Figure 1.
Nav channel localization at the AIS is disrupted by expression of FGF14F145S. A–R, Fluorescence images of rat hippocampal neurons expressing mRFP together with GFP (A–F), FGF14F145S-GFP (G–L), or FGF14-GFP (M–R) stained with a monoclonal anti-Nav α subunit-specific antibody, PanNav, visualized with an Alexa 647-conjugated secondary antibody (B, H, N), and a polyclonal anti-MAP2 antibody, visualized with an AMCA-conjugated secondary antibody (C, I, O). GFP fluorescence images are shown in A, G, and M, and mRFP fluorescence images are shown in D, J, and P. Overlay images of GFP and mRFP are shown in the green and red channels (E, K, Q). Overlay images of PanNav and mRFP are shown in cyan and red channels (F, L, R). The bottom panels (a–r) are high-magnification images of the boxed regions in each of the corresponding panels A–R, presented to highlight the AIS regions (arrows). Comparing B, H, and N, it is evident that the intensity of the Nav α subunit labeling in the AIS regions is lower in the FGF14F145S-expressing neuron (H) and is higher in the FGF14-expressing neuron (N) than in the cell expressing GFP (B). Scale bars, 10 μm. S–U, Representative examples of Nav α subunit immunofluorescence intensity line scans along the AIS regions in individual neurons expressing GFP (S), FGF14F145S-GFP (T), or FGF14-GFP (U). In each panel, the black line indicates the mean fluorescence intensity profile obtained from GFP-expressing (S; n = 20), FGF14F145S-GFP-expressing (T; n = 23), or FGF14-GFP-expressing (U; n = 28) neurons from one set of transfections. V, Mean ± SEM of Nav α subunit immunofluorescence intensities in GFP-expressing (n = 40), FGF14F145S-GFP-expressing (n = 47), and FGF14-GFP-expressing (n = 58) cells are plotted. Values were normalized to the mean control value determined in GFP-expressing cells (‡p < 0.05; ***p < 0.001).