA exposure. In comparison with DIDS, SITS showed more potent protective effect in a dose-dependent manner, whereas NPPB showed no significant neuroprotective effect. The results demonstrate that pretreatment with both SITS and DIDS have protective effect against neuronal apoptosis, which is achieved by blocking both NMDA receptor and chloride channel.

Objective: To study the localization of L-type calcium channel alpha1 subunits CaV1.2alpha1C and CaV1.3alpha1D in CA1 and CA3 regions of adult rat hippocampus., Methods: Immunohistochemical staining was employed for specific labeling of alpha1 subunits CaV1.2alpha1C and CaV1.3alpha1D., Results: CaV1.2alpha1C subunit was mainly located in the apical and basal dendrites in the CA1 area and the CA3 area, neuronal soma, basal dendrites and distal apical dendrites were all positively stained. In contrast to CaV1.2alpha1C, CaV1.3alpha1D displayed no obvious difference in immunostaining between CA1 and CA3, and was distributed mainly in the neuronal soma and proximal dendrites. At cellular level, CaV1.2alpha1C distribution showed a distinct clustered pattern while a uniform distribution was observed for CaV1.3alpha1D subunit., Conclusion: Different alpha1 subunits of L-type calcium channel have differential expression in adult hippocampal neurons.

Objective: To establish a method for analysis of the subproteomes following membrane protein enrichment, so as to investigate the differences in cytosolic and membrane protein/peptide contents among different hippocampal subregions (CA1, CA3 and dentate gyrus) of rats., Methods: The hippocampal subregions were isolated by microdissection, and after membrane protein enrichment with protein solubility-based sequential sample extraction, subproteome profiling was accomplished using modified two-dimensional electrophoresis and silver staining method compatible with high-performance mass spectrometry, which displayed cytosolic or membrane proteins/peptides separately. The efficacy of the electrophoresis was evaluated by image analyzing software., Results: The membrane proteins were enriched by 8 folds, and around 30% more spots were shown on the integrated dentate gyrus map than on the conventional map obtained by one-step protein extraction. Such improvement could also be seen in protein mapping of CA1 and CA3 regions., Conclusion: When analyzing hippocampal subregions sequential extraction and two-dimensional electrophoresis more effectively separate and display enriched membrane protein/peptides, which may play important roles in such cellular events as signal transduction.

Objective: To examine the neuroprotective effects of Naoxintong and Zhongfenghuichunwan on neuronal injury in CA1 region of rat hippocampus following transient forebrain ischemia., Methods: Transient rat forebrain ischemia for 15 min was induced by modified four-vessel occlusion method. The density of survived pyramidal cells (neurons per 1 mm linear length) in CA1 region of the hippocampus was measured under light microscope., Results: In Naoxintong or nimodipine-treated rats, the density of survived pyramidal cells in CA1 region was significantly greater than that of saline group (P<0.05, P<0.001, respectively), but oral administration of Zhongfenghuichunwan had no obvious effect on ischemia-induced neuronal damage in CA1 region., Conclusion: Naoxintong can protect CA1 neurons against ischemic insult in rats.

An improved method is described for fast and reliable isolation of neurons from hippocampus of adult rats by a combination of mechanical and enzymatic means. The procedure allows the isolation of neurons from 500-600-d-old rats (over 300 g), preserving the proximal dendritic structure without impairing the electrical characteristics of the cells. Morphologically distinct neurons can be recognized. Using cell-attached, inside-out and whole-cell configurations of patch clamp technique, it was shown that the enzymatically isolated neurons in hippocampus from rats weighing more than 300 g exhibited voltage-gated calcium, sodium and potassium currents, outwardly rectifying chloride channel and large conductance Ca(2+)-activated potassium channel currents. Approximately, 95% of healthy cells allowed the formation of giga-ohm seals.

Objective: To investigate the effects of potassium channel inhibitors on hypoxia/reoxygenation-induced death of cultured hippocampal neurons., Methods: Cultured 8 d in vitro, hippocampal neurons were exposed to hypoxia (in mixture of 95% N2 and 5% CO2) for 6 h, and then reoxygenated till the 72nd hour. Different potassium channel inhibitors were applied to the culture solution separately after reoxygenation. Neuron death was analyzed with cell counting and MTT assay., Results: Hypoxia/reoxygenation procedure induced a delayed death of cultured hippocampal neurons. Application of tetraethylammonium (TEA) offered concentration-dependent protection of the neurons against death. Selective high-conductance calcium-activated potassium channel (BK) inhibitor iberiotoxin (IbTX) showed significant neuron protection (P<0.001). However, A-type potassium channel inhibitor 4-aminopyridine presented no protection against neuron death (P>0.05)., Conclusion: Following hypoxia/reoxygenation, enhanced activity of potassium channel, especially BK channel, may induce neuron death.

Xiaoxuming decoction, an ancient traditional Chinese medical prescription for treatment of stroke, has somehow lost its clinical application since Tang Dynasty. Based on the historic record of this prescription, mechanisms of composition and clinical and experimental studies, the authors attempt to evaluate the standing of Xiaoxuming decoction as a remedy for stroke.