Histochemical studies on the distribution of thiamine pyrophosphatase and enzymes related to carbohydrate metabolism in the intercalated neurons of the rat supraoptic nucleus

Histochemical studies have been conducted by applying hexokinase (HK), aldolase (AD), glyceraldehyde-3-phosphate dehydrogenase (G3), succinate dehydrogenase (SDH), glucose-6-phosphate dehydrogenase (G6PD), and thiamine pyrophosphatase (TPPase) methods, as well as Nissl staining and Gomori's chrome-alum-hematoxylin-phloxine (CHP) methods to intercalated neurons of the supraoptic nucleus (SO) on Wistar strain rats. Intercalated neurons reacted weakly to the AD, G3, G6PD, and SDH tests, indicating that they belong to the category of ordinary neurons with low carbohydrate metabolism. Many fibrous astrocytes showing strong HK reactions surround neurosecretory neurons. However, they do not surround intercalated neurons with mild HK activity. These results indicate that the latter receive a poor supply of energy from glucose in the circulating blood in contrast to the former. Intercalated neurons are very rich in Nissl substance but lack CHP-positive material. They may have a high potential for synthesizing protein. The principal morphological features of the TPPase-positive Golgi material are peculiar and heterogeneous shape and poor development. These findings together with mild G6PD activity suggest that intercalated neurons are very likely to have poor synthesizing activity.     

Effects of 20-hydroxyecdysone and serotonin on neurite growth and survival rate of antennal lobe neurons in pupal stage of the silk moth Bombyx mori in vitro

Effects of 20-hydroxyecdysone and serotonin on the morphological development and the survival of antennal lobe neurons from day-2 pupal brains of the silk moth Bombyx mori were investigated in vitro. Four morphologically distinct neuronal types could be identified in the cultured antennal lobe neurons: unipolar, bipolar, multi-polar and projection neurons. Antennal lobe neurons in culture with 20-hydroxyecdysone and serotonin showed different patterns of the morphological development from those described in Manduca sexta. Projection neurons extend their neurites remarkably by 20-hydroxyecdysone in B. mori, but there is no extension from antennal lobe neurons in M. sexta. Multi-polar neurons conspicuously increase only formation of new branches from their primary neurites by serotonin in B. mori, but there are both extension and branching of the neurites in M. sexta. On day-5, antennal lobe neurons in lower titers of 20-hydroxyecdysone had significantly higher survival rates than those in higher titers. Neurons cultured for 7 days at different levels of 20-hydroxyecdysone generally showed significantly lower survival rates than neurons cultured for 5 days under the same conditions.     

Isoenzymes in two micromycetes species: taxonomic aspects

Strains of different origins belonging to the two species of fungi imperfecti Cylindrocarpon ianothele and Calcarisporium arbuscula exhibit difference in their morphological or physiological appearances. We have attempted to determine if these variabilities could also be observed at the level of other phenotypic markers, the enzymes glucose-6-phosphate dehydrogenase, alcohol dehydrogenase, octanol dehydrogenase and malate dehydrogenase. Isozymes were separated by polyacrylamide gel electrophoresis and demonstrated isozyme variability as a function of growth and physiological states of a given strain. At the same state of development, the different strains exhibited highly heterogeneous isozyme groups which appear difficult to use for taxonomic purposes.     

Histochemical studies on the morphology of the Golgi apparatus and on the enzymes of carbohydrate metabolism in various nuclei of the rat mesencephalon.

Detailed histochemical studies have been conducted on the distribution of various enzymes such as thiamine pyrophosphatase, alpha-glucan phosphorylase, hexokinase, glucose-6-phosphate dehydrogenase, aldolase, lactate dehydrogenase and succinate dehydrogenase in various components of the nucleus Edinger-Westphali, nucleus n. oculomotorii, nucleus ruber and nucleus niger of healthy adult male Wistar strain rats. The thiamine pyrophosphatase reaction showed the morphological patterns of the Golgi apparatus characteristic for each nucleus. The Golgi apparatus was well developed in the nucleus Edinger-Westphali, composing a network of highly fenestrated plates in the nucleus n. oculomotorii and nucleus ruber, and a simple network in the nucleus niger. These results indicate that the former three nuclei need a rich energy supply and argue against the possibility that the four nuclei have a secretory role. The neurons of the nucleus Edinger-Westphali may derive their energy mainly from glucose of the circulating blood, but glial cells may serve as energy donators to the neurons in the pars compacta of the nucleus niger, and the neurons of the other nuclei may derive energy from both sources. These conclusions are consistent with the morphological patterns of the Golgi apparatus.     

Enzyme heterozygosity,metabolism, and developmental stability

Developmental homeostasis, measured as either fluctuating asymmetry or variance of morphological characters, increases with enzyme heterozygosity in many, but not all, natural populations. These results have been reported forDrosophila, monarch butterflies, honeybees, blue mussels, side-blotched lizards, killifish, salmonid fishes, guppies, Sonoran topminnows, herring, rufous-collared sparrows, house sparrows, brown hares, white-tailed deer, and humans. Because heterozygosity at a few loci can not predict heterozygosity of the entiry genome, these loci must be detecting localized zones that influence the developmental environment. Studies of malate dehydrogenase in honeybees,Apis mellifera, and lactate dehydrogenase in killifish,Fundulus heteroclitus, revealed that developmental homeostasis varied with heterozygosity of individual loci. Heterozygotes differed from homozygotes in fluctuating asymmetry, morphological variance, and in correlations between morphological characters. The protein loci in these studies code for enzymes, and therefore do not directly influence morphological characters. However, some enzymatic loci substantially influence metabolism, and contribute to variation in the amount of energy available for development and growth. This argument can be made most convincingly for the LDH polymorphism in killifish. LDH genotypes differ in enzyme kinetic properties that measure differences in physiological efficiency, and these differences produce measurable and predictable differences in physiology and development. Under environmental conditions which impose a stress upon development, genotypes at these loci may have different amounts of energy available for development, and consequently exhibit different levels of developmental homeostasis.     

Histochemical studies on the morphology of the golgi apparatus and on the enzymes of carbohydrate metabolism in various nuclei of the rat mesencephalon

Summary Detailed histochemical studies have been conducted on the distribution of various enzymes such as thiamine pyrophosphatase, α-glucan phosphorylase, hexokinase, glucose-6-phosphate dehydrogenase, aldolase, lactate dehydrogenase and succinate dehydrogenase in various components of the nucleusEdinger-Westphali, nucleus n. oculomotorii, nucleus ruber and nucleus niger of healthy adult male Wistar strain rats. The thiamine pyrophosphatase reaction showed the morphological patterns of the Golgi apparatus characteristic for each nucleus. The Golgi apparatus was well developed in the nucleusEdinger-Westphali, composing a network of highly fenestrated plates in the nucleus n. oculomotorii and nucleus ruber, and a simple network in the nucleus niger. These results indicate that the former three nuclei need a rich energy supply and argue against the possibility that the four nuclei have a secretory role. The neurons of the nucleusEdinger-Westphali may derive their energy mainly from glucose of the circulating blood, but glial cells may serve as energy donators to the neurons in the pars compacta of the nucleus niger, and the neurons of the other nuclei may derive energy from both sources. These conclusions are consistent with the morphological patterns of the Golgi apparatus. It is suggested that the neurons of the nucleusEdinger-Westphali, nucleus n. oculomotorii, nucleus ruber and of the pars lateralis of the nucleus niger may be equipped almost equally with the Embden-Meyerhof pathway and with the hexose monophosphate shunt. But, the hexose monophosphate shunt is dominant in the pars compacta of the nucleus niger. It is also suggested that the pattern of distribution of succinate dehydrogenase may parallel that of lactate dehydrogenase. The nucleus n. oculomotorii, and nucleus ruber have a higher level of oxidative metabolism than the nucleusEdinger-Westphali and the nucleus niger. The nucleusEdinger-Westphali may be representative of autonomic nuclei with low oxidative metabolism whereas the nucleus n. oculomotorii may represent motor nuclei with high oxidative metabolism. Predominance of hexose monophosphate shunt, intense hexokinase reaction around the neurons, and weak activity of succinate dehydrogenase indicate that the pars compacta of the nucleus niger belongs to the category of “exceptional nuclei”.     

大鼠肠道内NOS与AChE、VIP阳性神经元的分布关系研究

应用一氧化氮合酶 (NOS)、乙酰胆碱酯酶 (ACh E)组织化学及血管活性肠肽 (VIP)免疫组织化学方法 ,光镜下比较观察大鼠肠道内 NOS、ACh E、VIP阳性神经元的形态学特征。结果显示 ,肠肌间丛 NOS阳性神经元胞体大小不等 ,形态不一 ,NOS、ACh E和 VIP阳性神经元的分布密度为 ACh E>NOS>VIP,在不同的肠段和层次分布密度有差异 ,NOS与 ACh E存在共染。在肌间丛和粘膜下丛 ,少数 VIP与 NOS共染。在粘膜下丛 ,三种阳性神经元的分布密度为 ACh E>VIP>NOS。在肌间丛和粘膜下丛 ,可见 VIP阳性末梢环抱 NOS阳性神经元胞体 ,两者呈终扣样接触。上述结果提示 NOS阳性神经元与 ACh E、 VIP阳性神经元有密切的形态学联系。在消化道功能调节上 ,它们可能起协调作用。     

Characterization of pancreas-projecting rat dorsal motor nucleus of vagus neurons

The electrophysiological and morphological properties of rat dorsal motor nucleus of the vagus (DMV) neurons innervating the pancreas were examined by using whole cell patch clamp recordings from brain stem slices and postfixation morphological reconstructions of Neurobiotin-filled neurons. Recordings were made from 178 DMV neurons whose projections had been identified by previous apposition of the fluorescent neuronal tracer DiI to the body of the pancreas. DMV neurons projecting to the pancreas had an input resistance of 434 +/- 14 M omega, an action potential duration of 3 +/- 0.1 ms, and an afterhyperpolarization of 18 +/- 0.4 mV amplitude and 108 +/- 7 ms time constant of decay; these electrophysiological properties resembled those of gastric-projecting neurons but were significantly different from those of intestinal-projecting neurons. Interestingly, 14 of 178 pancreas-projecting neurons showed the presence of a slowly developing afterhyperpolarization whose presence was not reported in DMV neurons projecting to any other gastrointestinal area. The morphological characteristics of pancreas-projecting neurons (soma area 274 +/- 12 microm2; soma diameter of 25 +/- 0.7 microm; soma form factor 0.74 +/- 0.01; segments 9.7 +/- 0.41), however, were similar to those of intestinal- but differed from those of gastric-projecting neurons. In summary, these results suggest that pancreas-projecting rat DMV neurons are heterogeneous with respect to some electrophysiological and morphological properties. These differences might underlie functional differences in the vagal modulation of pancreatic functions.     

Modifications in energy metabolism during the development of chick glial cells and neurons in culture

Developmental changes in lactate dehydrogenase (LDH), enolase, hexokinase (HK), malate dehydrogenase (MDH), and glutamate dehydrogenase (GDH) activities were measured in cultures of pure neurons and glial cells prepared from brains of chick embryos (8 day-old for neurons, 14 day-old for glial cells) as a function of cellular development with time in culture. The modifications observed in culture were compared to those measured in brain extracts during the development of the nervous tissue in the chick embryo and during the post-hatching period. A significant increase of MDH, GDH, LDH, and enolase activities are observed in neurons between 3 and 6 days of culture, whereas simultaneously a decrease of HK values occurs. In the embryonic brain between 11 and 14 days of incubation, which would correspond for the neuronal cultures to day 3 through 6, modifications of MDH, GDH, HK, and enolase levels are similar to those observed in neurons in culture. Only the increase of LDH activity is less pronounced in vivo than in cultivated cells. The evolution of the tested enzymatic activities in the brain of the chick during the period between 7 days before and 10 days after hatching is quite similar to that observed in cultivated glial cells (prepared from 14 day-old embryos) between 6 and 18 days of culture. All tested activities increased in comparable proportions. The modifications of the enzymatic profile indicate that some maturation phenomena affecting energy metabolism of neuronal and glial elements in culture, are quite similar to those occuring in the total nervous tissue. A relationship between the development of the energy metabolism of the brain and differentiation processes affecting neuroblasts and the glial-forming cells is discussed.     

Effects of the steroid hormone 20-hydroxyecdysone and prior sensory input on the survival and growth of moth central olfactory neurons in vitro

Neurons in the developing (antennal) olfactory lobe of the moth Manduca sexta undergo a period of extensive process outgrowth and branching that coincides temporally with both a rising titer of the steroid hormone 20-hydroxyecdysone and the ingrowth of sensory axons from receptors in the antenna. To evaluate the contribution of these two influences to the morphological development of antennal-lobe neurons, we placed the neurons in cell culture. Antennal-lobe neurons were dissociated from normal and chronically unafferented lobes at different stages of development and were exposed to different doses of hormone. Six neuronal cell types with distinctive and stable morphologies appeared in cultures from all stages of pupal development. Morphological changes in these neuronal types were examined quantitatively by comparison of the total length and number of branches. We found that 20-hydroxyecdysone had little direct effect on the morphological development of antennal-lobe neurons, but brief exposure to sensory axons in vivo prior to dissociation significantly enhanced subsequent outgrowth in culture. © 1993 John Wiley & Sons, Inc.     

Enzyme levels in cultured astrocytes,oligodendrocytes and Schwann cells,and neurons from the cerebral cortex and superior cervical ganglia of the rat

Data are presented for 16 enzymes from 8 metabolic systems in cell cultures consisting of approximately 95% astrocytes and 5% oligodendrocytes. Nine of these enzymes were also measured in cultures of oligodendrocytes, Schwann cells, and neurons prepared from both cerebral cortex and superior cervical ganglia. Activities, in mature astrocyte cultures, expressed as percentage of their activity in brain, ranged from 9% for glycerol-3-phosphate dehydrogenase to over 300% for glucose-6-phosphate dehydrogenase. Creatine phosphokinase activity in astrocytes was about the same as in brain, half as high in oligodendrocytes, but 7% or less of the brain level in Schwann cells and superior cervical ganglion neurons and only 16% of brain in cortical neurons. Three enzymes which generate NADPH, the dehydrogenases for glucose-6-phosphate and 6-phosphogluconate, and the NADP-requiring isocitrate dehydrogenase, were present in astrocytes at levels at least twice that of brain. Oligodendrocytes had enzyme levels only 30% to 70% of those of astrocytes. Schwann cells had much higher lactate dehydrogenase and 6-phosphogluconate dehydrogenase activities than oligodendrocytes, but showed a remarkable similarity in enzyme pattern to those of cortical and superior cervical ganglion neurons.Special issue dedicated to Dr. Lewis Sokoloff.     

Attenuation of neurotoxicity following anoxia or glutamate receptor activation in EGF- and hippocampal extract-treated neuronal cultures

Neurotoxicity following anoxia or glutamate receptor activation was studied in primary neuronal cultures grown in serum-free, chemically defined CDM R12 medium. Exposure to 1 mM KCN, 0.5 mM kainic acid and 0.5 mM N-methyl-D-aspartate led to progressive neuronal degeneration. This damage was quantified by measuring lactate dehydrogenase released in the culture medium. The toxic effects were observed early during the development of the neuronal culture (from 4 days in vitro on) and seemed to be neuron-specific since astrocyte cultures were not affected. Chronic treatment of the neuronal cultures with epidermal growth factor at 10 ng/ml and hippocampal extract at dil. 1/833 (w/v) induced morphological alterations, increased beta-adrenergic receptor coupled adenylate cyclase activity, increased level of total lactate dehydrogenase activity in the case of epidermal growth factor-treated cultures, and attenuation of lactate dehydrogenase release following exposure to KCN or glutamate receptor agonists. The alterations observed are probably due to the proliferation and differentiation of glial cells in these treated cultures. This suggests that glial cells protect neurons in vitro from degeneration induced by anoxia or glutamate receptor activation.     

ELECTROPHORETIC ANALYSIS OF SOLUBLE ENZYMES IN FIVE FRESHWATER DINOFLAGELLATE SPECIES

Electrophoretic analysis of 11 different clonal freshwater Peridinium isolates shows that currently used morphological taxonomic criteria are conservative measures of relatedness and that the presence or absence of an apical pore is an important taxonomic feature. Correlation of malate dehydrogenase and glutamate dehydrogenase isoenzyme patterns with chromosome number variations among our isolates lends no support to polyploidy as a speciation mechanism.     

温度对嗜冷酵母糖代谢途径某些关键酶的活性效应

对嗜冷酵母Y18和酿酒酵母细胞中EMP途径和TCA循环中一些关键酶的温度特性进行了比较研究。Y18细胞中,1,6二磷酸果糖醛缩酶、琥珀酸脱氢酶和己糖激酶对温度很敏感,符合Feller提出的冷活性的概念属于冷活性酶类。柠檬酸合成酶的温度特性类似于中温酶。Α酮戊二酸脱氢酶存在不同温度特性的同功酶。通过对嗜冷酵母和中温酶母细胞中琥珀酸脱氢酶的Km值进行比较,结果显示嗜冷酵母琥珀酸脱氢酶在20℃具有较低的Km值。另外还对嗜冷菌细胞中代谢酶类的一些特点进行了讨论。     

Light- and electron-microscopic study of electrophysiologically characterized neurons in the mediolateral part of the lateral septum of the guinea-pig

In slices of guinea-pig brains, 36 neurons located in the mediolateral part of the lateral septum were stained intracellularly with horseradish peroxidase (n=28) or biocytin (n=8) after electrophysiological characterization. These neurons belonged to class A neurons (n=23), which generated pronounced Ca⁺⁺-dependent high-threshold spikes in control medium, or to class C neurons (n=9), which were recognized by the occurrence of small-amplitude sodic spikes followed by slower larger calcic spikes. The present results demonstrate that, despite the variety of individual cell types, the major morphological population (30/36 cells) was composed of a homogeneous class of large-sized neurons that displayed thick primary dendrites and abundant dendritic appendages. The remaining 6 cells were small-sized, poorly-spiny neurons. Somatic spines were observed on 5 out of the 30 large cells and on one out of the six smaller cells. Labeled axons were mainly oriented to the anterior commissure. The axons of nine cells richly collateralized near the perikaryon. Ultrastructural examination of 3 horseradish peroxidase-injected cells showed indented nuclei, classic organelles and somatic spines. Terminal boutons established symmetric synapses with the injected cells. These results describe the morphological features of electrophysiologically identified neurons and indicate that class A and class C neurons are distributed among morphological populations differing in perikaryal size. This suggests that the different electrical properties of class A and class C neurons reflect recordings from different parts of the neuron rather than from neurons of different types. Furthermore, the present findings demonstrate that, in the guinea-pig, electrical and morphological characteristics of somatospiny neurons are comparable with those of non-somatospiny neurons. Somatospiny neurons have a recognized integrative role in the hippocampo-septo-hypothalamic complex.     

电磁脉冲辐照大鼠海马区细胞凋亡与形态学变化

以体外原代培养的大鼠海马神经元和Wistar大鼠为研究对象,探讨电磁脉冲(场强为6× 104 V/m)辐照后早期海马区细胞凋亡和病理形态学的变化.在照射后1h、6h、12h、24h和48h分别采用MTT法和流式细胞仪测定死亡细胞和凋亡细胞的比例,用光镜和电镜分别进行形态学观察.结果显示在电磁脉冲辐照后,海马神经细胞不仅发生快速的坏死,而且还发生凋亡,同时在早期即可见到血管、胶质细胞和神经元等组织的形态学异常.表明大鼠大脑受电磁脉冲辐照后早期海马区可发生神经细胞坏死和凋亡,以及各组织成分的病理形态学改变,上述变化可能与电磁脉冲致细胞DNA损伤有关.     

FATE OF l-GLUTAMATE IN THE BRAIN

Abstract— It is shown, using aminooxyacetate as metabolic inhibitor, that the process of oxidation of endogenous glutamate in incubated rat brain cortex slices follows a different course from that of exogenous l -glutamate. Whereas endogenous glutamate is largely oxidized by an initial reaction with glutamate dehydrogenase with release of ammonia, exogenous l -glutamate undergoes initial transamination to aspartate and α-oxoglutarate before oxidation occurs. In the presence of 2·5 m m l -glutamate, it is found that, of the total exogenous glutamate utilized, 49 per cent is converted to aspartate, 37 per cent is converted to glutamine and the rest is f uily oxidized through glutamate dehydrogenase. It is suggested that endogenous glutamate is normally oxidized in the neurons, and that glutamate released from neurons during excitation, and acting therefore as exogenous glutamate, is taken up by the glia where, besides conversion to glutamine, it largely undergoes initial transamination before oxidation takes place.     

大鼠尾壳核头部生长抑素mRNA的区域性表达

目的　观察尾壳核 (CP)头部背内侧区、背外侧区、腹内侧区和腹外侧区生长抑素 (SOM )mRNA阳性神经元分布特点。方法　采用原位杂交组织化学方法。结果　CP头部不同区域SOMmRNA阳性神经元的密度存在差异 ,腹内侧区SOMmRNA阳性神经元的密度明显高于其他区 (P <0 0 1)。结论　CP头部不同区域的SOMmRNA阳性神经元密度存在差异 ,这可能是CP头部不同区域机能差异的形态学基础之一。     

Developmental genetics of teleosts: a biochemical analysis of lake chubsucker ontogeny

Developing embryos of the lake chubsucker, Erimyzon sucetta, were analyzed with regard to both gross morphological changes and specific enzymatic changes from the unfertilized egg stage until some 3 weeks posthatching. Total activities of three enzymes—lactate dehydrogenase, glucose-6-phosphate dehydrogenase, and isocitrate dehydrogenase—were determined throughout the course of development. Each of these different enzymes exhibited a different pattern of change during ontogeny. Electrophoretic analysis of qualitative changes in isozyme patterns was accomplished for these three enzymes and for α-amylase, glucosephosphate isomerase, mannosephosphate isomerase, creatine kinase, esterase, glutamate dehydrogenase, alkaline phosphatase, aspartate aminotransferase, malate dehydrogenase, hexose diphosphatase, phosphoglucomutase, and phosphogluconate dehydrogenase. Many of the enzyme systems investigated exhibited rich patterns of ontogenetic change, while a few remained relatively unchanged throughout the interval studied. Several of the enzymes in particular metabolic pathways exhibited coincident changes suggestive of coordinate control. The appearance of several rather “tissue-specific” isozymes was closely correlated with the morphological and functional differentiation of these particular tissues or organs.     

Postnatal morphological changes in neurons and glial cells in neocortex of mice developing at the background of serotonin deficit

It has been shown that deficit of serotonin during embryogenesis in rodents is accompanied by changes of morphological characteristics of neurons and glial cells at the period of postnatal development. A characteristic peculiarity of these changes is cell vacuolization that is of different expression in various cortical layers. In the experimental animals as compared with control ones, neurons of all neocortex layers have changed nuclei and a reduced volume of the cytoplasm. In neurons of upper layers, nuclei and cytoplasm contain occasional small vacuoles. In deep layers, vacuolization both of nuclei and of the cytoplasm is expressed to the much greater degree and vacuoles of large size are predominant. Results of immunocytochemical study have shown that in animals developing at the background of serotonin deficit there takes place a delay of the rates of formation and differentiation of astrocytic glia.