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     

Immunohistochemical localization of gastrin-cholecystokinin-like material in the central nervous system of the migratory locust

Summary Brain, corpora cardiaca (CC)-corpora allata (CA) complex, suboesophageal ganglion, thoracic and abdominal ganglia of adults, larvae and embryos of Locusta migratoria have been immunohistochemically screened for gastrin cholecystokinin (CCK-8(s))-like material. In adult, numerous immunoreactive neurons and nerve fibres are located, with a marked symmetry, in various parts of the brain and throughout the ventral nerve cord. In the median part of the brain, cell bodies belonging neither to cellular type A1 nor A2 (following Victoria blue-paraldehyde fuchsin staining) are immunopositive; their processes terminate in the upper protocerebral neuropile. In lateral parts of the brain, external cell bodies send axons into CC and some up to CA, other internal have processes which terminate in the neuropile of the brain. Two of these latter cells react also with methionine-enkephalin antiserum. In the ventral nerve cord, in addition to numerous perikarya, immunore-active arborizations terminate in the neuropile or in close association with the sheath, at the dorsal part of all ganglia.This CCK-8(s) distribution pattern is observed only at the two last larval instars, but is precociously detected in the abdominal nerve cord of embryos, one day before hatching.     

Immunohistochemical localization of gastrin-cholecystokinin-like material in the central nervous system of the migratory locust

Brain, corpora cardiaca (CC)-corpora allata (CA) complex, suboesophageal ganglion, thoracic and abdominal ganglia of adults, larvae and embryos of Locusta migratoria have been immunohistochemically screened for gastrin cholecystokinin (CCK-8(s]-like material. In adult, numerous immunoreactive neurons and nerve fibres are located, with a marked symmetry, in various parts of the brain and throughout the ventral nerve cord. In the median part of the brain, cell bodies belonging neither to cellular type A1 nor A2 (following Victoria blue-paraldehyde fuchsin staining) are immunopositive; their processes terminate in the upper protocerebral neuropile. In lateral parts of the brain, external cell bodies send axons into CC and some up to CA, other internal have processes which terminate in the neuropile of the brain. Two of these latter cells react also with methionine-enkephalin antiserum. In the ventral nerve cord, in addition to numerous perikarya, immunoreactive arborizations terminate in the neuropile or in close association with the sheath, at the dorsal part of all ganglia. This CCK-8(s) distribution pattern is observed only at the two last larval instars, but is precociously detected in the abdominal nerve cord of embryos, one day before hatching.     

Neurogenesis in the adult central nervous system

Contrary to the long-held dogma, neurogenesis occurs throughout adulthood, and neural stem cells reside in the adult central nervous system (CNS) in mammals. The developmental process of the brain may thus never end, and the brain may be amenable to repair. Neurogenesis is modulated in a wide variety of physiological and pathological conditions, and is involved in processes such as learning and memory and depression. However, the relative contribution of newly generated neuronal cells to these processes, as well as to CNS plasticity, remains to be determined. Thus, not only neurogenesis contributes to reshaping the adult brain, it will ultimately lead us to redefine our knowledge and understanding of the nervous system.     

Immunohistochemical localization of low-affinity nerve growth factor receptor in the enteric nervous system of adult rats

The localization of low-affinity nerve growth factor receptor in the enteric nervous system of adult rats has been studied by immunohistochemistry using a monoclonal antibody (clone 192) against the rat receptor. Cryostat and whole-mount sections were stained. By light and confocal microscopy, positive staining in neural structures was found in every part of the gut. In the ganglionic plexus, dense staining was detected in the neuropil surrounding neuronal cell bodies that were themselves devoid of immunoreactivity. Immunoelectron microscopy revealed deposition of reaction products on the outer plasma membranes of both perikarya and processes of neuronal as well as glial cells. Such a selective localization of the receptor in the plasma membrane, but not the cytoplasm, suggests that the mechanisms of receptor-ligand interaction in the gut may differ from those in the brain, where internalization of the receptor is observed in cholinergic cells. The present study provides the morphological basis for future studies designed to elucidate the functional significance of this enteric nervous system receptor. Since it is found in both neuronal and glial cells, it is probably under the influence of a number of trophic factors, including nerve growth factor This revised version was published online in November 2006 with corrections to the Cover Date.     

Immunohistochemical mapping of galanin-like neurons in the rat central nervous system

Using an antiserum generated in rabbits against synthetic galanin (GA) and the indirect immunofluorescence method, the distribution of GA-like immunoreactive cell bodies and nerve fibers was studied in the rat central nervous system (CNS) and a detailed stereotaxic atlas of GA-like neurons was prepared. GA-like immunoreactivity was widely distributed in the rat CNS. Appreciable numbers of GA-positive cell bodies were observed in the rostral cingulate and medial prefrontal cortex, the nucleus interstitialis striae terminalis, the caudate, medial preoptic, preoptic periventricular, and preoptic suprachiasmatic nuclei, the medial forebrain bundle, the supraoptic, the hypothalamic periventricular, the paraventricular, the arcuate, dorsomedial, perifornical, thalamic periventricular, anterior dorsal and lateral thalamic nuclei, medial and central amygdaloid nuclei, dorsal and ventral premamillary nuclei, at the base of the hypothalamus, in the central gray matter, the hippocampus, the dorsal and caudoventral raphe nuclei, the interpeduncular nucleus, the locus coeruleus, ventral parabrachial, solitarii and commissuralis nuclei, in the A1, C1 and A4 catechaolamine areas, the posterior area postrema and the trigeminal and dorsal root ganglia. Fibers were generally seen where cell bodies were observed. Very dense fiber bundles were noted in the septohypothalamic tract, the preoptic area, in the hypothalamus, the habenula and the thalamic periventricular nucleus, in the ventral hippocampus, parts of the reticular formation, in the locus coeruleus, the dorsal parabrachial area, the nucleus and tract of the spinal trigeminal area and the substantia gelatinosa, the superficial layers of the spinal cord and the posterior lobe of the pituitary. The localization of the GA-like immunoreactivity in the locus coeruleus suggests a partial coexistence with catecholaminergic neurons as well as a possible involvement of the GA-like peptide in a neuroregulatory role.     

Stem cells in the adult mammalian central nervous system

Over the past year, evidence has accrued that adult CNS stem cells are a widespread progenitor cell type. These cells may normally replace neurons and/or glia in the adult brain and spinal cord. Advances have been made in understanding the signals that regulate stem cell proliferation and differentiation. A deeper understanding of the structure of germinal zones has helped us move towards identifying stem cells in vivo. Recent studies suggest that the fate of stem cell progeny in vivo may be linked to the complexity of the animal's environment.     

Infiltration of central nervous system in adult acute myeloid leukaemia.

Out of 64 consecutive unselected patients with acute myeloid leukaemia studied during 1973-6, five developed clinical evidence of spread to the central nervous system (CNS). Neuroradiological examination showed cerebral deposits in three, in whom rapid symptomatic relief was obtained with radiotherapy. In two of these patients who developed solid intracranial deposits haematological remission could be reinduced or maintained; they were still alive 86 and 134 weeks later. When patients presented with spread to the CNS complicating generalised uncontrolled leukaemia they had short survivals. CNS infiltration may respond dramatically to appropriate treatment provided that it is not associated with generalised uncontrolled leukaemia, which has a poor prognosis. In view of this, routine "prophylaxis" of the CNS in adult acute myeloid leukaemia does not seem justified at present.     

Reorganization of the human central nervous system

The key strategies on which the discovery of the functional organization of the central nervous system (CNS) under physiologic and pathophysiologic conditions have been based included (1) our measurements of phase and frequency coordination between the firings of alpha- and gamma-motoneurons and secondary muscle spindle afferents in the human spinal cord, (2) knowledge on CNS reorganization derived upon the improvement of the functions of the lesioned CNS in our patients in the short-term memory and the long-term memory (reorganization), and (3) the dynamic pattern approach for re-learning rhythmic coordinated behavior. The theory of self-organization and pattern formation in nonequilibrium systems is explicitly related to our measurements of the natural firing patterns of sets of identified single neurons in the human spinal premotor network and re-learned coordinated movements following spinal cord and brain lesions. Therapy induced cell proliferation, and maybe, neurogenesis seem to contribute to the host of structural changes during the process of re-learning of the lesioned CNS. So far, coordinated functions like movements could substantially be improved in every of the more than 100 patients with a CNS lesion by applying coordination dynamic therapy. As suggested by the data of our patients on re-learning, the human CNS seems to have a second integrative strategy for learning, re-learning, storing and recalling, which makes an essential contribution of the functional plasticity following a CNS lesion. A method has been developed by us for the simultaneous recording with wire electrodes of extracellular action potentials from single human afferent and efferent nerve fibres of undamaged sacral nerve roots. A classification scheme of the nerve fibres in the human peripheral nervous system (PNS) could be set up in which the individual classes of nerve fibres are characterized by group conduction velocities and group nerve fibre diameters. Natural impulse patterns of several identified single afferent and efferent nerve fibres (motoneuron axons) were extracted from multi-unit impulse patterns, and human CNS functions could be analyzed under physiologic and pathophysiologic conditions. With our discovery of premotor spinal oscillators it became possible to judge upon CNS neuronal network organization based on the firing patterns of these spinal oscillators and their driving afferents. Since motoneurons fire occasionally for low activation and oscillatory for high activation, the coherent organization of subnetworks to generate macroscopic function is very complex and for the time being, may be best described by the theory of coordination dynamics. Since oscillatory firing has also been observed by us in single motor unit firing patterns measured electromyographically, it seems possible to follow up therapeutic intervention in patients with spinal cord and brain lesions not only based on the activity levels and phases of motor programs during locomotion but also based on the physiologic and pathophysiologic firing patterns and recruitment of spinal oscillators. The improvement of the coordination dynamics of the CNS can be partly measured directly by rhythmicity upon the patient performing rhythmic movements coordinated up to milliseconds. Since rhythmic dynamic, coordinated, stereotyped movements are mainly located in the spinal cord and only little supraspinal drive is necessary to initiate, maintain, and terminate them, rhythmic, dynamic, coordinated movements were used in therapy to enforce reorganization of the lesioned CNS by improving the self-organization and relative coordination of spinal oscillators (and their interactions with occasionally firing motoneurons) which became pathologic in their firing following CNS lesion. Paraparetic, tetraparetic spinal cord and brain-lesioned patients re-learned running and other movements by an oscillator formation and coordination dynamic therapy. Our development in neurorehabilitation is in accordance with those of theoretical and computational neurosciences which deal with the self-organization of neuronal networks. In particular, jumping on a springboard 'in-phase' and in 'anti-phase' to re-learn phase relations of oscillator coupling can be understood in the framework of the Haken-Kelso-Bunz coordination dynamic model. By introducing broken symmetry, intention, learning and spasticity in the landscape of the potential function of the integrated CNS activity, the change in self-organization becomes understandable. Movement patterns re-learned by oscillator formation and coordination dynamic therapy evolve from reorganization and regeneration of the lesioned CNS by cooperative and competitive interplay between intrinsic coordination dynamics, extrinsic therapy related inputs with physiologic re-afferent input, including intention, motivation, supervised learning, interpersonal coordination, and genetic constraints including neurogenesis. (ABSTRACT TRUNCATED)     

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).     

Biochemical maturation of human central nervous system myelin.

—A developmental study of the lipid and protein composition of human CNS myelin was undertaken. The relative concentrations of the major lipid classes, cholesterol, glycolipids and phospholipids exhibited little change except for a modest decrease in the concentration of the phospholipids. In contrast to the total phospholipids, marked variations in the relative concentrations of individual phospholipids were found. Sphingomyelin increased over two-fold, and phosphatidyl choline decreased to almost half its original concentration. While the concentration of total myelin protein remained constant during maturation, variations in the concentrations of individual proteins were observed. Basic protein constituted 8·5 per cent of the total myelin proteins in the newborn brain and increased to about 30 per cent of the protein in the older ages. The concentrations of proteolipid protein and DM-20 seemed to increase with age, while the relative amounts of high molecular weight proteins decreased. The presence of myelin basic protein in newborn human brain was confirmed by electrophoretic studies involving several different polyacrylamide gel systems and by immunodiffusion experiments which showed a reaction of identity between a constituent present in the fraction containing the presumptive myelin basic protein and authentic myelin basic protein isolated from adult human brain.     

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.     

Extracellular regulators of axonal growth in the adult central nervous system

Robust axonal growth is required during development to establish neuronal connectivity. However, stable fibre patterns are necessary to maintain adult mammalian central nervous system (CNS) function. After adult CNS injury, factors that maintain axonal stability limit the recovery of function. Extracellular molecules play an important role in preserving the stability of the adult CNS axons and in restricting recovery from pathological damage. Adult axonal growth inhibitors include a group of proteins on the oligodendrocyte, Nogo-A, myelin-associated glycoprotein, oligodendrocyte-myelin glycoprotein and ephrin-B3, which interact with axonal receptors, such as NgR1 and EphA4. Extracellular proteoglycans containing chondroitin sulphates also inhibit axonal sprouting in the adult CNS, particularly at the sites of astroglial scar formation. Therapeutic perturbations of these extracellular axonal growth inhibitors and their receptors or signalling mechanisms provide a degree of axonal sprouting and regeneration in the adult CNS. After CNS injury, such interventions support a partial return of neurological function.     

Immunohistochemical localization of the luteinizing hormone releasing hormone (LHRH)-containing structures in the central nervous system of the domestic fowl

Summary The location of LHRH-containing neuronal elements was investigated in the domestic fowl by means of immunohistochemical techniques. LHRH antisera were raised against synthetic LHRH in the rabbit. The antiserum used in the present study cross-reacted with LHRH of mammalian and avian tissues.LHRH-immunoreactive perikarya are located in the preoptic and in the septal areas, and in the bulbus olfactorius; however, no LHRH-immuno-reactive perikarya were found in the tuberal part of the hypothalamus. LHRH-immunoreactive fibers course from these areas toward the median eminence mainly along the wall of the third ventricle in the form of a periventricular network. Originating from the same cell groups other fibers run caudally immediately above the optic chiasma, forming the median bundle of the tractus preoptico-infundibularis. The third bundle running toward the OVLT is named the tractus preoptico-terminalis. In addition to these structures, LHRH-containing fibers and terminals were also present in different regions of the limbic system, in the dorsal part of the hippocampus, in the tuberculum and bulbus olfactorius, as well as in the optic lobe, nuclei commissurales tectales, organon subcommissurale, periaqueductal area, and pars ventralis mesencephali.The general distribution of the LHRH system in the chicken corresponds principally to that described previously in rodents (Sétáló et al. 1976, 1978). However, some subtle differences were demonstrated between the location of the LHRH system in birds and mammals.     

Comparative immunocytochemical localization of putative opioid ligands in the central nervous system

Summary We report a detailed comparative immunocytochemical mapping of enkephalin, CCK and ACTH/gb-endorphin immunoreactive nerves in the central nervous system of rat and guinea pig. Enkephalin immunoreactivity was detected in many groups of nerve cell bodies, fibers and terminals in the limbic system, basal ganglia, hypothalamus, thalamus, brain stem and spinal cord. -endorphin and ACTH immunoreactivity was limited to a single group of nerve cell bodies in and around the arcuate nucleus and in fibers and terminals in the midline areas of the hypothalamus, thalamus and mesencephalic periaqueductal gray with lateral extensions to the amygdaloid area. Cholecystokinin immunoreactive nerve fibers and terminals displayed a distribution similar to that of enkephalin in many regions; but striking differences were also found. An immunocytochemical doublestaining technique, which allowed simultaneous detection of two different peptides in the same tissue section, showed that enkephalin-, CCK- and ACTH/-endorphin-immunoreactive nerves although closely intermingled in many brain areas, occurred separately. The distributions of nerve terminals containing these neuropeptides showed striking overlaps and also paralleled the distribution of opiate receptors. This may suggest that enkephalin, CCK, ACTH and -endorphin may interact with each other and with opiate receptors.Index of Abbreviations CA Commissura anterior - CAI Capsula interna - CO Chiasma opticum - CPF Cortex piriformis - CSDD Commissura supraoptica dorsalis, pars dorsalis (Ganser) - CSDV Commissura supraoptica dorsalis, pars ventralis (Meynert) - FMP Fasciculus medialis prosencephali - FOR Formatio reticularis - GD Gyrus dentatus - GP Glubus pallidus - H Habenula - HI Hippocampus - S Subiculum - SGCD Substantia grisea centralis, pars dorsalis - SGCL Substantia grisea centralis, pars lateralis - SGPV Substantia grisea periventricularis - SNC Substantia nigra, zona compacta - SNL Substantia nigra, pars lateralis - ST Stria terminalis - STP Stria terminalis, pars precommissuralis - TD Tractus diagonalis (Broca) - TO Tractus opticus - TSHT Tractus septohypothalamicus - TUOP Tuberculum olfactorium, pars corticalis - SUM Decussatio supramamillaris - a Nucleus accumbens - ac Nucleus amygdaloideus centralis - aco Nucleus amygdaloideus corticalis - am Nucleus amygdaloideus medialis - ar Nucleus arcuatus - cp Nucleus caudatus putamen - dcgl Nucleus dorsalis corporis geniculati lateralis - em Eminentia mediana - fm Nucleus paraventricularis, pars magnocellularis - fp Nucleus paraventricularis, pars parvocellularis - ha Nucleus anterior (hypothalami) - hd Nucleus dorsomedialis (hypothalami) - hl Nucleus lateralis (hypothalami) - hp Nucleus posterior (hypothalami) - hpv Nucleus periventricularis (hypothalami) - hv Nucleus ventromedialis (hypothalami) - ip Nucleus interpeduncularis - mcgm Nucleus marginalis corporis geniculatic medialis - mm Nucleus mammillaris medialis - ml Nucleus mammillaris lateralis - mh Nucleus medialis habenulae - p Nucleus pretectalis - pf Nucleus parafascicularis - pom Nucleus preopticus medialis - pop Nucleus preopticus periventricularis - posc Nucleus preopticus, pars suprachiasmatica - pt Nucleus paratenialis - pvs Nucleus periventricularis stellatocellularis - re Nucleus reuniens - sc Nucleus suprachiasmaticus - sl Nucleus septi lateralis - so Nucleus supraopticus - st Nucleus interstitialis striae terminalis - tad Nucleus anterior dorsalis thalami - tam Nucleus anterior medialis thalami - tav Nucleus anterior ventralis thalami - td Nucleus tractus diagonalis (Broca) - th Nuclei thalami - tl Nucleus lateralis thalami - tlp Nucleus lateralis thalami, pars posterior - tm Nucleus medialis thalami - tml Nucleus medialis thalami, pars lateralis - tmm Nucleus medialis thalami, pars medialis - tpo Nucleus posterior thalami - tr Nucleus reticularis thalami - tv Nucleus ventralis thalami - tvd Nucleus ventralis thalami, pars dorsomedialis - tvm Nucleus ventralis medialis thalami, pars magnocellularis     

Cellular and subcellular localization of 3H-diethylstilboestrol in the central nervous system

Summary The cellular and subcellular localization of radioactivity in the brain of immature female rats was determined by dry-mount autoradiography 2 h after iv injection of 1.0 g of (monethyl-³H) diethylstilboestrol per 100 g body weight. A specific topographic pattern of nuclear concentration of the synthetic oestrogen was obtained similar to that for ³H-oestradiol-17 in specific neurons of the basal hypothalamus, preoptic region and amygdala. In competition experiments, the nuclear concentration of radioactivity in all areas studied was inhibited by unlabeled oestradiol, while unlabeled testosterone had no effect. These data suggest that although oestradiol can bind to androgen receptors, the oestrogen receptor itself can account for the localization seen after the injection of ³H-oestradiol.This research was supported in part by US PHS Grant No. NS12933NIH Career Development Awardee No. NS00164The expert technical assistance of Ms. Riki Ison and Ms. Linda Furr is gratefully acknowledged.