05-08-08(18:28:29)
Authors:
Anderova M. 1,3
Prajerova I. 1,2,
Honsa P. 1
and Chvatal A. 1-3
Institutions:
1 Department of Cellular Neurophysiology - Laboratory of Neurobiology, Inst. Exp. Med., ASCR;
2 Dept. Neurosci. and
3 Center for Cell Therapy and Tissue Repair 2nd Medical Faculty, Charles University; Prague, Czech Republic
Title of abstract : The role of Sonic hedgehog and Wnt-7a in neural stem cell differentiation in vitro
Abstract text:
Sonic hedgehog (Shh) and Wnt-7a are secreted morphogens involved in ongoing neurogenesis of the adult brain. GFP-labeled P0 mouse neural stem cells expressing either Shh (Shh/GFP) or Wnt-7a (Wnt-7a/GFP) were used to study their effect on neural stem cell proliferation and differentiation in vitro; cells expressing only GFP (WT/GFP) were used as a control. Eight days after the onset of differentiation the cells were analyzed for expression of cell specific markers and receptors using immunohistochemical and Western blot analyses. The cell membrane properties were characterized using the patch-clamp technique. Both Shh and Wnt-7a increased the expression of neuronal marker MAP-2, however, only Wnt-7a also increased the expression of DCX and beta-III tubulin, when compared to control. In WT/GFP cells electrophysiological analysis revealed a neuron-like current pattern in MAP-2-, DCX- or beta-III tubulin-positive cells, with a mean Vm of -77 mV and IR of 876 MOhms, voltage-dependent KA, KDR and TTX-sensitive Na+ currents (INa). In Shh/GFP cells the number of cells expressing the neuron-like current pattern was significantly decreased when compared to control. In Wnt-7a/GFP cells the neuron-like current pattern was predominant and also the INa occurrence was increased Based on our immunohistochemical data we can conclude that both Shh and Wnt-7a increase the expression of neuronal markers in differentiated neural stem cells compared to WT/GFP cells. However, electrophysiological analysis revealed an increased incidence of neuron-like current patterns only in Wnt-7a/GFP cells.
Supported by GACR 305/06/1316, GAUK62/2006/C/2.LF, AVOZ50390512, LC554 and 1M0538.
