miR-429-3p mediates memory decline by targeting MKP-1 to reduce surface GluA1-containing AMPA receptors in a mouse model of Alzheimer's disease

Alzheimer's disease (AD) is a leading cause of dementia in the elderly. Mitogen-activated protein kinase phosphatase 1 (MKP-1) plays a neuroprotective role in AD. However, the molecular mechanisms underlying the effects of MKP-1 on AD have not been extensively studied. MicroRNAs (miRNAs) regulate gene expression at the post-transcriptional level, thereby repressing mRNA translation. Here, we reported that the microRNA-429-3p (miR-429-3p) was significantly increased in the brain of APP23/PS45 AD model mice and N2AAPP AD model cells. We further found that miR-429-3p could downregulate MKP-1 expression by directly binding to its 3'-untranslated region (3' UTR). Inhibition of miR-429-3p by its antagomir (A-miR-429) restored the expression of MKP-1 to a control level and consequently reduced the amyloidogenic processing of APP and Aβ accumulation. More importantly, intranasal administration of A-miR-429 successfully ameliorated the deficits of hippocampal CA1 long-term potentiation and spatial learning and memory in AD model mice by suppressing extracellular signal-regulated kinase (ERK1/2)-mediated GluA1 hyperphosphorylation at Ser831 site, thereby increasing the surface expression of GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). Together, these results demonstrate that inhibiting miR-429-3p to upregulate MKP-1 effectively improves cognitive and synaptic functions in AD model mice, suggesting that miR-429/MKP-1 pathway may be a novel therapeutic target for AD treatment.

Regulation of AMPA receptor on propofol induced hippocampal mitochondrial injury in neonatal rats / 中华危重病急救医学

Objective:To investigate whether propofol can cause injury to hippocampal mitochondria in neonatal rats and the regulation of excitatory amino acid receptor AMPA receptor.Methods:Forty-eight Sprague-Dawley (SD) rats aged 7 days were randomly divided into control group, propofol group, propofol+AMPA receptor agonist AMPA group (propofol+AMPA group) and propofol+AMPA receptor inhibitor CNQX group (propofol+CNQX group), with 12 rats in each group. The rats in the propofol groups were intraperitoneally injected with 30 mg/kg propofol, while in control group with 3 mg/kg normal saline. Each group was given 1/2 of the first dose every 20 minutes after the first administration, three times a day, for three consecutive days. The rats in the propofol+AMPA group and the propofol+CNQX group were injected with 1 g/L AMPA or CNQX 5 μL through left ventricle after the first administration. Three days after administration, the rats were sacrificed to obtain brain tissue. Western blotting was used to determine the expression of AMPA receptor glutamate receptors (GluR1, GluR2) subunit totally (T) and on membrane (M) in hippocampus. The expression of dynamin-related protein-1 (DRP-1) and phosphorylated-DRP-1 (p-DRP-1) and mitofusin 2 (Mfn2) related to mitochondrial fission and fusion were determined. The adenosine triphosphate (ATP) content and ATPase activity were determined.Results:Compared with the control group, GluR1 expression and its M/T ratio were significantly increased after treatment of propofol, GluR2 expression and its M/T ratio were significantly decreased, the ATP content and ATP-related enzyme activity were decreased significantly, while the expression of DRP-1 and its phosphorylation was significantly increased, and the expression of Mfn2 was significantly decreased. The changes indicated that repeated intraperitoneal injection of 30 mg/kg propofol leading to the injury of mitochondria in neural cells. Compared with the propofol group, the GluR1 expression and its M/T ratio further increased after AMPA agonist administration [T-GluR1 protein (T-GluR1/β-actin): 2.41±0.29 vs. 1.72±0.11, M-GluR1 protein (M-GluR1/β-actin): 1.18±0.15 vs. 0.79±0.09, M/T ratio: 0.78±0.12 vs. 0.46±0.08, all P < 0.01], GluR2 expression was significantly increased [T-GluR2 protein (T-GluR2/β-actin): 0.65±0.13 vs. 0.30±0.14, P < 0.01; M-GluR2 protein (M-GluR2/β-actin): 0.17±0.05 vs. 0.13±0.07, P > 0.05], but its M/T ratio was further decreased (0.27±0.10 vs. 0.41±0.08, P < 0.05). The ATP-related enzyme activity was further decreased, and the ATP content was further decreased (μmol/g: 0.32±0.07 vs. 0.70±0.10, P < 0.01). Mitochondria DRP-1 expression and its phosphorylation were further increased [DRP-1 protein (DRP-1/GAPDH): 2.75±0.36 vs. 1.70±0.19, p-DRP-1 protein (p-DRP-1/GAPDH): 0.99±0.14 vs. 0.76±0.15, both P < 0.05], and Mfn2 expression was further decreased (Mfn2/GAPDH: 0.23±0.12 vs. 0.54±0.12, P < 0.05). This indicated that the AMPA agonist increased the expression of the AMPA receptor GluR1 subunit on the cell membrane and shifted the GluR2 into the cell, thus increasing the mitochondrial injury caused by propofol. Compared with the propofol group, the GluR1 expression and its M/T ratio decreased significantly after AMPA inhibitor administration [T-GluR1 protein (T-GluR1/β-actin): 0.99±0.14 vs. 1.72±0.11, M-GluR1 protein (M-GluR1/β-actin): 0.21±0.07 vs. 0.79±0.09, M/T ratio: 0.21±0.07 vs. 0.46±0.08, all P < 0.01], the change of GluR2 expression was not significant, but its M/T ratio was significantly increased (0.59±0.09 vs. 0.41±0.08, P < 0.05). The ATP-related enzyme activity was increased significantly, and the ATP content was increased significantly (μmol/g: 0.87±0.12 vs. 0.70±0.10, P < 0.05). Mitochondria DRP-1 expression and its phosphorylation were significantly decreased [DRP-1 protein (DRP-1/GAPDH): 1.18±0.17 vs. 1.70±0.19, p-DRP-1 protein (p-DRP-1/GAPDH): 0.37±0.10 vs. 0.76±0.10, both P < 0.05], and Mfn2 expression was significantly increased (Mfn2/GAPDH: 0.78±0.10 vs. 0.54±0.12, P < 0.05). This indicated that AMPA inhibitor promoted the movement to the cell membrane of GluR2 subunits meanwhile inhibited the expression of GluR1 subunits, thus alleviating the injury of mitochondrial caused by propofol in the brain. Conclusions:Repeated intraperitoneal injection of 30 mg/kg propofol for 3 days can increase the expression of GluR1 subunits of AMPA receptor in 7-day neonatal rats hippocampus mainly distributing in the cell membrane, decrease the expression of GluR2 subunits moving into the cell, thus causing injury of mitochondrial function and dynamics, which can be aggravated by AMPA receptor agonist and alleviated by AMPA receptor inhibitors.

TAT-GluA2CT interferes with the protective effect of GluA2/TARPγ-8 coupling on nerve injury in status epilepticus model of rats / 中华实用儿科临床杂志

Objective:To investigate the protective effect of interfering peptide TAT-GluA2CT on hippocampal neurons in the Lithium chlorine-Pilocarpine status epilepticus model and the optimal time of administration.Methods:Male SD rats (72 cases) were induced to status epilepticus by using Lithium chlorine-Pilocarpine, while a control group ( n=12) was established.The 72 rats were divided into epilepsy group ( n=12), TAT-sham peptide group ( n=12), TAT-GluA2CT peptide group ( n=48) according to the random number table method, and the TAT-GluA2CT peptide group were further divided into the pre-1 h group ( n=12), the post-2 h group ( n=12), the post-4 h group( n=12), and the post-6 h group ( n=12) according to the administration time of the TAT-GluA2CT peptide.Nissl staining and terminal dUTP nick end labeling (TUNEL) assay were performed on 6 rats each from control group, epilepsy group, TAT-shampeptide group, pre-1 h group, post-2 h group, post-4 h group, and post-6 h group to observe the morphological changes and apoptosis of neurons in the CA1 region of the rat hippocampus.Western blot and co-immunopercipitation test were used to detect the expression of GluA2[second subunit of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) recepter] and the coupling of GluA2/transmembrane AMPA receptor regulatory protein (TARP γ-8) complex in control group, epilepsy group, pre-1 h group, post-2 h group, post-4 h group and post-6 h group.The t-test was used to compare the data differences between 2 groups, and one-way ANOVA was adopted to compare the differences between the groups. Results:Compared with the epilepsy group, the number of neurons in each TAT-GluA2CT peptide group increased significantly, and the difference was statistically significant( epilepsy group 20.07±3.51, pre-1 h group 39.40±2.39, post-2 h group 38.43±2.42, post-4 h group 30.30±2.55, and post-6 h group 27.93±3.20, F=235.28, P<0.05). Compared with the epilepsy group, the number of apoptotic cells in each TAT-GluA2CT peptide group was significantly reduced, and the difference was statistically significant(epilepsy group 31.47±3.19, pre-1 h group 7.30±3.45, post-2 h group 9.27±3.81, post-4 h group 12.86±3.08, and post-6 h group 14.43±3.13, F=248.60, P<0.05). Compared with the control group, the expression of hippocampal GluA2 decreased after epilepsy induction, and the difference was statistically significant(control group 21 626.53±2 700.58, epilepsy group 14 578.16±2 917.02, pre-1 h group 13 375.47±3 180.54, post-2 h group 15 244.10±1 390.41, post-4 h group 15 799.16±4 559.49, post-6 h group 15 722.95±1 756.01, F=3.83, P<0.05). No statistical difference was observed in the expression of GluA2 between the TAT-GluA2CT peptide group and the epilepsy group( F=0.45, P=0.77). Compared with the epilepsy group, GluA2/TARPγ-8 complex coupling was decreased in each TAT-GluA2CT peptide group, and the difference was statistically significant(epilepsy group 24 509.80±3 718.54, pre-1 h group 12 055.18±5 847.11, post-2 h group 9 630.51±5 805.17, post-4 h group 12 749.35±7 108.45, post-6 h group 11 092.98±7 330.08, F=10.68, P<0.05). Compared with the epilepsy group, the incubation period of seizures in the pre-1 h group was prolonged and the seizure rating was decreased, with statistically significant differences[epilepsy group (18.58±3.99) min, pre-1 h group (103.25±9.21) min, t=29.23, P<0.05]. Conclusions:TAT-GluA2CT peptide can attenuate the neuronal damage in hippocampus of epileptic rats.The neuroprotective effect of TAT-GluA2CT peptide was most obvious at 1 h before or 2 h after administration of Pilocarpine.

Cannabidiol improves cognitive impairment in dementia model rats by regulating the expression of AMPA receptor in the brain / 中风与神经疾病杂志

@#To observe the effects of cannabidiol（CBD）on cognitive function and AMPA receptor expression in sporadic Alzheimer’s disease（sAD）rats induced by bilateral intraventricular injection of streptozotocin（STZ）. MethodsForty adult male Sprague-Dawley（SD）rats were randomly divided into four groups：sham operation group（S），cannabidiol（CBD）+ sham operation group（CBD+S），model group（STZ），CBD+model group（CBD+STZ）. The cognitive function of rats was tested by Morris water maze（MWM）and step-down test. Western blot was used to detecte the expression of GluR1 and GluR2 in the cerebral cortex and hippocampus. ResultsIn the Morris water maze test，compared with the S group，the escape latency in the STZ group was significantly prolonged（P<0.01），and the escape latency in the CBD+STZ group was significantly shortened（P<0.01）. In the step-down test，compared with the latency and the number of errors in the S group，the latency in the STZ group was significantly shortened（P<0.01），and the number of errors was significantly increased（P<0.01）. Compared with the STZ group，the latency of CBD+STZ group was significantly prolonged（P<0.05），and the number of errors of CBD+STZ group was significantly reduced（P<0.05）. Compared with the STZ group，the contents of GluR1 in cortex and hippocampus of the CBD+STZ group were significantly（P<0.01）（P<0.05）decreased. Compared with STZ group，the contents of GluR2 in cortex and hippocampus of the CBD+STZ group were significantly（P<0.01）（P<0.01）decreased. ConclusionCannabidiol improves STZ-induced cognitive impairment in rats by regulating AMPA receptor expression in the brain.

Progress on the mechanism and intervention of glutamate AMPA receptor GluA2/TARPγ-8 in the pathogenesis of epilepsy / 国际儿科学杂志

Epilepsy occurs as a result of episodic abnormal synchronous discharges in cerebral neuronal networks.It is characterized by an imbalance between excitatory and inhibitory neurotransmission.Although various non-conventional mechanisms are implicated in epileptic synchronization, glutamate excitatory neurons play an essential role.AMPA receptors mediate fast synaptic excitation within and between brain regions relevant to epilepsy, and play a role in epileptogenesis and in seizure-induced brain damage.However, direct modulation of AMPA receptors may have undesirable consequences, given its wide expression within the central nervous system and critical roles on brain circuitry development.Hippocampal CA1 region, as the main site of epilepsy, selectively regulates the high expression of AMPA receptor GluA2 subunit and transmembrane AMPA receptor regulatory protein family(TARPs)γ-8 subtype and its complex GluA2/TARPγ-8, whether it can produce anti-epileptic and avoid adverse reactions.

Effect of AMPA receptor endocytosis inhibitor GluA2-3Y on cognitive function and postsynaptic protein expression in chronic cerebral hypoperfusion rats / 中华行为医学与脑科学杂志

Objective:To investigate the effect of GluA2-3Y which is an inhibitor of AMPA(α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid) receptor internalization on cognitive function and hippocampal postsynaptic protein expression in rats with chronic cerebral hypoperfusion.Methods:Forty-eight adult male SD rats were randomly divided into Sham group, 2VO group, high-dose GluA2-3Y group and low-dose GluA2-3Y group according to random number table, with 12 rats in each group.The chronic cerebral hypoperfusion model of rat was established by two vessel occlusion (2VO) while the Sham operation was performed in rats of Sham group.The rats in high dose GluA2-3Y group and low dose GluA2-3Y group were intraperitoneal injected with 3 μmol/kg and 0.03 μmol/kg GluA2-3Y respectively once a day for 2 weeks. Rats in 2VO group and Sham group were intraperitoneally injected with control peptide. Morris water maze test and new object recognition test were performed to evaluate the learning and memory ability of rats, and Western blot was used to evaluate the expression of Akt1、GSK3β、p-GSK3β、GluA2 and PSD-95 in rat hippocampus. The expressions of GluA2 and PSD-95 in rat hippocampus were evaluated by immunofluorescence. SPSS 23.0 software was used for data analysis. The comparison between multiple groups was analyzed by one-way ANOVA and repeated measurement ANOVA was used to analyze Morris water maze results. And independent-samples t-test was used for pairwise comparisons. Results:(1)In Morris water maze trials, the results of repeated measurement ANOVA showed that the interaction between group and time of escape latency of rats in each group was not significant ( F=0.79, P>0.05), and the group main effect and time main effect were significant ( F=24.44, 40.42, both P<0.05). On the 5th day of navigation trials, the escape latency of rats in 2VO group was longer than that in sham group ( t=5.87, P<0.05). The escape latency of rats in low dose GluA2-3Y group and high dose GluA2-3Y group were significantly shorter than that in 2VO group ( t=2.20, 3.41, both P<0.05), but there was no significant difference between low dose GluA2-3Y group and high dose GluA2-3Y group ( t=1.37, P>0.05). The target quadrant residence time and resolution coefficient ((14.57±1.40)s, (0.15±0.10)) in 2VO group were significantly lower than those in Sham group ((23.71±2.57)s, (0.40±0.06)) ( t=3.23, 2.24, both P<0.05), while the target quadrant residence time in high dose GluA2-3Y group ((20.19±1.53)s) and low dose GluA2-3Y group ((20.31±2.06)s) were longer than that in 2VO group( t=2.71, 2.35, both P<0.05). The discrimination coefficients in high dose GluA2-3Y group (0.47±0.10) and low dose GluA2-3Y group (0.59±0.06) were higher than that of 2VO group ( t=2.21, 3.94, both P<0.05). (2)The Western blot results showed that the expression of PSD-95 and GluA2 in hippocampus of rats in 2VO group were significantly lower than those in Sham group ( t=2.31, 2.20, both P<0.05), and the expression of PSD-95 in high dose GluA2-3Y group (1.026±0.056) was significantly higher than that in 2VO group ((0.760±0.061), t=2.49, P<0.05), while there was no significant difference between low-dose GluA2-3Y group and 2VO group( t=0.96, P>0.05). The expression of GluA2 in low-dose GluA2-3Y group was higher than that in 2VO Group ((1.130±0.087), (0.766±0.080), t=2.37, P<0.05), but there was no significant difference between high-dose GluA2-3Y group and 2VO group( t=1.06, P>0.05). (3) Immunofluorescence showed that compared with Sham group, the expression of PSD-95 and GluA2 in 2VO group decreased ( t=4.23, 2.57, P<0.05). Compared with 2VO group, the expression of PSD-95 and GluA2 in high dose GluA2-3Y group and low dose GluA2-3Y group increased significantly, and the differences were statistically significant (PSD-95: (7.757±0.578), (12.057±0.578), t=3.14, 6.96, both P<0.05; (9.721±0.950), (16.610±0.950), t=4.56, 9.34, both P<0.05). (4) The results of Western blot showed that the expression GSK3β in hippocampus of rats in each group were not statistically different( F=2.03, P>0.05). There were significant differences in the expression of Akt1, p-GSK3β and the percentage of p-GSK3β/GSK3β in hippocampus of rats in each group ( F=8.30, 4.76, 3.57, all P<0.05). Compared with Sham group, the levels of Akt1, p-GSK3β and the percentage of p-GSK3β/GSK3β in 2VO group were significantly lower ( t=3.00, 2.81, 3.17, all P<0.05). Compared with 2VO group, the levels of Akt1, p-GSK3β and p-GSK3β/GSK3β percentage in low dose GluA2-3Y group and high-dose GluA2-3Y group were significantly higher (Akt1: t=2.05, 5.20, both P<0.05; p-GSK3β: t=2.49, 4.15, both P<0.05; p-GSK3β/GSK3β percentage: t=2.30, 2.97, both P<0.05). Conclusion:GluA2-3Y, an AMPA receptor internalization inhibitor, can alleviate the cognitive impairment in rats with chronic cerebral hypoperfusion, which may be related to the increased expression of Akt1, p-GSK3β and postsynaptic proteins.

Effect of p-coumaric acid on olfactory bulbectomy induced depression-like behaviors and its AMPA receptor mechanism in mice / 中国药理学通报

Aim To investigated the effect of p-CA on depression-like behaviors of mice of olfactory bulbectomy and its possible mechanism. Methods The olfactory bulbectomy (OBX) model of mice was established by an operation of olfactory bulbectomy. The behaviors of the mice were detected by the forced swimming test and the tail suspension test. Results The depression-like behavior in the forced swimming and tail suspension test and the in the open field test significantly increased in OBX mice; however, p-CA improved the depres- sion-like behavior in the forced swimming and tail sus pension test and the hyper-locomotor activity in open field test in OBX mice. Moreover, treatment with AMPA receptor antagonist NBQX blocked this improving effect of p-CA. While, treatment with AMPA receptor agonist CX546 enhanced this improving effect of p-CA. Conclusions P-CA improves depression-like behaviors of OBX mice, and AMPA receptors may mediate this effect.

Chlorogenic acid alleviates chronic inflammatory pain via inhibiting synaptic expression of AMPA receptor / 中国药理学通报

Aim To investigate the effect of chlorogenic acid (CGA) on chronic inflammatory pain induced by complete Freund's adjuvant (CFA) and to observe the influence of CGA on CFA-induced synaptic expression of AMAP receptor in spinal dorsal horn. Methods CFA was injected intraplantarly into the hindpaws of mice to induce mechanical allodynia and thermal hyperalgesia. The changes of paw withdrawal threshold (PWT) and the paw withdrawal latency (PWL) were tested after intrathecal administration of CGA. Meanwhile, the synaptic expression and phosphorylation of AMPA receptor subunit were assessed by immunoblot-ting. Results The intrathecal injection of CGA produced dose-depended improvement of both PWT and PWL in CFA injected mice. A higher dose of CGA (200ng) did not influence the base thresholds of normal mice. The median dose of CGA (100 ng) effectively reversed the CFA-induced synaptic expression of GluAl; meanwhile, it suppressed the phosphorylation of GluAl at Ser845, with no influence on phosphorylation at Ser831. Conclusions CGA might exert its analgesic effect by specifically inhibiting the phosphorylation of GluAl -Ser845 and the synaptic incorporation of GluAl-containing AMPA receptor.

Distribution of PSD95 and GluA2 subunit during maturation of primary hippocampal neurons / 中国药理学通报

Aim To investigate the distribution of postsynaptic density protein 95 (PSD95) and AMPA receptor GluA2 subunit during the maturation of rat hippocampal neurons in vitro. Methods Cultured rat hippocampal neurons at 4(days in vitro, DIV), 7DIV, 14DIV and 20DIV were used in an immunofluorescence assay to test co-localization of PSD95 and GluA2 by laser confocal microscope at a magnification of 60 ×. Results PSD95 puncta were gradually distributed densely in dendrites and dendritic spines during neuron maturation progresses. GluA2 subunits were uniformly distributed in the cell body, axons, dendrites and dendritic spines. The Pearson' s correlation of PSD95 and GluA2 was 0. 830 ±0. 033 and 0. 734 ±0. 019 respectively at the dendrites of 4DIV and 7DIV neurons, which showed strong co-localization. While at the dendrites of 14DIV and 20DIV neurons, a moderate correlation was demonstrated by the Pearson correlation 0. 547 ± 0. 021 and 0. 574 ± 0. 024. Conclusions GluA2 has no specific intracellular distribution during neuron maturation, while the functional localization of PSD95 depends on the appearance of dendritic spines. The co-localization of GluA2 with PSD95 in dendritic spines suggests that there are stable synapses containing AMPA receptors.

The Effects of cerebrospinal fluid AMPA receptor levels on the clinical efficacy of lamotrigine in the treatment of epilepsy / 中风与神经疾病杂志

@#Objective　To explore the effects of cerebrospinal fluid AMPA receptor levels on the clinical efficacy of lamotrigine in the treatment of epilepsy. Methods　The 70 cases of epilepsy in our hospital were diagnosed and selected in this study from December 2017 to October 2019；The AMPA receptor content of patients in cerebrospinal fluid were determined by enzyme linked immunosorbent assay；The patients were paired into high AMPA group （n=34） and low AMPA group （n=36） according to the median value at 4.08 ng/ml. Clinical efficacy and the incidence of adverse reactions were compared between two groups. Results　Before treatment，there was no significant difference in seizure frequency between the two groups （P>0.05）；After treatment for 6 months and 12 months，the seizure frequencies of the two groups were gradually reduced （P<0.05）；Moreover，the number of seizures in the low AMPA group were significantly less than those in the high AMPA group （P<0.05）；The response rate of high AMPA group was 79.41%，which was significantly lower than that in the low group AMPA （χ2=6.055，P=0.048）；The improvement on the efficiency of electroencephalogram in the high AMPA group was 67.65%，which was significantly lower than that in the low AMPA group （χ2=4.686，P=0.030）；However，there was no significant on the incidence of adverse reactions between two groups （χ2=0.202，P=0.653）. Conclusion　AMPA receptor plays an important role in the development of epilepsy and the low AMPA receptor level patients were more efficacies to the treatment of lamotrigine.

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

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

AMPA receptor positive modulator, ampakines: research progress / 国际药学研究杂志

Ampakines are a class of pharmacological agents acting as positive modulators of AMPA receptors. Currently, clinical indications studied for ampakines involve many diseases including the respiratory depression and psychoneurosis, etc. The studies using CX516 and CX546 as tool compounds have shown that ampakines could be classified into "high impact ampakines"and "low impact ampakines". The two subclasses of ampakines differ in the chemical structures, influences on receptor dynamics, receptor-ligand bindings, synaptic transmissions, neurotrophin regulations and side effects. According to the available literature, the low impact ampakines have better clinical application prospects than high impact ampakines because of their high safety and good tolerance. The above different characteristics of the two subclasses of ampakines are reviewed in this paper.

Anti-fatigue activity screening of novel positive allosteric modulators on AMPA receptor / 中国药理学通报

Aim To explore the novel anti-fatigue a-gents targeting AMPA receptor. Methods Three ben-zoic acids of different methoxy substitutions were em-ployed, including piperonylic acid, 3,4,5-trimethoxy benzoic acid and 3,4-dimethoxy benzoic acid, which as mother nucleus and cyclic aliphatic amine or pheny-lpiperazine derivatives were introduced to modify the nitrogen atoms connecting to amino bonds. Forty-three compounds were synthesized and identified by 1 H-NMR. MTT assay, then pentobarbital induced hypno-sis experiment and mice burden swimming experiment were applied respectively to evaluate the new synthesized compounds’ cytotoxicity, CNS excitability and anti-fatigue activity. Results Compound 2j had low cytotoxicity,presenting certain central excitability and significant advantages on anti-fatigue. Conclusion The further development of compounds 2j with good anti-fatigue activities could be cultivated in further study.

Mechanism and impact of a novel allosteric AMPA receptor modulator on protection against respiratory depression / 中国药理学与毒理学杂志

OBJECTIVE Respiratory depression hinders the use of anaesthetics and sedative hyp-notics.To explore the mechanism of LCX001 on protection against respiratory depression,a novel AMPA receptor modulator LCX001,synthesized by our Institute of Medicinal Chemistry,is expected to relieve suppressed respiration. METHODS LCX001 was tested to alleviate respiratory depression triggered by opioid(fentanyl and TH-030418),propofol and pentobarbital in the plethysmography recording.The acetic acid writhing and hot-plate tests were conducted to evaluate analgesic effect of LCX001.Binding assay and whole-cell recording were used to analyze the property of LCX001 on positive modulation. The function of AMPA receptors were determined by location of receptors in the membrane and state of channel opening, and both processes were impressed by AMPA receptor regulatory proteins. Ac-cording to the theory,the effect of LCX001 on the expression of stargazin was measured firstly by west-ern blotting. The variation of receptor surface location was observed by live cell imaging. The regula-tion on neuronal Ca2+and cell function was investigated intensively by Ca2+imaging to clarify mecha-nism of LCX001. RESULTS LCX001 effectively rescued and prevented opioid (fentanyl and TH-030418), propofol, and pentobarbital-induced respiratory depression by strengthening respiratory fre-quency and minute ventilation in rats. The acetic acid writhing test and hot-plate test revealed potent anti-nociceptive efficacy of LCX001,in contrast to some ampakines that did not affect analgesia. Fur-thermore,LCX001 potentiated[3H]AMPA and L-glutamate binding affinity to AMPA receptors,and facili-tated glutamate-evoked inward currents in HEK293 cells stably expressing GluA2(R).Importantly,appli-cation of LCX001 generated a significant increase in GluA2(R) surface expression in a mechanism of stargazin up-regulation,and restrained opioid-induced abnormal intracellular Ca2+load,which might par-ticipate in breathing modulation. CONCLUSION The novel pharmacological effect and potential new mechanism of LCX001 might promote ampakines to be a therapeutic option for protection against respi-ratory depression.

The RAS/PI3K Pathway is Involved in the Impairment of Long-term Potentiation Induced by Acute Aluminum Treatment in Rats / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To explore the role of RAS/PI3K pathway in the impairment of long-term potentiation (LTP) induced by acute aluminum (Al) treatment in rats in vivo.</p><p><b>METHODS</b>First, different dosages of aluminum-maltolate complex [Al(mal)3] were given to rats via acute intracerebroventricular (i.c.v.) injection. Following Al exposure, the RAS activity of rat hippocampus were detected by ELISA assay after the hippocampal LTP recording by field potentiation technique in vivo. Second, the antagonism on the aluminum-induced suppression of hippocampal LTP was observed after the treatment of the RAS activator epidermal growth factor (EGF). Finally, the antagonism on the downstream molecules (PKB activity and the phosphorylation of GluR1 S831 and S845) were tested by ELISA and West-blot assays at the same time.</p><p><b>RESULTS</b>With the increasing aluminum dosage, a gradually decreasing in RAS activity of the rat hippocampus was produced after a gradually suppressing on LTP. The aluminum-induced early suppression of hippocampal LTP was antagonized by the RAS activator epidermal growth factor (EGF). And the EGF treatment produced changes similar to those observed for LTP between the groups on PKB activity as well as the phosphorylation of GluR1 S831 and S845.</p><p><b>CONCLUSION</b>The RAS→PI3K/PKB→GluR1 S831 and S845 signal transduction pathway may be involved in the inhibition of hippocampal LTP by aluminum exposure in rats. However, the mechanisms underlying this observation need further investigation.</p>

Effects of electroacupuncture on learning and memory in experimental Alzeheimer′s disease / 中华物理医学与康复杂志

Objective To observe the effects of electroacupuncture ( EA) on learning and memory ability and the expression of N-ethylmaleimide-sensitive fusion factor ( NSF) in experimental Alzheimer′s disease ( AD) and to explore the mechanism involved. Methods Sixty healthy, male Sprague-Dawley rats were randomly divided into a normal group, a sham operation group, a model group and an EA group, each of 15.The model group and the EA group were injected with Aβ25-35 bilaterally in the CA1 area of the hippocampus, while the sham operation group was injected with the same amount of normal saline in the same regions. From the 1stday after the AD model had been es-tablished successfully, EA was applied to the Shen Shu ( BL23) and Bai Hui ( GV20) acupoints in the EA group once a day,6 days a week for 2 weeks. No EA was given to the other 3 groups. After the treatment, Morris water maze tests were conducted to determine assess the rats′ learning and memory ability, and immunohistochemical methods were used to test the expression level of NSF in the CA1 areas of the rats′hippocampus. Results The average es-cape latent period of the EA group was significantly shorter than that of the model group and their average platform crossing time was significantly longer. The average expression of NSF in the EA group measured as integrated optical intensity was significantly higher than that in the model group. Conclusion EA can effectively improve learning and memory ability in AD, at least in rats. The mechanism may involve increasing the expression of NSF in the CA1 area of the hippocampus.

S-palmitoylation regulates AMPA receptors trafficking and function: a novel insight into synaptic regulation and therapeutics

Glutamate acting on AMPA-type ionotropic glutamate receptor (AMPAR) mediates the majority of fast excitatory synaptic transmission in the mammalian central nervous system. Dynamic regulation of AMPAR by post-translational modifications is one of the key elements that allow the nervous system to adapt to environment stimulations. S-palmitoylation, an important lipid modification by post-translational addition of a long-chain fatty acid to a cysteine residue, regulates AMPA receptor trafficking, which dynamically affects multiple fundamental brain functions, such as learning and memory. In vivo, S-palmitoylation is controlled by palmitoyl acyl transferases and palmitoyl thioesterases. In this review, we highlight advances in the mechanisms for dynamic AMPA receptors palmitoylation, and discuss how palmitoylation affects AMPA receptors function at synapses in recent years. Pharmacological regulation of S-palmitoylation may serve as a novel therapeutic strategy for neurobiological diseases.

Enhancement of GluN2B Subunit-Containing NMDA Receptor Underlies Serotonergic Regulation of Long-Term Potentiation after Critical Period in the Rat Visual Cortex

Serotonin [5-hydroxytryptamine (5-HT)] regulates synaptic plasticity in the visual cortex. Although the effects of 5-HT on plasticity showed huge diversity depending on the ages of animals and species, it has been unclear how 5-HT can show such diverse effects. In the rat visual cortex, 5-HT suppressed long-term potentiation (LTP) at 5 weeks but enhanced LTP at 8 weeks. We speculated that this difference may originate from differential regulation of neurotransmission by 5-HT between the age groups. Thus, we investigated the effects of 5-HT on apha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR)-, gamma-aminobutyric acid receptor type A (GABA(A)R)-, and N-methyl-D-aspartic acid receptor (NMDAR)-mediated neurotransmissions and their involvement in the differential regulation of plasticity between 5 and 8 weeks. AMPAR-mediated currents were not affected by 5-HT at both 5 and 8 weeks. GABA(A)R-mediated currents were enhanced by 5-HT at both age groups. However, 5-HT enhanced NMDAR-mediated currents only at 8 weeks. The enhancement of NMDAR-mediated currents appeared to be mediated by the enhanced function of GluN2B subunit-containing NMDAR. The enhanced GABA(A)R- and NMDAR-mediated neurotransmissions were responsible for the suppression of LTP at 5 weeks and the facilitation of LTP at 8 weeks, respectively. These results indicate that the effects of 5-HT on neurotransmission change with development, and the changes may underlie the differential regulation of synaptic plasticity between different age groups. Thus, the developmental changes in 5-HT function should be carefully considered while investigating the 5-HT-mediated metaplastic control of the cortical network.

Effects of exposure to aluminum on long-term potentiation and AMPA receptor subunits in rats in vivo / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To explore the effects of exposure to aluminum (Al) on long-term potentiation (LTP) and AMPA receptor subunits in rats in vivo.</p><p><b>METHODS</b>Different dosages of aluminum-maltolate complex [Al(mal)3] were given to rats via acute intracerebroventricular (i.c.v.) injection and subchronic intraperitoneal (i.p.) injection. Following Al exposure, the hippocampal LTP were recorded by field potentiation technique in vivo and the expression of AMPAR subunit proteins (GluR1 and GluR2) in both total and membrane-enriched extracts from the CA1 area of rat hippocampus were detected by Western blot assay.</p><p><b>RESULTS</b>Acute Al treatment produced dose-dependent suppression of LTP in the rat hippocampus and dose-dependent decreases of GluR1 and GluR2 in membrane extracts; however, no similar changes were found in the total cell extracts, which suggests decreased trafficking of AMPA receptor subunits from intracellular pools to synaptic sites in the hippocampus. The dose-dependent suppressive effects on LTP and the expression of AMPA receptor subunits both in the membrane and in total extracts were found after subchronic Al treatment, indicating a decrease in AMPA receptor subunit trafficking from intracellular pools to synaptic sites and an additional reduction in the expression of the subunits.</p><p><b>CONCLUSION</b>Al(mal)3 obviously and dose-dependently suppressed LTP in the rat hippocampal CA1 region in vivo, and this suppression may be related to both trafficking and decreases in the expression of AMPA receptor subunit proteins. However, the mechanisms underlying these observations need further investigation.</p>

Relationship of PTEN expression level with positioning of AMPA receptor subunit GluR2 in hippocampal neuron after stretch injury / 中华创伤杂志

Objective To explore the relationship of phosphatase and tensin homology deleted on chromosome ten ( PTEN ) expression with the positioning of AMPA recepter subunit GluR2 in hippocampal neuron after stretch injury. Methods The primary rat hippocampal neuron from neonate rats were cultured and the model of stretch injury established. Western blot and TUNEL staining were used to detect PTEN expression and the positioning of AMPA receptor subunit GluR2 in rat hippocampal neuron at different time intervals after stretch injury. Results The PTEN expression was increased obviously after stretch injury, while the expressions of AMPA receptors GluR1, GluR2 and GluR3 showed no evident differences at every time point. However, AMPA receptor subunit GluR2 located at cytomembrane was down-regulated obviously after stretch injury. Conclusion The intracellular PTEN expression and the expression of AMPA receptor subunit GluR2 located at cytomembrane are on the contrary, indicating that PTEN phosphatase may participate in the transport and reintegration process of AMPA receptor subunit GluR2 located at cytomembrane.