Sonic hedgehog (Shh) and Wnt-7a are morphogens involved in embryonic as well as ongoing adult neurogenesis. Their effects on the differentiation and membrane properties of neonatal neural stem/progenitor cells (NS/PCs) were studied in vitro using NS/PCs transduced with either Shh or Wnt-7a. Eight days after the onset of in vitro differentiation the cells were analyzed for the expression of neuronal and glial markers using immunocytochemical and Western blot analysis, and their membrane properties were characterized using the patch-clamp technique. Our results showed that both Shh and Wnt-7a increased the numbers of cells expressing neuronal markers; however, quantitative immunocytochemical analysis showed that only Wnt-7a enhanced the outgrowth and the development of processes in these cells. In addition, Wnt-7a markedly suppressed gliogenesis. The electrophysiological analysis revealed that Wnt-7a increased, while Shh decreased the incidence of cells displaying a neuron-like current pattern, represented by outwardly rectifying K(+) currents and tetrodotoxin-sensitive Na(+) currents. Additionally, Wnt-7a increased cell proliferation only at the early stages of differentiation, while Shh promoted proliferation within the entire course of differentiation. Thus we can conclude that Shh and Wnt-7a interfere differently with the process of neuronal differentiation and that they promote distinct stages of neuronal differentiation in neonatal NS/PCs.

Laboratory of Neurobiology Department of Cellular Neurophysiology Institute of Experimental Medicine The Academy of Sciences of the Czech Republic Videnska 1083 142 20 Prague 4 Czech Republic

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