OBJECTIVE: To observe the GluR2 expression in rat inferior colliculus (IC) in different developmental stages, and to investigate its developmental change and relationship with the synapse development. METHOD: The expression of GluR2 and synaptophysin(SYP) in IC were detected by double immunofluorescence method. RESULT: (1) All sorts of neurons in IC expressed GluR2 in every postnatal groups, and the GluR2 expression in P6w groups was higher than that in other groups. (2) The expression of GluR2 were different in different subnucleus of IC. (3) All sorts of neurons in IC expressed SYP in every postnatal groups, and the SYP expression in P6w groups was higher than others. (4) The expressions of GluR2 consistent with the expression of SYP in IC. CONCLUSION The developmental changes of GluR2 and SYP expression in the rat IC may be involved in the development and plasticity of auditory center.
Animals ; Hypothalamus ; cytology ; metabolism ; Inferior Colliculi ; growth & development ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, AMPA ; metabolism ; Synapses ; metabolism ; Synaptophysin ; metabolism

Extracellular recordings of field excitatory postsynaptic potential (fEPSP) is one of the most common ways for studies of synaptic plasticity, such as long-term potentiation (LTP) and paired-pulse plasticity (PPP). The measurement of the changes in the different components of fEPSP waveform, such as the initial slope, initial area, peak amplitude and whole area, were commonly used as criteria for the judgement of potentiation or depression of synaptic plasticity. However, the differences in the conclusions drawn from measuring different components of fEPSP waveform at the same recording have still been largely ignored. Here we compared high-frequency stimulation (HFS)-evoked synaptic plasticity, both LTP and PPP, by measuring different components of fEPSP waveform, including the initial slope, initial area, peak amplitude, whole area and time course. The results not only indicated the acceleration of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor kinetics underlies LTP in hippocampal CA1 region of mice, but also showed that different measurements of fEPSP waveform at the same recording result in different magnitudes of LTP and different forms of PPP in hippocampal CA1 region of mice. After HFS, the paired-pulse ratio was slightly decreased by measurement of the initial area, but obviously increased by measurement of the initial slope of the pair fEPSPs. These results might draw apparently contradictory conclusions. Therefore, careful and complete analysis of the data from different parts of fEPSP waveforms is important for reflection of the faithful changes in synaptic plasticity.
Animals ; CA1 Region, Hippocampal ; physiology ; Excitatory Postsynaptic Potentials ; Long-Term Potentiation ; Mice ; Neuronal Plasticity ; Receptors, AMPA ; metabolism

To explore novel antifatigue agents targeting with AMPA receptor, 10 compounds were synthesized and their structures were confirmed by 1H NMR, ESI-MS and elemental analysis. 1-BCP was treated as the leading compound. The antifatigue activities were evaluated by weight-loaded forced swimming test, and the AMPA receptor binding affinities were tested with radioligand receptor binding assays. The results unveiled that 5b appeared to possess potent antifatigue activities and high affinity with AMPA receptor, which deserved further studies.
Animals ; Benzamides ; chemistry ; pharmacology ; Dioxoles ; chemistry ; pharmacology ; Fatigue ; prevention & control ; Piperidines ; chemistry ; pharmacology ; Radioligand Assay ; Receptors, AMPA ; metabolism ; Swimming

Calcium is one of the most versatile intracellular second messengers, which plays crucial roles in many intracellular signaling pathways. Researches on intracellular calcium distribution, regulation and function are important for our understanding of cellular physiology. In this mini-review, the regulation of intracellular calcium signal in retinal horizontal cells and the relevant physiological functions were introduced based on the experiments carried out in our laboratory. Intracellular calcium dynamics following the activation of AMPA and NMDA receptors were introduced based on our experiments performed on carp retinal horizontal cells using calcium imaging technique and computational methods. An initial peak response was observed in both cases, which indicated an active participation of intracellular calcium store during the calcium dynamics initiated by AMPA/NMDA receptor activation. Intracellular recording experiments indicated that calcium signaling was crucial for the gradual enhancement of the retinal horizontal cell's responsiveness in exposure to repetitive red flashes. Possible roles of intracellular calcium signaling in the regulation of GABA transporter activity were also introduced based on our whole-cell recording experiments performed on isolated carp retinal horizontal cells.
Animals ; Calcium ; metabolism ; Calcium Signaling ; Carps ; Cells, Cultured ; Patch-Clamp Techniques ; Receptors, AMPA ; metabolism ; Receptors, N-Methyl-D-Aspartate ; metabolism ; Retinal Horizontal Cells ; physiology

OBJECTIVETo investigate the role of 83 site in interaction of GluR2 C-terminal and PICK1 PDZ domain.

METHODSDocking structure of PICK1 PDZ domain with GluR2 C terminal PDZ binding motif was built with computer software. After K83 site was substituted by other amino acid, the structure and binding energy were recalculated; meanwhile, site specific mutants were constructed using wild type full length cDNA as template. Mutants were co-transfected with GluR2 into HEK293T cells. After staining, the distribution of PICK1 and GluR2 were observed under confocal microscope.

RESULTSWild type PICK1 and GluR2 formed many co-clusters in HEK293T cells as reported by other research groups; but different K83 mutant had different distribution in HEK293T cells.

CONCLUSIONThe K83 site in PDZ domain of PICK1 is important for the interaction between PICK1 and GluR2. Altering lysine will probably change the hydrophobic interactions, the hydrogen bonds or the electrostatic interactions formed between PICK1 PDZ domain and GluR2 C terminal; accordingly, that will change the binding capacity between PICK1 and GluR2 in varying degrees.

Binding Sites ; Carrier Proteins ; chemistry ; metabolism ; Computer Simulation ; HEK293 Cells ; Humans ; Nuclear Proteins ; chemistry ; metabolism ; PDZ Domains ; Protein Binding ; Receptors, AMPA ; metabolism

OBJECTIVE: To explore whether sex difference exists in the performance in each arm of elevated plus maze (EPM) and GluR1 level in the hippocampus of female and male Sprague-Dawley rats. METHODS: Eleven male and 10 female SD rats were tested for 5 minutes in the EPM. These rats were decapitated 30 min after testing. The left and right hippocampus were dissected. Samples were stored at -80 degree for protein extracting. Western blot was used to detect the GluR1 levels in the hippocampus. RESULTS: Female rats exhibited less anxiety-like behaviors than male rats in the EPM (P<0.05).Female rats had lower GluR1 levels in total and left hippocampus than those of male rats (P<0.05). CONCLUSION Sex difference exists between female and male rats in the EPM and hippocampal GluR1.
Animals ; Anxiety ; metabolism ; physiopathology ; Behavior, Animal ; Female ; Hippocampus ; metabolism ; physiology ; Male ; Maze Learning ; physiology ; Rats ; Rats, Sprague-Dawley ; Receptors, AMPA ; metabolism ; physiology ; Sex Characteristics

Late-phase long-term potentiation (L-LTP) plays a very important role in the maintenance of long-term memory in hippocampus. However, studies have shown that L-LTP can be reversed by subsequent neuronal activity. The aim of the present study is to investigate whether the presynaptic mechanism and the change of AMPARs expressions are involved in the reversal of L-LTP in hippocampal CA1 area. Standard extracellular recording technique was used to record the potential change in the stratum radiatum of CA1 area of adult rat hippocampal slices. Two hours after LTP induction, which was induced by high-frequency stimulation (HFS), two episodes of high-intensity paired-pulse low-frequency stimulation (HI-PP-LFS) were delivered to induce L-LTP reversal. Paired-pulse ratios (PPR) were obtained before LTP induction, 2 h after LTP induction and 30 min after LTP reversal. On the other hand, immunofluorescence histochemistry was used to detect AMPARs expressions before and after L-LTP reversal. The results showed that, after 2 h of induction, L-LTP was partially reversed by two episodes of HI-PP-LFS, and the percentage of depotentiation was 61.79%+/-14.51%. PPR obtained before and after LTP induction, and as well that after LTP reversal, are all more than 1, showing paired-pulse facilitation (PPF). Multiple comparison indicated PPR before LTP induction was the greatest one, and PPR after LTP induction was the smallest. In addition, no significant difference was observed in the intensity of AMPAR/GluR2 immunoreactivity in CA1 area among control group, LTP group and LTP reversal group. These results suggest that the presynaptic mechanism is involved in both the maintenance and reversal of L-LTP and there is no change in AMPAR/GluR2 expression before and after the reversal of L-LTP.
Animals ; CA1 Region, Hippocampal ; metabolism ; physiology ; Electric Stimulation ; Long-Term Potentiation ; physiology ; Male ; Presynaptic Terminals ; physiology ; Rats ; Rats, Sprague-Dawley ; Receptors, AMPA ; metabolism

To study the effect of glutamate on the intracellular calcium signal of pure cultured rat astrocytes and the role of NMDA and AMPA receptors in the procedure, the change of calcium signal was investigated by monitoring the fluctuation of intracellular Ca2+ concentration ([Ca2+]i) on the basis of Fura-2 single cell fluorescent ratio (F345/F380). The changes in the effect of glutamate on the intracellular calcium signal were observed after blockage of NMDA and (or) AMPA receptors. It was found that L-glutamate could induce an increased [Ca2+]i in most of the cells in concentration- and time-dependent manner. D-(-)-2-amino-5-phosphonopentanoic acid (D-AP-5, a selective antagonist of the NMDA receptor) and 6-cyano-7-nitroquinoxaline-2, 3-dione (CNQX, a selective antagonist of the AMPA receptor) could abolish the effects of NMDA and AMPA respectively. The treatment of D-AP-5 and CNQX simultaneously or respectively could attenuate the effect of L-glutamate at varying degrees. All these indicated that glutamate could modulate intracellular Ca2+ of pure cultured rat astrocytes through different pathways. The activation of NMDA and AMPA receptors took part in the complex mechanisms.
Animals ; Animals, Newborn ; Astrocytes ; cytology ; metabolism ; Biological Transport ; Calcium ; metabolism ; Calcium Channel Blockers ; pharmacology ; Cells, Cultured ; Cytosol ; metabolism ; Glutamic Acid ; pharmacology ; Hippocampus ; cytology ; metabolism ; Rats ; Receptors, AMPA ; metabolism ; Receptors, N-Methyl-D-Aspartate ; metabolism ; Synaptic Transmission

The study was to investigate the role of homocysteine (Hcy) which was released by hippocampal glial cells and its relationship with NMDA receptor and AMPA receptor in depression induced by chronic unpredictable mild stress (CUMS), and explore the mechanism of changes of Glu/Glu receptor in glial cells and neurons. CUMS-induced depression model was established. The body weight of rats was weighed on the 1st, 7th, 14th, and 21st days during the experiment. The behavioral performances were observed by means of sucrose consumption test, open field test and tail suspension test. Intrahippocampal microinjection of Hcy, NMDA receptor antagonist MK-801 and AMPA receptor antagonist NBQX was performed under stereotaxic guide cannula. The concentration of Glu and the expression of its receptors' subunits were detected respectively by high performance liquid chromatography (HPLC) and Western blot. The Hcy content and the levels of phosphorylation of NMDA receptor and AMPA receptor in hippocampus were separately determined by enzyme linked immunosorbent assay (ELISA). The results showed that CUMS significantly induced the depression-like behaviors in rats, and the content of Glu and Hcy, the expression of NMDA receptors' subunits NR1/NR2B and the level of phosphorylation of NMDA receptor (p-NMDAR) in hippocampus increased significantly, while the expression of AMPA receptors' subunits GluR2/3 and the level of phosphorylation of AMPA receptor (p-AMPAR) decreased significantly. Microinjection of Hcy into hippocampus resulted in similar animal depression-like behaviors and increased Glu content compared to the CON/SAL group, the expression of NR1/NR2B/GluR2/3 and the level of p-NMDAR increased significantly, but the level of p-AMPAR reduced observably. Intrahippocampal injections of MK-801 effectively improved the depression-like behaviors induced by CUMS and Hcy, and attenuated the elevation of Glu content induced by Hcy in hippocampus, whereas NBQX could not improve the depression-like behaviors, but also decreased the Glu content induced by Hcy remarkably. These results suggest that CUMS may contribute to the production and release of Hcy via hippocampal astrocytes. Through the increase of expression of NR1/NR2B/GluR2/3 and level of p-NMDAR, and the decrease of level of p-AMPAR, Hcy results in elevation of Glu level, which leads to depression-like behaviors in the end. In a word, the Hcy released by astrocytes plays an important role in stress-induced elevation of Glu content and variation of NMDA/AMPA receptors in hippocampus.
Animals ; Behavior, Animal ; Depression ; metabolism ; Dizocilpine Maleate ; pharmacology ; Glutamic Acid ; metabolism ; Hippocampus ; metabolism ; Homocysteine ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Receptors, AMPA ; metabolism ; Receptors, N-Methyl-D-Aspartate ; metabolism ; Signal Transduction ; Stress, Psychological

Mutations in genes encoding amyloid precursor protein (APP) and presenilins (PSs) cause familial forms of Alzheimer's disease (AD), a neurodegenerative disorder strongly associated with aging. It is currently unknown whether and how AD risks affect early brain development, and to what extent subtle synaptic pathology may occur prior to overt hallmark AD pathology. Transgenic mutant APP/PS1 over-expression mouse lines are key tools for studying the molecular mechanisms of AD pathogenesis. Among these lines, the 5XFAD mice rapidly develop key features of AD pathology and have proven utility in studying amyloid plaque formation and amyloid β (Aβ)-induced neurodegeneration. We reasoned that transgenic mutant APP/PS1 over-expression in 5XFAD mice may lead to neurodevelopmental defects in early cortical neurons, and performed detailed synaptic physiological characterization of layer 5 (L5) neurons from the prefrontal cortex (PFC) of 5XFAD and wild-type littermate controls. L5 PFC neurons from 5XFAD mice show early APP/Aβ immunolabeling. Whole-cell patch-clamp recording at an early post-weaning age (P22-30) revealed functional impairments; although 5XFAD PFC-L5 neurons exhibited similar membrane properties, they were intrinsically less excitable. In addition, these neurons received smaller amplitude and frequency of miniature excitatory synaptic inputs. These functional disturbances were further corroborated by decreased dendritic spine density and spine head volumes that indicated impaired synapse maturation. Slice biotinylation followed by Western blot analysis of PFC-L5 tissue revealed that 5XFAD mice showed reduced synaptic AMPA receptor subunit GluA1 and decreased synaptic NMDA receptor subunit GluN2A. Consistent with this, patch-clamp recording of the evoked L23>L5 synaptic responses revealed a reduced AMPA/NMDA receptor current ratio, and an increased level of AMPAR-lacking silent synapses. These results suggest that transgenic mutant forms of APP/PS1 overexpression in 5XFAD mice leads to early developmental defects of cortical circuits, which could contribute to the age-dependent synaptic pathology and neurodegeneration later in life.
Mice ; Animals ; Alzheimer Disease/pathology* ; Amyloid beta-Peptides/metabolism* ; Receptors, N-Methyl-D-Aspartate/metabolism* ; Amyloid beta-Protein Precursor/metabolism* ; Mice, Transgenic ; Neurons/metabolism* ; Receptors, AMPA/metabolism* ; Disease Models, Animal