Myenteric nitrergic neurons along the rat esophagus: evidence for regional and strain differences in age-related changes

Several studies have suggested an age-related reduction in the number of myenteric neurons in the lower gastrointestinal (GI) tract linked to changes in GI neuromuscular functions. The present study, combining protein gene product 9.5 immunostaining and NADPH-diaphorase histochemistry, aimed at quantifying the proportion of nitrergic neurons compared to the overall number of enteric neurons in the esophagus of young (3-4.5 months) and aged (18-20 months) Sprague-Dawley and Wistar rats. In both strains, the neuron numbers per ganglion in the cervical region were almost twice as high as in the other esophageal regions. Irrespective of age or strain, the esophagus harbored a very high proportion of intrinsic nitrergic neurons (greater than approximately 65%). Both strains showed with aging an overall neuronal loss of approximately 27%. While a significant increase (young: 64-71%; aged: 82-89%) was observed in all esophageal regions in the Wistar strain, the proportion of nitrergic neurons remained stable with aging in the Sprague-Dawley strain (range: 72-82%). In conclusion, the age-related reduction in the overall number of myenteric, nitrergic, and non-nitrergic neurons observed in the rat esophagus, appears to be highly region- and strain-dependent. Therefore, a protective mechanism against neuronal cell loss, selectively present in specific (nitrergic) enteric subpopulations, as suggested in earlier reports, cannot be put forward as a general phenomenon throughout the entire GI tract.     

Immunohistochemical Evidence for the Presence of Synaptic Connections of Nitrergic Neurons in the Rat Rostral Migratory Stream

The rostral migratory stream (RMS) is a migration route for neuroblasts originating in the richest neurogenic niche of the adult mammalian brain—the subventricular zone. Most studies are focused on cellular dynamics of migrating neuroblasts and interactions between neuroblasts and astrocytes which both represent the major cellular component of the RMS. Our previous experiments have brought evidence about the existence of a small population of mature neurons in the adult rat RMS with capacity to produce nitric oxide (NO). In order to further support functional significance of nitrergic cells, the aim of the present study was to determine whether NO producing neurons could form synapses. Sagittal sections from the adult rat brain were processed for simultaneous immunohistochemical detection of neuronal nitric oxide synthase (nNOS), the enzyme present in NO producing cells and synaptophysin, a glycoprotein found in synaptic vesicles. Synaptophysin positivity in the RMS was significantly lower in comparison with other brain areas, but its colocalization with nNOS-positive neurons was obvious. Our results suggest that nitrergic neurons in the RMS could be involved in a neuronal circuitry with potential impact on regulation of neurogenesis in the RMS.     

Methylene Blue Protects Astrocytes against Glucose Oxygen Deprivation by Improving Cellular Respiration

Astrocytes outnumber neurons and serve many metabolic and trophic functions in the mammalian brain. Preserving astrocytes is critical for normal brain function as well as for protecting the brain against various insults. Our previous studies have indicated that methylene blue (MB) functions as an alternative electron carrier and enhances brain metabolism. In addition, MB has been shown to be protective against neurodegeneration and brain injury. In the current study, we investigated the protective role of MB in astrocytes. Cell viability assays showed that MB treatment significantly protected primary astrocytes from oxygen-glucose deprivation (OGD) & reoxygenation induced cell death. We also studied the effect of MB on cellular oxygen and glucose metabolism in primary astrocytes following OGD-reoxygenation injury. MB treatment significantly increased cellular oxygen consumption, glucose uptake and ATP production in primary astrocytes. In conclusion our study demonstrated that MB protects astrocytes against OGD-reoxygenation injury by improving astrocyte cellular respiration.     

Uptake,Metabolic Effects and Toxicity of Arsenate and Arsenite in Astrocytes

The inorganic arsenic species arsenate and arsenite are common environmental toxins which contaminate the drinking water in many countries. Chronic intoxication with arsenicals has been connected with various diseases, but causes also neurological complications and impairs cognitive development, learning and memory. In brain, astrocytes have a pivotal role as partners of neurons in homeostatic and metabolic processes. In addition, astrocytes are the first parenchymal brain cell type which encounters substances which cross the blood–brain barrier and are considered as first line of defence against the toxic potential of xenobiotics. Therefore, astrocytes are likely to play a prominent role in the metabolism and potential detoxification of arsenicals in brain. This article summarizes the current knowledge on the uptake and toxicity of arsenate and arsenite in astrocytes and discusses the modulation of the astrocytic glucose and glutathione metabolism by arsenicals.     

人胎大肠氮能神经元发育的研究

By using histochemical methed of NADPH-diaphorse, the development of the nitrergic neurons in the large intestine of human fetus were studied. The results showed: At the fifth month of gestation, weak positive reaction of nitric oxide synthase (NOS) appeared in part of the round cells of intermuscular ganglia. The round cells differentiated into the nitrergic nerve cells. At the sixth month, the bodies of nitrergic neurons were obviously enlarged, the processes of which were lengthened. The nitrergic nerve fibers were seen in the muscle layer, the submucosa and the base of the intestinal gland. The growth and development of nitrergic neurons reached its peak at the seventh month. The number of intermuscular ganglionic cells was increased. The density of nitrergic nerve fibers was increased in the inner circular muscle layer, and have bead-like structures. At the eighth to tenth month, the staining intensity of nitrergic neurons was increased. The myenteric plexus was densely distributed with nitrergic nerve cell bodies, whereas the submucosa and the inner circular muscle layers contained only a few neurons. The nitrergic nerve fibers were observed in all layer of large intestine, the density of the distribution of nitrergic nerve fibers was by far the highest in the inner circular muscle layer, less in the submucosa and outer longitudinal muscle layer, and only a few were found in the mucous layer. To our knowledge, it is the first time that the development of nitrergic neurons in the large intestine of human fetus was demonstrated.     

人胎大肠氮能神经元发育的研究

用NADPH-d组织化学法对人胎大肠氮能神经元的发育进行了观察.结果表明第5个月胎龄时,肌间神经节处圆形细胞中部分细胞出现一氧化氮合酶(NOS)阳性反应,并分化成氮能神经细胞.第6个月胎龄时,氮能神经元胞体增大,突起伸长,在肌层、粘膜下层和肠腺基部出现氮能神经纤维分布.第7个月胎龄时,氮能神经元生长发育达到高峰,肌间神经节细胞数目增多,环肌层神经纤维分布密度增加,膨体结构明显.第8-10个月胎龄时,氮能神经元染色强度加深,其胞体分布以肌间神经节最多,粘膜下层和内环肌层较少.氮能神经纤维的分布密度以内环肌层最高,粘膜下层和外纵肌层次之,粘膜层较低.本研究揭示了大肠氮能神经元发育的变化规律.     

NADPH-diaphorase, nitric oxide synthase, and tyrosine hydroxylase in the diencephalon of the Rhodeus sericeus (Cyprynidae:Teleostei)

The presence and distribution of nitric oxide synthase (NOS)-like neurons as well as tyrosine hydroxylase-immunoreactive (TH) neurons was studied in the diencephalon of the cypriniform teleost Rhodeus sericeus. The anatomical relationships between tyrosine hydroxylase (TH)- and nitric oxide synthase (NOS)-containing cells were visualized both by NOS-immunohistochemistry and NADPH-histochemistry. Immunohistochemical labeling and morphological studies were performed on the same sections. The results reported in this paper show that both a NOS and TH activity are present in the preoptic region, posterior tuberculum, paraventricular organ and hypothalamus of R. sericeus. Putative nitrergic neurons were identified in all major hypophysiotrophic nuclei of the R. sericeus brain using both NADPH-d histochemistry and nNOS immunohistochemistry. In the preoptic region, nitrergic neurons were found in both the parvocellular and the magnocellular nuclei. Within these nuclei, the distribution of NADPH-d reactivity was similar to that of nNOS immunoreactivity. However, we found no evidence of colocalization of NADPH-d and nNOS in consecutive sections. NOS- and TH-containing neurons were observed in all the nuclei under study (hypothalamus, posterior tuberculum, ventral thalamus) and telencephalon (preoptic region), although most neurons showing the coexistence of both substances were mainly located in the preoptic nucleus and hypothalamus, some labelled neurons were found in the posterior tuberculum. Most of the cerebrospinalliquor-contacting cells (LCNs) in diencephalic periventricular area of R. sericeus were TH-immunoreactive. Also, a large number ofnitrergic small LCNs distributed throughout the third ventricle were observed in these regions. The data obtained supports the existence of a nitrergic circumventricular system in teleost. LCNs in R. sericeus are thought to be involved in osmoregulatory functions as osmosensitive neurons. Due to their chemical properties, NO produced by these cells might play an important role in the maintenance and regulation of CSF homeostasis through the modulation of cerebral blood flow.     

Modulation of Nrf2/ARE Pathway by Food Polyphenols: A Nutritional Neuroprotective Strategy for Cognitive and Neurodegenerative Disorders

In recent years, there has been a growing interest, supported by a large number of experimental and epidemiological studies, for the beneficial effects of some phenolic substances, contained in commonly used spices and herbs, in preventing various age-related pathologic conditions, ranging from cancer to neurodegenerative diseases. Although the exact mechanisms by which polyphenols promote these effects remain to be elucidated, several reports have shown their ability to stimulate a general xenobiotic response in the target cells, activating multiple defense genes. Data from our and other laboratories have previously demonstrated that curcumin, the yellow pigment of curry, strongly induces heme-oxygenase-1 (HO-1) expression and activity in different brain cells via the activation of heterodimers of NF-E2-related factors 2 (Nrf2)/antioxidant responsive element (ARE) pathway. Many studies clearly demonstrate that activation ofNrf2 target genes, and particularly HO-1, in astrocytes and neurons is strongly protective against inflammation, oxidative damage, and cell death. In the central nervous system, the HO system has been reported to be very active, and its modulation seems to play a crucial role in the pathogenesis of neurodegenerative disorders. Recent and unpublished data from our group revealed that low concentrations of epigallocatechin-3-gallate, the major green tea catechin, induces HO-1 by ARE/Nrf2 pathway in hippocampal neurons, and by this induction, it is able to protect neurons against different models of oxidative damages. Furthermore, we have demonstrated that other phenolics, such as caffeic acid phenethyl ester and ethyl ferulate, are also able to protect neurons via HO-1 induction. These studies identify a novel class of compounds that could be used for therapeutic purposes as preventive agents against cognitive decline.     

Tetracycline derivatives and ceftriaxone, a cephalosporin antibiotic, protect neurons against apoptosis induced by ionizing radiation

DNA damage induced by low doses of ionizing radiation causes apoptosis, which is partially mediated via the generation of free radicals. Both free radicals and apoptosis are involved in the majority of brain diseases, including stroke, Alzheimer's disease and amyotrophic lateral sclerosis. Because previous studies have shown that tetracycline derivatives doxycycline and minocycline have anti-inflammatory effects and are protective against brain ischemia, we studied whether minocycline and doxycycline or ceftriaxone, a cephalosporin antibiotic with the potential to inhibit excitotoxicity, protect neurons against ionizing radiation in primary cortical cultures. A single dose of 1 Gy significantly increased lactate dehydrogenase release, induced DNA fragmentation in neurons and triggered microglial proliferation. Treatment with minocycline (20 nM), doxycycline (20 nM) and ceftriaxone (1 microM) significantly reduced irradiation-induced lactate dehydrogenase release and DNA fragmentation. The most efficient protection was achieved by minocycline treatment, which also inhibited the irradiation-induced increase in microglial cell number. Our results suggest that some tetracycline derivatives, such as doxycycline and minocycline, and ceftriaxone, a cephalosporin derivative, protect neurons against apoptotic death.     

Localization and fate of aluminium in the digestive gland of the freshwater snail Lymnaea stagnalis

The digestive gland of the freshwater snail Lymnaea stagnalis, exposed to water containing an elevated concentration of aluminium at neutral pH for up to 30 days, followed by a 20 day recovery period, was examined by light and electron microscopy and X-ray microanalysis. Aluminium was localized in the yellow granules present in the digestive and excretory cells and in the green and small granules present in the digestive cells. More aluminium, silicon, phosphorus and sulphur were present in all three granule types from aluminium exposed snails. The number of yellow and green granules from the digestive gland of aluminium exposed snails showed a progressive increase over the experimental period compared to controls. The number and aluminium content of the granules is likely to reflect the role of the digestive gland as a 'sink' for accumulated aluminium. We propose that intracellular monomeric silica is involved in the detoxification of aqueous aluminium which at neutral pH is largely in the form of an insoluble polyhydroxide. The increased amounts of sulphur and phosphorus in the granules are likely to be part of a broad response to metal loading but probably do not play a significant role in the storage and detoxification of aluminium.     

Postnatal development of nitrergic neurons in the myenteric plexus of rat stomach

The effect of age on the proportion of nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd)-positive neurons was investigated in the myenteric plexus of five different gastric areas of 1-day-, 1-week-, 2-week-, 1-month- and 2-month-old rats. Protein gene product 9.5 immunocytochemistry was used as a marker for the total enteric neuron population in order to establish the percentage of gastric nitrergic neurons in relation to age. The percentage of NADPHd-positive neurons in the proximal parts of the rat stomach (34–38%) is significantly higher than in the antral part (29%). This difference persists in all the age groups investigated. No significant relative increase with age of NADPHd-positive neurons could be observed in any of the areas studied. These findings imply that the increased nitrergic response in the rat proximal stomach as seen in pharmacological studies cannot be explained by an increased relative number of nitrergic neurons. Accepted: 31 March 1999     

The prevention of lung cancer induced by a tobacco-specific carcinogen in rodents by green and black Tea

A growing body of evidence from studies in laboratory animals indicates that green tea protects against cancer development at various organ sites. We have previously shown that green tea, administered as drinking water, inhibits lung tumor development in A/J mice treated with 4-(methylnitrosamino)-1-(3-pyridyl)-l-butanone (NNK), a potent nicotine-derived lung carcinogen found in tobacco. The inhibitory effect of green tea has been attributed to its major polyphenolic compound, epigallocatechin gallate (EGCG), and, to a lesser extent, to caffeine. We have also demonstrated that while levels of O6-methylguanine, a critical lesion in NNK lung tumorigenesis, were not affected in lung DNA. However, the levels of 8-hydroxydeoxyguanosine (8-OH-dG), a marker of oxidative DNA damage, were significantly suppressed in mice treated with green tea or EGCG. These studies underscore the importance of the antioxidant activity of green tea and EGCG for their inhibitory activity against lung tumorigenesis. Unlike green tea, the effect of black tea on carcinogenesis has been scarcely studied, even though the worldwide production and consumption of black tea far exceeds that of green tea. The oxidation products found in black tea, thearubigins and theaflavins, also possess antioxidant activity, suggesting that black tea may also inhibit NNK-induced lung tumorigenesis. Indeed, bioassays in A/J mice have shown that black tea given as drinking water retarded the development of lung cancer caused by NNK. However, data on the relationship of black tea consumption with the lung cancer risk in humans are limited and inconclusive. There is a need for additional tumor bioassays in animal models to better examine the protective role of black tea against lung cancer. The development of adenocarcinomas and adenosquamous carcinomas in F344 rats upon chronic administration of NNK provides an important and relevant model for lung carcinogenesis in smokers. Thus far, no information was previously available regarding the effects of tea on this model. We conducted a 2-year lifetime bioassay in F344 rats to determine whether black tea and caffeine are protective against lung tumorigenesis induced by NNK. Our studies in both mice and rats have generated important new data that support green and black tea and caffeine as potential preventive agents against lung cancer, suggesting that a closer examination of the roles of tea and caffeine on lung cancer in smokers may be warranted.     

Activation of spinally projecting and nitrergic neurons in the PVN following heat exposure

The present study investigated the effect of acute thermal stimulation in conscious rats on the production of Fos, a marker of increased neuronal activity, in spinally projecting and nitrergic neurons in the hypothalamic paraventricular nucleus (PVN). The PVN contains a high concentration of nitrergic neurons, as well as neurons that project to the intermediolateral cell column (IML) of the spinal cord that can directly influence sympathetic nerve activity (SNA). During thermal stimulation, the PVN is activated, but it is unknown whether spinally projecting PVN neurons and the nitrergic neurons are involved. Compared with controls, rats exposed to an environmental temperature of 39 degrees C for 1 h had a 10-fold increase in the number of cells producing Fos in the PVN (133 +/- 23 vs. 1,336 +/- 43, respectively, P < 0.0001). Of the spinally projecting neurons in the PVN of heated rats (98 +/- 10), over 20% expressed Fos. Additionally, of the nitrergic neurons (NADPH-diaphorase positive) located in the parvocellular PVN (723 +/- 17), 40% also expressed Fos (P < 0.0001 compared with controls). Finally, there was a significant increase in the number of spinally projecting neurons in the PVN that were nitrergic and expressed Fos after heat exposure (12%) compared with controls (0.1%) (P < 0.0001). These results suggest that spinally projecting and nitrergic neurons in the PVN may contribute to the central pathways activated by thermal stimulation.     

Model studies of the precipitation of silica in the presence of aluminium; implications for biology and industry

The unique chemical affinity between the oxides of silicon and aluminium has been cited as a potential route for the amelioration of the detrimental effects of aluminium in the environment and in biological systems. A greater understanding of silicon-aluminium interactions may assist in this endeavour and also provide a means of overcoming silica fouling problems encountered by industry which are exacerbated by the presence of aluminium. It is also conceivable that this increased knowledge may demonstrate a positive use for aluminium in the processing of the silicon dioxide phase. In this study we report the effect of aluminium ions, derived from aluminium chloride, on silicic acid species obtained from potassium catecholato complexes of silicon at circumneutral pH at the molar ratios 1000Si:Al, 100Si:Al and 50Si:Al. Silica and low levels of aluminium-rich silica materials were formed with Si:Al ratios of about 3.5:1 comparable with the element ratios detected in senile plaques and aluminium-rich scale. A kinetic study showed that aluminium in the reaction medium slowed down the rate of formation of one of the silica species formed early in the condensation process, e.g. trimers, but increased the rate at which silicic acid was removed from sub 1 nm diameter particles. The materials precipitated in the presence of aluminium were composed of smaller particles and aggregates with smaller pores (Si100:Al and Si50:Al systems) or larger pores (Si1000:Al) compared to the control. The nature of the interactions responsible for these differences is discussed. The effects described here demonstrate the ability of silica and aluminium to interact under conditions such as those found in biological systems. That silica reacts with aluminium in the presence of catechol supports the protective role assigned to silicon.     

吸氧预处理对大鼠脑缺血再灌注损伤的保护作用

目的探讨吸氧预处理对大鼠脑缺血再灌注损伤的保护作用。方法通过大鼠局灶脑缺血再灌注损伤模型,采用SOD、MDA测定、电镜及神经行为学检查的方法,观察吸氧预处理对大鼠脑缺血再灌注损伤后SOD、MDA、神经行为学评分及脑组织病理变化。结果吸氧预处理组SOD活力高于对照组(P<0.05),MDA含量、神经行为学评分均低于对照组(P<0.05),脑组织超微结构损伤均减轻。结论吸氧预处理对大鼠脑缺血再灌注损伤有保护作用。     

Ginkgolide B preconditioning on astrocytes promotes neuronal survival in ischemic injury via up-regulating erythropoietin secretion

Although ischemic preconditioning (IP) can provide powerful protection on brain against ischemic insult, it is rarely used in clinic to prevent the occurrence of ischemic stroke because of safety concerns. It is therefore necessary to seek the safer stimuli to initiate pharmacological preconditioning. Our previous work demonstrated that ginkgolide B (GB) could protect neurons against ischemia-induced apoptosis. Astrocytes are the most numerous cells in mammalian central nervous system and there is a close bi-directional communication between neurons and astrocytes in brain. Besides neurons, whether GB can exert the role of preconditioning on astrocytes through which to further improve neuronal survival under ischemic condition is not yet known. In the present study, primary cultured astrocytes were treated with GB for 24 h or short-term ischemia (ischemia for 3 h, as ischemic preconditioning/IP), and then cultured back to normoxia and normal medium for 24 h to induce the preconditioning response. Astrocyte-conditioned medium (ACM) was then collected and used to incubate the cultured neurons for 24 h before neurons were subjected to severe ischemia. Our results demonstrated that not only GB and IP increased astrocytic viability in ischemia, but also the conditioned medium from astrocytes treated with GB or IP increased cell viability and decreased the number of apoptosis of neurons in ischemia. We also found that GB and IP significantly stimulated astrocytes to express and secrete erythropoietin (EPO) into ACM, and the addition of anti-EPO antibody blocked the protective effect of GB or IP-treated astrocytes culture medium on neurons in ischemia. Further study of above protection revealed that ACM from astrocytes treated with GB or IP induced the inactivation of proapoptotic factor Bad by phosphorylation at serine 136 and 112 (¹³⁶p-Bad and ¹¹²p-Bad) in neurons. Together, our results suggest that GB is capable of preconditioning on astrocytes as IP and then protects neurons against ischemia-induced apoptosis, which is mediated by EPO.     

Evidence supporting a role for N-acetyl-L-aspartate as a molecular water pump in myelinated neurons in the central nervous system. An analytical review.

Molecular water pumps (MWPs) are characterized as biochemical systems existing at a compartmental boundary of living cells that can actively pump water against its gradient. A role for the observed intercompartmental transport of N-acetyl-L-aspartate (NAA), between neurons and oligodendrocytes in the CNS, as an efflux MWP for the removal of neuronal metabolic water has been proposed. In this review, accumulating evidence in support of such a role for NAA is presented, and the dynamics of the NAA cycle in myelinated neurons are considered. Based on the results of recent investigations, it is calculated that 1 mol of NAA is synthesized for every 40 mol of glucose (Glc) equivalent oxidized in the brain, and each mol of NAA may transport 121 mol of metabolic water out of neurons. In addition, turnover of total brain NAA is very rapid and appears to be only 16.7 h. Thus, the most important characteristic of NAA in the brain may not be its static level, but a dynamic aspect related to its rapid turnover. The relationship of NAA as a potential MWP to Canavan disease (CD), a genetic spongiform leukodystrophy in which the catabolic portion of the NAA cycle is deficient, and in a newly recognized brain disorder, hypoacetylaspartia, where the anabolic portion of the NAA cycle appears to be deficient, are discussed.     

Nitric oxide synthase in the glossopharyngeal and vagal afferent pathway of a teleost, Takifugu niphobles

To examine the presence of nitric oxide synthase (NOS) in the sensory system of the glossopharyngeal and vagus nerves of teleosts, nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd) activity and immunoreactivity for NOS were examined in the puffer fish Takifugu niphobles. The nitrergic sensory neurons were located in the ganglia of both the glossopharyngeal and the vagal nerves. In the vagal ganglion, positive neurons were found in the subpopulations for the branchial rami and the coelomic visceral ramus, but not for the posterior ramus or the lateral line ramus. In the medulla, nitrergic afferent terminals were found in the glossopharyngeal lobe, the vagal lobe, and the commissural nucleus. In the gill structure, the nitrergic nerve fibers were seen in the nerve bundles running along the efferent branchial artery of all three gill arches. These fibers appeared to terminate in the proximal portion of the efferent filament arteries of three gill arches. On the other hand, autonomic neurons innervating the gill arches were unstained. These results suggest that nitrergic sensory neurons in the glossopharyngeal and vagal ganglia project their peripheral processes through the branchial rami to a specific portion of the branchial arteries, and they might play a role in baroreception of this fish. A possible role for nitric oxide (NO) in baroreception is also discussed.     

Neuroprotective effect of quercetin against radiation‐induced endoplasmic reticulum stress in neurons

The endoplasmic reticulum (ER) plays an important role in the regulation and maintenance of cellular homeostasis. However, unresolved ER stress leads to deleterious effects by inducing the accumulation of unfolded proteins in the cell. Here we have demonstrated the protective aspects of quercetin against radiation‐induced ER stress and against inflammation in primary cultured dorsal root ganglion (DRG) neurons. The mature DRG neurons were pretreated with different concentrations of quercetin (5‐100 μM) for 24 hours before 2 Gy gamma radiation exposure and then subjected to a cytotoxicity assay, quantitative real‐time polymerase chain reaction and Western blot analysis. The results showed that quercetin decreased the expression of BiP and C/EBP‐homologous protein, the ER stress marker genes along with downregulation of tumor necrosis factor‐α, JNK in irradiated DRG neurons. Furthermore, quercetin pretreatment significantly increased the cytoskeletal protein Tuj1 and the neurotrophin brain‐derived neurotrophic factor in the neuron. These results indicate that quercetin plays a neuroprotective role against radiation‐mediated ER stress and inflammatory responses.     

Mitochondrial Malic Enzyme: Purification from Bovine Brain, Generation of an Antiserum, and Immunocytochemical Localization in Neurons of Rat Brain

Abstract: To elucidate the cellular location of mitochondrial malic enzyme in brain, immunocytochemical studies were performed. For this purpose, mitochondrial malic enzyme was purified to apparent homogeneity from bovine brain and used for the immunization of rabbits. Subjecting the antiserum to affinity purification on immobilized antigen as an absorbent yielded a purified immunoreactive antibody preparation, which was characterized by probing cytosolic and mitochondrial fractions of bovine and rat brain in western blotting. As neither crossreactivity with cytosolic malic enzyme nor immunoreactivity against other proteins could be observed, the antibody preparation was found suitable for immunocytochemistry. By using sections of perfusion-fixed rat brain, considerable resolution was achieved at the light-microscopic level. Distinct and specific staining of neurons was observed; in contrast, no staining of astrocytes and possibly unspecific staining within the nuclei of oligodendrocytes were obtained. From these data, it is concluded that mitochondrial malic enzyme is located in neurons; however, in astrocytes, the enzyme appears to be either lacking or present at a much lower level. A protective role against oxidative stress in neurons is proposed for mitochondrial malic enzyme.