Receptor Subunit GluR1 Research Articles

Neuroprotection Against Transient Focal Cerebral Ischemia and Oxygen–Glucose Deprivation by Interference with GluR6-PSD95 Protein Interaction

Previous studies have shown that KA receptor subunit GluR6 mediated c-Jun N-terminal protein kinase (JNK) signaling is involved in global ischemia injury. Our present study indicates that focal ischemic brain insult on rat middle cerebral artery occlusion (MACo) model enhances the assembly of the GluR6-PSD95-MLK3 module and facilitates the phosphorylation of JNK. Most importantly, a peptide containing the TAT protein transduction sequence, Tat-GluR6-9c, can perturb the assembly of the GluR6-PSD95-MLK3 signaling module and suppress the activation of MLK3, MKK7/4 and JNK. As result, the inhibition of JNK activation caused by Tat-GluR6-9c diminishes the phosphorylation of the transcription factor c-Jun, down-regulates FasL expression and attenuates bax translocation, release of cytochrome c and the activation of caspase-3. Furthermore, MCAo induced infract volume is reduced by intracerebroventricular injection of Tat-Glur6-9c. Oxygen-glucose-deprivation (OGD) cultured cortical neuronal cell also shows an improved cell viability by application of Tat-GluR6-9c. Taken together, our findings strongly suggest that GluR6-PSD95-MLK3 signaling module mediated activation of nuclear and non-nuclear pathways of JNK activation are involved in focal ischemia injury and OGD. Tat-GluR6-9c, the peptide we constructed, gives a new insight into the therapy for ischemic stroke.

The potential role of phrenic nucleus glutamate receptor subunits in mediating spontaneous crossed phrenic activity in neonatal rat

Cervical spinal cord hemisection rostral to the phrenic nucleus leads to paralysis of the ipsilateral hemidiaphragm in adult rats. Respiratory function can be restored to the paralyzed hemidiaphragm by activating a latent respiratory motor pathway. The latent pathway is called the crossed phrenic pathway. In adult rats, the pathway can be activated by drug-induced upregulation of NMDA receptor NR2A subunit and AMPA receptor GluR1 subunit in the phrenic nucleus following hemisection. In neonatal rats, this pathway is not latent as shown by the spontaneous expression of activity in the ipsilateral hemidiaphragm following hemisection. We hypothesized that the NR2A and GluR1 subunits may be highly expressed naturally on phrenic motoneurons of neonatal rats and may play a potential role in mediating the spontaneous expression of activity in the ipsilateral hemidiaphragm after hemisection. To test this hypothesis, the protein levels of NR2A and GluR1 in different age rats were assessed via Western blot analysis immediately following C2 hemisection and EMG recording of crossed phrenic activity. The protein levels of NR2A and GluR1 were transiently high in postnatal day 2 (P2) rats and then was significantly reduced in P7 and P35 animals. An immunofluorescence study qualitatively supported these findings. The present results indicate that the developmental downregulation of the phrenic nucleus glutamate receptor subunits correlates with the conversion of the crossed phrenic pathway in older postnatal animals from an active state to a latent state.

Effects of Memantine on NMDA Induced Long‐Term Depression in Mouse Models of Down Syndrome

The Ts65Dn mouse is the most complete animal model for Down syndrome. Recently, we have shown that the Ts65Dn mouse displays pharmacological responses consistent with a dysfunction in molecular pathways coupled to the gating of NMDA receptors; these include the finding that the uncompetitive NMDA receptor antagonist memantine rescues learning and memory deficits in these mice. In the present study, we used electrophysiological and biochemical techniques to probe potential mechanisms underlying these observations. NMDA mediated long‐term depression (LTD) was induced chemically (Chem‐LTD) in the CA1 region of acute hippocampal slices from Ts65Dn and euploid control mice. We found that Ts65Dn mice showed a greater synaptic depression when compared to euploid littermate controls. To determine whether memantine could rescue this phenotype in the Ts65Dn mice, half of the hippocampal slices were treated with memantine before Chem‐LTD treatment. We found that memantine‐treated hippocampal slices from Ts65Dn mice presented control levels of Chem‐LTD. Currently, phosphorylation of the AMPA receptor GluR1 subunit is being assessed on hippocampal slices from Ts65Dn and control mice, which were flash frozen under control and Chem‐LTD conditions, with or without drug. This study confirms our previous finding that molecular pathways coupled to the gating of NMDA receptors are altered in the Ts65Dn mouse.

Ultrastructural analysis of the glutamatergic system in the outer plexiform layer of zebrafish retina

l-Glutamate, the photoreceptor neurotransmitter, depolarizes horizontal cells and OFF-bipolar cells by ionotropic receptors and hyperpolarizes ON-bipolar cells by metabotropic receptors. Despite extensive light microscopy on the distribution of glutamate receptors in zebrafish retina, there are little ultrastructural data. Given the importance of zebrafish in studies on the genetic manipulation of retinal development and function, precise data on the synaptic neurochemical organization of the zebrafish retina is needed. Immunohistochemical techniques were used to determine the ultrastructural localization of glutamate receptor subunits GluR2, GluR4, NMDA2B (NR2B) and mGluR1α in zebrafish outer plexiform layer (OPL). These antibodies were chosen because of an apparent conservation of localization of GluR2, GluR4 and mGluR1α in the vertebrate OPL, while there is some support for NMDA receptors in the OPL. GluR2-immunoreactivity (IR) was in all horizontal cell dendrites that invaginated cone pedicles and rod spherules. Three arrangements of dendrites contained GluR-IR in rod spherules: classical-type with GluR2-IR on lateral horizontal cell dendrites, a butterfly-shaped horizontal cell dendrite, and a goblet-shaped dendrite, likely of bipolar cell origin. GluR4-IR was restricted to dendrites of OFF-bipolar cells that innervated rod and cone terminals. NR2B-IR was restricted to a subtype of cone ON-bipolar cell. mGluR1α-IR was restricted to ON mixed rod/cone (Mb) bipolar cells whose dendrites innervated rod and cone synaptic terminals. The presence of mGluR1α on Mb bipolar cell dendrites is consistent with a role in retrograde endocannabinoid suppression. The subunit composition of glutamate receptors should affect the kinetics and pharmacology of these cells to glutamate receptor activation.

Activation of Group I Metabotropic Glutamate Receptors Increases Serine Phosphorylation of GluR1 α-Amino-3-hydroxy-5-methylisoxazole-4-propionic Acid Receptors in the Rat Dorsal Striatum

Protein phosphorylation is an important mechanism for the post-translational modulation of ionotropic glutamate receptors. In this study, we investigated the regulation of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor GluR1 subunit phosphorylation by the stimulation of group I metabotropic glutamate receptors (mGluRs) in the rat dorsal striatum in vivo. Stimulation of group I mGluRs was found to increase GluR1 phosphorylation of Ser831 and Ser845 in phospholipase C (PLC)-coupled Ca(2+) cascades. Interactions of protein kinases activated by intracellular Ca(2+) release downstream to PLC modulate the phosphorylation state of GluR1 on Ser831 and Ser845: phosphorylation of GluR1 on Ser831 is up-regulated by the protein kinase C and calcium-calmodulin-dependent protein kinase (CaMK)/c-Jun N-terminal kinase (JNK) pathways, whereas phosphorylation of GluR1 on Ser845 is up-regulated by the protein kinase A (PKA), PKA/ERK1/2, and PKA/JNK pathways. The phosphorylation state of GluR1 on Ser831 and Ser845 and the activity of protein kinases are further regulated by protein phosphatases. These data suggest that GluR1 phosphorylation of Ser831 and Ser845 via stimulation of group I mGluRs is regulated by the interactions of PLC-coupled protein kinases and protein phosphatases in the dorsal striatum.

Chromatin structure and expression of the AMPA receptor subunit GluR2 in human glioma cells: role of REST and Sp1

Chromatin structure and expression of the AMPA receptor subunit GluR2 in human glioma cells: role of REST and Sp1

Chromatin structure and expression of the AMPA receptor subunit GluR2 in human glioma cells and the role of REST

The AMPA receptors are postsynaptic ion channels that open following stimulation with glutamate. The regulatory region of the gene encoding the GluR2 subtype of AMPA receptors contains a binding site for the transcriptional repressor REST. The cell-type specific microenvironment, in particular the cell type-specific structure of the chromatin, is crucial for the ability of REST to control target gene transcription. Using antibodies directed against methylated lysine residues 4 or 9 (H3K4 or H3K9) of histone H3, we show that the GluR2 gene has an open chromatin configuration in human U87MG glioma cells, with nucleosomes carrying di-and trimethylated H3K4. In contrast, the GluR2 gene is embedded into a repressed chromatin environment in non-neuronal hepatoma cells and keratinocytes. Chromatin immunoprecipitation experiments revealed binding of REST and histone deacetylase-1 to the GluR2 gene under physiological conditions. While overexpression of REST reduced GluR2 mRNA levels, expression of a mutant of REST that contained a transcriptional activation domain enhanced GluR2 gene transcription in U87MG glioma cells. Treatment of the cells with the histone deacetylase inhibitor trichostatin A (TSA) induced an upregulation of GluR2 expression, indicating that the transcription of the GluR2 gene is dependent of the balance between histone acetylation and deactylation. Transcription of the GluR2 gene was also inducible by TSA following inhibition of Sp1 target gene transcription, indicating that the inhibition of Sp1 is not required for the upregulation of GluR2 expression following histone deacetylase inhibition.

The function of glutamatergic synapses is not perturbed by severe knockdown of 4.1N and 4.1G expression

AMPA-type glutamate receptors mediate fast excitatory synaptic transmission in the vertebrate brain. Their surface expression at synapses between neurons is regulated in an activity-dependent and activity-independent manner. The protein machinery that regulates synaptic targeting, anchoring and turnover of AMPA receptors consists of several types of specialized scaffolding proteins. The FERM domain scaffolding proteins 4.1G and 4.1N were previously suggested to act jointly in binding and regulating synaptic trafficking of the AMPA receptor subunits GluR1 and GluR4. To determine the functions of 4.1G and 4.1N in vivo, we generated a mutant mouse line that lacks 4.1G entirely and expresses 4.1N at 22% of wild-type levels. These mice had combined 4.1G and 4.1N protein expression in the hippocampus at 12% of wild-type levels (equivalent to 8-10% of combined GluR1 and GluR4 expression levels). They show a moderate reduction in synaptosomal expression levels of the AMPA receptor subunit GluR1 at 3 weeks of age, but no change in basic glutamatergic synaptic transmission and long-term potentiation in the hippocampus. Our study indicates that 4.1G and 4.1N do not have a crucial role in glutamatergic synaptic transmission and the induction and maintenance of long-term plastic changes in synaptic efficacy.

Glutamate and reinstatement

The importance of glutamate in the reinstatement of cocaine-seeking behavior has been established. New molecular and neurochemical adaptations in the glutamatergic system which drive cocaine relapse have been identified, such as the ability of CB1 receptor stimulation to reduce basal glutamate levels and the involvement of the GluR1 receptor subunit in reinstatement. Furthermore, it is apparent that similar glutamatergic neuroadaptations arise after self-administration of cocaine, heroin, nicotine, and alcohol. For example, reinstatement to cocaine, nicotine, and alcohol can be prevented both by the stimulation of group II mGluR receptors and by the blockade of group I mGluR receptors. The similarities in the neurochemistry behind relapse to these varied drug classes indicate that drugs that target the glutamate system could be effective at treating relapse to multiple types of drugs.

Semaphorin3A drives AMPA receptor subunit GluR2 to dendrites

Semaphorin3A drives AMPA receptor subunit GluR2 to dendrites

Epac mediates PACAP‐dependent long‐term depression in the hippocampus

Extensive work has shown that activation of the cAMP-dependent protein kinase A (PKA) is crucial for long-term depression (LTD) of synaptic transmission in the hippocampus, a phenomenon that is thought to be involved in memory formation. Here we studied the role of an alternative target of cAMP, the exchange protein factor directly activated by cyclic AMP (Epac). We show that pharmacological activation of Epac by the selective agonist 8-(4-chlorophenylthio)-2'-O-methyl-cAMP (8-pCPT) induces LTD in the CA1 region. Paired-pulse facilitation of synaptic responses remained unchanged after induction of this LTD, suggesting that it depended on postsynaptic mechanisms. The 8-pCPT-induced LTD was blocked by the Epac signalling inhibitor brefeldin-A (BFA), Rap-1 antagonist geranylgeranyltransferase inhibitor (GGTI) and p38 mitogen activated protein kinase (P38-MAPK) inhibitor SB203580. This indicated a direct involvement of Epac in this form of LTD. As for other forms of LTD, a mimetic peptide of the PSD-95/Disc-large/ZO-1 homology (PDZ) ligand motif of the AMPA receptor subunit GluR2 blocked the Epac-LTD, suggesting involvement of PDZ protein interaction. The Epac-LTD also depended on mobilization of intracellular Ca(2+), proteasome activity and mRNA translation, but not transcription, as it was inhibited by thapsigargin, lactacystin and anisomycin, but not actinomycin-D, respectively. Finally, we found that the pituitary adenylate cyclase activating polypeptide (PACAP) can induce an LTD that was mutually occluded by the Epac-LTD and blocked by BFA or SB203580, suggesting that the Epac-LTD could be mobilized by stimulation of PACAP receptors. Altogether these results provided evidence for a new form of hippocampal LTD.

Modification of ionotropic glutamate receptor–mediated processes in the rat hippocampus following repeated, brief seizures

The seizure-induced molecular and functional alterations of glutamatergic transmission in the hippocampus have been investigated. Daily repeated epileptic seizures were induced for 12 days by intraperitoneal administration of 4-aminopyridine (4-AP; 4.5 mg/kg) in adult Wistar rats. The seizure symptoms were evaluated on the Racine's scale. One day after the last injection, the brains were removed for in vitro electrophysiological experiments and immunohistochemical analysis. The glutamate receptor subunits NR1, NR2A, NR2B, GluR1, GluR1flop, GluR2, and KA-2 were studied using the histoblotting method. The semi-quantitative analysis of subunit immunoreactivities in hippocampal layers was performed with densitometry. In the hippocampus, increase of GluR1, GluR1flop and NR2B immunostaining was observed in most of the areas and layers. The significant decrease of GluR2 staining intensity was observed in the CA1 and dentate gyrus. Calcium permeability of hippocampal neurons was tested by a cobalt uptake assay in hippocampal slices. The uptake of cobalt increased in the CA1 area and dentate gyrus, but not in the CA3 region following 4-AP treatment. Effects of AMPA and NMDA (N-methyl-d-aspartate) glutamate receptor antagonists (1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride (GYKI 52466) and D-APV respectively) were measured in hippocampal slices using extracellular recording. Analysis of the population spikes revealed the reduced effectiveness of the AMPA receptor antagonist GYKI 52466, while the effect of the NMDA receptor antagonist d-(2R)-amino-5-phosphonovaleric acid was similar to controls. The results demonstrated that repeated convulsions induced structural and functional changes in AMPA receptor–mediated transmission, while NMDA and kainate receptor systems displayed only alterations in receptor subunit composition.

Extinction of morphine‐dependent conditioned behavior is associated with increased phosphorylation of the GluR1 subunit of AMPA receptors at hippocampal synapses

In abstinent opiate addicts, relapse can be triggered by exposure to environmental cues associated with drug use; thus, the disruption of these learned associations may be an effective approach for reducing relapse. Interestingly, glutamatergic systems are thought to be involved in opiate-induced behavioral plasticity. In this study, changes in expression and phosphorylation levels of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor subunits (GluR1, GluR2) in the hippocampus were investigated in rats showing a conditioned response (CR) to an opiate-paired environment as well as in animals in which this conditioned behavior was extinguished. Additionally, another set of animals went through a drug-unpaired paradigm (without conditioning) in order to examine the effects of the pharmacology of the drug itself. Subcellular fractionation techniques were used to analyse the local distribution of AMPA glutamate subunits within the synapse, especially at the postsynaptic density (PSD). Results showed that morphine-dependent CRs did not alter expression or redistribution of GluR1 or GluR2; however, the unpaired administration of morphine resulted in an increase in the phosphorylation of the GluR1 subunit at extrasynaptic sites. Interestingly, the extinction of the CR significantly increased phosphorylation of the GluR1 subunit at the PSD. Therefore we propose that, within the synapse, the phosphorylation of the GluR1 subunit at the PSD may be a key mechanism in the extinction of opiate-associated CRs.

Spinal anterior horn cells in sporadic amyotrophic lateral sclerosis show ribosomal detachment from, and cisternal distention of the rough endoplasmic reticulum

Sporadic amyotrophic lateral sclerosis (ALS) is a progressive and invariably fatal disease involving the upper and lower motor neurones of adult humans. Among the neuropathological features of the disease, abnormalities in the protein-synthesizing system in motor neurones of the brainstem and spinal cord, such as a decrease of cytoplasmic RNA and rough endoplasmic reticulum (rER) (chromatolysis), defective editing of the Q/R site of the glutamate receptor subunit GluR2 mRNA, fragmentation of the Golgi apparatus and accumulation of ubiquitinated inclusions and abnormal TdP-43 protein have been reported to be essential for the degeneration. In relation to these features, although the possibility of ER stress has been reported in motor neurones of the brainstem and spinal cord of ALS patients, the rER itself has not been a main target of ultrastructural investigation. The present study examined the rER, ultrastructurally and quantitatively in the spinal anterior horn cells (AHCs) of 21 Japanese patients with sporadic ALS and eight Japanese control subjects. It was found that: (i) the rER cisternae in AHCs showing central chromatolysis were fragmented, but retained their width and had normally attached ribosomes, and (ii) the rER cisternae in shrunken AHCs were irregularly distended with detachment of the ribosomes, thus suggesting that (iii) ribosomal detachment was related to rER distention.

Chronic fluoxetine treatment increases expression of synaptic proteins in the hippocampus of the ovariectomized rat: Role of BDNF signalling

Antidepressant drugs have been suggested to regulate synaptic transmission and structure. We hypothesised that antidepressant-induced changes in synapses and their associated proteins might become more apparent if they were measured under conditions of reduced synapse density. Therefore, in the present study, we examined whether chronic treatment with the antidepressant, fluoxetine alters expression of synaptic proteins in the hippocampus of rodents that underwent ovariectomy, a procedure which reportedly decreases synapse density in the CA1 region of the rat hippocampus. Using Western blotting, we measured changes in hippocampal expression of proteins associated with synapse structure, strength and activity namely, postsynaptic density protein 95 (PSD-95), the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPA-R) subunit GluR1 and phosphosynapsin (Ser9), respectively. We found that fluoxetine treatment increased expression of phosphosynapsin, PSD-95 and synaptic GluR1 (but not total GluR1) in the hippocampus of ovariectomized but not sham rats. Since BDNF and signalling at its receptor, TrkB, can mediate behavioural responses to antidepressants and induce neuronal plasticity, we investigated the contribution of TrkB signalling to fluoxetine-induced changes in synaptic protein expression by using a transgenic mouse model overexpressing a truncated form of the TrkB receptor (TrkB.T1). Fluoxetine produced a small but significant increase in hippocampal PSD-95 in ovariectomized wildtype mice but not in ovariectomized TrkB.T1 mice or sham mice. In contrast to rats, fluoxetine did not alter expression of synaptic GluR1 and did not reverse ovariectomy-induced decreases in hippocampal phosphosynapsin in either genotype. Taken together, these results suggest that chronic fluoxetine treatment can increase synaptic protein expression in the hippocampus and at least some of these effects require TrkB signalling. Moreover, these effects were only observed in ovariectomized animals, thus suggesting that fluoxetine-induced increases in synaptic protein levels might only occur or become detectable when hippocampal synaptic connectivity is perturbed.

The changes of AMPA receptors subunit GluR2 and mEPSCs in visual cortex of monocular deprivation rats

To observe the change of expression in quantity of AMPA receptor subunit GluR2 and mEPSCs from monocular derivated rats. The preparative work was conducted for the further study on mechanism involved in regulation and control of the visual development plasticity. It was an experimental study. Forty 14 d healthy Wistar rats were divided into two groups randomly; twenty in each group which were divided into two groups (I, II) and derivated monocular in experiment group (group I), and then were killed at a week later. ForIA and IIA 5 microm thick section was cut from tissue of visual cortex in coronary side, then the expression of GluR2 was observed by microscope and image analyze of GluR2 was conducted after immunohistochemical staining; for IB and IIB, mEPSCs was investigated which was used Voltage-clamp technique with whole-cell mode. The level of GluR2 in visual cortex of normal rats was higher than experiment group. The mean amplitude and frequency of mEPSCs increased in the derivated monocular group. The level of GluR2 has changed association with the visual stimulation, which suggests that GluR2 should have the important physiological function on the postnatal development of visual system of rats.

Long Term Synaptic Depression That Is Associated with GluR1 Dephosphorylation but Not α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid (AMPA) Receptor Internalization

Long lasting changes in the strength of synaptic transmission in the hippocampus are thought to underlie certain forms of learning and memory. Accordingly, the molecular mechanisms that account for these changes are heavily studied. Postsynaptically, changes in synaptic strength can occur by altering the amount of neurotransmitter receptors at the synapse or by altering the functional properties of synaptic receptors. In this study, we examined the biochemical changes produced following chemically induced long term depression in acute hippocampal CA1 minislices. Using three independent methods, we found that this treatment did not lead to an internalization of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. Furthermore, when the plasma membrane was separated into synaptic membrane-enriched and extrasynaptic membrane-enriched fractions, we actually observed a significant increase in the concentration of AMPA receptors at the synapse. However, phosphorylation of Ser-845 on the AMPA receptor subunit GluR1 was significantly decreased throughout the neuron, including in the synaptic membrane-enriched fraction. In addition, phosphorylation of Ser-831 on GluR1 was decreased specifically in the synaptic membrane-enriched fraction. Phosphorylation of these residues has been demonstrated to control AMPA receptor function. From these data, we conclude that the decrease in synaptic strength is likely the result of a change in the functional properties of AMPA receptors at the synapse and not a decrease in the amount of synaptic receptors.

Increased AMPA receptor GluR1 subunit incorporation in rat hippocampal CA1 synapses during benzodiazepine withdrawal

Prolonged benzodiazepine treatment leads to tolerance and increases the risk of dependence. Flurazepam (FZP) withdrawal is associated with increased anxiety correlated with increased alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptor (AMPAR)-mediated synaptic function and AMPAR binding in CA1 pyramidal neurons. Enhanced AMPAR synaptic strength is also associated with a shift toward inward rectification of synaptic currents and increased expression of GluR1, but not GluR2, subunits, suggesting augmented membrane incorporation of GluR1-containing, GluR2-lacking AMPARs. To test this hypothesis, the postsynaptic incorporation of GluR1 and GluR2 subunits in CA1 neurons after FZP withdrawal was examined by postembedding immunogold quantitative electron microscopy. The percentage of GluR1 positively labeled stratum radiatum (SR) synapses was significantly increased in FZP-withdrawn rats (88.2% +/- 2.2%) compared with controls (74.4% +/- 1.9%). In addition, GluR1 immunogold density was significantly increased by 30% in SR synapses in CA1 neurons from FZP-withdrawn rats compared with control rats (FZP: 14.1 +/- 0.3 gold particles/mum; CON: 10.8 +/- 0.4 gold particles/mum). In contrast, GluR2 immunogold density was not significantly different between groups. Taken together with recent functional data from our laboratory, the current study suggests that the enhanced glutamatergic strength at CA1 neuron synapses during benzodiazepine withdrawal is mediated by increased incorporation of GluR1-containing AMPARs. Mechanisms underlying synaptic plasticity in this model of drug dependence are therefore fundamentally similar to those that operate during activity-dependent plasticity.

C-Type natriuretic peptide modulates pre- and postsynaptic properties in hippocampal area CA1 in vitro

The cGMP producing natriuretic peptide receptor B (NPR-B) and its ligand C-type natriuretic peptide (CNP) are widely distributed in the brain and are highly expressed in the hippocampal regions CA1–CA3. To date only limited functional data is available concerning the physiological effects of the peptide hormone in the hippocampus. Therefore, we were interested in how bath application of the peptide hormone might influence synaptic plasticity following high frequency stimulation (HFS). We found that CNP application decreased the population spike (PS) amplitude after HFS, thereby affecting long-term potentiation (LTP) in acute hippocampal slices. To investigate the molecular consequences of CNP application leading to a decrease in PS amplitude, we further analyzed the impact of the hormone on the number of presynaptic synapsin I clusters and number of postsynaptic AMPA receptor subunit GluR1 clusters as well as their co-localization in a primary hippocampal cell culture system. The observed pre-and postsynaptic effects after CNP stimulation of the cGMP pathway in hippocampal cell cultures may underlie the effect of the peptide hormone on LTP.

DECREASED AMPA GLuR2, BUT NOT GLuR3, mRNA EXPRESSION IN RAT AMYGDALA AND DORSAL HIPPOCAMPUS FOLLOWING MORPHINE‐INDUCED BEHAVIOURAL SENSITIZATION

1. Repeated administration of psychostimulants and micro-opioid receptor agonists elicits a progressive enhancement of drug-induced behavioural responses, a phenomenon termed behavioural sensitization. These changes in behaviour may reflect plastic changes requiring regulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) receptor function. 2. In the present study, rats were treated for 7 days with saline or morphine (10 mg/kg). After a washout period of either 24 h or 7 days, locomotion, oral stereotypy and state-dependent memory in a passive avoidance test were measured in the presence or absence of 6-cyano-7-nitroquinoxaline-2,3-dione disodium salt (CNQX; 3 mg/kg), an AMPA receptor antagonist. In order to evaluate the mechanism underlying the behavioural responses, quantitative real-time reverse transcription-polymerase chain reaction was used to evaluate mRNA expression of the AMPA receptor subunits GluR2 and GluR3 in the striatum, prefrontal cortex, hippocampus, hypothalamus and amygdala of animals treated repeatedly with morphine. 3. The results indicate that repeated morphine treatment followed by 7 days (but not 24 h) washout produces behavioural sensitization, as determined by locomotion, oral stereotypy and state-dependent memory. Blockade of AMPA receptors with CNQX on the test day did not alter these behavioural responses. In addition, repeated morphine treatment followed by 7 days (but not 24 h) washout decreased GluR2 mRNA expression in both the amygdala (by 50%) and hippocampus (by 35%). Repeated morphine treatment did not alter GluR3 mRNA expression in any brain area assessed. 4. These data imply that AMPA receptors are involved in the development (but not expression) phase of behavioural sensitization. The decreases in GluR2 mRNA expression in the amygdala and hippocampus may result in the formation of calcium-permeable AMPA receptors, which are believed to play an important role in behavioural sensitization.