Receptor NR1 Research Articles

Association of anti-NR2 and U1RNP antibodies with neurotoxic inflammatory mediators in cerebrospinal fluid from patients with neuropsychiatric systemic lupus erythematosus

Autoantibodies (auto Abs) and inflammatory mediators (IMs) in cerebrospinal fluid (CSF) may be involved in the pathogenesis of neuropsychiatric systemic lupus erythematosus (NPSLE). It is suggested that anti-N-methyl D-aspartate receptor NR2 subunit (NR2) Ab can develop NP manifestation after blood-brain barrier (BBB) abruption. We also reported the association between NPSLE and CSF anti-U1RNP Ab. In the present study, combined effects of CSF anti-NR2 and anti-U1RNP Abs on IMs in patients with NPSLE were examined. CSF samples were collected from 69 patients with NPSLE and 13 non-NPSLE controls. CSF anti-NR2 and anti-U1RNP Abs were determined using ELISA. Levels of IL-6, IL-8, and monokine induced by IFN-γ (MIG) in CSF were measured by quantitative multiplex cytokine analysis. CSF IL-6 levels were higher in CSF anti-NR2-positive than in CSF anti-NR2-negative patients (p = 0.003) and non-NPSLE controls (p = 0.015) and were positively correlated with anti-NR2 titer (r = 0.42). CSF IL-8 levels were higher in CSF anti-U1RNP-positive than in CSF anti-U1RNP-negative patients (p = 0.041). CSF MIG levels were more elevated in CSF anti-NR2-positive (p = 0.043) and anti-U1RNP-positive patients (p = 0.029) than in non-NPSLE controls. Additionally, in double positive (DP; both anti-NR2 and U1RNP Ab positive) group, CSF IL-6 and MIG levels were significantly higher than in the double negative (DN; both anti-NR2 and U1RNP Ab negative) group. However, combined effect of both Abs on IM elevation and clinical manifestation was not clear. CSF anti-NR2 and anti-U1RNP Abs have different effects on the elevation of CSF IM levels in patients with NPSLE. Additional effect of anti-U1RNP Abs on anti-NR2 Ab-mediated NP manifestation, however, was not recognized in our study.

Tyrosine Kinase Inhibitors Reduce NMDA NR1 Subunit Expression, Nuclear Translocation, and Behavioral Pain Measures in Experimental Arthritis.

In the lumbar spinal cord dorsal horn, release of afferent nerve glutamate activates the neurons that relay information about injury pain. Here, we examined the effects of protein tyrosine kinase (PTK) inhibition on NMDA receptor NR1 subunit protein expression and subcellular localization in an acute experimental arthritis model. PTK inhibitors genistein and lavendustin A reduced cellular histological translocation of NMDA NR1 in the spinal cord occurring after the inflammatory insult and the nociceptive behavioral responses to heat. The PTK inhibitors were administered into lumbar spinal cord by microdialysis, and secondary heat hyperalgesia was determined using the Hargreaves test. NMDA NR1 cellular protein expression and nuclear translocation were determined by immunocytochemical localization with light and electron microscopy, as well as with Western blot analysis utilizing both C- and N-terminal antibodies. Genistein and lavendustin A (but not inactive lavendustin B or diadzein) effectively reduced (i) pain related behavior, (ii) NMDA NR1 subunit expression increases in spinal cord, and (iii) the shift of NR1 from a cell membrane to a nuclear localization. Genistein pre-treatment reduced these events that occur in vivo within 4 h after inflammatory insult to the knee joint with kaolin and carrageenan (k/c). Cycloheximide reduced glutamate activated upregulation of NR1 content confirming synthesis of new protein in response to the inflammatory insult. In addition to this in vivo data, genistein or staurosporin inhibited upregulation of NMDA NR1 protein and nuclear translocation in vitro after treatment of human neuroblastoma clonal cell cultures (SH-SY5Y) with glutamate or NMDA (4 h). These studies provide evidence that inflammatory activation of peripheral nerves initiates increase in NMDA NR1 in the spinal cord coincident with development of pain related behaviors through glutamate non-receptor, PTK dependent cascades.

Age-related shifts in gut microbiota contribute to cognitive decline in aged rats.

Cognitive function declines during the aging process, meanwhile, gut microbiota of the elderly changed significantly. Although previous studies have reported the effect of gut microbiota on learning and memory, all the reports were based on various artificial interventions to change the gut microbiota without involvement of aging biological characteristics. Here, we investigated the effect of aged gut microbiota on cognitive function by using fecal microbiota transplantation (FMT) from aged to young rats. Results showed that FMT impaired cognitive behavior in young recipient rats; decreased the regional homogeneity in medial prefrontal cortex and hippocampus; changed synaptic structures and decreased dendritic spines; reduced expression of brain-derived neurotrophic factor (BDNF), N-methyl-D-aspartate receptor NR1 subunit, and synaptophysin; increased expression of advanced glycation end products (AGEs) and receptor for AGEs (RAGE). All these behavioral, brain structural and functional alterations induced by FMT reflected cognitive decline. In addition, FMT increased levels of pro-inflammatory cytokines and oxidative stress in young rats, indicating that inflammation and oxidative stress may underlie gut-related cognitive decline in aging. This study provides direct evidence for the contribution of gut microbiota to the cognitive decline during normal aging and suggests that restoring microbiota homeostasis in the elderly may improve cognitive function.

Modulation of NR1 receptor by CaMKIIα plays an important role in chronic itch development in mice

Intractable scratching is the characteristic of chronic itch, which represents a great challenge in clinical practice. However, the mechanism underlying chronic itch development is largely unknown. In the present study, we investigated the role of NMDA receptor in acute itch and in development of chronic itch. A mouse model was developed by painting DNFB to induce allergic contact dermatitis (ACD). We found that the expression of pNR1, which is a subunit of NMDA receptor, was significantly increased in the dorsal root ganglion in the DNFB model. The DNFB-evoked spontaneous scratching was blocked by the NMDA antagonist D-AP-5, the calcium-calmodulin-dependent protein kinase (CaMK) inhibitor KN-93, a CaMKIIα siRNA and the PKC inhibitor LY317615. Moreover, activation of PKC did not reverse the CaMKIIα knockdown-induced decrease in scratching, suggesting that PKC functions upstream of CaMKIIα. Thus, our study indicates that modulation of NR1 receptor by CaMKIIα plays an important role in the development of chronic itch.

Muscone reduced the hypnotic and analgesic effect of ketamine in mice.

The aim of this study was to determine the effects of different concentrations of muscone on the ketamine requirement for hypnosis and analgesia and possible mechanism in mice. In the hypnotic response experiment, muscone (0.5, 1.0, 2.0, 4.0, and 8.0 mg/kg) was administered 15 minutes before ketamine by intraperitoneal injection. The hypnotic response was evaluated by loss of righting reflex (LORR). In the analgesia experiment, muscone (0.5, 1.0, 2.0, and 4.0 mg/kg) was administered 15 minutes before 50 mg/kg ketamine injection. Pain threshold was assessed by measuring the tail-flick latency induced by heat radiation. Twenty minutes after ketamine injection, the mRNA expression of N-methyl-D-aspartate receptors (NR) subunits, γ-aminobutyric acid (GABA) receptors subunits, opioid receptors subunits, and some Na and Ca channels were detected by qPCR in the hippocampus of mice. The 50% effective dose (ED50) with 95% confidence interval of ketamine-induced LORR was 49.2 (43.4-56.4) mg/kg. About 4.0 or 8.0 mg/kg muscone increased ED50 of ketamine-induced hypnosis, which was 82.7 (70.0-98.4) mg/kg or 72.0 (65.4-85.7) mg/kg, respectively. In the analgesic experiment, ketamine alone caused an obvious analgesic effect, whereas different dose of muscone decreased pain threshold in the presence of ketamine; 4.0 mg/kg muscone up-regulated the mRNA expression of NR1 and inhibited ketamine-induced increase of δ-opioid receptor mRNA level. Muscone also inhibited Cav2.1 mRNA expression in the presence of ketamine. Muscone reduced the hypnotic and analgesic effect of ketamine in dose-independent manner in mice, which may be related to the changes of NR1 and δ-opioid receptor.

個性別学習能力差とラット脳海馬NR1受容体の左右非対称性の検討

個性別学習能力差とラット脳海馬NR1受容体の左右非対称性の検討

Testosterone modulates structural synaptic plasticity of primary cultured hippocampal neurons through ERK - CREB signalling pathways

Although hippocampus-derived androgens play an important role in hippocampal synaptic plasticity, studies at the cellular level have received relatively less attention. Furthermore, the underlying signalling pathways associated with synaptic plasticity remain unclear. Results of the present study demonstrated that testosterone treatment of primary cultured rat hippocampal neurons resulted in a rapid increase in spine density, accompanied by the elevation of protein and messenger RNA levels of synaptophysin, developmentally regulated brain protein (Drebrin), and the N-methyl-D-aspartate receptor NR1 subunit. Testosterone treatment also increased the phosphorylation levels of extracellular-regulated protein kinase (ERK)1/2 and cAMP-responsive element binding protein (CREB), rather than p38 and Jun N-terminal kinase (JNK). U0126 significantly reversed the testosterone-mediated phosphorylation of CREB. Importantly, the increase in spine density was not induced by testosterone under U0126 treatment. These findings suggest that the ERK1/2-CREB signalling pathway plays an important role in testosterone-mediated rapid spinogenesis of cultured rat hippocampal neurons. Results of this study will be helpful in further clarifying the physiological function of testosterone and related signalling pathways in vitro.

Nitric Oxide and Cyclic Guanosine Monophosphate Signaling Mediates the Antidepressant Effects of Acupuncture in the Rat Model of Chronic Unpredictable Mild Stress.

BackgroundDepression is a major mood disorder. Some patients have been reported to improve following acupuncture. This study aimed to investigate the effects of acupuncture on behaviors associated with depression in the chronic unpredictable mild stress (CUMS) rat model. The expression of signaling pathway components of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) in the rat hippocampus and plasma were also measured.Material/MethodsMale Sprague-Dawley rats (N=40) were divided into the control group (N=10), the model group (N=10), the acupuncture group (N=10), and the non-acupuncture group (N=10). The rat model was established by orphaning combined with chronic unpredictable mild stress (CUMS) for six weeks. The acupuncture group was given 21 days of treatment using acupoints (AP) or non-acupoints (NP). Rat behaviors associated with depression were tested using the sucrose preference test (SPT), the open field test (OFT), and the elevated plus maze (EPM) test. Enzyme-linked immunoassay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) were used to detect the expression of inducible nitric oxide synthase (iNOS), neuronal nitric oxide synthase (nNOS), and the N-methyl-D-aspartate (NMDA) receptor subunits, NR1, NR2A, and NR2B in the rat plasma and hippocampus.ResultsAcupuncture reversed the behaviors associated with depression in the CUMS rat model and reduced the expression of components of the NO and cGMP pathway in the rat hippocampus and plasma.ConclusionsIn the CUMS rat model, treatment with acupuncture reduced behaviors associated with depression, and these effects were associated with changes in the NO and cGMP signaling pathway.

Effect of Lactobacillus casei on the depressive behavior and the expression of NR1 and NR2A receptors in hippocampus of rats with postpartum depression induced by maternal separation

Objective To investigate the effect of Lactobacillus casei on depressive behavior and the expression of NR1, NR2A receptor in postpartum depression rats induced by maternal separation. Methods Twenty-four Sprague-Dawley female rats were randomly divided into control group(CON group) (n=8), postpartum depression group(PPD group) (n=8), postpartum depression with lactobacillus casei group(PPD+ Lactobacillus casei group) (n=8). Rats of the control group were given no interventions.Rats in PPD group and PPD+ Lactobacillus casei group were given maternal separation to establish depression model.And then the rats in PPD+ Lactobacillus casei group were treated with Lactobacillus casei(8×108CFU/(kg·d)) for 4 weeks after 14 days of maternal separation.The forced swimming test (FST) was employed to detect the depressive behaviors.Lactobacillus, Bifidobacterium, Enterococcus faecalis and Escherichia coli in cecum of rats and the expression of NR1 and NR2A mRNA in hippocampus of rats were detected by real-time fluorescence quantitative PCR. Results Compared with CON group ((157.50±8.13) s), the immobility time of PPD group((200.00±10.35) s) was significantly longer (t=-3.23, P<0.05). Compared with PPD group, the immobility time of PPD+ Lactobacillus casei group ((153.25±7.41) s) was significantly shortened (t=3.67, P<0.05), and the depressive behavior was improved.Compared with CON group, the mRNA expression of Lactobacillus ((1.47±0.08), t=-5.87, P<0.01), Enterococcus faecalis ((1.23±0.08), t=-2.85, P<0.05) and Escherichia coli( (1.33±0.07), t=-4.96, P<0.01) in caecum were significantly increased in PPD group, while that of Bifidobacterium decreased significantly((0.65±0.07), t=5.18, P<0.01). Compared with PPD group, the mRNA expression of Lactobacillus( (1.05±0.05), t=3.67, P<0.01), Enterococcus faecalis ((0.97±0.05), t=2.74, P<0.05) and Escherichia coli ((1.06±0.05), t=3.31, P<0.01) were significantly decreased in PPD+ Lactobacillus casei group, while that of Bifidobacterium increased significantly ((0.98±0.04), t=-4.26, P<0.01). Compared with CON group, the mRNA expression of NR1 receptor ((0.57±0.04), t=9.65, P<0.01) and NR2A receptor ((0.60±0.05), t=8.64, P<0.05) in hippocampus of rats in PPD group were significantly decreased.Compared with PPD group, the expression of NR1 receptor ((1.01±0.05), t=-5.39, P<0.01) and NR2A receptor ((0.98±0.07), t=-3.91, P<0.05) in hippocampus were increased significantly in PPD+ Lactobacillus casei group. Conclusion Lactobacillus casei can improve the depressive behavior of postpartum depression in rats, and improve the intestinal flora, which affect the expression of NR1 and NR2A in hippocampus. Key words: Postpartum depression; Hippocampus; Intestinal flora; Lactobacillus casei; NR1 receptor; NR2A receptor; Rat

Molecular Modeling of the antagonist compound esketamine and its molecular docking study with non-competitive N-methyl-D-aspartate (NMDA) receptors NR1, NR2A, NR2B and NR2D

The main purpose of the present article is reactivity and stability properties study of the antagonist compound esketamine and analyzing of its binding to the non-competitive N-methyl-D-aspartate receptor subunits (NR1, NR2A, NR2B and NR2D). In first step, the molecular structure of esketamine was optimized using density functional theory (DFT) method at B3YP/6-311++G(d,p) level of theory. The reactivity and stability properties of the title medicinal compound were studied by global reactivity indices. The computational data showed the molecule is stable and has low tendency to interact with residues of the biomolecules like receptors and proteins. Secondly, the molecule binding to the receptors were analyzed by molegro virtual docker (MVD) program. Our computations indicated that the compound asserts its pharmacological effects mainly through interactions with NR2B receptors and the NR2B residues containing Gly [A] 128, His [A] 127, Gly [A] 264, Tyr [A] 282, Ser [A] 131, Asp [A] 265, Ser [A] 260 and Met [A] 132 are the main amino acids involved in the ligand-receptor complex formation.

Neurotoxicity Biomarkers as Potential Indicators of Spinal Cord Ischemia

Objectives. To study the concentration of antibodies to glutamate receptors in the serum and CSF of patients with vascular myelopathy in parallel with conventional diagnostic approaches. Materials and methods. The study group included 40 patients with vascular myelopathy: 10 patients with spinal stroke and 30 with subacute and chronic myelopathy. After application of exclusion criteria, the study group consisted of 27 patients. The reference group consisted of 30 patients with acute ischemic stroke and 30 with spondylogenic radiculopathy. The control group consisted of 15 healthy subjects. Patients underwent neurological investigations and MRI scans of the spine and spinal cord. Concentrations of antibodies to glutamate receptors (NMDA receptor NR2 fragments and AMPA/kainate receptors) were assessed by immunoenzyme analysis. Results and conclusions. Increases in titers of antibodies to NMDA receptor NR2 fragments were seen in the serum (p = 0.0001) and CSF (p = 0.0005) of patients with spinal cord ischemia as compared with the reference and control groups. Data on NR2 antibodies reliably detected myeloischemia as compared with analysis of antibodies to AMPA/kainate receptors and S100β protein. On the other hand, increased levels of antibodies to AMPA/kainate receptors were seen in patients with more severe disease, with more extensive lesions to the white matter. The authors take the view that determination of antibodies to glutamate receptors can be used in the diagnosis of vascular myelopathy and in the identification of disease severity.

Effect of Sedentary Conditions on the Rostrocaudal Expression of the NMDA NR1 Receptor Subunit in Bulbospinal C1 and non‐C1 Neurons of the Rat Rostral Ventrolateral Medulla (RVLM)

Bulbospinal RVLM neurons directly innervate sympathetic preganglionic cells and therefore play important roles in the regulation of sympathetic nerve activity and blood pressure. RVLM neurons are divided phenotypically into catecholaminergic (C1) and non‐catecholaminergic (non‐C1).Previous studies from our laboratory reported that the RVLM of sedentary compared to physically active rats demonstrates greater structural and functional neuroplasticity in a subregion‐specific manner. For example, RVLM neurons from sedentary versus physically active rats exhibit greater dendritic branching and elicit greater sympathoexcitation upon direct stimulation with glutamate in rostral versus caudal RVLM. Bulbospinal RVLM neurons also express NMDA receptors and could contribute to inactivity‐induced neuroplasticity and enhanced sympathetic activation.The purpose of the present study was to test the hypothesis that sedentary conditions increase NMDA NR1 subunit expression in the RVLM in a subregion‐dependent manner. Male Sprague‐Dawley rats were sedentary (no running wheels) or physically active (free access to running wheels) for 10–12 wks (n=6 ea). Spinal cord injections of cholera toxin B (CTB)‐labeled bulbospinal RVLM neurons. We examined coronal sections for triple‐label immunofluorescence by laser confocal microscopy, and quantified CTB‐positive C1 and non‐C1 neurons expressing the NR1 receptor subunit.As expected from previous work, significantly more bulbospinal C1 neurons were positive for NR1 in the caudal RVLM compared to rostral RVLM in both experimental groups (p&lt;0.001). Also as expected, more non‐C1 neurons immunoreactive for NR1 were in the rostral compared to caudal RVLM (p&lt;0.001). Interestingly, sedentary rats demonstrated a strong trend for a higher percentage of C1 cells and significantly lower percentage of non‐C1 cells positive for NR1 in rostral RVLM when compared to active animals (p=0.067 and p=0.0048, respectively). In contrast, there were no overt differences between sedentary versus active rats in the percentage of C1 and non‐C1 cells positive for NR1 in caudal RVLM.Preliminary western blotting data showed lower expression of PSD95 (an excitatory post‐synaptic density protein) in sedentary versus active animals (0.89±0.15 vs. 1.64±0.24, PSD95/GAPDH, respectively), suggesting differential synaptic/extrasynaptic localization of glutamate receptors, including NR1, in sedentary and active groups.The greater number of NR1‐positive, bulbospinal C1 neurons in rostral RVLM of sedentary rats is consistent with our hypothesis and previous data showing enhanced SNA following activation of glutamate receptors in sedentary vs. active rats. Sedentary conditions may also enhance excitation of RVLM neurons by altering the composition of NMDAR subunits, their synaptic/extrasynaptic localization, trafficking, or expression of synapse‐related proteins and/or transporters.Support or Funding InformationHL096787‐08; AHA25810010This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Central blockade of the AT1 receptor attenuates pressor effects via reduction of glutamate release and downregulation of NMDA/AMPA receptors in the rostral ventrolateral medulla of rats with stress-induced hypertension.

Glutamatergic activity in the rostral ventrolateral medulla (RVLM), which is an important brain area where angiotensin II (Ang II) elicits its pressor effects, contributes to the onset of hypertension. The present study aimed to explore the effect of central Ang II type 1 receptor (AT1R) blockade on glutamatergic actions in the RVLM of stress-induced hypertensive rats (SIHR). The stress-induced hypertension (SIH) model was established by electric foot shocks combined with noises. Normotensive Sprague-Dawley rats (control) and SIHR were intracerebroventricularly infused with the AT1R antagonist candesartan or artificial cerebrospinal fluid for 14 days. Mean arterial pressure (MAP), heart rate (HR), plasma norepinephrine (NE), glutamate, and the expression of N-methyl-D-aspartic acid (NMDA) receptor subunit NR1, and α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors in the RVLM increased in the SIH group. These increases were blunted by candesartan. Bilateral microinjection of the ionotropic glutamate receptor antagonist kynurenic acid, the NMDA receptor antagonist D-2-amino-5-phosphonopentanoate, or the AMPA/kainate receptors antagonist 6-cyano-7-nitroquinoxaline-2,3-dione into the RVLM caused a depressor response in the SIH group, but not in other groups. NR1 and AMPA receptors expressed in the glutamatergic neurons of the RVLM, and glutamate levels, increased in the intermediolateral column of the spinal cord of SIHR. Central Ang II elicits release of glutamate, which binds to the enhanced ionotropic NMDA and AMPA receptors via AT1R, resulting in activation of glutamatergic neurons in the RVLM, increasing sympathetic excitation in SIHR.

Inhibition of MyD88 Signaling Skews Microglia/Macrophage Polarization and Attenuates Neuronal Apoptosis in the Hippocampus After Status Epilepticus in Mice

Inflammation is implicated in epileptogenesis. Activated microglia and macrophages (MG/MΦ) are found in the brains of patients with epilepsy-related diseases and animal models of epilepsy. It is not yet known how the MG/MΦ activation phenotype affects pathological changes in the brain after a single seizure. In this study, we had 2 main purposes: first, to characterize post-status epilepticus (SE) inflammation by tracking MG/MΦ polarization, and, second, to explore the role of an innate immune receptor adaptor protein, namely, myeloid differentiation primary response gene 88 (MyD88), in the induction of SE in a mouse model. A lithium–pilocarpine model of seizure conditions was generated in C57BL/6 mice. The intensity and distribution of MG/MΦ polarization were tracked by fluorescent immunohistochemistry and Western blotting for the polarization markers inducible nitrogen oxygenized synthase, arginase-1, CD163, and mannose receptor. We observed steadily increasing M1 MG/MΦ along with MyD88 signal upregulation after SE in the hippocampi of mice, whereas the M2 marker arginase-1 was localized mainly in astrocytes rather than in MG/MΦ. Inhibition or gene knockout of MyD88 reduced M1 MG/MΦ and gliosis although increasing M2 MG/MΦ in the hippocampi of SE mice. MyD88 inhibition also augmented glutamate transporter 1 expression and reduced N-methyl-D-aspartate receptor NR1 subunit expression in the hippocampus to protect pyramidal neurons from apoptosis. These data suggest that MG/MΦ polarization after SE impacts the pathological outcome of the hippocampus via MyD88 signaling and point to MyD88 as a potential neuroprotective target for epilepsy therapy.

Chronic Adolescent CDPPB Treatment Alters Short-Term, but not Long-Term, Glutamatergic Receptor Expression.

Dysfunction of the glutamatergic system is believed to underlie many neurodevelopmental disorders including autism, Rett syndrome and schizophrenia. Metabotropic glutamate receptor (mGluR5) positive allosteric modulators (PAM) potentiate glutamatergic signaling, particularly indirectly via the NMDA receptor. Preclinical studies report mGluR5 PAMs can improve schizophrenia-relevant behaviours. Furthermore, adolescent administration has shown to prevent cognitive induced deficits in adult rodents. However, there is limited understanding of the short- and long-term neurochemical effects of mGluR5 PAMs, which may underlie their therapeutic effects. We examined the effect of 7-day adolescent (PN28-34) treatment with the mGluR5 PAM, CDDPB (30mg/kg), on glutamatergic receptor expression at adolescence (PN35) and adulthood (PN96). Immunoblot analysis revealed that 7-day adolescent CDPPB treatment increased protein expression of glutamatergic receptors including the NMDA receptor subunits, NR1 and NR2A and the AMPA subunits (GluA1 and GluA2) in the adolescent hippocampus, changes that did not extend to adulthood. In contrast, there were no changes in the adolescent frontal cortex, however elevated mGluR5 protein expression was observed at adulthood following adolescent CDPPB treatment. The present study indicates adolescent CDPPB treatment may cause brain region dependent effects on the glutamatergic system, which do not persist into adulthood. These findings may have implications for the preclinical development of mGluR5 PAMs for the treatment of neurodevelopmental disorders.

Ketamine exacerbates cortical neuroapoptosis under hyperoxic conditions by upregulating expression of the N-methyl-D-aspartate receptor subunit NR1 in the developing rat brain

BackgroundKetamine and hyperoxia are widely used in obstetric and pediatric settings. Either ketamine or hyperoxia has been reported to cause neuroapoptosis in the developing brain, and ketamine-induced neuronal apoptosis may involve a compensatory upregulation of the N-methyl-D-aspartate (NMDA) receptor NR1 subunit. This study investigated the impact of ketamine administration under hyperoxic conditions on cortical neuroapoptosis and NR1 subunit expression in the infant rat brain.MethodsMale, 7-day-old rats were randomly allocated to four groups: control, ketamine, hyperoxia, and ketamine + hyperoxia (n = 18 per group). Rats in the control and ketamine groups received subcutaneous injections of either vehicle (saline) or ketamine (50 mg/kg) in room air (21% oxygen). The hyperoxia and ketamine + hyperoxia groups were exposed to 60% oxygen for 2 h after receiving saline or ketamine. Physiological parameters and arterial oxygen saturation were observed. Neuronal apoptosis and the expressions of NR1 mRNA and protein in the frontal cortex were also examined by transferase dUTP nick end labeling (TUNEL) assays, qPCR and Western blot, respectively.ResultsKetamine alone had no effect on paO2 (P > 0.05), but pups exposed to hyperoxia or hyperoxia + ketamine had significantly greater paO2 values compared to control animals (P < 0.01). Animals exposed to ketamine and ketamine + hyperoxia showed higher apoptotic scores, mRNA and protein expression levels of NR1 than control animals (P < 0.01), and ketamine + hyperoxia caused a significantly greater increase than ketamine alone (P < 0.01).ConclusionsThese data suggest that ketamine administration under hyperoxic conditions exacerbates cortical neuroapoptosis in the developing brain, which may be closely associated with an enhancement in NMDA receptor NR1 subunit expression.

The Midline Axon Crossing Decision Is Regulated through an Activity-Dependent Mechanism by the NMDA Receptor.

Axon guidance in vertebrates is controlled by genetic cascades as well as by intrinsic activity-dependent refinement of connections. Midline axon crossing is one of the best studied pathfinding models and is fundamental to the establishment of bilaterally symmetric nervous systems. However, it is not known whether crossing requires intrinsic activity in axons, and what controls that activity. Further, a mechanism linking neuronal activity and gene expression has not been identified for axon pathfinding. Using embryonic zebrafish, we found that the NMDA receptor (NMDAR) NR1.1 subunit (grin1a) is expressed in commissural axons. Pharmacological inhibition of grin1a, hypoxia exposure reduction of grin1a expression, or CRISPR knock-down of grin1a leads to defects in midline crossing. Inhibition of neuronal activity phenocopies the effects of grin1a loss on midline crossing. By combining pharmacological inhibition of the NMDAR with optogenetic stimulation to precisely restore neuronal activity, we observed rescue of midline crossing. This suggests that the NMDAR controls pathfinding by an activity-dependent mechanism. We further show that the NMDAR may act, via modulating activity, on the transcription factor arxa (mammalian Arx), a known regulator of midline pathfinding. These findings uncover a novel role for the NMDAR in controlling activity to regulate commissural pathfinding and identify arxa as a key link between the genetic and activity-dependent regulation of midline axon guidance.

High and low nightly running behavior associates with nucleus accumbens N-Methyl-d-aspartate receptor (NMDAR) NR1 subunit expression and NMDAR functional differences

The extent to which N-Methyl-d-aspartate (NMDA) receptors facilitate the motivation to voluntarily wheel-run in rodents has yet to be determined. In so, we utilized female Wistar rats selectively bred to voluntarily run high (HVR) and low (LVR) nightly distances in order to examine if endogenous differences in nucleus accumbens (NAc) NMDA receptor expression and function underlies the propensity for high or low motivation to voluntarily wheel-run. 12–14 week old HVR and LVR females were used to examine: 1.) NAc mRNA and protein expression of NMDA subunits NR1, NR2A and NR2B; 2.) NMDA current responses in isolated medium spiny neurons (MSNs) and 3.) NMDA-evoked dopamine release in an ex vivo preparation of NAc punches. Expectedly, there was a large divergence in nightly running distance and time between HVR and LVR rats. We saw a significantly higher mRNA and protein expression of NR1 in HVR compared to LVR rats, while seeing no difference in the expression of NR2A or NR2B. There was a greater current response to a 500 ms application of 300 μM of NMDA in medium-spiny neurons isolated from the NAc HVR compared to LVR animals. On average, NMDA-evoked punches (50 μM of NMDA for 10 min) taken from HVR rats retained ∼54% of the dopamine content compared to their bilateral non-evoked sides, while evoked punches from LVR animals showed no statistical decrease in dopamine content compared to their non-evoked sides. Collectively, these data suggest a potential link between NAc NR1 subunit expression as well as NMDA function and the predisposition for nightly voluntary running behavior in rats. In light of the epidemic rise in physical inactivity, these findings have the potential to explain a neuro-molecular mechanism that regulates the motivation to be physically active.

Memory and Learning Dysfunction Following Copper Toxicity: Biochemical and Immunohistochemical Basis.

The prototype disease of Cu toxicity in human is Wilson disease, and cognitive impairment is the presenting symptom of it. There is no study correlating Cu-induced excitotoxicity, apoptosis, and astrocytic reaction with memory dysfunction. We report excitotoxicity, apoptosis, and astrocytic reaction of the hippocampus and frontal cortex with memory dysfunction in rat model of Cu toxicity. Thirty-six rats were divided into group I (control) and group II (100mg/kgBwt/day CuSO4 orally). Y-maze was performed for memory and learning at 0, 30, 60, and 90days. Frontal and hippocampal free Cu concentration, oxidative stress markers [glutathione (GSH), total antioxidant toxicity (TAC), and malondialdehyde (MDA)], and glutamate were measured by atomic absorption spectroscopy, spectrophotometry, and ELISA, respectively. N-methyl-D-aspartate receptors (NMDARs) NR1, NR2A, and NR2B were done by real-time polymerase chain reaction. Immunohistochemistry for caspase-3 and glial fibrillary acidic protein (GFAP) were done and quantified using the ImageJ software. The glutamate level in hippocampus was increased, and NMDAR expression was decreased at 30, 60, and 90days in group II compared to group I. In the frontal cortex, glutamate was increased at 90days, but NMDARs were not significantly different in group II compared to group I. Caspase-3 and GFAP expressions were also higher in group II compared to group I, and these changes were more marked in hippocampus than frontal cortex. These changes correlated with respective free tissue Cu, oxidative stress, and Y-maze attention score. Cu toxicity induces apoptosis and astrocytosis of the hippocampus and frontal cortex through direct or glutamate and oxidative stress pathways, and results in impaired memory and learning.

Crack cocaine inhalation induces schizophrenia-like symptoms and molecular alterations in mice prefrontal cortex

Crack cocaine (crack) addiction represents a major social and health burden, especially seeing as users are more prone to engage in criminal and violent acts. Crack users show a higher prevalence of psychiatric comorbidities – particularly antisocial personality disorders – when compared to powder cocaine users. They also develop cognitive deficits related mainly to executive functions, including working memory. It is noteworthy that stimulant drugs can induce psychotic states, which appear to mimic some symptoms of schizophrenia among users. Social withdraw and executive function deficits are, respectively, negative and cognitive symptoms of schizophrenia mediated by reduced dopamine (DA) tone in the prefrontal cortex (PFC) of patients. That could be explained by an increased expression of D2R short isoform (D2S) in the PFC of such patients and/or by hypofunctioning NMDA receptors in this region. Reduced DA tone has already been described in the PFC of mice exposed to crack smoke. Therefore, it is possible that behavioral alterations presented by crack users result from molecular and biochemical neuronal alterations akin to schizophrenia. Accordingly, we found that upon crack inhalation mice have shown decreased social interaction and working memory deficits analogous to schizophrenia's symptoms, along with increased D2S/D2L expression ratio and decreased expression of NR1, NR2A and NR2B NMDA receptor subunits in the PFC. Herein we propose two possible mechanisms to explain the reduced DA tone in the PFC elicited by crack consumption in mice, bringing also the first direct evidence that crack use may result in schizophrenia-like neurochemical, molecular and behavioral alterations.