N-methyl-D-aspartate Receptor Subunit 2B mRNA Research Articles

GLYX-13 pretreatment ameliorates long-term isoflurane exposure-induced cognitive impairment in mice.

Accumulating evidence indicates that inhalation anesthetics induce or increase the risk of cognitive impairment. GLYX-13 (rapastinel) acts on the glycine site of N-methyl-D-aspartate receptors (NMDARs) and has been shown to enhance hippocampus-dependent learning and memory function. However, the mechanisms by which GLYX-13 affects learning and memory function are still unclear. In this study, we investigated these mechanisms in a mouse model of long-term anesthesia exposure. Mice were intravenously administered 1 mg/kg GLYX-13 at 2 hours before isoflurane exposure (1.5% for 6 hours). Cognitive function was assessed using the contextual fear conditioning test and the novel object recognition test. The mRNA expression and phosphorylated protein levels of NMDAR pathway components, N-methyl-D-aspartate receptor subunit 2B(NR2B)-Ca2+/calmodulin dependent protein kinase II (CaMKII)-cyclic adenosine monophosphate response element binding protein (CREB), in the hippocampus were evaluated by quantitative RT-PCR and western blot assay. Pretreatment with GLYX-13 ameliorated isoflurane exposure-induced cognitive impairment and restored NR2B, CaMKII and CREB mRNA and phosphorylated protein levels. Intracerebroventricular injection of KN93, a selective CaMKII inhibitor, significantly diminished the effect of GLYX-13 on cognitive function and NR2B, CaMKII and CREB levels in the hippocampus. Taken together, our findings suggest that GLYX-13 pretreatment alleviates isoflurane-induced cognitive dysfunction by protecting against perturbation of the NR2B/CaMKII/CREB signaling pathway in the hippocampus. Therefore, GLYX-13 may have therapeutic potential for the treatment of anesthesia-induced cognitive dysfunction. This study was approved by the Experimental Animal Ethics Committee of Drum Tower Hospital affiliated to the Medical College of Nanjing University, China (approval No. 20171102) on November 20, 2017.

Correlation between the epigenetic modification of histone H3K9 acetylation of NR2B gene promoter in rat hippocampus and ethanol withdrawal syndrome.

Patients with alcohol use disorder may develop acute ethanol withdrawal syndrome (EWS). Previous studies showed that an epigenetic modification of the N-methyl-D-aspartate (NMDA) receptor, especially NMDA receptor 2B subunit (NR2B), was involved in the pathological process of EWS. However, the relationship between the epigenetic regulation of the NR2B gene in the rat hippocampus region and EWS were inconsistent. The purpose of this study was to explore the role of the histone H3K9 acetylation of the NR2B gene in the rat hippocampus region in EWS. A rat model of chronic ethanol exposure was established. EWS score and the behavioral changes were recorded at different time points. The NR2B expression levels and the histone H3K9 acetylation level in the NR2B gene promoter region were measured using qRT-PCR, Western blot, immunofluorescence, and chromatin immunoprecipitation, respectively. Finally, the relationship between the epigenetic modification of histone H3K9 acetylation of NR2B gene promoter and EWS were examined. Our ultimate results showed that the EWS score was increased at 2h, peaked at 6h after withdrawal of ethanol, and reduced to the level parallel to the normal control group at day 3 after ethanol withdrawal. The NR2B mRNA expression and protein levels showed similar patterns. Further correlation analyses indicted that both histone H3K9 acetylation in NR2B gene promoter and the expression levels of NR2B were positively associated with EWS. Our results suggest that chronic ethanol exposure may result in epigenetic modification of histone H3K9 acetylation in NR2B gene promoter in rat hippocampus, and the expression levels of NR2B were found to be positively correlated with ethanol withdrawal syndrome.

Monosialotetrahexosylganglioside Inhibits the Expression of p-CREB and NR2B in the Auditory Cortex in Rats with Salicylate-Induced Tinnitus.

This study investigated the effects of monosialotetrahexosylganglioside (GM1) on the expression of N-methyl-D-aspartate receptor subunit 2B (NR2B) and phosphorylated (p)-cyclic AMP response element-binding protein (CREB) in the auditory cortex of rats with tinnitus. Tinnitus-like behavior in rats was tested with the gap prepulse inhibition of acoustic startle paradigm. We then investigated the NR2B mRNA and protein and p-CREB protein levels in the auditory cortex of tinnitus rats compared with normal rats. Rats treated for 4 days with salicylate exhibited tinnitus. NR2B mRNA and protein and p-CREB protein levels were upregulated in these animals, with expression returning to normal levels 14 days after cessation of treatment; baseline levels of NR2B and p-CREB were also restored by GM1 administration. These data suggest that chronic salicylate administration induces tinnitus via upregulation of p-CREB and NR2B expression, and that GM1 can potentially be used to treat tinnitus.

Effect of intraperitoneal WSLP/NR2B siRNA compound on neuropathic pain in rats

Objective To evaluate the effect of intraperitoneal water soluble lipopolymer (WSLP)/ N-methyl-D-aspartate receptor subunit 2B (NR2B) siRNA compound on the neuropathic pain (NP) in rats.Methods Eighty-four healthy male Sprague-Dawley rats,aged 6 weeks,weighing 180-220 g,were randomly divided into 7 groups (n =12 each) using a random number table:control group (group C),sham operation group (group S),NP group,WSLP/NR2B siRNA group (siWSLP group),WSLP/negative control siRNA group (ncWSLP group),PEI/NR2B siRNA group (PEI group) and WSLP group (WSLP group).NP was produced by ligation of the left L5 spinal nerve.In group S,the left L5 spinal nerve was only exposed,but not ligated.In group C,the rats underwent no treatment.Groups siWSLP,ncWSLP,PEI and WSLP received single intraperitoneal injection of WSLP/NR2B siRNA,WSLP/negative control siRNA,PEI/NR2B siRNA and WSLP compound 2 ml,respectively,at 10 days after NP.At 1 day before operation,7 days after operation,and 3,7,14 and 21 days after intraperitoneal injection,6 rats in each group were chosen randomly to measure mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL).At 3 days after intraperitoneal injection,the left 6 rats in each group were sacrificed and the spinal cord was removed for detection of NP2B mRNA expression (using PCR) and NR2B expression (by Western blot).Results Compared with group C,MWT was significantly decreased,and TWL was shortened on 7 days after operation and 3,7 and 21 days after intraperitoneal injection,and the expression of NR2B mRNA and protein was down-regulated on 3 days after administration in the other groups.Compared with group NP,MWT was significantly increased,and TWL was prolonged on day 3 and 7 after intraperitoneal injection,and the expression of NR2B mRNA and protein was down-regulated on day 3 after administration in siWSLP group.Conclusion Intraperitoneal WSLP/NR2B siRNA compound can effectively relieve the NP in rats. Key words: Gene therapy; Transfection ; RNA, small interfering; Receptors, N-methyl-D-aspartate ; Neuralgia; Injections,intraperitoneal

Role of endogenous hydrogen sulfide of the spinal cord in the maintenance of diabetic neuropathic pain in rats

Objective To evaluate the role of endogenous hydrogen sulfide in the maintenance of diabetic neuropathic pain (DNP) in rats.Methods The DNP models were established by intraperitoneal injection of stretopzin (STZ) in 24 male SD rats aged 4 weeks (150-170 g),and other 24 rats were intraperitoneally injected with the same volume of normal saline as normal group.All the rats were implanted intrathecal catheter.The rats with DNP were randomly divided into 2 subgroups (n =12 each):saline group (group D) and methemoglobin group (group DM).The rats in normal group were randomly divided into 2 subgroups (n =12 each):saline group (group C) and methemoglobin group (group CM).Three weeks after intraperitoneal injection,the rats were separately intrathecally administered methemoglobin 10 μ mol/L (10 μ1) (group DM),saline 10 μl (group D),methemoglobin 10 μmol/L (10 μl) (group CM) and saline 10 μl (group C),respectively,once a day for 4 days.The paw withdrawal mechanical threshold (PWT) of the rats were measured before and after injection at 1 (T1),2 (T2),3 (T3),4 (T4) days,then the rats were sacrificed at T4 point after the measurement and the lumbar segments of the spinal cord were removed for determination of N-methyl-D-aspartate receptor subunit 2B (NR2B) protein expression (by Western blotting) and NR2B mRNA expression [by reverse transcriptase-polymerase chain reaction (RT-PCR)].Results As compared with group C and group CM,the PWT in group D and group DM was significantly reduced (P ＜ 0.05),while the expression of NR2B protein and mRNA was significantly increased (P ＜ 0.05).As Compared with group D [NR2B mRNA (0.89 ± 0.09),NR2B protein (1.28 ± 0.17)],the PWT in group DM was significantly increased at T2-T4 point before the injection and T1-T4 point after the injection,while the expression of NR2B mRNA (0.69 ±0.11) and NR2B protein (0.98 ± 0.14) in group DM was significantly decreased (P ＜ 0.05).As compared with the point before the injection (3.8 ± 0.9 and 6.1 ± 1.4),the PWT in group DM were significantly increased at T1-T2 point after the injection (7.5 ± 1.8 and 8.7 ± 1.6) (P ＜ 0.05).Conclusion Endogenous hydrogen sulfide in the spinal cord is involved in the maintenance of DNP in rats and the possible mechanisms were related with up-regulation of NR2B in the spinal dorsal horn. Key words: Hydrogen sulfide; N-methyl-D-aspartate receptor subunit 2B; Spinal dorsal horn; Diabete ; Pain

Protective effect of L-3-n-Butylphthalide on the brain of ethanol dependence rats

This study was carried out to explore the protective effect of L-3-nbutylphthalide (NBP) on the brain of ethanol dependence rats. With the administration of different dosage of NBP, the variation of withdrawal scores, hydrogen sulfide (H2S) contents, cystathionine β-synthase (CBS) activities and expression N-methyl-D-aspartate (NMDA) receptor 2B subunit (NR2B) in the hippocampus tissue of ethanol dependent rats were discussed. Results showed that most significant difference in withdrawal symptom score, H2S content, CBS activity and the expression of NR2B mRNA between normal and ethanol dependence model group rats (P 0.05). However, withdrawal symptom score, H2S content, CBS activity and NR2B mRNA expression in both middle and high dose NBP group rats were much lower than those of experiment control group (P<0.05). With increasing concentrations of NBP, it could reduce withdrawal symptoms of ethanol dependence rats to different extent. The alleviation of ethanol dependence symptoms mediated by NBP may be related to down-regulation in NR2B mRNA expression. Key words: Ethanol dependence, L-3-nbutylphthalide, hydrogen sulfide (H2S), cystathionine β-synthase activity, N-methyl-D-aspartate (NMDA) receptor 2B subunit (NR2B).

Effect of isoflurane anesthesia during early pregnancy on cognitive function of offspring rats

Objective To investigate the effect of isoflurane anesthesia during early pregnancy on the cognitive function of offspring rats.Methods Thirty SD rats at 5-7 day gestation were randomly divided into 3 groups ( n =10 each):control group (group C) and 2 isoflurane groups (groups Ⅰ1,Ⅰ2).Groups Ⅰ1 and Ⅰ2 inhaled 1.4％ isoflurane in O2 for 4 and 8 h respectively while group C inhaled 95 ％ O2 for 8 h.At 20 and 30 days after birth,offspring rats from 5 pregnant rats were tested for learning and memory abilities using Morris water maze.The offsprings were sacrificed at 7 days after test and their hippocampi were isolated for determination of N-methyl-D-aspartate receptor subunit 2B (NR2B) mRNA and protein expression.Results There were no significant differences in the results of Morris water maze test and NR2B mRNA and protein expression among the three groups.Conclusion Isoflurane anesthesia during early pregnancy has no effect on the cognitive function of the offspring rats. Key words: Isoflurane; Pregnancy trimester,first; Cognition disorders

Effect of inhalation of enflurane in early pregtancy on the expression of NR2B in the hippocampus of offsprings of rats

Objective To investigate the effect of inhalation of enflurane on the expression of N-methyl-D-aspartate receptor subunit 2B (NR2B) in the hippocampus of the offsprings of rats.Methods Thirty SD rats pregnancy 8-10 day weighing 200-250 g were randomly divided into 3 groups ( n =10 each):control group (Group C),4 h inhalation of enflurane group ( group E1 ) and 8 h inhalation of enflurane group ( group E2 ).Group E1 and E2inhaled 1.7 ％ enflurane (in O2 2 L/min) for 4 and 8 h respectively,while group C inhaled oxygen 2 L/rin for 8 h.The learning and memory functions of the offsprings were assessed at 20 and 30 days after birth by Morris maze test.The expression of the NR2B mRNA were examined by RT-PCR,NR2B protein were examined by mmunohistochemistry.Results Compared with group C,the escape latency was significantly prolonged,the frequency of crossing the original platform was significantly decreased,the staying time at the original platform quadrant was significantly shortened at 3-5 days after the test in group E1 and E2 (.P ＜ 0.05 ),the expression levels of NR2B mRNA and protein were significantly decreased at 20 and 30 days after birth in group E1 and E2 ( P ＜ 0.05).There were no significant differences in the indexes mentioned above between groups E1 and F2 ( P ＞ 0.05).Conclusion Inhalation enflurane in the early pregnancy can result in cognition dysfunction through inhibiting NR2B expression in the hippocampus of the offsprings of the rats. Key words: Anesthetics,inhalation; Pregnancy trimester, first; Receptors, N-methyl- D-aspartate

Rhynchophylline down-regulates NR2B expression in cortex and hippocampal CA1 area of amphetamine-induced conditioned place preference rat

N-methyl-D-aspartate receptor 2B subunit (NR2B) has an important role in the development of conditioned place preference (CPP) and psychostimulant abuse. Rhynchophylline is presently used to treat central nervous systems diseases and has a non-competitive antagonistic effect on NMDA receptors. In this study, amphetamine was administered in rats (2 mg/kg, s.c., once each day for 4 consecutive days), during which they were treated with rhynchophylline (60 mg/kg, i.p., once each day for the next 3 days). NR2B mRNA and protein expression were examined by in situ hybridization and immunohistochemistry. CPP was induced by amphetamine (2 mg/kg, s.c.) by 4th day in rats. Rhynchophylline effectively reversed the expression of amphetamine-induced CPP and itself did not produce a CPP. Amphetamine-CPP rats showed a significantly increased NR2B mRNA and protein expression in medial prefrontal cortex and hippocampal CA1 areas as compared to the control group. Rhynchophylline reversed NR2B mRNA and protein levels induced by amphetamine but rhynchophylline by itself had no effect on NR2B expression in control rats. These results indicate that rhynchophylline inhibits the expression of amphetamine-induced rewarding effect, and this action might be related to down-regulation of NR2B expression in medial prefrontal cortex and hippocampal CA1 area.

トルエン曝露したマウスの空間学習行動と記憶機能関連遺伝子発現へのD-シクロセリンの影響

D-cycloserine (DCS), a partial N-methyl-D-aspartate (NMDA) receptor agonist, is a well-known cognitive enhancer. To investigate the effect of DCS in cognitive function following toluene exposure, eight-week-old male C3H/HeN mice were exposed to filtered air (0 ppm) or 50 ppm toluene for 6 h a day on 5 consecutive days a week for 6 weeks. The day after the day of final exposure, a spatial learning task was performed using a Morris water maze apparatus. During the learning task, some mice were treated with DCS intraperitoneally (20 mg/kg) 30 min before the first trial of the acquisition phase and probe trial. After completion of the spatial learning task, the hippocampus was collected from each mouse to examine memory function-related gene expression using the real-time RT-PCR method. During the acquisition phase, on day 3 and 4, toluene-exposed mice with DCS treatment showed significantly better learning performance than corresponding saline treated groups. Moreover, the toluene-exposed mice with DCS treatment also showed significantly improved memory retention during the probe trial and up-regulation of hippocampal NMDA receptor subunit 2B mRNA expression compared with the saline treated groups. Our findings indicate that a subunit-specific modulation of hippocampal NMDA receptor mRNA expression by DCS contributes to improvement of spatial learning performance in mice following toluene exposure.

Effects of different concentrations of sevoflurane on cognitive function in gerontal rats

Objective To investigate the changes in cognitive function and glutarnatergie N-methyl-D-aspartate receptor-2B subunit (NR2B) expression in hippocarnpus after sevoflurane anesthesia in gerontal rats.Methods Forty 18 month old male SD rats weighing 500-650 g were randomly divided into 3 groups: group control (C n=8);group Ⅱ and Ⅲ inhaled 1.5% and 3.0% sevoflurane for 2 h respectively (S1, S2 n=16each). Y-maze was used to assess the cognitive function at 1 and 7 day after 2 h sovoflurane inhalation respectively (n=8 at each time point). The animals were killed at 12 h after the test and bilateral hippocarnpi were isolated for determination of expression of NR2B mRNA and protein using RT-PCR and immuno-histochemistry respectively. Results 3.0% sevoflurane inhalation induced cognitive decline and increased expression of NR2B mRNA and protein at 1 d in group S2 as compared with group C. There was no significant change in cognitive function after 2 h 1.5 % sevoflurane inhalation. Conclusion Anesthesia with high concentration of sevoflurane can induce cognitive decline in gerontal rats possibly through up-regulation of NMDA receptor expression in hippocampus. Key words: Anesthetics, inhalation; Cognition disorders; Aged; Receptors, N-methy-D-aspartate

Developmental regulation of NMDA receptor 2B subunit mRNA and ifenprodil binding in the zebra finch anterior forebrain

In passerine songbirds, song learning often is restricted to an early sensitive period and requires the participation of several discrete regions within the anterior forebrain. Activation of N-methyl-D-aspartate (NMDA) receptors is implicated in song learning and in one forebrain song region, the lateral magnocellular nucleus of the anterior neostriatum (IMAN), NMDA receptors decrease in density, their affinity for the antagonist MK-801 increases, and their currents decay more quickly as young male zebra finches lose the ability to imitate new song elements. These developmental changes in NMDA receptor pharmacology and physiology suggest that the subunit composition of NMDA receptors changes developmentally. Here, we have used in situ hybridization and [3H]ifenprodil receptor autoradiography to study the developmental regulation of the NMDA receptor 2B subunit (NR2B) within the anterior forebrain of male zebra finches. NR2B mRNA expression within the IMAN was twice as great in 30-day-old males (early in the sensitive period for song learning) as in adult males, and this developmental decrease in NR2B mRNA expression was mirrored by a decrease in high-affinity (NR2B-associated) [3H]ifenprodil binding within this song region. In another anterior forebrain song region, Area X, NR2B mRNA also declined significantly after 30 days posthatch, but this decline was not accompanied by a significant decrease in [3H]ifenprodil binding. The results are consistent with the hypothesis that developmental changes in NMDA receptor function mediated by regulation of subunit composition contribute to the sensitive period for vocal learning in birds.