NMDA受体在正常大鼠颈动脉体的表达

应用免疫组织化学方法观察了离子型谷氨酸受体——NMDA受体的NR1及NR2A亚单位在正常成年雄性SD和Wistar大鼠颈动脉体的表达,并对阳性产物的表达强度进行了灰度分析及统计学处理。结果表明:正常成年SD大鼠及Wistar大鼠的颈动脉体内均存在NMDA NR1免疫反应阳性细胞,从阳性细胞的形态和分布特点判断这些阳性细胞为主细胞,两种大鼠之间无明显差异(P>0.05);而两种大鼠的颈动脉体内几乎不存在NMDA NR2A免疫反应阳性细胞。本实验的结果提示:在正常成年大鼠颈动脉体的主细胞上有NMDA受体的分布,并且该受体的二聚体构成不同于经典受体,这可能与谷氨酸在颈动脉体发挥的特殊功能有关。     

NR2B反义寡核苷酸对脑缺血海马CA1区NR2BmRNA表达的影响

为研究NMDA受体2B亚单位(NR2B)反义寡核苷酸(antisense oligonucleotide to NR2B,ANR2B)对短暂性脑缺血后海马CA1区NR2B mRNA表达的影响,分别向成年SD大鼠海马CA1区内立体定位注射ANR2B、NR2B正义寡核苷酸(SNR2B)、无菌生理盐水(NS),或者插针不注射(NSNO),24h后行四动脉阻断前脑缺血手术(缺血15min、再灌注24h),经心冲灌固定取脑,连续冰冻切片,原位杂交组织化学方法染色,光镜下观察各组每侧鼠脑NR2B mRNA的表达变化,并用LEICAQWin进行图像分析。结果显示,单纯缺血组海马各区的NR2B mRNA显色强度明显增加;缺血再灌组、假手术组和正常组海马CA1区内ANR2B注射点及其周围的NR2B mRNA显色明显下降;而在注射SNR2B、NS或NSNO的各组海马切片上,NR2B mRNA显色均无明显变化。结果表明ANR2B可以特异性地在体局部防止缺血后NR2B mRNA的高水平表达。     

Are glutamate receptors specifically implicated in some forms of memory processes?

Convergent data indicate that certain substances that interact with N-methyl-d-aspartate (NMDA) receptors or metabotropic glutamate receptors (mGluRs) do not affect acquisition processes per se, or retrieval, but interfere specifically with the formation of memory traces. This action differs widely in its amplitude and time-course according to the learning task used. We showed that systemic injection of the competitive NMDA receptor antagonists, γ-l-glutamyl-l-aspartate (γ-LGLA) and 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonate (CPP), or intracerebroventricular infusion of d-2-amino-5-phosphonovalerate (D-AP5), immediately following acquisition of a Y-maze avoidance learning task in mice, deeply impaired retention of the temporal component of the task (leaving the start alley within the first 5 s of a trial), which significantly improved in controls during the hours following acquisition. In contrast the same substances had no or only slight effects on retention of the discrimination component (choice of the correct alley), which did not improve over time in control animals. This retention deficit did not appear to be due to an action on acquisition, retrieval and/or forgetting processes, or to state-dependent effects. Moreover, γ-LGLA, CPP or AP5, when administered immediately after partial acquisition of a food-reinforced bar-press task, suppressed the spontaneous improvement in post-training performance observed in control mice 24 h after the training session. (R,S)-α-methyl-4-carboxyphenylglycine (MCPG), an antagonist of mGluRs, also suppressed the post-training performance increment and its effects were antagonized by the co-administration of trans-ACPD, an agonist of mGluRs. Post-training improvement of performance over time is thought to reflect an active and dynamic process, leading to the organization of memory traces. According to this hypothesis, our results suggest that synaptic plasticity mediated by NMDA receptors and/or mGluRs activation is involved in mechanisms underlying long-term consolidation of memory traces.     

Modification of Redox Sites of N-Methyl-D-Aspartate Receptors Affects Anoxia-Induced Changes in the Bioelectrical Activity of Rat Brain Olfactory Cortex Slices

Living slices of Wistar-Kyoto rat brain olfactory cortex were used to study the effects of the thiol-oxidizing agent 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB), which inhibits NMDA receptor activity, on changes in the generation of evoked focal potentials (NMDA and non-NMDA EPSP) in response to long-term and short-term anoxia, which induces functional damage and facilitates increases in the resistance of neurons to severe hypoxia respectively. These studies showed that DTNB (200 'M) efficiently prevented the suppression of focal EPSP generation due to long-term anoxia in most slices. In addition, DTNB partially reversed the protective effect of preconditioning with short-term anoxia on the impairment of focal EPSP generation induced by long-term anoxia. This affected the NMDA component of the EPSP to a greater extent than the non-NMDA component. The possible role of changes in the state of modulatory redox sites of NMDA receptors in the mechanisms of functional damage and increases in neuron resistance due to hypoxia is discussed.     

Early-life epileptiform discharges exert both rapid and long-lasting effects on AMPAR subunit composition and distribution in developing neurons

The perinatal period of brain is characterized by dynamic changes in structure and high propensity for epilepsy. Animal models have shown that alterations of AMPA receptor (AMPAR) assembly or function may be related to seizure-induced cell damage, long-lasting impairments in brain development and seizure threshold. However, effects of earlier epileptiform discharges on AMPAR composition and sub-cellular distribution remain understudied. In this study, we analyzed age-dependent variation of relative GluR1 and GluR2 protein levels in primary cultured rat cortical neurons at 7 DIV, 12 DIV, 17 DIV and 21 DIV. By inducing a single event of epileptiform activity at 6 DIV, we tested the effects of early-life seizure-like insults on AMPAR subunit distribution. We found a significant increase in synaptosomal membrane GluR1 expression in magnesium-free (MGF) medium-treated neurons at each time point detected (p < 0.05), while GluR2 expression increased at 7 DIV, and declined at 17 DIV and 21 DIV respectively (p < 0.05). That is, a trend of high GluR1 with much lower GluR2 expression on the surface membrane of epileptiform discharges experienced neurons over time in culture was presented. These findings in an in vitro model of early-life seizure may inform rodent models of epilepsy, as well as the cellular mechanism involved in epilepsy-associated brain dysfunction.     

NMDA receptor mediated dendritic plasticity in cortical cultures after oxygen-glucose deprivation

Dendrites and spines undergo dynamic changes in physiological and pathological conditions. Dendritic outgrowth has been observed in surviving neurons months after ischemia, which is associated with the functional compensation. It remains unclear how dendrites in surviving neurons are altered shortly after ischemia, which might reveal the mechanisms underlying neuronal survival. Using primary cortical cultures, we monitored the dendritic changes in individual neurons after oxygen-glucose deprivation (OGD). Two to four hours of OGD induced approximately 30–50% cell death in 24 h. However, the total dendritic length in surviving neurons was significantly increased after OGD with a peak at 6 h after re-oxygenation. The increase of dendritic length after OGD was mainly due to the sprouting rather than the extension of the dendrites. The dendritic outgrowth after 2 h of OGD was greater than that after 4 h of OGD. Application of NMDA receptor blocker MK-801 abolished OGD-induced dendritic outgrowth, whereas application of AMPA receptor antagonist CNQX had no significant effects. These results demonstrate a NMDA receptor-dependent dendritic plasticity shortly after OGD, which provides insights into the early response of surviving neurons after ischemia.     

5-HT mechanisms of median raphe nucleus in the conditioned freezing caused by light/foot-shock association

We have shown that 5-HT mechanisms of the median raphe nucleus (MRN) are involved in contextual fear-conditioning processes as electrolytic or neurotoxic lesions with N-methyl-D-aspartate (NMDA) or injections of 8-hydroxy-2-(di-n-propilamino)-tetralin (8-OH-DPAT) into this structure inhibit freezing behavior in a contextual fear paradigm. In this work, we extend these studies by analyzing the behavioral responses in a classical fear-conditioning paradigm (light or tone/foot-shock association) in rats with either neurochemical lesion with NMDA or injected with 8-OH-DPAT into the MRN. The animals received NMDA or 8-OH-DPAT or saline microinjections into the MRN and were submitted to conditioning trials in an experimental chamber, where they received 10 foot-shocks (0.6 mA, 1 s, variable interval between 10 and 50 s) paired with tone or light (CS). On the next day, they were tested in a different experimental chamber, with or without CS presentation, where the duration of freezing and the number of rearing episodes were recorded. Light or tone alone caused a significant amount of freezing. NMDA lesions or 8-OH-DPAT injections into the MRN clearly inhibited freezing behavior in rats conditioned to light/foot-shock association, but not in the conditioning sessions with tones. Besides the proposed role in contextual fear conditioning, these results clearly show that MRN is involved in the fear conditioning with light as conditioned stimuli. Distinct neural substrates seem to subserve conditioning fear with acoustic stimuli.     

The Involvement of Dopamine in Strengthening Cortical Signals Activating NMDA Receptors in the Striatum (a hypothetical mechanism)

A possible mechanism is proposed for the enhancement/weakening of those cortical signals in the cortex-basal ganglia-thalamus-cortex neural network which induce/do not induce opening of NMDA channels in the spiny neurons of the striatum and which can be regarded as strong/weak in terms of this measure. The mechanism is based on the modulatory influences of dopamine on changes in the efficiency of corticostriatal inputs. In the absence of dopamine, relative increases in the intensity of strong (weak) cortical signals can lead to the induction of long-term potentiation (depression) of corticostriatal synapses. In this case, because of the differently directed influences on thalamic cells of signals passing via strionigral and striopallidal cells, strong signals at the output of the thalamus are weakened, while weak signals are strengthened. Activation of dopamine D₁ (D₂) receptors on strionigral (striopallidal) neurons may facilitate increases in the extent of long-term potentiation/depression (decreases in the extent of long-term potentiation/depression or induction of long-term potentiation/depression). The consequence of this is that strong signals at the output of the thalamus can be strengthened synergistically, while weak signals cab be weakened synergistically. Background cortical signals evoking tonic release of dopamine in the striatum can decrease strengthening because of weakening of the modulatory influence of dopamine on the modification of corticostriatal synapses.     

Activity of the enantiomers of 2-amino-5-phosphono-valeric acid as stereospecific antagonists of excitatory aminoacids

The (+) and (?) enantiomers of 2-amino-5-phosphono-valeric acid have been separated and tested as antagonists of aminoacid excitation of neurones in rat cerebral cortex. The compounds were applied by microiontophoresis. The (?)-isomer was about 8–10 times more active than the racemate in blocking responses to N-methyl-d-aspartate, and was better able to distinguish between N-methyl-d-aspartate and glutamate.The results support the concept of a distinct population of receptors for N-methyl-d-aspartate.