PubMed · 7711846
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G L Bratthauer. 1994. The avidin-biotin complex (ABC) method.. https://doi.org/10.1385/0-89603285-x%3A175
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The purpose of the present report was to investigate to what extent the new peroxidase substrate Vector VIP (V-VIP) can be used in combination with DAB chromogen for the unequivocal and permanent detection of colocalising antigens within a single neurone, according to a two-colour paradigm. With this aim, retrograde tract-tracing with cholera toxin B subunit (CTB) or fluoro-gold (FG) was performed to disclose individual, identified subpopulations of neurones in the primate substantia nigra projecting to the caudate nucleus or to the putamen, respectively. Each tracer was detected by means of a PAP procedure and finally stained brown using DAB as a chromogen. Subsequently, both series of sections were processed for the immunocytochemical detection of tyrosine hydroxylase (TH). TH-immunoreactive neurones were stained purple with the peroxidase substrate V-VIP. As a result of the present procedure, several cell bodies of projection neurones, stained brown, can easily be identified within the primate substantia nigra. Some of these neurones additionally displayed purple TH immunoreaction product located in the neuronal dendrites. By contrast, CTB- or FG-unlabelled neurones only show the typical purple precipitate that belongs to V-VIP substrate, both in the cell body as well as in the dendrites.
The sensory input to the neostriatum from groups of cortical cells related to individual facial vibrissae has been investigated at both light- and electron-microscopic resolution. The purpose of the study was to establish the extent to which corticostriatal input maintains the anatomical coding of spatial information that is present in cortex. A double anterograde tracing method was used to identify the output projections from groups of adjacent neurons in different barrel columns, so that the anatomical relationships between two groups could be studied throughout their length. Adjacent whiskers are represented in adjoining cortical barrels and an examination of corticostriatal projections from these reveals two patterns of projection. In one, the anatomical topography is partially preserved; the barrels are represented in adjoining, discrete, areas of the somatosensory neostriatum. In the second projection pattern, the neostriatal innervation is diffuse and adjacent barrels are represented in overlapping regions of the neostriatum. Moreover, the fibres are thinner, have smaller boutons, and are present in both the ipsilateral and contralateral neostriatum. The two systems also enter the neostriatal neuropile separately. The discrete topographic system enters the adjacent neostriatum as collaterals which leave the descending corticofugal fibres at right angles, while the diffuse system enters directly from the corpus callosum at an acute angle. Examination of the neostriatal terminal fields by correlated light and electron microscopy, shows that characteristic axospinous terminals on spiny neurons are made by both groups of cortical fibres, although they differ in their size and morphology. It is concluded that at least two corticostriatal pathways arise from the barrel cortex. One connection maintains some of the anatomical code implicit in the barrel pattern of primary somatosensory cortex, but another, more diffuse, system is overlaid upon it which may carry different information from this complex area of cortex.
A simple technique for the morphological characterization of pre-labelled neurons in fixed brain slices is described. Neurons are retrogradely-labeled with a carbocyanine dye and the tissue is fixed and sliced. Individual labeled cells from the interior of a slice are then visualized on an upright fluorescence microscope and impaled with a micropipet containing rhodamine or fluorescein-peroxidase conjugates. The cells are filled by iontophoresis, postfixed, and the peroxidase is oxidized into a permanent, opaque reaction product. The dendritic morphology of the neurons is then reconstructed under camera lucida and quantified.