Opposite modulation of cortical N-methyl-D-aspartate receptor-mediated responses by low and high concentrations of dopamine.

To examine whether dopamine modulates cortical N-methyl-D-aspartate receptor-mediated glutamate transmission, whole-cell recordings were made from identified pyramidal cells located in layers V and VI of the medial prefrontal cortex of the rat using a slice preparation. In the presence of tetrodotoxin and the absence of Mg2+, a brief local application of N-methyl-D-aspartate evoked an inward current which was blocked by the N-methyl-D-aspartate antagonist dizocilpine maleate but not affected by the non-N-methyl-D-aspartate antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)quinoxaline, suggesting that the observed current is mediated by N-methyl-D-aspartate receptors located on recorded cells. Bath application of dopamine produced opposite effects on the N-methyl-D-aspartate current depending on the concentrations of dopamine applied. At low concentrations (<50 microM), dopamine enhanced the N-methyl-D-aspartate current, whereas at higher concentrations, dopamine suppressed the current. The same concentrations of dopamine did not significantly affect the inward current induced by the non-N-methyl-D-aspartate agonist alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid. The enhancing effect of dopamine on the N-methyl-D-aspartate response was mimicked by the D1 agonist SKF38393 and blocked by the D1 antagonist SCH31966, whereas the suppressing effect was mimicked by the D2 agonist quinpirole and blocked by the D2 antagonist eticlopride. The above results suggest that dopamine at low concentrations acts preferentially on D1-like receptors to promote N-methyl-D-aspartate receptor-mediated transmission, while at high concentrations dopamine also activates D2-like receptors, leading to a suppression of the N-methyl-D-aspartate function. This differential modulation of N-methyl-D-aspartate function may have significant implications for understanding behaviors and disorders involving both cortical dopamine- and glutamate-mediated neurotransmission.     

Hippocampal N-methyl-D-aspartate receptor-mediated encoding and retrieval processes in spatial working memory: delay-interposed radial maze performance in rats

In order to clarify the role of hippocampal N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors in different stages of spatial working memory, we first assessed the rats' performance in a delay-interposed eight-arm radial maze task (experiment 1). When a delay was interposed after the first four correct choices, rats showed more errors in the second-half performance depending on the length of delay; however, they did not show any significant increase of error choices until the delay was beyond 2 h. We then tested the effect of 2-amino-5-phosphonopentanoic acid (AP5), a competitive NMDA receptor antagonist, and 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide (NBQX)-disodium, an AMPA receptor antagonist, on a standard (no delay-interposed) radial maze task (experiment 2). The drug effect was maintained 15-30 min but it completely disappeared 60 min after dorsal hippocampal microinjection. Based on these findings we finally investigated the effects of hippocampal AP5 and NBQX administered at different stages of 2 h delay-interposed radial maze task on the second-half performance (experiment 3). AP5 immediately before the first-half and before the second-half performance significantly impaired the correct choices, but the treatment immediately after the first-half performance did not, while NBQX impaired them in all three conditions. Results suggest that hippocampal NMDA receptors play an important role in encoding and retrieval processes of spatial working memory, while AMPA receptor activation is necessary not only in these processes but also in consolidation/retention process.     

Contributions of mGlu1 and mGlu5 receptors to interactions with N-methyl-D-aspartate receptor-mediated responses and nociceptive sensory responses of rat thalamic neurons

The nociceptive responses of rat ventrobasal thalamus neurons can be reduced by N-methyl-D-aspartate antagonists and by selective metabotropic glutamate receptor mGlu1 antagonists. The recent development of the mGlu5-selective antagonist 6-methyl-2-(phenylethynyl)-pyridine now allows the direct probing of the possible involvement of mGlu5 receptors in thalamic nociceptive responses. Extracellular recordings were made from single neurons in the ventrobasal thalamus and immediately overlying dorsal thalamic nuclei of adult urethane-anaesthetized rats using multi-barrel electrodes. Responses of neurons to iontophoretic applications of the mGlu5-selective agonist (R,S)-2-chloro-5-hydroxyphenylglycine were selectively reduced during continuous iontophoretic applications of 6-methyl-2-(phenylethynyl)-pyridine. Similar applications of 6-methyl-2-(phenylethynyl)-pyridine reduced neuronal responses to noxious thermal stimuli to 53+/-9.5% of control responses. Co- application by iontophoresis of N-methyl-D-aspartate and metabotropic glutamate receptor agonists resulted in a mutual potentiation of excitatory responses. This effect could be reduced by either 6-methyl-2-(phenylethynyl)-pyridine or the mGlu1 antagonist LY367385.These results, taken together with previous data, suggest that acute thalamic nociceptive responses are mediated by a combination of mGlu1, mGlu5 and N-methyl-D-aspartate receptor activation, and that co-activation of these receptors produces a synergistic excitatory effect. Thus blockade of any of these receptor types would have a profound effect on the overall nociceptive response.     

Retinal influences induce bidirectional changes in the kinetics of N-methyl-D-aspartate receptor-mediated responses in striate cortex cells during postnatal development

Development of the visual callosal projection in rodents goes through an early critical period, from postnatal day (P) 4 to P6, during which retinal input specifies the blueprint for normal topographic connections, and a subsequent period of progressive pathway maturation that is largely complete by the time the eyes open, around P13. This study tests the hypothesis that these developmental stages correlate with age-related changes in the kinetics of synaptic responses mediated by the N-methyl-D-aspartate subclass of glutamate receptors (NMDARs). We used an in vitro slice preparation to perform whole-cell recordings from retrogradely-labeled visual callosal cells, as well from cortical cells with unknown projections. We analyzed age-related changes in the decay time constant of evoked as well as spontaneous excitatory postsynaptic currents mediated by N-methyl-D-aspartate subclass of glutamate receptors (NMDAR-EPSCs) in slices from normal pups and pups enucleated at different postnatal ages. In normal pups we found that the decay time constant of NMDAR-EPSCs increases starting at about P6 and decreases by about P13. In contrast, these changes were not observed in rats enucleated at birth. However, by delaying the age at which enucleation was performed we found that the presence of the eyes until P6, but not until P4, is sufficient for inducing slow NMDAR-EPSC kinetics during the second postnatal week, as observed in normal pups. These results provide evidence that the eyes exert a bidirectional effect on the kinetics of NMDARs: during a P4-P6 critical period, retinal influences induce processes that slow down the kinetics of NMDAR-EPSCs, while, near the age of eye opening, retinal input induces a sudden acceleration of NMDAR-EPSC kinetics. These findings suggest that the retinally-driven processes that specify normal callosal topography during the P4-P6 time window also induce an increase in the decay time constant of NMDAR-EPSCs. This increase in response kinetics may play an important role in the maturation of cortical topographic maps after P6. Using ifenprodil, a noncompetitive NR2B-selective blocker, we obtained evidence that although NR1/NR2B diheteromeric receptors contribute to evoked synaptic responses in both normal and enucleated animals, they are not primarily responsible for either the age-related changes in the kinetics of NMDAR-mediated responses, or the effects that bilateral enucleation has on the kinetics of NMDAR-EPSCs.     

Complex influence of the L-type calcium-channel agonist BayK8644(+/-) on N-methyl-D-aspartate responses and neuronal survival

Past studies have implicated calcium influx through the N-methyl-D-aspartate class of ionotropic glutamate receptors as a key factor in excitotoxicity. Here, primary cultures of hippocampal neurons were exposed to N-methyl-D-aspartate with or without the L-type calcium channel agonist BayK8644(+/-). Calcium influxes were monitored with Fura-2 microfluorescent imaging and 45Ca measurements, and survival was assayed through cell counts. While 100 microM BayK8644 alone evoked a moderate elevation of intraneuronal calcium concentrations ([Ca2+]i), it dramatically attenuated the larger calcium influxes triggered by 500 microM N-methyl-D-aspartate. This attenuation was non-competitive and reversible; it was not inhibited by charybdotoxin or cyclosporin A. In spite of this attenuation of [Ca2+]i responses, 5-min exposures to BayK8644 produced much greater neurotoxicity 24 h later than did doses of N-methyl-D-aspartate evoking larger [Ca2+]i increases. This neurotoxicity was not observed with potassium-mediated depolarization or cobalt; indeed, both reversed the neurotoxicity of BayK8644. The relevant conclusions are two-fold: BayK8644 inhibits influx of calcium through a ligand-gated glutamate receptor, and BayK8644 exhibits considerable neurotoxicity. The former effect does not appear to depend upon the major metabolic pathways that modulate N-methyl-D-aspartate channels and thus may involve a direct allosteric interaction with the N-methyl-D-aspartate receptor. The toxicity of BayK8644 depends, at least partially, upon its activation of voltage-gated (cobalt-sensitive) calcium channels. However, the reversal of this toxicity by depolarization suggests that depolarization can be beneficial to neuronal survival through mechanisms other than calcium influx through voltage-gated calcium channels.     

Peripheral administration of an N-methyl-D-aspartate receptor antagonist (MK-801) changes dorsal horn neuronal responses in rats

Due to the discovery of peripheral N-methyl-D-aspartate (NMDA) receptors, the effects of peripherally administrated MK-801, a non-competitive NMDA receptor antagonist, and phosphate buffered saline were tested by using the response changes of wide-dynamic range cells in the lumbar enlargement of the spinal cord in Sprague-Dawley rats. MK-801 (1 microM, 50 microl) administered directly into the subcutaneous tissue of the receptive field (n = 7), produces a reversible reduction of responses to noxious and innocuous stimuli by a peripheral action. There was no change in the responses to cutaneous stimuli following injection of phosphate buffered saline (n = 7) or following administration of MK-801 into the contralateral foot (n = 7). The present study suggests that MK-801 produces a local anesthetic like effect in the peripheral tissue.     

ApoE genotype does not affect plasma tPA and PAI-1 antigen levels

The presence of one or two apoliprotein E4 (apoE4) alleles constitutes a major risk factor for Alzheimer's disease (AD) and coronary heart disease (CHD). Numerous observations have suggested that misregulation of proteases may be instrumental in both diseases. Tissue-type plasminogen activator (tPA) has been recently demonstrated to play a key role in neuronal plasticity and in experimental neurodegeneration. One receptor for the ApoE protein is the LRP/α2 macroglobulin receptor, which also binds to and endocytoses tPA and plasminogen activator inhibitor I (PAI-1). Here we tested whether the apoE genotype has an influence on the plasma levels of these proteins. We demonstrate that there is no difference in plasma levels of tPA- and PAI-1-antigens between middled-aged individuals with one apoE4 allele and those having none. This suggests that the impact of apoE4 on Alzheimer's disease is not the result of altered clearance of tPA or PAI-1 by the LRP receptor. Am. J. Med. Genet. 74:172–175, 1997. © 1997 Wiley-Liss, Inc.     

Magnesium removal induces paroxysmal neuronal firing and NMDA receptor-mediated neuronal degeneration in cortical cultures

Removal of extracellular Mg2+ triggered the onset of repetitive excitatory discharges in cultured murine cortical neurons, detected by recording with patch electrodes in the whole cell configuration. The discharges were suppressed by 100 microM D-2-amino-5-phosphonovalerate. Over the next 24-72 h substantial numbers of neurons, but not glia, degenerated, releasing lactate dehydrogenase to the bathing medium. The neuronal death induced by removal of extracellular Mg2+ could be attenuated by either 3 microM tetrodotoxin or 50 microM dextrorphan, and thus likely reflects excessive activation of N-methyl-D-aspartate receptors triggered by excitatory discharges. This Mg2+ removal model may be a useful model in which to study certain aspects of epileptic neocortical injury.     

Leptin enhances NR2B-mediated N-methyl-D-aspartate responses via a mitogen-activated protein kinase-dependent process in cerebellar granule cells

It is well documented that the hormone leptin regulates energy balance via its actions in the hypothalamus. However, evidence is accumulating that leptin plays a key role in numerous CNS functions. Indeed, leptin receptors are expressed in many extrahypothalamic brain regions, with high levels found in the hippocampus and cerebellum. In the hippocampus leptin has been shown to facilitate N-methyl-D-aspartate receptor function and modulate synaptic plasticity. A role for leptin in cerebellar function is also indicated as leptin-deficient rodents display reduced mobility that is unrelated to obesity. Here we show that leptin receptor immunolabeling can be detected in cultured cerebellar granule cells, being expressed at the somatic plasma membrane and also concentrated at synapses. Furthermore, leptin facilitated NR2B N-methyl-D-aspartate receptor-mediated Ca2+ influx in cerebellar granule cells via a mitogen-activated protein kinase-dependent pathway. These findings provide the first direct evidence for a cellular action of leptin in cerebellar neurons. In addition, given that N-methyl-D-aspartate receptor activity in the cerebellum is crucial for normal locomotor function, these data also have important implications for the potential role of leptin in the control of movement.     

Chronic nicotine exposure reduces N-methyl-D-aspartate receptor-mediated damage in the hippocampus without altering calcium accumulation or extrusion: evidence of calbindin-D28K overexpression

Neuronal accumulation of excess Ca2+ has been implicated in cellular death following several forms of physical and chemotoxic insult. Recent studies have suggested that exposure to agonists at brain nicotinic acetylcholine receptors reduces cytotoxic consequences of increased intracellular Ca2+ following some insults. In the present study, the ability of chronic exposure to (-)-nicotine to reduce cytotoxicity and attenuate increases in intracellular Ca2+ caused by exposure to N-methyl-D-aspartate were examined in organotypic cultures of rat hippocampus. Cultures were exposed to nicotine (0.1-10.0 microM) for five days prior to excitotoxic insult with N-methyl-D-aspartate. Exposure to N-methyl-D-aspartate produced concentration-dependent increases in both accumulation of 45Ca and in early and delayed cell death in the CA1, CA3 and dentate gyrus regions of cultures. The CA1 region of the hippocampus displayed the greatest sensitivity to cytotoxic effects of N-methyl-D-aspartate exposure; however, this regional difference was not associated with increased accumulation of 45Ca. Prior exposure to nicotine markedly attenuated N-methyl-D-aspartate-induced early and delayed cell death in each hippocampal region at concentrations as low as 0.1microM. However, nicotine did not alter the initial N-methyl-D-aspartate-stimulated influx of 45Ca or enhance extrusion of accumulated 45Ca measured at several time-points after insult. Five days of exposure to nicotine markedly increased immunoreactivity of the Ca2+ binding protein calbindin-D28K in each region of hippocampal cultures, effects reduced by mecamylamine co-exposure.These findings suggest that the potent protective effects of chronic nicotine exposure against neuronal overexcitation are not likely attributable to attenuations of Ca2+ accumulation, but are likely related to increased buffering of accumulated Ca2+.     

Selective neuronal vulnerability and the distribution of N-methyl-D-aspartate (NMDA) receptors

N-Methyl-D-Aspartate (NMDA) receptors are believed to play a critical role in excitotoxic cell death in the CNS. The distribution of NMDA-preferring binding sites is compared here with the patterns of selective neuronal death observed in Alzheimer's disease and following transient ischemia. The distribution of NMDA receptors, by itself, is unable to account for the characteristic patterns of selective neuronal vulnerability observed in conjunction with these types of neuropathology.     

The mechanisms of neuronal death produced by mitochondrial toxin 3-nitropropionic acid: the roles of N-methyl-D-aspartate glutamate receptors and mitochondrial calcium overload

Previous studies showed that 3-nitropropionic acid, an irreversible inhibitor of succinate dehydrogenase, produced neuronal death secondary to perturbed intracellular calcium homeostasis. However, the response of intramitochondrial calcium ([Ca(2+)](m)) to 3-nitropropionic acid remains unknown. In this study, we investigated the roles of and relationships among [Ca(2+)](m) overload, mitochondrial reactive oxygen species, and mitochondrial membrane depolarization in 3-nitropropionic acid-induced neuronal death. Following 1 mM 3-nitropropionic acid treatment on primary rat neuronal cultures, there was a gradual increase of [Ca(2+)](m) beginning at 2-4 h post 3-nitropropionic acid application, and a twofold increase of mitochondrial reactive oxygen species at 4 h. These were followed by mitochondrial membrane depolarization at 6-8 h post-treatment. By inhibiting [Ca(2+)](m) uptake, Ruthenium Red attenuated the production of reactive oxygen species, and prevented the 3-nitropropionic acid-induced mitochondrial membrane depolarization and 70% of apoptotic neuronal death (P<0.001). Inhibition of caspase activation attenuated the elevation of [Ca(2+)](m) (P<0.001), indicating that caspase activation plays a role in the elevation of [Ca(2+)](m). MK-801, an antagonist of N-methyl-D-aspartate (NMDA) glutamate receptors, prevented 3-nitropropionic acid-induced [Ca(2+)](m) elevation, caspase-3 activation, mitochondrial depolarization, and neuronal death.We conclude that the activation of NMDA glutamate receptor contributes to mitochondrial alterations induced by 3-nitropropionic acid. Inhibition of its activation and [Ca(2+)](m) overload with subsequent mitochondrial membrane depolarization can therefore attenuate the neuronal death induced by 3-nitropropionic acid.     

分子开关Rho族GTP酶与神经突起生长

生长端是神经唯一的生长点 ,它将寻址 ,选择靶位 ,最后形成突触。生长端如何正确破译导向信号、产生形状及运动变化 ,一直是神经科学的研究焦点。本文重点对分子开关 Rho族 GTP酶在神经发育中的调节作用及其 Rho族 GTP酶的上、下游信号联系机制等进行综述     

Evolution of neuronal and glial tau isoforms in chronic traumatic encephalopathy

Chronic traumatic encephalopathy (CTE) is a neurodegenerative tauopathy characterized by accumulation of hyperphosphorylated tau (p‐tau) in perivascular aggregates in neurons and glia at the depths of neocortical sulci and progresses to diffuse neocortical, allocortical and brainstem structures. The strongest risk factor is exposure to repetitive head impacts acquired most commonly through contact sports and military service. Given that CTE can only be definitively diagnosed after death, a better understanding of the cellular and molecular changes in CTE brains may lead to identification of mechanisms that could be used for novel biomarkers, monitoring progression or therapeutic development. Disruption of alternative pre‐mRNA splicing of tau mRNA plays a pathogenic role in tauopathy, with multiple characteristic patterns of isoform accumulation varying among tauopathies. Limited data are available on CTE, particularly at early stages. Using biochemical and histological approaches, we performed a detailed characterization of tau isoform signatures in post‐mortem human brain tissue from individuals with a range of CTE stages (n = 99). In immunoblot analyses, severity was associated with decreased total monomeric tau and increased total oligomeric tau. Immunoblot with isoform‐specific antisera revealed that oligomeric tau with three and four microtubule binding domain repeats (3R and 4R) also increased with CTE severity. Similarly, immunohistochemical studies revealed p‐tau accumulation consisting of both 3R and 4R in perivascular lesions. When the ratio of 4R:3R was analyzed, there was mixed expression throughout CTE stages, although 4R predominated in early CTE stages (I‐II), a 3R shift was observed in later stages (III‐IV). While neurons were found to contain both 3R and 4R, astrocytes only contained 4R. These 4R‐positive cells were exclusively neuronal at early stages. Overall, these findings demonstrate that CTE is a mixed 4R/3R tauopathy. Furthermore, histologic analysis reveals a progressive shift in tau isoforms that correlates with CTE stage and extent of neuronal pathology.     

Upregulation of purinergic P2Y receptor-mediated calcium responses in glial cells during experimental detachment of the rabbit retina

To investigate injury-induced alterations of purinergic P2Y receptor-mediated calcium responses in glial (Müller) cells of the rabbit retina, neural retinae were experimentally detached from the pigment epithelium. The ATP-evoked calcium responses were recorded in the endfeet of glial cells at the vitread surface of retinal wholemounts. In control retinae, approximately 7% of the glial cells investigated showed ATP-evoked calcium responses. Within 24 h of detachment, significantly more retinal glial cells (42%) showed calcium responses, and glial ATP responsiveness increased further in retinae which were detached for 48 (44%) or for 72 h (64%). The results indicate that in the detached retina, glial cells upregulate their responsiveness to extracellular ATP within 24 h of injury. Thus, P2Y receptor-mediated signalling may be involved in the early steps of glial response to retinal injury.     

Brain-derived neurotrophic factor attenuates mouse cerebellar granule cell GABAA receptor-mediated responses via postsynaptic mechanisms

In addition to exerting long-term neurotrophic influences on developmental process such as neuronal survival and neuritic outgrowth, brain-derived neurotrophic factor (BDNF) has been reported to modulate synaptic transmission in the short-term. Considerable evidence indicates that BDNF acutely modulates NMDA receptor-mediated synaptic activity. However, whether BDNF modulates inhibitory synaptic transmission remains to be firmly established. In the present study, we examined the effect of acute BDNF exposure on GABA-evoked whole-cell responses as well as GABAergic synaptic activity in cultured mouse cerebellar granule cells. GABA-evoked responses were reduced by 39.5 ± 4.7 % upon acute and focal application of BDNF (100 ng ml⁻¹). The reduction of the GABA response recovered only partially even minutes after removal of BDNF. TrkB-IgG and K252a, but not K252b, prevented the BDNF-induced attenuation of the GABA response. BDNF exposure shifted the cumulative peak amplitude distribution leftward for both spontaneous IPSCs (sIPSCs) and miniature IPSCs (mIPSCs) without affecting the rise time and decay time constants. Acute exposure to BDNF also resulted in internalization of GABA_A receptors in cultured cerebellar granule cells, as reflected by diminished immunostaining with an antibody against the GABA_A receptor β_2/3 subunit. Although the BDNF-induced GABA_A receptor internalization was sensitive to K252a, it did not become manifest until 5 min after exposure to BDNF. Therefore, receptor internalization alone cannot account for the prompt BDNF-induced attenuation of GABA-mediated activity. We conclude that BDNF modulates GABA_A receptor-mediated activity through TrkB receptor signalling that triggers a kinase-dependent short latency effect and a delayed longer latency effect hallmarked by receptor internalization.     

The effects of temperature and scopolamine on N-methyl-D-aspartate antagonist-induced neuronal necrosis in the rat

The effects of temperature and scopolamine on dizocilpine maleate-induced neuronal necrosis in the rat cingulate/retrosplenial cortex, entorhinal/olfactory cortices and the dentate gyrus were studied. Mild, protracted hypothermia (48 h at a brain temperature of 34 degrees C), induced by a servo-controlled "exposure technique" in the awake female rat, significantly reduced dizocilpine maleate (5.0 mg/kg, i.p.)-induced neuronal death in the cingulate/retrosplenial and entorhinal/olfactory cortices seven days following drug administration. Scopolamine (0.25 mg/kg, i.p.), putatively neuroprotective [Olney J. W. et al. (1991) Science 254, 1515-1518], did not reduce injury in the cingulate/retrosplenial cortex of female rats following one injection, but did following two and three doses. Scopolamine had no significant effect in the other brain regions. A temperature elevation of only 1 degree C above baseline for 48 h in awake female rats increased dizocilpine maleate-induced damage. Finally, the sex differences in N-methyl-D-aspartate antagonist toxicity were replicated and extended to other structures, and found not to be due to temperature differences. Our data show that dizocilpine maleate neurotoxicity is temperature sensitive. Scopolamine treatment needed to be prolonged in order to reduce injury, and even then was only efficacious in one of three brain regions. The results underscore the importance of using neuronal necrosis in several brain regions as the endpoint and for the use of prolonged therapeutic interventions. Furthermore, given the potential hypothermic action of other putative neuroprotective drugs, a mechanistic re-evaluation of N-methyl-D-aspartate antagonist-induced injury is needed, with precise brain temperature measurement.