The purpose of the work outlined here is to see if all DRG cells within a particular functional class express a single member of the trk family of tyrosine kinase neurotrophin receptors, and whether the particular trk expressed determines which neurotrophin (or neurotrophins) regulate the functional class during development. In order to assess this idea, four specific hypotheses will be tested: I. All DRG cells express a single, functional tyrosine kinase neurotrophin receptor, II. DRG cell classes with identified functions can be correlated with expression of particular trks, III. Neurotrophins appropriate for the trk expressed by particular DRG cell classes are expressed in both their central and peripheral target fields during development, and IV. Neurotrophins regulate spinal axonal arborizations of DRG cells and do so selectively. The developmental patterns of expression of mRNA for the trk family of tyrosine kinase neurotrophin receptors trkA, trkB, trkC, and of the neurotrophins, NGF, BDNF, NT3, and NT4/5 will be assessed by in situ hybridization. Functional DRG classes which express particular members of the trk family will be identified by combining in situ hybridization with retrograde tracing from characteristic peripheral and central target fields and with immunohistochemistry. Finally, the functional effects of different neurotrophins on specific DRG cell populations will be assessed by staining characteristic spinal arborizations of different classes of DRG cells with DiI after neurotrophin administration in utero. All of the work proposed here will require interaction with CORE resources. Administration of neurotrophins and in situ hybridization will require interaction with the Growth Factor Support CORE (F). Quantitative analysis of in situ hybridization histochemistry will require creation of video montages and automated grain counting to be performed with the Scanning Stage Image Processing Computer Microscope CORE (B). Finally, quantitation of branching of fluorescently-stained dorsal root axons will require the Confocal Microscope with Volume Rendering And Object Segmentation Core (C). Determination of the biological influences of neurotrophins on DRG cells is fundamental to the prospects of using neurotrophins as treatments for peripheral neuropathies and as agents to promote regeneration of sensory axons in the spinal cord.
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