大鼠睁眼前初级视皮层2/3层锥体神经元突触自身稳态可塑性的变化特征

目的：通过对Wistar大鼠睁眼前初级视皮层2/3层锥体神经元突触AMPA（α-氨基-3-羧基-5-甲基异恶唑-4-丙酸）介导的微小兴奋性突触后电流 (mEPSCs)的测定分析,研究突触自身稳态可塑性在生后早期初级视皮层的作用特点。方法: 采用红外可视膜片钳技术全细胞模式记录生后4-11(d)（P4-11）Wistar大鼠初级视皮层脑片2/3层锥体神经元AMPA介导的mEPSCs,钳制电位-70 mV。人工脑脊液中加入河豚毒素（TTX）、荷包牡丹碱（BMI）及2-氨基-5-磷酸基戊酸（AP-5）分离出AMPA介导的mEPSCs,加入阻断剂6-氰基-7-硝基喹喔啉-2,3二酮 (CNQX）可消除mEPSCs。使用Clampfit 9.0进行数据分析。结果: P4至P11,大鼠初级视皮层2/3层锥体神经元AMPA介导的mEPSCs的波幅呈现上升趋势,频率自P7至P11逐渐增加,上升时间常数及下降时间常数均呈缩短趋势,以下降时间常数变化为著。P4至P7可见“单通道样”电流形态。结论: 在大鼠睁眼前初级视皮层2/3层锥体神经元亦存在突触自身稳态可塑性调节机制,其作用特点不同于睁眼后。     

The buoyant density of synaptic plasma membranes from the cerebral cortex of neonatal rats

Summary A fraction enriched in synaptic plasma membranes was prepared from neonatal (5–6 day old) rat cerebral cortex. The procedure was based on a method used to prepare synaptic plasma membranes from adult cerebral cortex. Critical steps were monitored by electron microscopy. Synaptic plasma membranes from neonatal cerebral cortex sedimented as a broad peak between 0.9 M and 1.1 M sucrose. In contrast, the majority of adult synaptic plasma membranes have been reported to sediment to 1.2 M sucrose. The activities of various enzyme markers were determined in subfractions of neonatal preparations in order to estimate contamination. The specific activities of these markers indicated substantial contamination of the neonatal synaptic plasma membrane fractions by microsomes and glia.     

Loss of dendritic spines in aging cerebral cortex

Summary Previous work has shown that the dendritic spines of pyramidal neurons of the cerebral cortex are sensitive to a wide variety of environmental and surgical manipulations. The present study shows that the normal aging process also affects these spines. The spines were studied with the light microscope in Golgi preparations from rats ranging in age from 3 to 29.5 months. Visible spines were counted on either 25 or 50 segments of the basal dendrites, apical dendrites, oblique branches, and terminal tufts of layer V pyramidal cells in area 17. A progressive loss of spines occurred at each of these loci. The smallest observed spine loss (24%) occurred on the dendrites of the terminal tuft, and the largest (40%) on the oblique branches. Age-related spine loss appears to affect all animals, and for animals of any one age the overall loss is similar. However, the cell-to-cell variability within an individual animal is pronounced, some cells with high spine densities being present at every age examined. As a general rule, there is a positive relationship between visible spine density along the apical dendrite as it traverses layer IV and the thickness of the dendrite. With advancing age, the relatively thick dendrites decrease in number so that the thinner dendrites make up an increasingly larger proportion of the total apical dendrite population. Questions that remain for the future include the genesis of the spine loss, its relation to other aging changes, and its functional significance for the neuron.Supported by United States Public Health Service Program Project Grant HDO-5796-03 and Research Grant NB-07016     

The effect of aging on the neuronal population within area 17 of adult rat cerebral cortex

The brains of Sprague-Dawley rats in various age groups from 3 to 33 months were fixed by perfusion with standard aldehyde solutions in order to determine the effects of aging on neuronal numbers. Several indices of cortical volume were then measured to determine whether neuronal packing densities were affected by age-related change in cortical volume. The lengths, heights and widths of individual hemispheres for 160 animals ranging in age from 1 day to 36 months were first determined, after which blocks of tissue were removed from area 17 of some of the brains. These blocks were osmicated, embedded in Araldite and sectioned at 1 micrometer to ascertain, in the vertical plane, the thickness of area 17 and, in the tangential plane, the packing density of the clusters of apical dendrites extending from layer V pyramidal neurons. Results indicate the overall dimensions of the cerebral hemispheres increased until 3 months of age, after which there was no further increase in size. Between 3 and 33 months of age there was no age-related change in either the thickness of area 17 or in the separation between dendritic clusters, indicating the volume of area 17 did not change after 3 months of age. Within individual age groups the amount of variation present is greater than that among age groups. Since the number of nucleus-containing neuronal profiles per unit area of layers II/III, IV, V, VIa and VIb was similar in two groups of three animals at 3 and 33 months of age and the diameters of neuronal nuclei were unchanged, there seems to be no significant change in the number of neurons contained in these layers of rat visual cortex between 3 and 33 months of age. It is therefore concluded that no neurons are lost from area 17 as the mature cerebral cortex ages.     

The effect of aging on the neuronal population within area 17 of adult rat cerebral cortex

The brains of Sprague-Dawley rats in various age groups from 3 to 33 months were fixed by perfusion with standard aldehyde solutions in order to determine the effects of aging on neuronal numbers. Several indices of cortical volume were then measured to determine whether neuronal packing densities were affected by age-related change in cortical volume. The lengths, heights and widths of individual hemispheres for 160 animals ranging in age from 1 day to 36 months were first determined, after which blocks of tissue were removed from area 17 of some of the brains. These blocks were osmicated, embedded in Araldite and sectioned at 1 micrometer to ascertain, in the vertical plane, the thickness of area 17 and, in the tangential plane, the packing density of the clusters of apical dendrites extending from layer V pyramidal neurons. Results indicate the overall dimensions of the cerebral hemispheres increased until 3 months of age, after which there was no further increase in size. Between 3 and 33 months of age there was no age-related change in either the thickness of area 17 or in the separation between dendritic clusters, indicating the volume of area 17 did not change after 3 months of age. Within individual age groups the amount of variation present is greater than that among age groups. Since the number of nucleus-containing neuronal profiles per unit area of layers II/III, IV, V, VIa and VIb was similar in two groups of three animals at 3 and 33 months of age and the diameters of neuronal nuclei were unchanged, there seems to be no significant change in the number of neurons contained in these layers of rat visual cortex between 3 and 33 months of age. It is therefore concluded that no neurons are lost from area 17 as the mature cerebral cortex ages.     

脑缺血大鼠大脑皮质NMDA受体的早期表达

目的　探讨脑缺血和缺血再灌注早期大脑皮质NMDA受体的变化规律。方法　健康雄性SD大鼠 4 8只 ,随机分为 6组 :假手术对照组、单纯脑缺血 1 5min、2 0min和 30min组及脑缺血 1 5min再灌流 30min和 2 4h组 ;4血管脑缺血动物模型 ;连续冰冻冠状切片观察 ,NR1 免疫组化染色特异性地显著NMDA受体的变化 ,图象分析及计算阳性单位PU值。结果　①单纯脑缺血各组PU值分别为 7 5 5± 1 81、6 91± 2 5 3和 6 0 9± 1 5 0 ,与对照组PU值 9 0 4± 1 5 7相比呈下降趋势 (P≤ 0 0 5 ) ;②再灌流 30min和 2 4h组PU值分别为 5 76± 2 2 4和 4 73± 1 34,与对照组PU值相比明显减少 36 2 9%～ 4 7 6 8% ,有统计学意义(P <0 0 5 ) ;③再灌流 0min、30min和 2 4h组两两比较 ,除 2 4h和 0min组相比NR1 阳性反应物减少有统计学意义 (P <0 0 5 )外 ,其它组别间NMDA受体下降的变化无显著性差异 (P >0 0 5 )。结论　脑缺血和缺血再灌流早期NMDA受体都存在下行调节 ,这种调节可能是一种保护性作用     

Synaptic interactions involving acetylcholine,glutamate, and GABA in rat auditory cortex

Using electrophysiological techniques in the in vitro rat auditory cortex, we have examined how spontaneous acetylcholine (ACh) release modifies synaptic potentials mediated by glutamate and -aminobutyric acid (GABA). Single stimulus pulses to lower layer VI elicited in layer III a four-component (A-D) extracellular field response involving synaptic potentials mediated by glutamate and GABA. The cholinesterase inhibitor eserine (10–20 M) or the cholinergic agonist carbachol (25–50 M) depressed by 10–50% the glutamatergic components A and C, and the GABAergic components B and D. Atropine reversed the depressive effects of eserine and carbachol. A novel finding was that the degree of depression of component A varied inversely with stimulus intensity. However, during partial pharmacological antagonism of GABA_A receptors, depression of A varied directly, not inversely, with stimulus intensity. Normally, then, depression of A is offset by reduced GABAergic inhibition of A. We also tested for differential depression of responses mediated by N-methyl-d-aspartate (NMDA) versus non-NMDA glutamate receptors. Following physiological and pharmacological isolation of the responses, eserine depressed the non-NMDA, but not the NMDA, receptor-mediated potential. Since the isolated NMDA potential still could be depressed by carbachol, the data suggested that activation of NMDA receptors may reduce spontaneous ACh release. In support of this, preincubation of slices in NMDA (10–20 M) largely prevented eserine's, but not carbachol's, depression of components A and B.These results permit three conclusions of relevance to cortical information processing: (1) spontaneous ACh release tonically depresses synaptic potentials mediated by glutamate and GABA; (2) ACh depresses responses to weak inputs to a greater degree than responses to strong inputs; (3) activation of NMDA receptors may feed-back to reduce ACh release, a mechanism that could place regulation of local ACh release under glutamatergic afferent control.     

Effects of alcohol administration during adulthood on parvalbumin and glial fibrillary acidic protein immunoreactivity in the rat cerebral cortex

The pathology of brain atrophy mediated by alcohol was investigated in all parts of the cerebral cortex (the frontal, parietal, temporal lobes and occipital cortex) by using two markers: parvalbumin (PV) and glial fibrillary acidic protein (GFAP). Three-month old male Wistar rats were divided into control (C) and alcohol-exposed groups. The control group received distilled water, whereas the alcohol-exposed groups received either a low dose (2 g/kg body wt) or a high dose (5 g/kg) of ethanol for periods of 21 days, 3 or 6 months. The brains of the animals were processed for immunohistochemistry using anti-parvalbumin and anti-GFAP antibodies and the number of PV immunoreactive (PV-ir) neurons and GFAP immunoreactive (GFAP-ir) astrocytes were counted per unit area. Results showed that all groups exposed to ethanol had significantly reduced numbers of PV-ir neurons in all parts of the cerebral cortex compared to those of the control group (p<0.05). In contrast, the numbers of GFAP-ir astrocytes were increased in all parts of the cerebral cortex following the exposure to a high dose of ethanol after 21-days (but not a low dose) and both high and low doses of ethanol after 3-months or 6-months treatment compared to those of age-matched control groups (p<0.05). This indicated that in young rats (21-days), PV-ir neurons in all cerebral cortex areas seemed to be more sensitive to alcohol than GFAP-ir astrocytes. Moreover, the change in densities of both PV-ir neurons and GFAP-ir astrocytes became more apparent after exposure to prolonged and high doses of ethanol. The decrease of PV-ir neurons and the increase of GFAP-ir astrocytes indicated that alcohol may induce pathology in broad areas of the cerebral cortex. This may explain the underlying mechanism of brain atrophy and other impairments found in alcoholics. For investigations of the effects of alcohol on mediating brain pathology, we recommend the use of the two markers (PV and GFAP).     

Synaptic development in the rabbit superior colliculus and visual cortex

Summary The development of synapses in the visual cortex (VC) and superior colliculus (SC) of the rabbit has been examined with the electron microscope. In both areas, the number of synapses reaches adult levels by 20–25 days of postnatal age, but the development in the visual cortex is delayed in comparison to that in the superior colliculus. When S synapses (spheroidal vesicles, asymmetric thickening) are compared with F synapses (flattened vesicles, symmetric thickening), even greater differences are seen. In both the VC and SC, S synapses develop earlier than F synapses, though there is considerable overlap. Of interest is the fact that synapses in the visual cortex seem to overshoot their adult levels late in development, suggesting that an excess of synapses may be formed in this system. Multiple synapses, probably of retinal origin, increase in the first 3 weeks of synaptic development in the SC, but never are present in significant proportions in the VC.Synapse formation most often is characterized by formation of a junction and a postsynaptic thickening, followed by acquisition of synaptic vesicles. After 15 days, there is only a small number of such non-vesicle synapses in either the SC or VC.     

Tyrosine hydroxylase-immunoreactive intrinsic neurons in the rat cerebral cortex

Summary Using specific antisera against the catecholamine synthesizing enzyme, tyrosine hydroxylase (TH), in combination with the peroxidase-antiperoxidase method and/or the avidin-biotin complex method, we have found a new group of TH immunoreactive (TH-I) neurons in the rat cerebral cortex. Numerous TH-I cells were observed all over the isocortex, that is, frontal, temporal, parietal and occipital regions, and in some parts of the allocortex such as the anterior cingulate cortex, the retrosplenial cortex and anterior part of the insular cortex. In contrast, they were rare in the perirhinal cortex, posterior part of the insular cortex, piriform cortex, entorhinal cortex and hippocampal formation. TH-I cells were situated throughout all cortical layers, but were most concentrated in layer II/III. Although TH-I cells were heterogeneous in shape, the majority were bipolar. All TH-I cells so far examined appeared to be of the nonpyramidal type. The majority of these intrinsic TH-I neurons also contained the GABA-like immunoreactivity and thus could be regarded as a subpopulation of cortical GABAergic neurons.     

Spatial correlation between sensory regions and the drainage fields of pial veins in rat cerebral cortex

Summary Visual and somatosensory evoked potentials were mapped in the cerebral cortex of adult rats and, after filling the cerebral arteries and veins with dye, the mappings were then compared to the distribution of pial veins. A close relationship was found between the position, size and shape of the occipital venous drainage field and the distribution of visual evoked potentials with high amplitudes and short latencies. Accordingly, such potentials evoked by stimulation of the forepaw and the tailroot were confined to the fronto-parietal drainage field. In the case of individual variations in the expansion and shape of sensory areas, the medial and lateral borders of the occipital drainage field and the medial border of the fronto-parietal drainage field covaried. Only at the common border between these two drainage fields, visual evoked potentials with small amplitudes and long latencies extended into the parietal drainage field and overlapped with somatosensory evoked potentials. This overlapping area corresponds in position to the anterior part of the peristriate cortex. A comparison between the vascular organization and cytoarchitectonic maps of the rat cortex indicates that other parts of the characteristic pattern of venous drainage fields may also correlate with the cytoarchitectonic and functional organization of the cerebral cortex. These observations suggest that during morphogenesis the formation of sensory projections to the cerebral cortex may interact with the angiogenesis, mainly with the development of veins.Fellow of the Alexander von Humboldt Stiftung     

An early development defect in the cerebral cortex of the reeler mouse

Summary Brains of reeler and normal mouse embryos have been studied on semi-thin sections and with Golgi impregnations. No change can be seen in the neuroepithelium or in the primary cortical organization. The first evidence of a morphological abnormality appears at E 14, in the cortical plate. Instead of being closely packed and radially oriented, nerve cells are loosely arranged and show quite variable orientations of their long axis and apical dendrite. The axons run obliquely through the cortical plate and do not display the characteristic angular course seen in the normal animal.It is suggested that the primary defect in reeler mice may be in the plasma membrane of cortical plate cells, resulting in a loss of their capacity for mutual recognition and binding. This could account for the cytoarchitectonic disorganization in this mutant, especially the absence of a molecular layer and the inversion of the histogenetic gradient in the developing cerebral cortex.Aspirant au Fonds National de la Recherche Scientifique de Belgique     

Flattened cortical maps of cerebral function in the rat: a region-of-interest approach to data sampling, analysis and display

We describe a method for the measurement, analysis and display of cerebral cortical data obtained from coronal brain sections of the adult rat. In this method, regions-of-interest (ROI) are selected in the cortical mantle in a semiautomated fashion using a radial grid overlay, spaced in 15 degrees intervals from the midline. ROI measurements of intensity are mapped on a flattened two-dimensional surface. Topographic maps of statistical significance at each ROI allow for the rapid viewing of group differences. Cortical z-scores are displayed with the boundaries of brain regions defined according to a standard atlas of the rat brain. This method and accompanying software implementation (Matlab, Labview) allow for compact data display in a variety of autoradiographic and histologic studies of the structure and function of the rat brain.     

不同雌激素对大鼠海马和脑皮质区域雌激素受体α和β表达的差异性调节研究

目的:探讨不同种类雌激素药物对雌性去势大鼠海马和脑皮质区域雌激素受体(estrogenreceptor,ER)的调节差异。方法:雌性去势大鼠分别给予倍美力或补加乐持续灌胃3个生理周期,采用半定量RT-PCR法检测雌激素受体ERα、ERβ的mRNA表达,免疫组化法检测ERα、ERβ蛋白的分布及表达。结果:倍美力组海马和脑皮质ERαmRNA表达均显著低于去势对照组,ERα蛋白在海马中相对表达量也低于对照组,但在脑皮质中表达无改变;该组上述两个区域中ERβmRNA以及脑皮质ERβ蛋白的表达无显著改变,仅海马ERβ蛋白表达升高。补加乐组脑皮质和海马ERβmRNA和蛋白表达均显著高于去势对照组,海马ERαmRNA表达也高于对照组,但脑皮质ERαmRNA及脑皮质和海马ERα蛋白的表达水平均无显著改变。结论:两种不同种类雌激素药物对雌性去势大鼠认知区域中ERα、ERβ表达的调节水平存在显著差异,这可能是选用不同的雌激素药物进行激素替代治疗产生的保护认知疗效存在差别的机制之一。     

Acetylcholine release from rat cortical slices during postnatal development and aging

Acetylcholine release from cortical slices superfused with choline-enriched Krebs solution containing physostigmine was investigated at birth, at 7, 20 and 30 days, and at 3 and 24 months of age, in order to assess age influence on the functional efficiency of the cortical cholinergic network. The slices were electrically stimulated at frequencies from 1 to 10 Hz for 5 min periods, preceded and followed by rest periods. The superfusate was collected every 5 min and acetylcholine content quantified by bioassay. In the newborn and 7 day-old pups acetylcholine release was approximately 50% lower than that of the 3 month-old rats at all frequencies tested. The highest release was elicited in the 30 day-old rats. Beginning with this age the evoked ACh release underwent a decline which in the 24 month-old rats brought it back to the same level as in the newborn ones. The blockade of the muscarinic autoreceptors by atropine 1.5 × 10^?8 M caused an increase in acetylcholine release at 20 day, 3 and 24 months of age but not in the newborn and 7 day-old pups. Adenosine 3 × 10^?5 M decreased acetylcholine output in newborn and adult but had no effect in the senescent rats.     

Exchange and efflux of tryptophan from superfused cerebral cortex slices

Summary The efflux and exchange of L-tryptophan (Trp) from rat cerebral cortex slices were studied in a superfusion system. The substrate specificity of Trp exchange was assessed by measuring the stimulation of [³H]Trp exit provoked by other extracellular amino acids. Large neutral amino acids were the most potent, but also glutamic acid, lysine and glycine had some effect. The stimulation caused by extracellular Trp and phenylalanine persisted also at 0 ° C though severalfold attenuated. Only intracellular histidine provoked slight inhibition of [³H]Trp efflux and intracellular Trp, phenylalanine and lysine had a small stimulatory effect. The results suggest an involvement of carrier-mediated processes in the exchange and efflux of Trp. The substrate specificities of the exchange and efflux are not apparently identical.     

多巴胺受体蛋白在大鼠心肌肥厚时的表达变化

目的:观察多巴胺受体（DR）1和2 mRNA和蛋白质在大鼠病理性心肌肥厚时的表达情况。 方法:应用肾动脉缩窄术复制Wistar大鼠心肌肥厚的动物模型。于术后35 d取心脏,测定心肌肥大指数，左室内压，V-G染色观察胶原含量。应用心肌原位杂交和RT-PCR，免疫荧光，Western blotting结合图像分析系统分别检测心肌组织中多巴胺受体D1、D2 mRNA 和蛋白质的表达变化。 结果:DR1、DR2 mRNA在正常大鼠心肌组织有表达，其中血管平滑肌细胞内DR2的分布多于心室肌和心房肌细胞。模型组左心肥大明显，表现为室内压显著升高，胶原含量增多;模型组心室肌DR1和 DR2 mRNA和蛋白质的含量均明显低于假手术组。 结论:正常大鼠心肌组织存在DR1和 DR2 mRNA和蛋白质的表达，心肌肥厚时其表达明显减低，两者的关系及可能机制有待于进一步研究。     

Influence of maternal adrenalectomy and glucocorticoid administration on the development of rat cerebral cortex

In order to determine the incidence of maternal glucocorticoids on morphological parameters in fetal development, we performed optic and electron microscopic analysis of the cerebral cortex of fetuses of 16 and 20 days of gestation, from control (C) and pregnant rats bilaterally adrenalectomized on day 1 of gestation (ADX). We also studied fetuses 20 days old from pregnant rats betamethasone-injected on days 15, 16 and 17 (BET), and adrenalectomized on day 1 and betamethasone-injected on days 15, 16 and 17 (ADX+BET). Absence of maternal glucocorticoids during gestation caused, in fetuses 16 and 20 days old, a marked increase of cellular density, laxity of tissue and lower cellular maturation in comparison with the control group. Beta-methasone injected into sham-operated animals (BET) caused a slight advance in relation to controls in developmental parameters such as cellular density, maturation and synapse formation. Betamethasone injection into adrenalectomized animals prevented the lower degree of maturation characteristic of the adrenalectomized group, although an increase of cellular density could be detected. The cerebral cortex from fetuses of 16 days of gestation from adrenalectomized mothers also showed an increase of cellular density as compared with the control group. These results show that glucocorticoids participate in prenatal rat brain in control mechanisms of cellular division and maturation.Abbreviations ADX adrenalectomized animals - ADX+BET adrenalectomized, betamethasone-injected animals - BET betamethasone-injected animals - C control animals - CORT corticosterone - CP cortical plate - GC glucocorticoids - GR glucocorticoid receptor - I cortical layer I - IZ intermediate zone - LV lateral ventricle - NE neuroepithelium - Sp subplate - SV subventricular layer