Fig 2. shRNA-mediated knockdown of Nav1.1.
Lentiviral vectors were generated for two separate shRNAs targeting the rat Scn1a gene (sh-1 and sh-2). (A-B) Lentiviral vectors were tested in B50 neuroblastoma cells for efficient knockdown of the target gene. (A) Both shRNAs reduced Scn1a expression by approximately 70–80% compared to a control vector, (B) but did not affect expression of a related sodium channel gene, Scn2a. Mean +/- SEM, **p<.01. (C) Example of the MSDB injection site showing expression of the GFP reporter sequence in infected cells. Scale bar, 200 μm. (D) Quantification of mean fluorescent intensity per cell for Nav1.1 signal in GFP-positive and GFP-negative MSDB cells. Grey bars show mean +/- SEM, **p<.01. (E) Example of Nav1.1 immunofluorescence in the MSDB of a control and shScn1a injected rat showing a reduction in Nav1.1 fluorescent signal in neurons infected with the shScn1a, but not control, vector. Scale bar, 100 μm.