Patterned barrel-like maps of the whiskers or fingers exist in the trigeminal (V) subnuclei principalis (PrV) and interpolaris (SpVi), the dorsal column nuclei (DCN), thalamus and barrel cortex. Little is known of how these maps form. Proposed studies will address this issue by revealing mechanisms controlling development of V afferents and their brainstem target cells. Our general hypothesis is that Drg11, a paired homeodomain transcription factor, is required for development of the whisker-barrel neuraxis. Initial studies in Drg11-/- mice show that such patterns fail to develop in the PrV, thalamus, and whisker cortex; but that they do develop in the SpVi, DCN and limb cortex. Thus, Drg11 may have a specific role in PrV pattern formation. Drg11-/- mutants also display excessive prenatal cell death in PrV. These preliminary data prompt the following specific hypotheses and experiments: 1. In Drg11-/-s, somatotopic patterns fail to develop in the PrV, and consequently in the VPM thalamus and whisker barrel cortex; and that such patterns develop normally in the Sp Vi, DCN, VPL thalamus and limb barrel cortex. Wildtype mice will be analyzed to assess where and when Drg11 is expressed in the developing barrel neuraxis. Pattern markers will then be applied to brains from Drg11-/-s to determine where, when, how, and if somatotopic patterns develop in the barrel neuraxes. Transient, delayed or morphologically altered pattern formation will be of particular interest. 2. Drg11 is necessary for normal morphogenesis and neuronal survival in the PrV, but not in the V ganglion, SpVi, or DCN. Neurons will be assessed and counted in the PrV, V ganglion, SpVi, and DCN from Drg11-/- and wildtype mice during prenatal intervals shown to be the most prolific epochs for apoptosis. Then, to assess contributing effects of altered input and output pathways upon PrV cell survival in Drg11-/-s, V ganglion and PrV projections will be analyzed. 3. In Drg11-/- mice, PrV cell death is sufficient to explain pattern alterations there and in high-order targets of the PrV. Drg11+/- and Bax-/- mice will be crossed so as to potentially dissociate PrV patterning anomalies from exuberant apoptotic cell death in Drg11-/-mice. Normal PrV and higher-order patterns in these double knockouts would indicate that Drg11 is necessary for PrV cell survival, not pattern formation. 4. In Drg11 -/-mice, exo utero electroporation-induced overexpression of Drg11 in the embryonic PrV will rescue PrV cells from the -/- phenotype, therein preserving normal cell numbers and pattern formation there. Drg11 will be augmented in the PrV of Drg11-/- embryos by intrauterine injection of DNA in the brainstem followed by electroporation mediated gene transfer. PrV cell morphogenesis, numbers, inputs, transcription factor expression, and pattern formation will be assessed as above. Control groups consist of wild-type embryos subjected to identical electroporation parameters, as well as Drg11+ embryos receiving control plasmid injections and identical Drg11 electroporation parameters.
