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Biomedical subjects

D Dahl

Publications and source records attributed to D Dahl.

At least 73 records · Page 4Linked to original sources
PubMed1989

Vimentin-GFAP transition in primary dissociated cultures of rat embryo spinal cord.

Primary dissociated cultures derived from 15-day-old rat embryo spinal cord with or without dorsal root ganglia (DRG) were grown on polylysine, Primaria and laminin substrates. On polylysine and Primaria substrates, spinal cord neurons formed aggregates connected by bundles of neurites in a distinctive pattern similar to that observed in cultures derived from embryonal rat brain and neonatal rat cerebellum. After 2 days in culture, the number of cells stained with GFAP antibodies progressively increased within the vimentin-positive monolayer surrounding the neuronal aggregates. These astrocytes had the typical appearance of astrocytes in primary dissociated cultures derived from late fetal or early neonatal murine brain, i.e. large flat or stellate cells with thick processes staining equally well with GFAP and vimentin antibodies. Astrocytes found within the neuronal aggregates in 4-5 day cultures were markedly different, i.e. small stellate cells with slender processes forming a delicate mesh throughout the aggregate. These GFAP-positive cells stained only weakly with vimentin antibodies. Spinal cord neurons formed aggregates on laminin substrates but failed to extend neurites and rapidly degenerated. The large flat cells in the surrounding monolayer gradually invaded the aggregates. These cells stained with both GFAP and vimentin antibodies. DRG neurons developed equally well on Primaria and laminin substrates, extending their neurites on the vimentin-positive flat cells forming the monolayer regardless of their reactivity with GFAP antibodies.

Animals↗
PubMed1989

Effects of glycosaminoglycans and proteinase inhibitors on astroglia-induced detachment of cultured rat cerebellar neurons.

Neurons in mixed primary embryonic CNS cultures degenerate secondary to their detachment from the substratum. The present study demonstrates that in primary cultures of postnatal cerebellum, detachment of neurons can be prevented by antiproliferative drugs which inhibit the growth of astroglia. Several types of proteinase inhibitors did not affect the process of detachment. However, among several types of glycosaminoglycans, heparan sulfate and to a lesser degree heparin, could reversibly inhibit neuron detachment without causing morphological changes of astroglia. The enzymes heparitinase and heparinase caused neuron detachment but only within the first 24-48 hr after plating and not in older cultures. We conclude: (1) cerebellar interneurons in culture are not dependent on astroglia for their survival; (2) astroglia are most probably responsible for neuron detachment via a membrane associated activity and (3) heparan sulfate-like glycosaminoglycans are important in neuron-substratum attachment.

Animals↗
PubMed1989

Morphological and electrophysiological studies of human hippocampal transplants in the anterior eye chamber of athymic nude rats.

Human fetal hippocampal tissue from normal women was obtained following elective abortion in the 8th to the 11th week of gestation. The hippocampal tissue was transplanted to the anterior chamber of the eye of adult athymic nude rats, where it was allowed to develop for up to 9 months before histological and electrophysiological evaluation. The transplants were revascularized from the host iris and many grew extensively in oculo. Large neurons were present in all transplants. Immunohistochemical studies revealed glutamic acid decarboxylase-containing terminals and clusters of gamma-aminobutyric acid-positive nerve cell bodies within the transplants, as well as scattered tyrosine hydroxylase-positive and acetylcholinesterase-containing fibers. Single neurons recorded extracellularly from transplants 4-9 months in oculo showed a slow spontaneous discharge, with both complex and single action potentials. Stimulation of the transplant surface evoked a small initial wave followed by a larger and longer-lasting field potential, similar to that seen in hippocampus in situ. A conditioning-testing paradigm was used to evaluate the presence of inhibitory circuitry in the hippocampal transplants. Significant suppression of the evoked test response was seen with interstimulus intervals ranging from 20 to 500 ms. Superfusion of enkephalin (100-300 nM) or penicillin (1600 U/ml) increased slow-wave activity, as did tetanic electrical stimulation. These treatments appeared to generate ictal-like activity, which in some cases persisted as interictal spikes. Illumination of the retina also increased neuronal activity, presumably by reflex activation of cholinergic afferents from the parasympathetic innervation of the iris. Taken together, our data suggest that fragments of hippocampus from aborted first trimester human fetuses, grafted to the eye chamber of rodent hosts, develop many organotypic histological and physiological features. This preparation may provide a unique means for the study of neurobiological properties of human brain in both normal and disease states.

Animals↗
PubMed1989

Norepinephrine induces pathway-specific long-lasting potentiation and depression in the hippocampal dentate gyrus.

The study presented here indicates that norepinephrine (NE) selectively induces long-lasting modifications of synaptically mediated responses in the dentate gyrus of the rat hippocampal slice. A low concentration of NE (1.0 microM; in the presence of 50 microM phentolamine, an alpha-adrenergic antagonist) or a 1.0 microM concentration of the specific beta-adrenergic agonist isoproterenol induced long-lasting pathway-specific alterations of granule cell electrophysiological responses. Excitatory postsynaptic potentials and population spikes evoked by stimulation of the medial perforant pathway (PP) were potentiated for more than 45 min. In contrast, responses to lateral PP stimulation were depressed for the same period. Both potentiation and depression were blocked by the beta-adrenergic antagonist propranolol (1.0 microM). These results indicate that NE can act differentially on projections to the dentate gyrus arising in the entorhinal cortex. Such selective persistent modifications of cortical circuits may be involved in processes in the mammalian brain underlying attention, learning, and memory.

Animals↗
PubMed1989

Glial hyaluronate-binding protein in polar spongioblastoma.

Glial hyaluronate-binding (GHA) protein is a 60 kDa glycoprotein isolated from human white matter by affinity chromatography on immobilized hyaluronate. It is localized in white matter astrocytes by immunofluorescence with monoclonal antibodies. Amino acid sequences have not revealed similarities with other proteins except cartilage extracellular matrix proteins, the region of similarity being located within the hyaluronate-binding region. Cryostat sections of 13 intracranial neoplasms removed at surgery were tested for the presence of GHA protein by indirect immunofluorescence with monoclonal antibodies. These included seven astrocytomas, one oligodendroglioma, one medulloblastoma and one spinal cord ependymoma. All tumors were negative with the exception of one astrocytoma in which the GHA protein-positive areas had the typical appearance of polar spongioblastoma, i.e. small cells palisading around blood vessels and very delicate glial fibrillary acidic (GFA) protein-positive fibrils. Conversely, neoplastic as well as reactive GFA protein-positive astrocytes were GHA protein-negative. We suggest that polar spongioblastoma derives from a GHA protein-positive glial precursor and pertinent to this suggestion is the observation that the periventricular germinal layer was found GHA protein-positive in a 22-week human fetus.

Antibodies, Monoclonal↗
PubMed1988

Astroglia-induced detachment of central neurons but astroglia-dependent growth of peripheral neurons in rat embryonic spinal cord primary cultures.

In mixed primary cultures, intrinsic neurons from embryonic mammalian brains degenerate secondary to their detachment from the substratum and this is caused by the under-growing co-cultured astroglia. In the present study we sought to find out whether or not peripheral neurons, sensory and motor neurons which reside and/or only project outside the CNS respectively, interact with astroglia similarly as their central counterparts do. Mixed primary cultures prepared from dissociated embryonic rat spinal cord and dorsal root ganglia were examined by phase and immunofluorescence microscopy after labeling with antibodies to neurofilaments (neuronal markers) and to glial fibrillary acidic protein and vimentin (astroglia markers). Acetylcholinesterase staining served as a marker for motor neurons. In this system astroglia grew exclusively under intrinsic neurons of the spinal cord and with time (about 8 days) all these neurons detached and disappeared. In contrast, astroglia were intimately associated with perikarya of peripheral neurons, sometimes growing over them. Furthermore, the neuritic processes of these neurons were attached to the undergrowing astroglia. Central neurons could be rescued by treatment of cultures with the antimitotic drug cytosine arabinoside which led to the elimination of astroglia. However, this treatment resulted in death of all peripheral neurons. We conclude: (1) survival of intrinsic CNS neurons in culture is independent of astroglia; (2) astroglia are responsible for the detachment of these neurons from their growth substratum; (3) survival of peripheral sensory and motoneurons is dependent on co-cultured astroglia and (4) the differences in neuron-astroglia interactions between central and peripheral neurons are membrane-associated and probably independent of soluble factors.

Animals↗
PubMed1988

Neurofilament phosphorylation in axons and perikarya: immunofluorescence study of the rat spinal cord and dorsal root ganglia with monoclonal antibodies.

Rat dorsal root ganglia and spinal cord were stained with 12 monoclonal antibodies reacting with phosphorylated epitopes of two neurofilament proteins (NF 150K and NF 200K). Three monoclonal antibodies were axon-specific in both locations; neuronal perikarya were not stained. Nine monoclonal antibodies stained a subpopulation of neurofilament-positive sensory neurons, as indicated by double labeling experiments with polyclonal antibodies reacting with phosphorylated and dephosphorylated forms of the neurofilament protein triplet. Of these nine antibodies, two stained motor neuron perikarya in the spinal cord, while the remaining seven antibodies were axon-specific in this location. Subpopulations of stained and unstained motor neurons were not observed. With all 12 antibodies, the staining pattern in the lumbar dorsal root ganglia and spinal cord remained unchanged following sciatic nerve crush and ligature. The findings suggest that, in the neurofilament, some phosphorylated epitopes are axon specific, while other phosphorylated epitopes are present in both axons and perikarya. Furthermore, they suggest that differences exist between neuronal populations as to the presence of phosphorylated epitopes in perikaryal neurofilaments. It remains to be seen whether phosphorylation events in perikarya and axons have similar or different effects on neurofilament structure and function.

Antibodies, Monoclonal↗
PubMed1988

2,5-Hexanedione-induced accumulations of neurofilament-immunoreactive material throughout the rat autonomic nervous system.

In rats intoxicated with 2,5-hexanedione, nerve fibres supplying virtually all visceral organs showed large numbers of densely immunoreactive accumulations of neurofilament-like material, of fusiform, elongated, smoothly tapering morphology. In the gut, round to oval, morphologically different lesions were also present, and abnormal neurofilament-immunoreactive accumulations were revealed in oesophageal terminal end-plates. An extensive damage to autonomic nerve fibres, which are largely non-myelinated, was thus revealed in 2,5-hexanedione intoxication. The observed diversity in lesion morphology may suggest heterogeneity in cytoskeletal and/or associated proteins among autonomic neurons.

Animals↗
PubMed1988

Early and late appearance of neurofilament phosphorylated epitopes in rat nervous system development: in vivo and in vitro study with monoclonal antibodies.

Neurofilament phosphorylation in rat nervous system development was studied by indirect immunofluorescence with monoclonal antibodies reacting with phosphorylated epitopes in tissue sections and in primary dissociated cultures. The antibodies either decorated neurofilaments shortly after their appearance or after a considerable delay (from 4 to 9 days in vivo and from 12 to 27 days in vitro), thus suggesting the existence of at least two classes of phosphorylated epitopes. With most antibodies there was a good correlation between in vivo and in vitro findings as to the early or late appearance of phosphorylated epitopes. Monoclonal NE14 was the main exception in that immunoreactivity with this antibody was present in 1-day cultures, while it only occurred 4 days after the first appearance of neurofilaments in vivo. The effect of phosphorylation on neurofilament structure and function remains to be determined. Neurofilament expression is an early phenomenon in ontogeny coinciding with neuronal differentiation. It is possible that late phosphorylation events may stabilize the axonal cytoskeleton following the massive loss of axons that occurs in several fiber tracts during late fetal and neonatal life.

Aging↗
PubMed1988

Expression of brain-specific hyaluronectin (BHN), a hyaluronate-binding protein, in dog postnatal development.

Monoclonal antibodies reacting with the brain-specific form of hyaluronectin, a hyaluronate-binding protein, were used in conjunction with antibodies to the glial fibrillary acidic protein (GFAP), the subunit of astrocyte-specific intermediate filaments, to study the postnatal development of spinal cord and cerebral white matter in the dog. As previously reported, the distributions of brain-specific hyaluronectin (BHN) and GFAP in adult dog spinal cord white matter were similar. Both antigens formed a mesh surrounding individual myelinated axons. Furthermore, the glia limitans on the surface of the spinal cord and the glial septa were stained by both antibodies. In newborn dog spinal cord, hyaluronectin immunoreactivity was confined to the glia limitans on the surface. At this time GFAP-positive fibers formed a dense mesh throughout the myelinated white matter. Staining of spinal cord white matter with BHN antibodies first appeared on day 15 and reached its mature appearance in the fully myelinated spinal cord on day 21. In cerebral white matter BHN immunoreactivity was first observed on day 21. With GFAP antibodies astrocytes were extremely few in the nonmyelinated cerebral white matter of 1- and 3-day-old dogs. GFAP-positive astrocytes in cerebral white matter had markedly increased on day 9 before the onset of myelination on day 15. On day 21, myelination was confined to deep cerebral white matter and myelin sheaths were still very few in subcortical white matter. We conclude that BHN expression by white matter astrocytes is an extremely late event in brain development, first occurring after the onset of myelination.

Aging↗
PubMed1987

Immunological study of a neurofilament protein variant (S150) with polyclonal and monoclonal antibodies.

Ten polyclonal neurofilament antibodies were tested for domain specificity with immunoblots of chymotrypsin digests of a neurofilament protein of 150 kDa (NF 150K). In contrast to most monoclonal antibodies previously reported, the five polyclonal antibodies which showed domain specificity reacted with the 40 kDa alpha-helical rod domain of the molecule. (With one exception, monoclonal antibodies reacted with the 100 kDa carboxy-terminal peripheral domain). Of these ten polyclonal antibodies only two reacted with an isoelectric variant of NK 150K (S150) isolated by Liem and collaborators (Wong, J., Hutchison, S.B. and Liem, R.K.H. (1984) J. Biol. Chem. 259, 10867-10874) from bovine brain. 13 monoclonal antibodies were also tested for reactivity with S150 protein. With one exception, none of these antibodies reacted with this variant, not even a monoclonal antibody which we have previously shown to react with a non-phosphorylated epitope located in the rod domain of NF 150K. We suggest that either there are modifications other than dephosphorylation in the S150 isoelectric variant or, alternatively, that it is not derived from NF 150K.

Animals↗
PubMed1987

Buffers and H2O2 reduce neuronal death and/or enhance differentiation of neurons and astrocytes in dissociated mouse brain cultures.

Tissue of the mammalian central nervous system (CNS) undergoes complex and uncoordinated pathological responses upon injury. Efforts to develop pharmacological approaches to achieve functionally meaningful regeneration largely have been unsuccessful. Assuming that anoxia and drop in tissue H are initiating factors in most pathological sequences consequent to CNS injury, we studied the effects of increasing the buffering and oxygenating capacities of the medium on dissociated embryo brain cultures. The presence of H2O2 in the medium led to greatly enhanced neuronal survival and/or differentiation. Increased buffering capacity favored enhanced neurite outgrowth with remarkable elongation of fibers. A combination of the two gave a synergistic effect in which both of the above responses were seen. Both buffers and H2O2 enhanced astrocytic differentiation and extension of processes while reducing DNA synthesis. The results favor the view that attempts to encourage self-repair in CNS tissue or to enhance repair of CNS damage with potential therapeutic agents or procedures should be carried out in the context of a near optimal environment in which, at the least, pH and pO2 values are stably maintained within normal operational limits.

Animals↗
PubMed1987

Maturation of a large neurofilament protein (NF 150K) in rat postnatal development.

The mammalian neurofilament is made of three neuron-specific proteins with approximate molecular weights of 70 kilodaltons (kDa) (NF 70K), 150 kDa (NF 150K), and 200 kDa (NF 200K) by SDS-PAGE. As previously reported in the rat by Strocchi et al (J Neurochem 39:1132-1141, 1982) and Nixon et al (J Cell Biol 94:150-58, 1982), NF 150K comprises three molecular weight variants with the same isoelectric point. A fourth lower molecular weight and slightly less acidic variant was identified by monoclonal and polyclonal antibodies reacting with the alpha-helical middle domain of NF 150K. With few exceptions, this lower molecular weight variant did not stain with monoclonal antibodies reacting with the peripheral carboxy-terminal domain. Staining with these antibodies was abolished or markedly reduced following neurofilament dephosphorylation. The distribution of the NF 150K variants varied in different regions of the nervous system. The higher molecular weight variant (component a) was less prominent in brain compared to spinal cord, optic nerve, and sciatic nerve. Furthermore, the lower molecular weight variant (component d) was not identified in optic nerve and sciatic nerve. All four variants were identified in brain and spinal cord extracts of newborn rats with monoclonal and polyclonal antibodies reacting with the alpha-helical middle domain of NF 150K. As a general (see Results for exceptions) monoclonal antibodies reacting with the carboxy-terminal region of NF 150K did not stain the variants in newborn rat brain extracts until day 10 when immunoreactivity of component a first appeared. The adult pattern was first observed on postnatal day 15.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1987

Neurofilament-like and glial fibrillary acidic protein-like immunoreactivities in rat and guinea-pig sympathetic ganglia in situ and after perturbation.

The presence of neurofilament (NF)-like and glial fibrillary acidic protein (GFAP)-like immunoreactivities was studied in sympathetic ganglia of adult rats and guinea pigs during normal conditions and after perturbation. In the superior cervical ganglion (SCG) of normal rats, many ganglion cells and nerve fibers show NF immunoreactivity. Some of these nerve fibers disappear after preganglionic decentralization of SCG; this indicates the presence of a mixture of pre- and postganglionic NF-positive nerves in the ganglion. Cuts in both pre- and postganglionic nerves result in a marked increase in GFAP immunoreactivity in SCG, whereas NF immunoreactivity increases in nerve cell bodies after preganglionic cuts. Only a few ganglion cells show NF immunoreactivity in the normal SCG of guinea pig. All intraganglionic NF-positive nerves are of preganglionic origin; decentralization abolishes NF immunoreactivity in these nerve fibers. The inferior mesenteric ganglion, the hypogastric nerves and colonic nerves in guinea pigs contain large numbers of strongly NF-immunoreactive nerve fibers. When the SCG of adult rat is grafted to the anterior eye chamber of adult rat recipients, both ganglionic cell bodies and nerve fibers, forming on the host iris from the grafted ganglion, are NF-positive. As only the perikarya of these neurons normally exhibit NF immunoreactivity, and the terminal iris arborizations are NF-negative, it appears that the grafting procedure causes NF immunoreactivity to become more widespread in growing SCG neurons.

Animals↗
PubMed1987

Glial fibrillary acidic protein (GFAP)-like immunoreactivity in normal and transected rat olfactory nerve.

Normal and transected rat olfactory nerves were stained immunohistochemically using a monoclonal antibody previously shown to selectively detect GFAP-like immunoreactivity in central astrocytes but not in peripheral Schwann cells. Low levels of "central" type GFAP were found in the olfactory nerves, presumably in ensheathing cells. The levels of GFAP increased dramatically after nerve transection. A population of strongly GFAP-positive cells was detected at the junction between the olfactory epithelium and initial part of the nerves, of possible relevance to the regenerative abilities of this pathway.

Animals↗
PubMed1987

Sustained seizures cause circumscribed cerebral changes in glial fibrillary acidic protein, neurofilament and laminin immunofluorescence.

Sustained experimental seizures in rats have previously been shown to cause an extensive necrosis in pars reticulata of substantia nigra (SNPR) and globus pallidus (GP). In the present paper we have studied the effects of hexafluorodiethyl ether-induced seizures on the immunoreactivity seen with antibodies directed against glial fibrillary acidic protein, GFA, used to visualize astrocytes, antibodies to the glycoprotein laminin as a marker for blood vessel walls and neurofilament (NF) antibodies to monitor neuronal disturbances. Already 12 h after a 20-min seizure period a reduction in GFA immunofluorescence intensity was observed in SNPR. After 3 days, marked lesions were noted in SNPR and GP as seen with cresyl violet staining. The lesions contained almost no GFA-positive structures. In the proximity of the lesions, an increase in GFA-immunoreactivity was noted. Such an increase, although less pronounced, was also seen in the major projection areas of SNPR. Two months post-seizure, the gliotic reaction had disappeared, and only a thin and elongated gliotic scar was observed. In spite of the development of a profound central necrosis especially evident in SNPR, both laminin- and NF-immunoreactivity was slightly increased within the lesioned areas. NF-immunoreactivity was also increased in the superior colliculus and in the reticular formation. Two months post-experiment NF-immunofluorescence was normalized but the former lesion sites showed signs of hypervascularization. We conclude that hexafluorodiethyl ether-induced 20-min seizures lead to rapid, localized glial and neuronal changes in the rat brain as evidenced by GFA and NF immunohistochemistry, while the vascular network remains intact.

Animals↗