Morphological and functional features of the organization of perineuronal satellites of the optic nerve of Pholidapus dybowskii

Glial cells in the optic nerve of Pholidapus dybowskii were observed by the Golgi silver impregnation method. Subtypes of protoplasmatic astroglia (free strongly ramified glia), perivascular macroglia (connected with vessels, ramified glia), and oligodendroglia were selected in concordance with the position of cell plexus. By correlative analysis of cell morphological characteristics, obtained by the Leontovich method of cell measuring, some cell features were discovered. The features positively correlated with the capacity of cell branch territory. Correlation coefficients of these features varied from 0.40 to 0.98. Groups of features with a high correlation coefficient were assumed as a basis for recognition of the detailed glial subtypes. Valid differences (P = 0.95) between the detailed glial subtypes were found. The main feature has a reliable difference equal to P = 0.99. Group of constant features were discovered, which define the overall performance of glial cells. Among glial cells, modification of capacities of cell branch territory was measured.     

嗜银细胞在菲牛蛭消化管中的形态与分布特征

运用Grimelius银染法对菲牛蛭(Poecilobdella manillensis)消化管内嗜银细胞的分布和形态进行观察,并根据嗜银细胞的分布特点统计其分布密度。消化管除食道外各部位均有嗜银细胞分布,细胞染棕色或黑色;嗜银细胞基本位于上皮细胞之间和固有层内,主要呈圆形、椭圆形、锥体形和长条形等多种形态;部分细胞胞突明显,细胞内可见黑色分泌颗粒。嗜银细胞分布密度为颚片最高,嗉囊次之,直肠最低,食道内未检测出嗜银细胞。消化管各段嗜银细胞的形态特征和分布规律可能与其食性和消化生理活动密切相关。     

Nitric Oxide Modulates the Temporal Properties of the Glutamate Response in Type 4 OFF Bipolar Cells

Nitric oxide (NO) is involved in retinal signal processing, but its cellular actions are only partly understood. An established source of retinal NO are NOACs, a group of nNOS-expressing amacrine cells which signal onto bipolar, other amacrine and ganglion cells in the inner plexiform layer. Here, we report that NO regulates glutamate responses in morphologically and electrophysiologically identified type 4 OFF cone bipolar cells through activation of the soluble guanylyl cyclase-cGMP-PKG pathway. The glutamate response of these cells consists of two components, a fast phasic current sensitive to kainate receptor agonists, and a secondary component with slow kinetics, inhibited by AMPA receptor antagonists. NO shortened the duration of the AMPA receptor-dependent component of the glutamate response, while the kainate receptor-dependent component remained unchanged. Application of 8-Br-cGMP mimicked this effect, while inhibition of soluble guanylate cyclase or protein kinase G prevented it, supporting a mechanism involving a cGMP signaling pathway. Notably, perfusion with a NOS-inhibitor prolonged the duration of the glutamate response, while the NO precursor L-arginine shortened it, in agreement with a modulation by endogenous NO. Furthermore, NO accelerated the response recovery during repeated stimulation of type 4 cone bipolar cells, suggesting that the temporal response properties of this OFF bipolar cell type are regulated by NO. These results reveal a novel cellular mechanism of NO signaling in the retina, and represent the first functional evidence of NO modulating OFF cone bipolar cells.     

大鼠老化过程大脑皮层及小脑神经元染色质构象分析

 本文在前文~[2]的基础上进一步以MCN和DNaseⅠ为探针研究大鼠脑神经元终末分化后不同生理时期染色质构象,结果表明:MCN酶解DNA产物PAGE显示脑老化过程大脑皮层及小脑神经元染色质核小体单体DNA分别保持在176bp和215bp水平,核小体连接DNA长度存在组织差异,但不受老化影响;<2>DNaseⅠ酶解DNA产物PAGE显示各年龄组大脑皮层及小脑神经元染色质DNA存在10bp间隔重复结构和相同的泳动区带分布特征,提示脑老化中染色质具有稳定的B型双螺旋结构和一致的螺线管卷曲形式。染色质DNaseⅠ降解率随年龄增加而降低,提示老化导致活性染色质区域减少,老化过程脑神经元染色质构象改变成为其转录功能减退的结构基础。     

MDMA (Ecstasy) Decreases the Number of Neurons and Stem Cells in Embryonic Cortical Cultures

Ecstasy, 3,4-methylenedioxymetamphetamine (MDMA), is a recreational drug used among adolescents, including young pregnant women. MDMA passes the placental barrier and may therefore influence fetal development. The aim was to investigate the direct effect of MDMA on cortical cells using dissociated CNS cortex of rat embryos, E17. The primary culture was exposed to a single dose of MDMA and collected 5 days later. MDMA caused a dramatic, dose-dependent (100 and 400 μM) decrease in nestin-positive stem cell density, as well as a significant reduction (400 μM) in NeuN-positive cells. By qPCR, MDMA (200 μM) caused a significant decrease in mRNA expression of the 5HT3 receptor, dopamine D₁ receptor, and glutamate transporter EAAT2-1, as well as an increase in mRNA levels of the NMDA NR1 receptor subunit and the 5HT_1A receptor. In conclusion, MDMA caused a marked reduction in stem cells and neurons in embryonic cortical primary cell cultures, which was accompanied by changes in mRNA expression of specific receptors and transporters for glutamatergic and monoaminergic neurotransmitters.     

Downregulation of Map Kinase Activity Signalled by HIV-1-gp120 Coat Protein in Granular Neurons and Glial Cells from Rat Cerebellum

We have studied the effect of gp120 coat protein from HIV-1 on tyrosine phosphorylation processes in primary cultures of granular neurons or glial cells from the cerebellum of neonatal rats. The extracellular application of recombinant gp120 (200 pM) was able to reduce the phosphotyrosine content and the immunoreactivity for active form-specific antibodies of MAP kinase. Whereas in neurons MAP kinase appeared to be the only protein whose phosphotyrosine content was decreased, in glial cultures the inhibitory effect of gp120 on tyrosine phosphorylation processes appeared to be more widespread. In neuronal cultures, the effect of the viral protein was prevented by the concomitant treatment with depolarizing agents.     

Endothelins Induce Fos Expression in Neurons and Glia in Organotypic Cultures of Rat Cerebellum

Abstract: Endothelins (ETs) and their receptors are present in high levels in the brain and have been proposed to act as neuromodulators or neurotransmitters. However, neither their role nor their precise mechanism of action in the brain is understood. In this study, c- fos expression was used as a marker of neuronal activation in organotypic cultures of rat cerebellum. ETs induced Fos protein expression in both granule cells and glia but not in Purkinje cells. Granule cells and glia were both very sensitive to ETs, but different receptor subtypes appeared to be involved, because granule cells did not respond to ET-3. However, they did respond to the ET_B-selective agonist BQ3020, suggesting the possible existence of a novel neuronal ET_B-like receptor. The induction of Fos in granule cells was independent of extracellular calcium ion concentration, but the ryanodine receptor antagonist dantrolene significantly inhibited the response to ETs, suggesting that the mobilisation of calcium ions from intracellular stores is important. These data support previous evidence that ETs act directly on neurones and show that the intracellular pathways after ET receptor activation are complex. It appears likely that ETs play an important neuromodulatory role in the cerebellum.     

Distribution of Neuropeptide Y-Immunoreactive Neurons in the Human Brainstem, Cerebellum, and Cortex During Development

1. Neuropeptide Y is found throughout the central nervous system where it appears to play a wide range of often poorly understood functions. In this study, the distribution of neuropeptide Y immunoreactive (NPY-ir) neurons in the brainstem, cerebellum, and cerebral cortex of human fetuses ranging in age from 11 gestational weeks to term was investigated by immunohistochemistry. 2. The NPY-ir cells were detected in the dorsal and ventral rostral midbrain and the interpeduncular nucleus by 21 weeks and 32 weeks of gestation, respectively. Although no positive cells were found in the pons, the NPY-ir fibers were detected there at 32 gestational weeks. 3. The vagal, hypoglossal, and olivary nuclei of the medulla oblongata contained immunoreactive cells by week 21 and the medullary reticular formation by week 25 of gestation. In most of these locations, both the number and size of neuropeptide Y positive cells were greater at birth and reached maximal values of 100-400 cells per 1 mm2 and 2-5 microm in diameter, respectively. 4. In the cerebellum, numerous NPY-ir horizontal and granule cells, as well as the cells within the dentate nucleus were observed as early as 21 weeks of gestation. 5. The NPY-ir cells were also detected in the developing cerebral cortex, with the earliest activity observed within the temporal cortex at 14 weeks of gestation. By week 21, positive cells appeared in the visual, frontal, sensory, and motor cortices. Most of these cells were bipolar or multipolar in morphology but their numbers at birth were relatively low. 6. Our results show a wide distribution of the NPY-ir cells in the developing human brain and offer supporting evidence for the important modulatory role of NPY in both the fetus and adult.     

The Output Signal of Purkinje Cells of the Cerebellum and Circadian Rhythmicity

Measurement of clock gene expression has recently provided evidence that the cerebellum, like the master clock in the SCN, contains a circadian oscillator. The cerebellar oscillator is involved in anticipation of mealtime and possibly resides in Purkinje cells. However, the rhythmic gene expression is likely transduced into a circadian cerebellar output signal to exert an effective control of neuronal brain circuits that are responsible for feeding behavior. Using electrophysiological recordings from acute and organotypic cerebellar slices, we tested the hypothesis whether Purkinje cells transmit a circadian modulated signal to their targets in the brain. Extracellular recordings from brain slices revealed the typical discharge pattern previously described in vivo in single cell recordings showing basically a tonic or a trimodal-like firing pattern. However, in acute sagittal cerebellar slices the average spike rate of randomly selected Purkinje cells did not exhibit significant circadian variations, irrespective of their specific firing pattern. Also, frequency and amplitude of spontaneous inhibitory postsynaptic currents and the amplitude of GABA- and glutamate-evoked currents did not vary with circadian time. Long-term recordings using multielectrode arrays (MEA) allowed to monitor neuronal activity at multiple sites in organotypic cerebellar slices for several days to weeks. With this recording technique we observed oscillations of the firing rate of cerebellar neurons, presumably of Purkinje cells, with a period of about 24 hours which were stable for periods up to three days. The daily renewal of culture medium could induce circadian oscillations of the firing rate of Purkinje cells, a feature that is compatible with the behavior of slave oscillators. However, from the present results it appears that the circadian expression of cerebellar clock genes exerts only a weak influence on the electrical output of cerebellar neurons.     

Isolation and characterization of microsatellite markers from the Hypoptychus dybowskii (Pisces,Gasterosteiformes)

Hypoptychus dybowskii (Gasterosteiformes) exhibits allopaternal care frequently caused by various types of male reproductive tactics (sneaking, egg desertion and taking over). In order to understand this interesting reproductive system, we isolated microsatellites loci from H. dybowskii. Five microsatellites showed 2–10 alleles and expected heterozygosities ranged from 0.15 to 0.84. These were not significant deviations from Hardy–Weinberg expectations. These results suggest that these novel polymorphic loci should be useful for parentage analysis of H. dybowskii.     

Pyramidal Cells Make Specific Connections onto Smooth (GABAergic) Neurons in Mouse Visual Cortex

One of the hallmarks of neocortical circuits is the predominance of recurrent excitation between pyramidal neurons, which is balanced by recurrent inhibition from smooth GABAergic neurons. It has been previously described that in layer 2/3 of primary visual cortex (V1) of cat and monkey, pyramidal cells filled with horseradish peroxidase connect approximately in proportion to the spiny (excitatory, 95% and 81%, respectively) and smooth (GABAergic, 5% and 19%, respectively) dendrites found in the neuropil. By contrast, a recent ultrastructural study of V1 in a single mouse found that smooth neurons formed 51% of the targets of the superficial layer pyramidal cells. This suggests that either the neuropil of this particular mouse V1 had a dramatically different composition to that of V1 in cat and monkey, or that smooth neurons were specifically targeted by the pyramidal cells in that mouse. We tested these hypotheses by examining similar cells filled with biocytin in a sample of five mice. We found that the average composition of the neuropil in V1 of these mice was similar to that described for cat and monkey V1, but that the superficial layer pyramidal cells do form proportionately more synapses with smooth dendrites than the equivalent neurons in cat or monkey. These distributions may underlie the distinct differences in functional architecture of V1 between rodent and higher mammals.     

活性氧参与-氧化氮诱导的神经细胞凋亡

采用激光共聚焦成像技术,用氧化还原敏感的特异性荧光探针(DCFH-DA和DHR123)直接研究了一氧 化氮供体S-亚硝基-N-乙酰基青霉胺(SNAP)诱导未成熟大鼠小脑颗粒神经元凋亡过程中的细胞胞浆、线粒体 中活性氧水平的变化,发现神经细胞经0.5mmol/LSNAP处理1h后,细胞胞浆及线粒体中活性氧水平大大增 加.一氧化氮清除剂血红蛋白能够有效抑制细胞胞浆、线粒体中活性氧的产生,防止细胞凋亡.外源性谷胱甘 肽对细胞也具有良好的保护作用,而当细胞中谷胱甘肽的合成被抑制后,一氧化氮的神经毒性大大增强.实验 结果表明一氧化氮通过促进神经细胞产生内源性活性氧而启动细胞凋亡程序,而谷胱甘肽可能是重要的防止一 氧化氮引发神经损伤的内源性抗氧化剂     

Ocular morphology of the Baikal sculpin-oilfishes,Comephorus baicalensis and C. dybowskii (Comephoridae)

Synopsis Baikal sculpin-oilfishes (Comephorus baicalensis and C. dybowskii) were captured from Lake Baikal by mid-water trawl. External morphology was assessed using camera lucida drawings of whole heads. Retinal morphology was examined in material prepared for light and electron microscopy. A single rod-like photoreceptor is present in the retina, and this is at low density compared with marine species of similar depth distribution. C. baicalensis displays tubular eyes, with a dorsally displaced spherical lens and pupil, creating a dorsally directed visual axis. This corresponds to an area with a high density of photoreceptors in the ventral retina. Retinal structure is very similar in C. dybowskii with the exception that regional variation of cell densities within the retina is less marked than in C. baicalensis. In contrast to C. baicalensis, C. dybowskii has a caudal aphakic space and highest photoreceptor density in the rostral retina. Both species display changes in body attitude in association with diurnal vertical migration. We suggest that this may facilitate inspection of the water column from which predator strikes are most likely to occur. Comephorus in general display less specialization for vision in low light than marine counterparts with similar depth distributions.     

MAH Cells and the Development of Rat Sympathetic Neurons

The generation of cell lines in the sympathoadrenal lineage has greatly facilitated our understanding of how precursor cells that do not respond to NGF give rise to mature NGF-dependent neurons. The neuronal developmental pathway in this lineage has been worked out by studying both primary precursor cells in culture and the v-myc-immortalized MAH cell line. MAH cells were established by retroviral infection of immunoisolated rat sympathoadrenal precursor cells. These cells have many of the characteristics of primary progenitor cells including neural precursor morphology, antigenic profile, and response to growth factors. MAH cells are able to recapitulate sympathetic development, giving rise to mature, postmitotic, NGF-dependent neurons. These cells have provided a model system for studying the factors, receptors, and modulating influences that play a role in the development of sympathetic neurons.     

中国黄羽鹌鹑消化道嗜银细胞胚后发育的分布及形态学观察

采用Grimelius银染法,对中国黄羽鹌鹑(Coturnix aponica)消化道嗜银细胞胚后发育的分布规律和形态结构进行了观察。结果显示,口腔、食管、嗉囊和泄殖腔中未发现嗜银细胞,其余部位均有不同数量的分布,其分布呈波浪形,大多数日龄段在腺胃和结直肠中存在2个分布高峰,回肠次之,十二指肠、空肠、盲肠较少,随着日龄增加,不同部位嗜银细胞数量均先增加,后减少,在100 d时达高峰。嗜银细胞主要分布在腺胃腺叶内细胞之间、肠黏膜上皮细胞之间及固有层内,形态多呈圆形、椭圆形、锥形及梭形等。结合嗜银细胞形态与功能间的联系,发现消化道内广泛分布着4种类型的嗜银细胞。我们认为中国黄羽鹌鹑消化道的嗜银细胞具有内分泌、外分泌及旁分泌3种功能,其分布特点可能与动物的食性及生活环境有关。     

循环饥饿投喂对大鼠消化道嗜银细胞形态及分布的影响

为了研究循环饥饿投喂对大鼠(Rattus norvegicus)消化道内嗜银细胞的分布密度及形态功能的影响,本实验采用Grimelius银染法观察和测定循环饥饿投喂(饥饿1 d,投喂1 d,周期为14 d)期间大鼠消化道嗜银细胞形态功能及密度分布。结果表明,实验组(即循环饥饿投喂组)和对照组(正常喂食组)大鼠消化道嗜银细胞除食管外均有分布,形态上对照组嗜银细胞以椭圆形和锥体形为主,实验组嗜银细胞则主要以锥体形为主;两组大鼠消化道嗜银细胞的分布密度高峰都位于胃,密度低谷对照组位于空肠、盲肠、回肠和直肠,实验组位于空肠到直肠各段;实验组大鼠消化道嗜银细胞分布密度在贲门和幽门部极显著低于对照组(P <0.01),结肠显著低于对照组(P <0.05),说明循环饥饿投喂会明显改变消化道嗜银细胞的形态并降低细胞数量,这可能与内分泌细胞的功能改变有关。     

Involvement of protein kinase C in the regulation of oocyte maturation in amphibians (Rana dybowskii)

Ovarian oocytes of Rana dybowskii, isolated early in the hibernation period (late autumn), failed to mature, i.e., germinal vesicle breakdown (GVBD), in response to progesterone during in vitro follicle culture. Oocytes collected during the middle hibernation period matured in response to progesterone, whereas those collected late during the hibernation period (close to the breeding season) underwent spontaneous maturation without added hormone (Kwon et al., '89). The maturational response (GVBD) of oocytes, collected at the three stages of hibernation, to protein kinase C (PKC) activation was investigated and compared to that of progesterone stimulation. A phorbol ester, phorbol 12-myristate 13-acetate (TPA) was used for PKC activation. TPA addition to cultured follicles collected during the early or middle period of hibernation induced oocyte GVBD. The incidence of maturation (% GVBD) induced by TPA varied markedly between animals. TPA (10 microM) induced oocyte maturation in the presence or absence of follicle cells. The time course of the TPA-induced maturation was similar to that of progesterone-stimulated maturation (ED50, 7-9 h). TPA also accelerated the onset of maturation of the follicular oocytes exhibiting spontaneous in vitro maturation. Both TPA- and progesterone-stimulated maturation was blocked by treatment with cycloheximide (1 microgram/2 ml), forskolin (9 microM) (an adenylate cyclase stimulator), and verapamil (0.27 mM) (a calcium transport blocker). Treatment of oocytes with a calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7) (100 microM) or a PKC inactivator 1-(5-isoquinolinylsulfonyl)-2-methyl-piperazine (H-7) (50 microM) likewise suppressed TPA- or progesterone-induced maturation.(ABSTRACT TRUNCATED AT 250 WORDS)