NMDA Receptor Subunit NR2B Research Articles

Bicuculline Reduces Neuroinflammation in Hippocampus and Improves Spatial Learning and Anxiety in Hyperammonemic Rats. Role of Glutamate Receptors.

Patients with liver cirrhosis may develop minimal hepatic encephalopathy (MHE) with mild cognitive impairment. Hyperammonemia is a main contributor to cognitive impairment in MHE, which is mediated by neuroinflammation. GABAergic neurotransmission is altered in hyperammonemic rats. We hypothesized that, in hyperammonemic rats, (a) enhanced GABAergic tone would contribute to induce neuroinflammation, which would be improved by reducing GABAergic tone by chronic bicuculline treatment; (b) this would improve spatial learning and memory impairment; and (c) modulation of glutamatergic neurotransmission would mediate this cognitive improvement. The aim of this work was to assess the above hypotheses. Bicuculline was administrated intraperitoneally once a day for 4 weeks to control and hyperammonemic rats. The effects of bicuculline on microglia and astrocyte activation, IL-1β content, on membrane expression of AMPA and NMDA glutamate receptors subunits in the hippocampus and on spatial learning and memory as well as anxiety were assessed. Treatment with bicuculline reduces astrocyte activation and IL-1β but not microglia activation in the hippocampus of hyperammonemic rats. Bicuculline reverses the changes in membrane expression of AMPA receptor subunits GluA1 and GluA2 and of the NR2B (but not NR1 and NR2A) subunit of NMDA receptors. Bicuculline improves spatial learning and working memory and decreases anxiety in hyperammonemic rats. In hyperammonemia, enhanced activation of GABAA receptors in the hippocampus contributes to some but not all aspects of neuroinflammation, to altered glutamatergic neurotransmission and to impairment of spatial learning and memory as well as anxiety, all of which are reversed by reducing activation of GABAA receptors with bicuculline.

Involvement of NMDA-AKT-mTOR Signaling in Rapid Antidepressant-Like Activity of Chaihu-jia-Longgu-Muli-tang on Olfactory Bulbectomized Mice.

Background: Fast-onset antidepressants are urgently needed. Chaihu-jia-Longgu-Muli-tang (CLM), a classic Chinese herbal medicine, has been used for antidepressant treatment with long history. Olfactory bulbectomization (OB) model is validated for identification of rapid antidepressant efficacy. Here we used OB model for investigating the rapid onset activity of CLM in mice, and also tested the involvement of prefrontal Akt-mTOR and associated AMPA/NMDA receptors as well as hippocampal BDNF in the rapid antidepressant-like effect of CLM.Methods: The OB model was first characterized with depression-like behaviors and the time course changes of the behaviors. The fast onset of antidepressant effect of CLM was evaluated using sucrose preference test, tail suspension test and forced swim test in OB mice after a single administration. The expression of synaptic proteins of AMPA and NMDA subunits as well as Akt/mTOR signaling in the prefrontal cortex, and hippocampal BDNF was evaluated with the immunoblotting method.Results: A single dose of CLM significantly improved the deficiency in the sucrose preference and decreased the immobility time in the tail suspension test in OB mice. In the prefrontal cortex (PFC) in OB mice, there was lower expression level of the AMPA receptor subunit GluR1, rescued by a single dose of CLM. Additionally, the expression of NMDA subunit NR1 was up-regulated in OB mice, whereas mTOR and its upstream Akt signalings were both down-regulated. These deficiencies were reversed by a single dose of CLM. The CLM treatment also attenuated the expressions of NMDA receptor subunits NR2A and NR2B, which did not change in OB mice. In the hippocampus, expressions of GluR1 and brain derived neurotrophic factor (BDNF) were both up-regulated in OB mice, although CLM increased GluR1, but not BDNF.Conclusion: CLM elicited rapid antidepressant-like effects in the OB model mice, and CLM reversal of the abnormality in PFC expression of AMPA and NMDA receptors and associated Akt-mTOR signaling may underlie the effects.

Inhibition of CB1 receptor ameliorates spatial learning and memory impairment in mice with traumatic brain injury

Traumatic brain injury (TBI) is an increasingly prevalent condition affecting people of all ages and genders. The impairment of spatial learning and memory is one of the most common effects of TBI. Unfortunately, it currently lacks effective therapeutic interventions. The endocannabinoid (EC) system regulates a diverse array of physiological processes. Here, we found a 6.7-fold increase of 2-AG levels at 1 d post-TBI, declining thereafter. After 5 d, the levels were still 3.3-fold higher than in the controls. AM281, a CB1 receptor antagonist, reversed the TBI-reduced NMDA receptor subunits NR2B in the hippocampus and ameliorate the spatial learning and memory impairment at 7 d post-TBI, suggesting CB1 receptor is involved in the TBI-induced hippocampal-dependent spatial learning and memory impairment.

Prenatal propofol exposure downregulates NMDA receptor expression and causes cognitive and emotional disorders in rats

The impacts of prenatal propofol on cognition and emotion of offspring remain elusive. In the present study, pregnant rats in the second trimester were anesthetized with propofol. Neuronal apoptosis and proliferation was determined in fetuses and postnatal rats by detecting caspase-3 and BrdU expression. The offspring were subjected to several behavior tests. Then the pyramidal neurons in hippocampus and the expression of NR1, NR2A, and NR2B subunits of NMDA receptor and PSD-95 in frontal cortex were examined. Propofol exposure significantly increased the number of caspase-3+ cells in lateral ganglionic eminence and hippocampus compared with control group (P < 0.001). The number of BrdU+ cells in the subventricular zone (SVZ) and dentate gyrus (DG) was also reduced in propofol group (P < 0.001). In propofol group, the swimming distance and time in the quadrant were shorter,but longer out of the quadrant. The number of escapes was less than those of the control group. Prenatal propofol exposure also decreased the sucrose preference, reduced the numbers of grooming, crossing, and rearing of the pups, and increased the fecal particle numbers and immobility time (P < 0.05). There were fewer and shorter dendritic branches of pyramidal neurons in the CA1 and CA3 of offspring. The expression of NR1, NR2A, NR2B, and PSD-95 in the frontal cortex was downregulated by propofol exposure (P < 0.001). These data suggested that prenatal propofol exposure impairs neuronal development, which may be associated with depression- and anxiety-like behaviors and cognitive disorders in offspring.

EphrinB/EphB forward signaling in M\xfcller cells causes apoptosis of retinal ganglion cells by increasing tumor necrosis factor alpha production in rat experimental glaucomatous model

It was previously shown that EphB/ephrinB reverse signaling in retinal ganglion cells (RGCs) is activated and involved in RGC apoptosis in a rat chronic ocular hypertension (COH) model. In the present work, we first show that ephrinB/EphB forward signaling was activated in COH retinas, and RGC apoptosis in COH retinas was reduced by PP2, an inhibitor of ephrinB/EphB forward signaling. We further demonstrate that treatment of cultured Müller cells with ephrinB1-Fc, an EphB1 activator, or intravitreal injection of ephrinB1-Fc in normal rats induced an increase in phosphorylated EphB levels in these cells, indicating the activation of ephrinB/EphB forward signaling, similar to those in COH retinas. The ephrinB1-Fc treatment did not induce Müller cell gliosis, as evidenced by unchanged GFAP expression, but significantly up-regulated mRNA and protein levels of tumor necrosis factor-α (TNF-α) in Müller cells, thereby promoting RGC apoptosis. Production of TNF-α induced by the activation of ephrinB/EphB forward signaling was mediated by the NR2B subunit of NMDA receptors, which was followed by a distinct PI3K/Akt/NF-κB signaling pathway, as pharmacological interference of each step of this pathway caused a reduction of TNF-α production, thus attenuating RGC apoptosis. Functional analysis of forward and reverse signaling in such a unique system, in which ephrin and Eph exist respectively in a glial element and a neuronal element, is of theoretical importance. Moreover, our results also raise a possibility that suppression of ephrinB/EphB forward signaling may be a new strategy for ameliorating RGC apoptosis in glaucoma.

Changes in synaptic AMPA receptor concentration and composition in chronic temporal lobe epilepsy

Excitotoxicity caused by excessive stimulation of glutamate receptors, resulting in pathologically increased Ca2+-concentrations, is a decisive factor in neurodegenerative diseases. We investigated long-term changes in synaptic contents of AMPA receptor subunits that play important roles in calcium regulation in chronic epilepsy. Such plastic changes may be either adaptive or detrimental. We used a kainic acid (KA)-based rat model of chronic temporal lobe epilepsy (TLE). Using hippocampal synaptosomes, we found significant reductions in the concentration of the AMPA receptor subunits GluA1 and GluA2, and the NMDA receptor subunit NR2B. The relative size of GluA1 and GluA2 reductions were almost identical, at 28% and 27%, respectively. In order to determine whether the synaptic reduction of the AMPA receptor subunits actually reflected the pool of receptors present along the postsynaptic density (PSD), as opposed to cytoplasmic or extrasynaptic pools, we performed postembedding immunogold electron microscopy (EM) of GluA1 and GluA2 in Schaffer collateral synapses in the hippocampal CA1 area. We found significant reductions, at 32% and 52% of GluA1 and GluA2 subunits, respectively, along the PSD, indicating that these synapses undergo lasting changes in glutamatergic neurotransmission during chronic TLE. When compared to the overall concentration and composition of AMPA receptors expressed in the brain, there was a relative increase in GluA2-lacking AMPA receptor subunits following chronic epilepsy. These changes in synaptic AMPA receptor subunits may possibly contribute to further aggravate the excitotoxic vulnerability of the neurons as well as have significant implications for hippocampal cognitive functions.

Neuroprotective effect of zolpidem against glutamate-induced toxicity is mediated via the PI3K/Akt pathway and inhibited by PK11195

Excitotoxicity is a pathological process in which neuronal dysfunction and death are induced by excessive glutamate stimulation, the major fast excitatory neurotransmitter in the mammalian brain. Excitotoxicity-induced neurodegeneration is a contributing factor in ischemia-induced brain damage, traumatic brain injury, and various neurodegenerative diseases. It is triggered by calcium overload due to prolonged over-activation of ionotropic N-methyl-d-aspartate (NMDA) receptors. Enhanced Ca2+ release results in neuronal vulnerability through several intertwined mechanisms, including activation of proteolytic enzymes, increased production of reactive oxygen species (ROS), mitochondrial dysfunction and modulation of intracellular signalling pathways. We investigated the neuroprotective effect of hypnotic zolpidem, a drug that exerts its central effects at the GABAA receptor complex, against glutamate-induced toxicity in P19 neurons. Zolpidem prevented death of P19 neurons exposed to glutamate, and abolished the glutamate-induced increase in ROS production, p53 and Bax expression, and caspase-3/7 activity. Zolpidem effects were mediated by marked over-activation of Akt kinase. The pro-survival effect, as well as the pAkt induction, were prevented in the presence of wortmannin, an inhibitor of phosphatidylinositol-3-kinase (PI3K) that functions upstream of Akt. The beneficial effect of zolpidem on neuronal survival was not prevented by flumazenil, a GABAA receptor antagonist. PK11195, a drug that modulates the mitochondrial translocator protein 18 kDa (TSPO) and F0F1-ATPase, prevented the beneficial effect of zolpidem, indicating that the mechanism of zolpidem action involves preservation of mitochondrial function and integrity. Zolpidem effects were further mediated by prevention of glutamate-induced increase in the expression of the NR2B subunit of NMDA receptor. The obtained results suggest the promising therapeutic potential of zolpidem against excitotoxic insults and highlight the importance of mitochondria and the Akt pathway as valuable targets for therapeutic interventions in glutamate-mediated neuropathological conditions.

Docosahexaenoic Acid Helps to Lessen Extinction Memory in Rats.

Memory extinction is referred to as a learning process in which a conditioned response (CR) progressively reduces over time as an animal learns to uncouple a response from a stimulus. Extinction occurs when the rat is placed into a context without shock after training. Docosahexaenoic acid (DHA, C22:6, n-3) is implicated in memory formation in mammalian brains. In a two-way active shuttle-avoidance apparatus, we examined whether DHA affects the extinction memory and the expression of brain cognition-related proteins, including gastrin-releasing peptide receptor (GRPR), brain-derived neurotrophic factor receptor (BDNFR) tyrosine kinase receptor B (TrKB), and N-methyl-d-aspartate receptor (NMDAR) subunits NR2A and NR2B. Also, the protein levels of GRP, BDNF, postsynaptic density protein-95 (PSD-95), and vesicular acetylcholine transporter (VAChT), and the antioxidative potentials, in terms of lipid peroxide (LPO) and reactive oxygen species (ROS), were examined in the hippocampus. During the acquisition phase, the rats received a conditioned stimulus (CS-tone) paired with an unconditioned stimulus (UCS foot shock) for three consecutive days (Sessions S1, S2, and S3, each consisting of 30-trials) after 12 weeks of oral administration of DHA. After a three-day interval, the rats were re-subjected to two extinction sessions (S4, S5), each comprising 30 trials of CS alone. During the acquisition training in S1, the shock-related avoidance frequency (acquisition memory) was significantly higher in the DHA-administered rats compared with the control rats. The avoidance frequency, however, decreased with successive acquisition trainings in sessions S2 and S3. When the rats were subjected to the extinction sessions after a break for consolidation, the conditioned response (CR) was also significantly higher in the DHA-administered rats. Interestingly, the freezing responses (frequency and time) also significantly decreased in the DHA-administered rats, thus suggesting that a higher coping capacity was present during fear stress in the DHA-administered rats. DHA treatments increased the mRNA levels of GRPR, BDNF receptor TrKB, and NMDAR subunit NR2B. DHA also increased the protein levels of GRP, BDNF, PSD-95, and VAChT, and the antioxidative potentials in the hippocampus. These results suggest the usefulness of DHA for treating stress disorders.

Antagonism at the NR2B subunit of NMDA receptors induces increased connectivity of the prefrontal and subcortical regions regulating reward behavior

Evidence indicates that ketamine's rapid antidepressant efficacy likely results from its antagonism of NR2B-subunit-containing NMDA receptors (NMDAR). Since ketamine equally blocks NR2A- and NR2B-containing NMDAR, and has affinity to other receptors, NR2B-selective drugs might have improved therapeutic efficiency and side effect profile. We aimed to compare the effects of (S)-ketamine and two different types of NR2B-selective antagonists on functional brain networks in rats, in order to find common circuits, where their effects intersect, and that might explain their antidepressant action. The experimental design comprised four parallel groups of rats (N = 37), each receiving (S)-Ketamine, CP-101,606, Ro 25-6981 or saline. After compound injection, we acquired resting-state functional magnetic resonance imaging time series. We used graph theoretical approach to calculate brain network properties. Ketamine and CP-101,606 diminished the global clustering coefficient and small-worldness index. At the nodal level, all compounds induced increased connectivity of the regions mediating reward and cognitive aspects of emotional processing, such as ventromedial prefrontal cortex, septal nuclei, and nucleus accumbens. The dorsal hippocampus and regions involved in sensory processing and aversion, such as superior and inferior colliculi, exhibited an opposite effect. The effects common to ketamine and NR2B-selective compounds were localized to the same brain regions as those reported in depression, but in the opposite direction. The upregulation of the reward circuitry might partially underlie the antidepressant and anti-anhedonic effects of the antagonists and could potentially serve as a translational imaging phenotype for testing putative antidepressants, especially those targeting the NR2B receptor subtype.

Plantar mechanical stimulation prevents neurochemical alterations in the hippocampus induced by stimulated microgravity

Plantar mechanical stimulation prevents neurochemical alterations in the hippocampus induced by stimulated microgravity

Differential Modulation of GABAA and NMDA Receptors by an α7-nicotinic Acetylcholine Receptor Agonist in Chronic Glaucoma.

Presynaptic modulation of γ-aminobutyric acid (GABA) release by an alpha7 nicotinic acetylcholine receptor (α7-nAChR) agonist promotes retinal ganglion cell (RGC) survival and function, as suggested by a previous study on a chronic glaucomatous model from our laboratory. However, the role of excitatory and inhibitory amino acid receptors and their interaction with α7-nAChR in physiological and glaucomatous events remains unknown. In this study, we investigated GABAA and N-methyl-D-aspartate (NMDA) receptor activity in control and glaucomatous retinal slices and the regulation of amino acid receptor expression and function by α7-nAChR. Whole-cell patch-clamp recordings from RGCs revealed that the α7-nAChR specific agonist PNU-282987 enhanced the amplitude of currents elicited by GABA and reduced the amplitude of currents elicited by NMDA. The positive modulation of GABAA receptor and the negative modulation of NMDA receptor (NMDAR) by PNU-282987-evoked were prevented by pre-administration of the α7-nAChR antagonist methyllycaconitine (MLA). The frequency and the amplitude of glutamate receptor-mediated miniature glutamatergic excitatory postsynaptic currents (mEPSCs) were not significantly different between the control and glaucomatous RGCs. Additionally, PNU-282987-treated slices showed no alteration in the frequency or amplitude of mEPSCs relative to control RGCs. Moreover, we showed that expression of the α1 subunit of the GABAA receptor was downregulated and the expression of the NMDAR NR2B subunit was upregulated by intraocular pressure (IOP) elevation, and the changes of high IOP were blocked by PNU-282987. In conclusion, retina GABAA and NMDARs are modulated positively and negatively, respectively, by activation of α7-nAChR in in vivo chronic glaucomatous models.

Developmental Exposure to Pesticides Alters Motor Activity and Coordination in Rats: Sex Differences and Underlying Mechanisms.

It has been proposed that developmental exposure to pesticides contributes to increasing prevalence of neurodevelopmental disorders in children, such as attention deficit with hyperactivity (ADHD) and to alterations in coordination skills. However, the mechanisms involved in these alterations remain unclear. We analyzed the effects on spontaneous motor activity and motor coordination of developmental exposure to a representative pesticide of each one of the four main chemical families: organophosphates (chlorpyrifos), carbamates (carbaryl), organochlorines (endosulfan), and pyrethroids (cypermethrin). Pesticides were administered once a day orally, in a sweet jelly, from gestational day 7 to post natal day 21. Spontaneous motor activity was assessed by an actimeter and motor coordination using the rotarod, when rats were adults. The effects were analyzed separately in males and females. Extracellular GABA in cerebellum and NMDA receptor subunits in hippocampus were assessed as possible underlying mechanisms of motor alterations. Motor coordination was impaired by developmental exposure to endosulfan, cypermethrin, and chlorpyrifos in females but not in males. The effect of endosulfan and cypermethrin would be due to increased extracellular GABA in cerebellum, which remains unaltered in male rats. Chlorpyrifos increased motor activity in males and females. Cypermethrin decreased motor activity mainly in males. In male rats, but not in females, expression of the NR2B subunit of NMDA receptor in hippocampus correlated with motor activity. These results show sex-specific effects of different pesticides on motor activity and coordination, associated with neurotransmission alterations. These data contribute to better understand the relationship between developmental exposure to the main pesticide families and motor disorders in children.

Effects of Immune Activation during Early or Late Gestation on N-Methyl-d-Aspartate Receptor Measures in Adult Rat Offspring.

Glutamatergic receptor [N-methyl-d-aspartate receptor (NMDAR)] alterations within cortex, hippocampus, and striatum are linked to schizophrenia pathology. Maternal immune activation (MIA) is an environmental risk factor for the development of schizophrenia in offspring. In rodents, gestational timing of MIA may result in distinct behavioral outcomes in adulthood, but how timing of MIA may impact the nature and extent of NMDAR-related changes in brain is not known. We hypothesize that NMDAR-related molecular changes in rat cortex, striatum, and hippocampus are induced by MIA and are dependent on the timing of gestational inflammation and sex of the offspring. Wistar dams were treated the with viral mimic, polyriboinosinic:polyribocytidylic acid (polyI:C), or vehicle on either gestational day 10 or 19. Fresh-frozen coronal brain sections were collected from offspring between postnatal day 63-91. Autoradiographic binding was used to infer levels of the NMDAR channel, and NR2A and NR2B subunits in cortex [cingulate (Cg), motor, auditory], hippocampus (dentate gyrus, cornu ammonis area 3, cornu ammonis area 1), and striatum [dorsal striatum, nucleus accumbens core, and nucleus accumbens shell (AS)]. NR1 and NR2A mRNA levels were measured by in situ hybridization in cortex, hippocampus, and striatum in male offspring only. In the total sample, NMDAR channel binding was elevated in the Cg of polyI:C offspring. NR2A binding was elevated, while NR2B binding was unchanged, in all brain regions of polyI:C offspring overall. Male, but not female, polyI:C offspring exhibited increased NMDAR channel and NR2A binding in the striatum overall, and increased NR2A binding in the cortex overall. Male polyI:C offspring exhibited increased NR1 mRNA in the AS, and increased NR2A mRNA in cortex and subregions of the hippocampus. MIA may alter glutamatergic signaling in cortical and hippocampal regions via alterations in NMDAR indices; however, this was independent of gestational timing. Male MIA offspring have exaggerated changes in NMDAR compared to females in both the cortex and striatum. The MIA-induced increase in NR2A may decrease brain plasticity and contribute to the exacerbated behavioral changes reported in males and indicate that the brains of male offspring are more susceptible to long-lasting changes in glutamate neurotransmission induced by developmental inflammation.

Pre-existing weakness is critical for the occurrence of postoperative cognitive dysfunction in mice of the same age.

Occurrence of postoperative cognitive dysfunction (POCD) is age-dependent and heterogenous. Factors deciding the occurrence of POCD in patients of the same age undergone same surgeries remain unclear. Here we investigated the effects of pre-existing weakness on the occurrence of POCD in mice of the same age. Pre-existing weakness of mice was induced by intraperitoneal injection of lipopolysaccharide (8mg/kg) and was evaluated by physical frailty index (by open field test), neuroinflammation level (by Iba1 immunostaining and inflammatory factors TNF-α and IL-1β), and neuronal activity (by p-CREB immunostaining). POCD was induced by partial hepatolobectomy and was evaluated by puzzle box test and Morris water maze test. The brains were collected to detect the levels of neuroinflammation, synaptophysin and NMDA receptor subunits NR2A, NR2B and NR1 (by western blot), and oxidative stress (by Dihydroethidium). Compared to the normal adult mice of the same age, LPS pretreated mice had increased physical frailty index, higher levels of neuroinflammation, and lower neuronal activity. Partial hepatolobectomy induced obvious impairments in executive function, learning and memory in LPS pretreated mice after surgery, but not in normal mice of the same age. Partial hepatolobectomy also induced heightened neuroinflammation, obvious loss of NMDA receptor subunits, strong oxidative stress in LPS pretreated mice on the 1st and 3rd postoperative day. However, the POCD-associated pathological changes didn’t occur in normal mice of the same age after surgery. These results suggest that pre-existing weakness is critical for the occurrence of POCD in mice of the same age.

Effects of ERK1/2 kinases inactivation on the nigrostriatal system of Krushinsky-Molodkina rats genetically prone to audiogenic seizures

Recently, we demonstrated that inhibition of ERK1/2 activity by SL-327 treatment blocks seizure behavior in Krushinsky-Molodkina (KM) rats, which was mediated by altering of GABA and glutamate release mechanism in the hippocampus. Basal ganglia representing various subcortical cell groups play a significant role in the regulation of motor activity, including epileptiform seizures.Objectives: To verify if nigrostriatal system could be also affected by SL-327 treatment we analyzed the expression of tyrosine hydroxylase, D1 and D2 dopamine receptors, NR2B subunit of NMDA receptor as well as vesicular glutamate transporter VGLUT2 and glutamic acid decarboxylases GAD65/67 in the striatum and substantia nigra of KM rats.Methods: Animals were injected i.p. with SL-327 (50 mg/kg) 60 min before audio stimulation. After audiogenic stimulation the brains of control and SL 327 treated rats were removed for further immunohistochemical and biochemical analysis.Results: Obtained results demonstrated a decrease activity in synapsin I, and accumulation of VGLUT2 in the striatum after blockade of audiogenic seizure (AGS) by SL 327 that could lead to inhibition of glutamate release. While in the striatum GAD65/67 level was diminished, in the substantia nigra GAD65/67 was increased showing enhanced inhibitory output to the compact part of the substantia nigra. Analysis of dopaminergic system showed a significant reduction of tyrosine hydroxylase activity and expression in the substantia nigra, and decreased D1 and D2 receptor expression in the striatum.In summary, we propose that changes in the nigrostriatal system could be mediated by inhibitory effect of SL 327 on AGS expression.

Anti-androgenic effects of bisphenol-A on spatial memory and synaptic plasticity of the hippocampus in mice

Bisphenol-A (BPA) is a common environmental endocrine disruptor. Our recent studies found that exposure to BPA in both adolescent and adulthood sex-specifically impaired spatial memory in male mice. In this study, 11-week-old gonadectomied (GDX) male mice daily received subcutaneous injections of testosterone propionate (TP, 0.5mg/kg), TP and BPA (0.4 and 4mg/kg), or vehicle for 45days. The results of Morris water maze task showed that exposure to BPA did not affect the spatial memory of GDX mice but impaired that of sham (4mg/kg/day) and TP-treated GDX mice (0.4mg/kg/day). In addition, BPA reduced the level of testosterone (T) in the serum and brain of sham and TP-treated GDX mice. Exposure to BPA decreased the synaptic density and had an adverse effect on the synaptic interface of the hippocampus in sham and TP-treated GDX mice. The results of western blot analysis further showed that BPA (4mg/kg) reduced the levels of synaptic proteins (synapsin I and PSD-95) and NMDA receptor subunit NR2B in sham and TP-treated GDX mice. BPA decreased the phosphorylation of ERK1/2 but increased the phosphorylation of p38 in sham and TP-treated GDX mice. These results suggest that impairment of spatial memory and adverse effects on synaptic remodeling of hippocampal neurons in males after long-term BPA exposure is related to the anti-androgen effect of BPA. These effects of BPA may be associated with downregulated synaptic proteins and NMDA receptor through inhibiting ERKs and promoting the p38 pathways.

Myeloid differentiation factor 88 is up-regulated in epileptic brain and contributes to experimental seizures in rats

Accumulating evidence supports that activation of inflammatory pathways is a crucial factor contributing to the pathogenesis of seizures. In particular, the activation of interleukin-1 beta (IL-1β) system exerts proconvulsant effects in a large variety of seizure models. Myeloid differentiation factor 88 (MyD88) is a critical adaptor protein in the signaling cascade elicited by IL-1β. The present study aimed to investigate the expression pattern of MyD88 in rat models of seizures and in patients with refractory temporal lobe epilepsy (TLE), and to study the role of MyD88 in epileptic seizures. Our results revealed that MyD88 was up-regulated in the hippocampus of rats in the lithium-pilocarpine model of acute seizures. Importantly, MyD88 overexpression was also significantly present in the brain from chronic epileptic rats and the temporal neocortex specimens from drug-resistant TLE patients. In the acute seizure model, both the behavioral and electrographic seizure activities were record and analyzed in rats for 90min, starting immediately after pilocarpine injection. ST2825, a synthetic MyD88 inhibitor, was administered intracerebroventricularly (2.5–5.0–10μg in 2μl) 20min before pilocarpine injection. We found that ST2825 at doses of 5μg and 10μg significantly inhibited the pilocarpine-induced behavioral and electrographic seizures. Moreover, 10μg ST2825 prevented the proconvulsant actions of IL-1β. As previous evidence suggested that IL-1β proconvulsant effects was mediated by enhancing the phosphorylation level of the NR2B subunit of N-methyl-d-aspartate (NMDA) receptor, we then probed whether this molecular was involved in the effect of the pharmacological inhibition. Our results revealed that 10μg ST2825 markedly reversed the increased Tyr1472-phosphorylation of the NR2B subunit of NMDA receptor observed in the proconvulsant conditions of IL-1β and in seizures induced by pilocarpine alone. These findings indicate that altered expression of MyD88 might contribute to the pathogenesis of seizures and targeting of this adaptor protein might represent a novel therapeutic strategy to suppress seizure activities.

NMDA receptor subunit composition controls dendritogenesis of hippocampal neurons through CAMKII, CREB-P, and H3K27ac.

Dendrite arbor growth, or dendritogenesis, is choreographed by a diverse set of cues, including the NMDA receptor (NMDAR) subunits NR2A and NR2B. While NR1NR2B receptors are predominantly expressed in immature neurons and promote plasticity, NR1NR2A receptors are mainly expressed in mature neurons and induce circuit stability. How the different subunits regulate these processes is unclear, but this is likely related to the presence of their distinct C-terminal sequences that couple different signaling proteins. Calcium-calmodulin-dependent protein kinase II (CaMKII) is an interesting candidate as this protein can be activated by calcium influx through NMDARs. CaMKII triggers a series of biochemical signaling cascades, involving the phosphorylation of diverse targets. Among them, the activation of cAMP response element-binding protein (CREB-P) pathway triggers a plasticity-specific transcriptional program through unknown epigenetic mechanisms. Here, we found that dendritogenesis in hippocampal neurons is impaired by several well-characterized constructs (i.e., NR2B-RS/QD) and peptides (i.e., tatCN21) that specifically interfere with the recruitment and interaction of CaMKII with the NR2B C-terminal domain. Interestingly, we found that transduction of NR2AΔIN, a mutant NR2A construct with increased interaction to CaMKII, reactivates dendritogenesis in mature hippocampal neurons in vitro and in vivo. To gain insights into the signaling and epigenetic mechanisms underlying NMDAR-mediated dendritogenesis, we used immunofluorescence staining to detect CREB-P and acetylated lysine 27 of histone H3 (H3K27ac), an activation-associated histone tail mark. In contrast to control mature neurons, our data shows that activation of the NMDAR/CaMKII/ERK-P/CREB-P signaling axis in neurons expressing NR2AΔIN is not correlated with increased nuclear H3K27ac levels.

Prenatal exposure to lambda-cyhalothrin impairs memory in developing rats: Role of NMDA receptor induced post-synaptic signalling in hippocampus

Effect of prenatal exposure to lambda-cyhalothrin (LCT) has been assessed on the integrity of NMDA receptors and associated post-synaptic signalling in hippocampus of developing rats. Decrease in the binding of [3H]-MK 801, known to label NMDA receptors was observed in hippocampus of rats prenatally exposed to LCT (1 and 3mg/kg body weight) on PD22, compared to controls. Consistent with this, decrease in the mRNA and protein expression of NR1 and NR2B subunits of NMDA receptors was evident in rats prenatally exposed to LCT (1 and 3mg/kg body weight) on PD22. There was no change in mRNA and protein expression of NR2A subunit of NMDA receptors. Prenatal exposure to LCT (1 and 3mg/kg body weight) decreased the expression of positive regulators (PSD95, pERK1/2, CaMKIIα & pCREB) and increased the expression of negative regulators (Cdk5 & SynGAP) associated with NMDA receptor dependent synaptic plasticity in hippocampus and impaired learning and memory of rats on PD22. The neurobehavioral changes continued to persist in rats exposed to LCT at high dose (3mg/kg body weight) while exhibited trend of recovery in those exposed at moderate dose (1mg/kg body weight) on PD45, compared to controls. No change in any of the neurobehavioral endpoint was observed in developing rats prenatally exposed to LCT at low dose (0.5mg/kg body weight) on PD22 and PD45. The results exhibit that alterations in NMDA receptors on prenatal exposure to LCT may affect postsynaptic signalling associated with impaired learning and memory in developing rats.

Electroacupuncture restores spatial learning and downregulates phosphorylated N-methyl-D-aspartate receptors in a mouse model of Parkinson's disease.

Parkinson's disease (PD) is a degenerative disorder of the central nervous system. PD can be classified as idiopathic, acquired or hereditary and may be caused by various factors such as oxidative stress, loss of mitochondrial function, neuronal excitotoxicity or calcium imbalance. We hypothesised that electroacupuncture (EA) at KI3 would reduce neuronal excitotoxicity by regulating N-methyl-D-aspartate (NMDA) receptor function and may represent a novel therapeutic approach for PD. Our results showed that deficits in spatial learning (reflected by the escape latency time in the Morris water maze task) and long-term potentiation (LTP) caused by systemic 6-hydroxydopamine (6-OHDA) administration (that damages dopaminergic neurons) could be rescued by EA on day 3. In PD mice, phosphorylated NMDA receptor subunits NR1 and NR2B were elevated (134.03±10.17% and 123.46±3.47% of baseline levels, respectively) but total NR1 and NR2B was unaffected (101.37±3.87% and 102.61±4.22% of baseline, respectively). Elevated levels of pNR1 and pNR2B, and phosphorylated forms of protein kinase A, protein kinase C, α Ca2+/calmodulin-dependent protein kinase extracellular signal-regulated kinases (pERK), and cAMP response element-binding protein were also reduced following EA. These novel findings suggest that EA can rescue learning and LTP deficits in a rodent model of PD. The results point to a possible role for EA-based approaches in the clinical treatment of learning deficits associated with PD.