Fig. 5. F-actin staining in isolated MG neurons. (A) The control shows a representative example of the so called migratory-phenotype, with F-actin staining in the neurite and only a single fiber bundle in the soma (red). When the embryos were cultured in ODQ, the MG neurons showed a stationary-phenotype with multiple actin fiber bundles in the somata (ODQ). Co-staining with anti-HRP antiserum (green) reveals the neuronal identity of the cell (anti-HRP), whereas non-neuronal cells are unlabelled. Scale bars: 10 µm. (B) Quantitative evaluation of the actin cytoskeleton. After incubation in neurochemicals for 24 hours, a high percentage of MG neurons show the normal migratory phenotype in cultured control embryos (control). Conversely, a significantly smaller number of neurons with migratory phenotype was found in the presence of NOS inhibitor 7NI (500 µM). The disruptive effect of 7NI on the actin cytoskeleton could be rescued by the addition of 1 mM protoporphyrin IX free acid (7NI + protop.). Similarly, the number of MG neurons showing the migratory phenotype was reduced after incubation in the sGC inhibitor ODQ (200 µM). This disruptive effect of ODQ on the actin cytoskeleton organization could be rescued by the addition of 500 µM 8Br-cGMP (ODQ + 8Br-cGMP). (B) Neurochemicals that affect the cAMP/PKA pathway also had an effect on the actin organization. A significantly smaller number of neurons from embryos cultured in the AC activator forskolin (100 µM) or the PKA activator SPcAMPS (50 µM) showed the migratory phenotype as compared with controls. The PKA inhibitor RPcAMPS (50 µM) had no significant effect on the actin cytoskeleton organization. ^**P<0.005; ^***P<0.001.

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