Immunocytochemical localization of acyl-CoA oxidase in the rat central nervous system

Peroxisomal β-oxidation, consisting of four steps catalysed by an acyl-CoA oxidase, a multifunctional protein and a thiolase, is responsible for the shortening of a variety of lipid compounds. The first reaction of this pathway is catalysed by a FAD-containing acyl-CoA oxidase, three isotypes of which have been so far recognised. Among these, straight-chain acyl-CoA oxidase (ACOX) acts on long and very long chain fatty acids, prostaglandins and some xenobiotics. We investigated ACOX localisation by means of a sensitive, tyramide based, immunocytochemical technique, thus obtaining a complete distribution atlas of the enzyme in adult rat CNS. Granular immunoreaction product was found in the cytoplasm of neuronal and glial cells, both in the perikarya and in the cell processes. ACOX immunoreactive neurons were present to variable extent, in either forebrain or hindbrain areas. Specifically, the strongest signal was detected in the pallidum, septum, red nucleus, reticular formation, nuclei of the cranial nerves, and motoneurons of the spinal cord. We then compared the ACOX immunoreactivity pattern with our previous distribution maps of other peroxisomal enzymes in the adult rat brain. While ACOX appeared to colocalise with catalase in the majority of cerebral regions, some differences with respect to d-amino acid oxidase were noted. These observations support the hypothesis of heterogeneous peroxisomal populations in the nervous tissue. The wide distribution of the enzyme in the brain is consistent with the severe and generalised neurological alterations characterising the peroxisomal disorder caused by ACOX deficiency (pseudo-neonatal adrenoleukodystrophy).     

Immunohistochemical localization of minor gangliosides in the rat central nervous system

We previously described the differential distribution of majorgangliosides (GM1, GD1a, GD1b, GT1b and GQ1b) in adult rat braindetected by specific antibodies (Kotani,M., Kawashima,I., Ozawa,I.,Terashima,T. and Tai,T. Glycobiology, 3, 137–146, 1993).We report here the distribution of minor gangliosides in theadult rat brain by an immunofluorescence technique with mousemonoclonal antibodies (MAbs). Ten MAbs (GMR6, GMB28, GMR11,GMR19, GMR2, GMR7, GGR51, AMR10, NGR54 and NGR53) that specificallyrecognize GM3, GM2, GT1a, GD3, O-Acdisialoganglioside, GD2,GM1b, GM4, IV³NeuAc     

Immunohistochemical localization of Bis protein in the rat central nervous system

We studied the distribution of Bis (Bcl-2 interacting death suppressor) protein in the adult rat brain and spinal cord using immunohistochemistry. Immunoreactivity was observed in specific neuronal populations in distinct nuclei. The most intensely labeled cells were associated with the motor system, including most cranial nerve motor nuclei, Purkinje cells of the cerebellum, the red nucleus, and the ventral motor neurons of the spinal cord. Bis protein was also expressed in several structures associated with the ventricular system, including the subventricular zone of the lateral ventricle and its rostral extension, in the subcommissural organ, and in tanycytes, radial glial cells in the hypothalamus. Using double-labeling techniques, Bis-immunoreactive cells in the rostral migratory stream, coexpressing Bcl-2, were confirmed as glial fibrillary acidic protein-positive astrocytes comprising the glial tubes. The widespread distribution of Bis suggests that this protein has broader functions in the adult rat central nervous system than previously thought, and that it could be associated with a particular role in the rostral migratory system.J.-H. Lee and M.-Y. Lee contributed equally to this study. This work was supported by the KOSEF through the Cell Death Disease Research Center of MRC at the Catholic University of Korea (R13-2002-005-01001-0) and the Catholic Medical Center Research Foundation grant made in the program year of 2002     

On the ultrastructural localization of noradrenaline in the central nervous system of the rat

Summary Isolated cells were prepared from microdissected pancreatic islets of guinea-pigs. Phase-contrast microscopy of the fresh islet cells suspended in a balanced salt solution displayed a number of cellular details, including cytoplasmic secretion granules. There was morphologic evidence of the survival of islet cells in monolayer cultures for up to 3 weeks. A moderate proliferation of cells occurred during the first 2 weeks after explantation. Different types of islet cells could not be distinguished in the phase-contrast microscope.Research carried out with the financial support of the Swedish Medical Research Council (B 67-12 X-109-03), the Medical Faculty of Uppsala University and the U. S. Public Health Service (Grant AM-05759-05). We thank Assistant Professor Björn Sandström, M. D., for much advice on the tissue-culture method.     

Immunocytochemical localization of the precursor protein for beta-amyloid in the rat central nervous system

Two rabbit polyclonal antibodies generated against different portions of the amyloid precursor protein were used to localize this protein in normal rat brain. Light and electron microscopic immunohistochemical localizations demonstrate that the protein is widely distributed throughout the neuraxis, with the highest concentrations of immunoreactive neurons occurring in the olfactory bulb, cerebral cortex, septum-diagonal band, globus pallidus, cerebellum, and hippocampus. Immunoreactive astrocytes are also present in the cerebral cortex in relation to both neurons and capillaries. However, immunoreactivity was not observed within the endothelium of the cerebral vasculature. These data demonstrate that the beta-amyloid precursor is widely distributed in the CNS and provide further insight into the cellular elements that may be involved in the neuropathological changes associated with Alzheimer's disease.     

Elastic properties of the rat respiratory system related to age

The experiments were performed on male rats of the Wistar strain under urethane anaesthesia (1.3 g/kg i.p.). Changes of oesophageal and tracheal pressures were registered in a group of 30 spontaneously breathing, supine rats, of 295 +/- 13 g average body weight during lung inflations with 1-5 ml of air. In another group of 25 rats of 70 +/- 6 g average body weight (young rats) we made the measurements during inflation with volumes 0.5-2 ml. The measurements were also performed in a group of 10 paralyzed, ventilated rats with 347 +/- 24 g average body weight and inflations 1-5 ml. Compliance of the lungs (CL), chest wall (CW) and of the respiratory system (Crs) was calculated from the linear part of the pressure-volume curve during inflation. The results indicate: 1. Cw is significantly (p less than 0.001) higher in young (134.7 ml.kPa-1.kg-1) than in adult rats (44.1 ml.kPa-1.kg-1). CL (related to body weight) is not significantly different in young and adult rats. 2. Cw is significantly (p less than 0.001) higher than CL. 3. No difference was observed in CLs Cw and Crs between paralyzed and spontaneously breathing animals.     

Ultrastructural localization of D-amino acid oxidase in microperoxisomes of the rat nervous system.

A recently developed procedure for the localization of D-amino acid oxidase (D-AAO) has been used to investigate the distribution of this enzyme in rat nervous tissue. Initial studies were carried out on kidney to validate the methods. The cytochemically demonstrable enzyme in kidney is inhibited by kojic acid, a known competitive D-AAO inhibitor. Omission of the catalse inhibitor, aminotriazole, from the cytochemical medium produces a marked diminution of D-AAO reaction product in kidney peroxisomes. This would be expected if catalase and D-AAO are present in the same particles. In brain, kojic acid-inhibitable D-AAO is demonstrable in numerous bodies within astrocytes especially in the cerebellum, a brain region known from biochemistry to contain particularly high levels of the oxidase. In preparations incubated for catalase, far fewer positive bodies are seen in the cerebellum. Moreover, omission of aminotriazole has little evident effect on the D-AAO reaction. Thus, the oxidase-containing cerebellar bodies may be relatively poor in catalse. In contrast, several nervous system cell types that contain relatively numerous catalase-positive bodies, contain none with detectable D-AAO. Such heterogeneity of peroxisome enzyme content is in accord with reports from biochemical studies of brain.     

New data on the immunocytochemical localization of thyrotropin-releasing hormone in the rat central nervous system

An antiserum raised against the synthetic tripeptide pyroglutamyl-histidyl-proline (free acid) was used to localize thyrotropin-releasing hormone (TRH) in the rat central nervous system (CNS) by immunocytochemistry. The distribution of TRH-immunoreactive structures was similar to that reported earlier; i.e., most of the TRH-containing perikarya were located in the parvicellular part of the hypothalamic paraventricular nucleus, the suprachiasmatic portion of the preoptic nucleus, the dorsomedial nucleus, the lateral basal hypothalamus, and the raphe nuclei. Several new locations for TRH-immunoreactive neurons were also observed, including the glomerular layer of the olfactory bulb, the anterior olfactory nuclei, the diagonal band of Broca, the septal nuclei, the sexually dimorphic nucleus of the preoptic area, the reticular thalamic nucleus, the lateral reticular nucleus of the medulla oblongata, and the central gray matter of the mesencephalon. Immunoreactive fibers were seen in the median eminence, the organum vasculosum of the lamina terminalis, the lateral septal nucleus, the medial habenula, the dorsal and ventral parabrachial nuclei, the nucleus of the solitary tract, around the motor nuclei of the cranial nerves, the dorsal vagal complex, and in the reticular formation of the brainstem. In the spinal cord, no immunoreactive perikarya were observed. Immunoreactive processes were present in the lateral funiculus of the white matter and in laminae V-X in the gray matter. Dense terminal-like structures were seen around spinal motor neurons. The distribution of TRH-immunoreactive structures in the CNS suggests that TRH functions both as a neuroendocrine regulator in the hypothalamus and as a neurotransmitter or neuromodulator throughout the CNS.     

Presence and localization of molecules related to the cholinergic system in developing rat testis

The presence of cholinergic molecules was recently found in male gametes of different animal species; very little is known from this point of view about the somatic component of the gonad. In the present study, a contribution is given to the localization of some cholinergic-like molecules, such as acetylcholinesterase (AChE), nicotinic acetylcholine and muscarinic acetylcoline (nAchR and mAChR, respectively) receptors during postnatal development of the testis. Maturation stages were checked by use of histochemical stainings, such as DAPI for nuclear shape changes, and PSA binding to reveal acrosomal maturation. The distribution of cholinergic-like molecules, revealed by immunocytochemical methods in both gametes and somatic cells, varied with development. Generally, during early stages, molecules immunologically related to AChE, and to mAChR's were mainly found in the interstitial and peritubular compartment, while, during maturation, they were found in Sertoli cells and in differentiating germ cells. nAChR's were not found in the somatic compartment, except in the blood vessel walls, and were distributed in mature germ cells, mainly in the flagella. The presence of cholinergic molecules in somatic as well as germ cells could play a role in cell-to-cell communications affecting testicular cell differentiation and function.     

Heavy metal accumulation in plants grown in heavily polluted soils

In a polluted area, samples of soils and plants were collected and analyzed in order to assess the mobility and accumulation of Cd, Pb and Zn in plants as influenced by the of different metal forms in soil. Metals were accumulated in roots. Metal mobility in plants decreased in the order Zn>Cd>Pb; it was generally lower in heavily polluted areas. The content of all the three metals in plant roots correlated with topsoil concentration of their 2 mol/L HNO₃-extractable forms. Besides, Cd and Zn correlated with exchangeable and Zn also with the organic form content. The concentration factor as a measure of metal mobility from soil to plants decreased with increasing level of contamination. Presented at the 4th Mini-Symposium on Biosorption and Microbial Degradation, Prague, Czech Republic, November 26–29, 1996.     

Heavy metal accumulation in rice (Oryza sativa) near electronic waste dumps and related human health risk assessment

This research was conducted to assess heavy metal contamination in the environment and within Oryza sativa. The translocation factors (TFs) and bioaccumulation factors (BAFs) for heavy metals in O. sativa and estimated daily intake (EDI) and health risk index (HRI) were measured. The samples were analyzed for heavy metals using inductively coupled plasma optical emission spectrometry (ICP-OES). Pb and Cr concentrations in water samples within and near the electronic-waste dumping area exceeded water quality standards for surface water sources from the Pollution Control Department in the Ministry of Natural Resources and Environment of Thailand (PCD). The Pb concentration in soil samples within the area also exceeded soil quality standards for habitat and agriculture from PCD. Most of the metals were highly concentrated in roots, except for Mn which has the highest concentration in leaves. Pb concentrations in rice grains exceeded the FAO/WHO standard (0.2 mg/kg). The average TF values for heavy metals from the soil to roots, roots to stems, stems to leaves, and stems to grains were Mn > Pb > Ni > Cr, Mn > Cr > Ni > Pb, Ni > Pb > Mn > Cr, and Pb > Ni > Cr > Mn, respectively. The average BAF values in O. sativa were Mn > Ni > Pb > Cr. The EDI for Cr, Pb, Mn, and Ni via O. sativa consumption were 6.19, 6.02, 370.57, and 3.80 µg/kg/day, respectively. The HRI for Cr, Pb, Mn, and Ni via O. sativa consumption were 0.30, 1.50, 2.60, and 0.002, respectively.     

Distribution and localization of pro-brain-derived neurotrophic factor-like immunoreactivity in the peripheral and central nervous system of the adult rat

The precursors for neurotrophins are proteolytically cleaved to form biologically active mature molecules which activate their receptors p75NTR and trks. A recent study showed that the precursor for nerve growth factor (NGF) can bind to p75NTR with a high affinity and induces apoptosis of neurons in vitro. Mutation in Val66Met of brain-derived neurotrophic factor (BDNF) results in reduction in hippocampal function in learning and in the dysfunction of intracellular BDNF sorting and secretion. To examine the functions of pro-neurotrophins in vivo, it is essential to know where they are expressed in the nervous system. In the present study, we have raised and characterized rabbit polyclonal antibodies against a peptide coding for the precursor region of the BDNF gene. The antibody specifically recognizes the precursor for BDNF by western blot. With the affinity purified precursor antibody, we have mapped the distribution and localization of the precursor for BDNF. The results showed that, like mature BDNF, pro-BDNF is localized to nerve terminals in the superficial layers of dorsal horn, trigeminal nuclei, nuclei tractus solitarius, amygdaloid complex, hippocampus, hypothalamus and some peripheral tissues. These results suggest that pro-BDNF, like mature BDNF, is anterogradely transported to nerve terminals and may have important functions in synaptic transmission in the spinal cord and brain.     

超富集植物遏蓝菜对重金属吸收、运输和累积的机制

遏蓝菜Thlaspi caerulescens可以在其地上部累积大量重金属如锌、镉等,是公认的超富集植物。由于该植物生物量小,不宜直接用于重金属污染的土壤植物修复,而被广泛作为一种模式植物来进行重金属富集机制研究。遏蓝菜对重金属离子的累积大致经过螯合剂解毒、地上部长距离运输以及在液泡中的储存等生理过程。已经发现的植物体内的金属螯合剂——有机酸、氨基酸、植物络合素(PCs)、金属硫蛋白(MT)和尼克烟酰胺NA等,区室化以及长距离运输相关的转运蛋白——ZIP(ZRT/IRTlike protein)、CDF(Cation diffusion facilitator)、Nramp(Natural resistance and macrophage protein)和HMA(Heavy metal ATPase)等家族,以上各种基因、多肽与蛋白等共同参与了植物对金属累积与耐受过程并发挥各自重要的作用。以下主要介绍了遏蓝菜重金属超富集相关的基因、多肽和蛋白,以及它们在重金属螯合作用和运输过程中的功能。     

Immunocytochemical localization of glycogen phosphorylase in primary sensory ganglia of the peripheral nervous system of the rat

Neuronal localization was investigated of glycogen phosphorylase (GP) in ganglia of the peripheral nervous system of the rat. Immunofluorescence and immunoenzymatic procedures were applied with a monoclonal anti-bovine brain GP antibody on paraformaldehyde-fixed, paraffin-embedded tissues. Immunoreactivity was only present in the somatic neurons of the mesencephalic trigeminal nucleus in the brain stem and in dorsal root ganglia (DRG), but not in the autonomic neurons of the superior cervical ganglia or in the sensory nuclei of the spinal cord. GP immunoreactivity was present as early as day 1 after birth. In the adult rat, staining was present in neurons of different sizes, and to varying intensities. No relationship was apparent between the staining intensities and morphologically distinguishable types of neurons. In DRG, the type of reactivity was the same from cervical to sacral ganglia. The selected occurrence of GP in specific neurons of the peripheral nervous system in contrast to the ubiquitous occurrence in all astrocytes of the central nervous system may indicate a different role of neuronal glycogen compared to astrocytic glycogen.