Michelle Southard-Smith Group (GUDMAP1)
Basic developmental neuro-urology studies are needed to gain insight into mechanisms underlying the pathogenesis of neurogenic bladder disorders and innervation deficiencies of external genitalia. Fundamental mechanistic knowledge of how autonomic innervation develops in these structures is lacking. Our long-term goals are to understand the migration processes, neural crest (NC) cell derivatives and essential signaling pathways that contribute to innervation of the developing bladder and external genitalia. We have established a series of transgenic mouse lines that drive expression of histological and fluorescent reporters in NC-derived cells as they migrate into the urogenital tract during development. The aims of this proposal are: 1) to derive a three-dimensional reconstruction of NC derivatives migrating into the developing bladder and genital tubercle based on transgene expression in murine embryos 2) to catalog cell phenotypes of NC derivatives within the bladder and genital tubercle by immunohistochemical labeling of transgene expressing cells with lineage markers and 3) to define the transcriptional profiles of NC-derived cell types in the bladder and genital tubercle during a focused developmental window.
To visualize migration of NC derivatives relative to the morphological ontogeny of the bladder and genital tubercle we are implementing 3-D reconstructions during the temporal window from embryonic day 11 to 16 (E11-E16). Immunohistochemistry on paraffin and cryo-sections of transgenic embryos will be applied to identify lineages including neuronal, glial, and smooth muscle cell types within the population of NC derivatives specifically at E14. Finally NC-derived cells will be isolated from E14 and E15 sub-dissected tissue by enzymatic dissociation and flow cytometry of transgene positive cells. RNA from sorted cells will be isolated, amplified and hybridized to microarray gene chips for identification of genes that are expressed as these structures develop. This analysis will define the migration pathways, cell type derivatives, and signaling pathways of NC progenitors that give rise to innervation during organogenesis of the bladder and external genitalia.
Relationship to the Overall Goals of GUDMAP
The primary goals of GUDMAP are to identify cell specific markers for key lineages, define morphogenetic events during organogenesis and assimilate temporal and spatial gene expression patterns during normal development. Our contribution to these goals will be to illuminate processes of NC derivatives that give rise to the innervation of the bladder and external genitalia. This analysis will significantly expand the current knowledge base of developmental neurobiology in these structures and may reveal innervation mechanisms that may be shared among other aspects of the urogenital tract.