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C J Barnstable

Publications and source records attributed to C J Barnstable.

At least 73 records · Page 4Linked to original sources
PubMed1990

Monoclonal antibody HNK-1 selectively stains a subpopulation of GABAergic neurons containing the calcium-binding protein parvalbumin in the rat cerebral cortex.

Monoclonal antibody HNK-1 is shown to outline selectively a subpopulation of GABAergic neurons containing a specific calcium-binding protein parvalbumin (PV) in the adult rat parietal cortex, using pre- and postembedding immunocytochemistry at light microscopic level. About 98% of HNK-1 stained cells in the rat parietal cortex were PV immunoreactive. About 95% of HNK-1 immunoreactive cells were also shown to be stained with a lectin, Vicia villosa agglutinin (VVA), with a specific affinity for terminal N-acetylgalactosamine, which has been previously shown to stain selectively a subpopulation of PV-containing GABAergic neurons in this region. Furthermore almost all HNK-1 immunoreactive cells were also stained with a monoclonal antibody, 3B3, which is specific for chondroitin sulfate proteoglycan. 3B3 was shown in the present study to stain selectively a subpopulation of PV-immunoreactive neurons in the adult rat parietal cortex. In addition, a direct comparison of two monoclonal antibodies HNK-1 and VC1.1 revealed that these two were identical in their staining properties and that they defined the same subset of PV-containing GABAergic neurons in the rat parietal cortex.

Animals↗
PubMed1990

Expression of the cell surface antigens RET-PE2 and N-CAM by rat retinal pigment epithelial cells during development and in tissue culture.

Invagination of the optic vesicle to form the optic cup results in the formation of two apposed layers of neuroepithelium which follow divergent developmental pathways. Changes in the expression of cell surface molecules may be either the cause or result of important inductive signals during this process. We have used immunological reagents to study the expression of two molecules in the rat: the neural cell adhesion molecule N-CAM, and a cell membrane-associated protein which is specific for pigment epithelium in the adult, RET-PE2. Both N-CAM and RET-PE2 are present in both layers of the optic cup at embryonic age E13, but they become restricted to inner retina and pigment epithelium, respectively, by E17 and maintain that pattern of expression in the normal adult. Culture of pigment epithelial cells results in the reexpression of N-CAM and the continued expression of RET-PE2. Western blotting reveals that the size and relative proportions of the 180- and 140-kDa N-CAM molecules synthesized by rat pigment epithelial cells in vitro differ from that made by rat brain, retina and liver. Embryonic RPE N-CAM contains the sulfated carbohydrate recognized by the HNK-1 antibody, but this epitope was not present on N-CAM synthesized by cultured RPE cells. The reexpression of an embryonic antigen when placed in culture suggests that pigment epithelial cells retain some degree of plasticity in the adult state.

Animals↗
PubMed1990

Neuronal subsets express multiple high-molecular-weight cell-surface glycoconjugates defined by monoclonal antibodies Cat-301 and VC1.1.

Cat-301 and VC1.1 are monoclonal antibodies that recognize surface-associated molecules on subsets of mammalian CNS neurons. Earlier work demonstrated that Cat-301 recognizes a 680-kDa chondroitin sulfate proteoglycan (PG). VC1.1 has been shown to recognize 3 polypeptide bands on Western blot analysis; a major band at 95-105 kDa and additional bands at 145 kDa and 170 kDa. In the present report, we show that VC1.1 also reacts with a high-molecular-weight glycoconjugate. Immunoprecipitation experiments and biochemical characterizations indicate that Cat-301 and VC1.1 define at least 3 distinct high-molecular-weight antigens. The VC1.1 antigens react with antikeratan sulfate antibodies, while the Cat-301 antigens do not. By immunodepletion, we show that some VC1.1 antigens are Cat-301 positive, while others are Cat-301 negative. In addition, Cat-301-reactive proteoglycans are heterogeneous with respect to the presence or absence of VC1.1 epitopes. Double-label immunofluorescence studies with these 2 antibodies are consistent with the biochemical results and show that there are 3 classes of immunoreactive neurons in the cat CNS:Cat-301+/VC1.1+, Cat-301-/VC1.1+, and Cat-301+/VC1.1-. These results indicate that structural microheterogeneity exists among Cat-301 and VC1.1 high-molecular-weight glycoconjugates. This heterogeneity may be a reflection of the diverse neuronal phenotypes that are recognized by Cat-301 and VC1.1 in the mammalian CNS.

Animals↗
PubMed1989

Expression of the growth cone specific epitope CDA 1 and the synaptic vesicle protein SVP38 in the developing mammalian cerebral cortex.

CDA 1 is a novel antigen that within the brain is present specifically in neuronal growth cones. Electron microscope immunohistochemistry and subcellular fractionation showed the CDA 1 epitope to be on a cytosolic molecule. In cultured neurons, it is abundant in growth cones and not detectable in neurites or cell bodies. The development of the rat cerebral cortex was investigated by using the monoclonal antibody to CDA 1 and an antibody to SVP38, the synaptic vesicle glycoprotein. CDA 1 immunoreactivity in the rat cerebral cortex peaks just before birth and disappears by postnatal day 12, a few days before the major increase in the number of mature synapses. In contrast, SVP38 is expressed in parallel with the appearance of mature synapses. CDA 1 and SVP38 thus are markers of growth cones and synapses, respectively. Their expression during development reflects some of the structural and functional changes that occur during synapse formation.

Aging↗
PubMed1989

Monoclonal antibody VC1.1 selectively stains a population of GABAergic neurons containing the calcium-binding protein parvalbumin in the rat cerebral cortex.

Monoclonal antibody VC1.1 is shown to stain selectively a subpopulation of GABAergic neurons in the rat cerebral cortex. Almost all VC1.1 immunoreactive cells were also GABA-like immunoreactive (GABA-LI) and parvalbumin (PV) immunoreactive, whereas they were about 30% and 65% of GABA-LI and PV-positive cells in the parietal cortex and about 13% and 32% in the occipital cortex, respectively. Although a few VC1.1 positive cells showed somatostatin-like and/or cholecystokinin-like immunoreactivities, they were exceptional (less than 1% of VC1.1 positive cells). Furthermore about 90% of VC1.1 positive cells were also stained with a lectin, Vicia villosa agglutinin, with a specific affinity for terminal N-acetylgalactosamine.

Animals↗
PubMed1989

Immunoelectron microscopical examination of the surface distribution of opsin in rat rod photoreceptor cells.

The distribution of the visual pigment protein opsin in postnatal day 8 and adult rat retinal photoreceptor cells was studied using several well characterized monoclonal antibodies against rhodopsin and a sensitive, modified colloidal gold technique. Distribution of opsin was studied in intact tissue, explant cultures and freshly dissociated cell preparation. At PN8 labeling was seen in the plasma membrane overlying the cell body, inner and outer segment. Adult retinas still showed heavy labeling of the cell body and outer segment, but reduced labeling of the inner segment surface. Semi-quantitative estimates of colloidal gold particles bound indicated a ratio of 3-20 times more particles per unit length in the outer vs. the inner segment, depending on tissue age, preparation and antibody. Together with the subcellular localization of other photoreceptor plasma membrane proteins, these data reveal a complex system of membrane domains into which different protein components are segregated.

Animals↗
PubMed1989

Molecular determinants of GABAergic local-circuit neurons in the visual cortex.

Golgi impregnation, intracellular marking techniques and immunocytochemistry have led to the identification of several distinct GABAergic cell types. Co-localization of neuropeptides or calcium-binding proteins has provided additional markers for GABAergic cells. Recently, immunological or lectin probes have helped to identify additional subsets of GABAergic neurons. In combination with other immunocytochemical and anatomical approaches, these probes are now being used to link molecular composition to cellular architecture in the visual cortex.

Animals↗
PubMed1988

Coexpression of opsin- and VIP-like-immunoreactivity in CSF-contacting neurons of the avian brain.

Cerebrospinal fluid-contacting (CSF) cells in both the septal and the tuberal areas in the brain of the ring dove are labeled by RET-P1, a monoclonal antibody to opsin that reacts with inner and outer segment membranes of rod photoreceptors in a variety of vertebrates. Immunoblot analysis of proteins from diverse brain regions, however, revealed bands of anti-RET-P1 immunoreactivity that did not correspond to opsin. Binding of RET-P1 to opsin-containing membranes, was not inhibited by membranes rich in muscarinic and beta-adrenergic receptor proteins (red blood cells, heart, lung) taken from doves. RET-P1-immunoreactive CSF-contacting cells emit a dendritic process that penetrates the ependyma and ends in a knob-like terminal suspended in the ventricle. These cells also possess other processes that penetrate more or less deeply into the neuropil. Additionally, a band of labeled fibers occurs in the external layer of the median eminence. A double-label technique demonstrated that RET-P1-positive cells coexpress VIP-like immunoreactivity. VIP-positive cells in other brain areas are not RET-P1-positive.

Animals↗
PubMed1988

Opsin expression in the rat retina is developmentally regulated by transcriptional activation.

The gene for rhodopsin, the primary light sensor of the visual system, is specifically expressed in the rod photoreceptor cells of the retina. We show here that in the rat, opsin RNA first accumulates to detectable levels at postnatal day 2 (PN2) and that nascent transcripts can be detected at PN1; this is the time when peak numbers of photoreceptor cells are generated by the final division of their neuroepithelial precursors. Accumulated opsin RNA then increases to reach the adult level, 0.06% of total retinal RNA, at about PN10. The transcription rate of the opsin gene increases to a similar extent over the same time course between PN3 and adulthood, suggesting that transcriptional activation is responsible for the increase in opsin expression. We used the antibody RET-P1 to show that rhodopsin protein is also detectable at PN2 and that the number of cells expressing the protein increases with time in a central-to-peripheral gradient in the retina. This increase in the number of differentiating photoreceptors in the tissue appears to account for much of the increase in opsin gene transcription and RNA accumulation. In situ hybridization to opsin RNA shows that it is restricted to the photoreceptor layer from the time it can first be detected, at PN7. Later in development, when RET-P1 staining shifts to the photoreceptor outer segments, opsin RNA becomes localized to the inner segments, suggesting that the distributions of opsin protein and RNA are related.

Aging↗
PubMed1988

Selective staining of a subset of GABAergic neurons in cat visual cortex by monoclonal antibody VC1.1.

VC1.1 is a monoclonal antibody generated against cat area 17, which selectively outlines subsets of cortical neurons (Arimatsu et al., 1987). This study was conducted to determine the ultrastructural distribution of the VC1.1 antigen and to identify the particular subclasses of cortical neurons that were labeled. In the light microscope, VC1.1 delineated the surfaces of neurons located mainly in layer IV but also in other layers. The staining surrounded neuronal cell bodies and dendrites in a periodic or meshwork pattern but did not label axons. VC1.1-labeled neurons were morphologically heterogeneous and included multipolar, bipolar, and bitufted classes. In the electron microscope, VC1.1 immunoreactivity surrounded presynaptic membranes of terminal boutons and intersynaptic sections of postsynaptic membranes, but was not present within terminal boutons or synaptic clefts. Both asymmetric and symmetric synapses were immunoreactive. Labeling was also observed intracellularly on VC1.1-outlined neurons, associated with perisynaptic portions of plasma membranes. Tract-tracing methods were used in conjunction with immunocytochemistry to determine whether VC1.1 identified projection neurons, local circuit neurons, or a combination of both types. Layer V and VI corticogeniculate and corticotectal projection neurons were retrogradely labeled with rhodamine fluorescent latex microspheres. In a large sample of retrogradely labeled neurons, none were VC1.1-positive, suggesting that VC1.1 stained a population of local circuit neurons. Additional immunocytochemical double-labeling studies with an antiserum to GABA and VC1.1, revealed that VC1.1-positive neurons were immunoreactive to GABA. These were a major subset of the GABAergic neurons in area 17 and tended to have medium to large cell bodies. It is concluded that VC1.1 identifies a new, immunologically distinct subset of GABAergic neurons in area 17. The restricted distribution of this antigen on perisynaptic portions of GABA-containing cells and surrounding terminal boutons onto these cells suggests that this antigen may play an important role in inhibitory cortical circuits.

Animals↗
PubMed1988

A developmentally regulated antigen associated with neural cell and process migration.

The distribution of an epitope recognized by the monoclonal antibody JONES has been studied immunohistochemically in the developing nervous system of the rat. In the present report, we survey selected regions of the fetal, postnatal, and adult rat nervous system to test the hypothesis that JONES binding is invariably associated with neural cell migration and axon growth in the developing rat. A series of selected developmental stages extending from embryonic day (E) 9 to adult were used in this investigation. The distribution of JONES binding was examined using indirect immunofluorescence, as well as the immunogold procedure. Particular attention was paid to regions where the positions and timing of cell and axon migrations have been well described for the rat. JONES immunoreactivity first appears at E11-12, when it is localized to the lamina terminalis, the telencephalic-diencephalic junction, the midbrain, and the rhombic lip regions of the cytologically undifferentiated neural tube. In all the regions studied, during embryonic and early postnatal life, the labeling is very intense in the ventricular zone and shows a radial array in the adjacent intermediate and marginal zones. The expression of JONES epitope correlates particularly with times of cell migration in the retina, superior colliculus, cerebellum, and telencephalon and in regions undergoing neurite extension, such as the developing optic tract, the white matter of the cerebellum, the dorsal roots, the trigeminal system, and olfactory nerve. JONES binding becomes progressively restricted in the postnatal period. In the adult brain, immunoreactivity is present only in the retina and cerebellum. In the retina, JONES labeling is present in the outer plexiform layer and optic fiber layer. The labeling in the optic fiber layer extends to the optic nerve head and stops abruptly outside the orbit. In the cerebellum, JONES shows a radially oriented pattern throughout the molecular layer and delineates the cell bodies in the Purkinje cell layer. The only non-neural regions that show JONES immunoreactivity are the adrenal medulla and the kidney glomeruli. We conclude that the antigens recognized by the JONES monoclonal antibody are associated with the migration of subsets of cells and axons within the developing rat nervous system and, consequently, may play a role in conveying selectivity to these processes.

Animals↗
PubMed1988

Developmental regulation of ganglioside antigens recognized by the JONES antibody.

The JONES monoclonal antibody has been immunocytochemically associated with regions of the developing rat brain where cell and axon migrations are occurring (Mendez-Otero et al., 1986, 1988). In the present study the antigens recognized by the JONES antibody were analyzed in a variety of brain regions and at developmental ages selected to correspond to the preceding immunocytochemical observations. In accordance with earlier results from retina, JONES binding could not be detected in SDS gels from developing brain. Binding of the antibody was, however, prominent in chloroform/methanol extracts of the same tissues, and it was completely removed from tissue sections by brief chloroform/methanol form/methanol treatment. Enzymatic analyses of chloroform/methanol extracts indicated that the JONES epitope was sensitive to neuraminidase but insensitive to proteases. Overlay assays on developed high-performance thin-layer chromatographic plates (HPTLC) indicate that in all regions the JONES epitope resides on 2 or 3 ganglioside bands, depending on the age examined. These bands migrate between ganglioside standards GD 1a and GM2 on HPTLC plates and have been designated GJ1, GJ2, and GJ3, with the higher number designating the more rapidly migrating species. Occasionally, additional bands migrating in the range of polysialogangliosides were observed. The pattern of expression of GJ species was studied in forebrain, retina, and cerebellar tissue taken from embryonic day 18 (E18), postnatal day 0 (P0), P7, P14, and adult animals. Both region-specific differences in the relative prominence of each band and stage-specific differences in the total amount of the JONES gangliosides were detected. The stage-specific differences in the amount of JONES antigens are well correlated with the developmental periods of maximal cell migration in each region. While the JONES gangliosides are most prominent in forebrain before birth, in they are most prominent during the first 2 postnatal weeks. In cerebellum, JONES antigen expression is more pronounced during the 2 periods of cell migration in this tissue. In retina, the more rapidly migrating GJ3 band was the most prominent band at all stages examined, and this same band is retained in the adult. In cerebellum and forebrain GJ3 is also the most pronounced band during development. However, in contrast to the retina, the more slowly migrating GJ1 band is retained in adult forebrain and cerebellum. A variety of non-brain tissues have also been examined for the presence of the JONES antigens.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Histiotypic organization and cell differentiation in rat retinal reaggregate cultures.

Reaggregate cultures have been formed from cell suspensions of neonatal rat retinas. Histological sections of the reaggregates showed evidence of lamination with central rosettes formed around a lumen, a clear neuropil layer and an outer cellular layer. Each of the major retinal cell types, except ganglion cells, could be positively identified using cell type-specific antibodies to label cryostat sections. Many of these were found to occupy positions within the reaggregates similar to those found in the intact retina. Electron microscopic observations showed abundant immature and mature synaptic endings within the neuropil layer, including a number of ribbon synapses. Examination of the rosettes showed an arrangement of Müller glia and photoreceptors that closely resembled that of the intact retina. Within the lumen of rosettes, photoreceptors were found to contain stacks of disc-like membranes bounded by a plasma membrane, analogous to immature outer segments. The photoreceptors within rosettes also underwent molecular differentiation and expressed an outer segment specific marker. The findings suggest that retinal cells have intrinsic properties that allow them to organize themselves into a correctly laminated structure and that particular cell interactions are necessary for continued differentiation of at least rod photoreceptors and Müller cells.

Animals↗
PubMed1987

Identification and characterisation of cell types accumulating GABA in rat retinal cultures using cell type specific monoclonal antibodies.

Monolayer and reaggregate cultures have been established from neonatal rat retina. After 7 days in culture, 60 nM [3H]gamma-aminobutyric acid (GABA) was used to identify cells with a high affinity uptake mechanism for GABA. Approximately 80% of the process-bearing cells were found to be labelled. These cells were identified as amacrine cells by double-labelling experiments combining [3H]GABA uptake with immunocytochemical labelling with monoclonal antibody HPC-1 which in retina is specific for amacrine cells. The ability of cultures to synthesize GABA from glutamate was investigated at various times. Little synthesis was observed during the first few days in culture. This lag was followed by an increase in the amount of synthesis until 3 weeks of culture. When clumps and reaggregate cultures of retinal cells were examined by [3H]GABA uptake, a time-dependent redistribution of labelled cells was observed. After 20 h in culture, GABA-positive cells were distributed over the whole cell mass. Over the next few days, the labelled cells became more common on the outer edge of the aggregates and less common in inner regions. By 7 days of culture, no labelled cell bodies were found on the inside of the aggregates, although such cells could be labelled by [3H]D-aspartate. The results provide positive identification of a subclass of retinal cells in culture, and show that at least one aspect of retinal histogenesis is not dependent upon extra-retinal tissues or the position imposed by the temporal order of retinal cell birth.

Animals↗
PubMed1987

Selective localization of glycine-accumulating cells in reaggregate culture of rat retina.

Uptake of [3H]glycine into the cells either in monolayer or reaggregate cultures of retina from two-day-old rat pups was studied. The glycine-accumulating cells (glycine cells) had short processes with several branches. Only 5% of process-bearing cells were labelled in the monolayer cultures. The major cell type, previously identified as photoreceptor cells, was unlabelled. In reaggregate cultures, the glycine cells were localized mainly in the outermost layer of the reaggregate. But the proportion of positive cells among all the cells in that layer was not so large. Although the cell type of the glycine cells has not yet been unambiguously identified, these results demonstrate a possible example of selective sorting out of a group of biochemically distinct cells from a cell mixture.

Animals↗
PubMed1987

O-acetylation of a cell-surface carbohydrate creates discrete molecular patterns during neural development.

The cell-surface antigen detected by the monoclonal antibody JONES is expressed in the retina and a number of other central nervous system regions of the rat during the latter part of embryonic development and the early postnatal period. In addition to the expression on certain neuroblast populations it is found on some but not all axons and is also expressed at high levels on the end feet of radial glia in regions through which axons actively grow. In the perinatal rat retina, almost all the antigenic activity was carried on a ganglioside migrating between GM1 and GM2. The epitope recognized by antibody JONES was base labile and treatment with 0.1 M sodium carbonate or ammonia vapor converted the antigen into GD3. Resistance to oxidation by sodium periodate and reformation of the epitope by chemical acetylation of base-treated gangliosides with N-acetylimidazole identify the antigen as 9-O-acetyl GD3. The acetylation of GD3 seems to be regulated independently from GD3 expression itself since acetylated and nonacetylated GD3 do not have identical immunocytochemical distributions in the developing central nervous system. In addition, five independent human melanoma cell lines varied substantially in their expression of 9-O-acetyl GD3, even though they all expressed high levels of GD3. Acetylation of ganglioside-linked sialic acid provides a mechanism for generating unique patterns of surface carbohydrates, which may influence cell interactions in development.

Acetylation↗