Whole-mount LacZ staining of control embryos crossed with Rosa26LacZ demonstrates recombination within the proximal peripheral nerve by E13.5 and most of the sciatic nerve by E14.5 (data not shown). E15.5 Erk1/2CKO(Dhh)

Rosa26LacZ embryos showed a morphologically similar pattern of recombination and peripheral nerve patterning as littermate controls ( Figures 5A and 5B). The distribution of Schwann cells in the mature, P20 Erk1/2CKO(Dhh) Z/EG phrenic nerve projections appeared similar to controls, with Schwann cells present up to the NMJ ( Figures S5B and S5C). These finding suggest Schwann cells take up relatively normal positions in Erk1/2CKO(Dhh) embryos. Although Schwann click here cells appeared to be present within the nerve, gross dissection of P18, Erk1/2CKO(Dhh) sciatic nerves revealed markedly decreased nerve caliber and increased translucency ( Figure 5C). Electron microscopy revealed a clear, striking reduction in the number of myelinated axons and an increase in the number of unmyelinated axons in Erk1/2CKO(Dhh) mice ( Figures 5D–5F). There was no change in the number of Schwann cell nuclei within the sciatic nerve and dying, pyknotic nuclei were not detected at this stage ( Figure 5G). These data show that loss of ERK1/2 in Schwann cell progenitors clearly inhibits myelination. The development of myelinating

Schwann cells involves the upregulation of numerous factors, including Egr2/Krox-20, S100β, and various myelin components (Jessen and Mirsky, 2005). Immunohistochemical analysis of P18 Erk1/2CKO(Dhh) sciatic nerves revealed a 77.8% ± 8.1% decrease in the number of Egr2/Krox-20 positive cells science and the expression of OSI-906 concentration S100β was nearly absent ( Figures 5H–5K). GFAP immunolabeling of non-myelinating Schwann cells appeared normal at this stage (data not shown). These findings suggest that ERK1/2 is required for the progression of the myelinating Schwann cell lineage after initial specification. In order to better understand the potentially diverse developmental mechanisms underlying ERK1/2 regulation of PNS development, we performed

microarray analysis on RNA extracts derived from E12.5 Erk1/2CKO(Wnt1) and wild-type DRGs. We did not detect overt changes in DRG neuron number at this developmental stage, suggesting the profile is a reflection of ERK1/2 regulated genes and not the loss of any particular cell type. 209 distinct genes met our inclusion criteria, which included 98 downregulated and 111 upregulated genes in Erk1/2CKO(Wnt1) samples ( Figures 6A and S6). Functional annotation of regulated genes revealed significant changes in mediators of transcriptional regulation, cell-cell signaling, intracellular signaling, and cell-cycle/division ( Figures 6A and S6). A number of genes involved in transcriptional regulation were modified that have been shown to regulate glial development (Figure 6B). Microarray changes were validated by qPCR of DRG samples from E12.