Fig. 3
Immunolabeling of different Nav subtypes in hESC-derived and mouse RPE, mass-spectrometry studies of Nav expression, and patch clamp recordings with selective Nav blockers. a, b The specific pattern of Nav subtypes was studied by immunolabeling. Laser scanning confocal microscopy Z-maximum intensity projections (xy-MIP) and yz cross-sections of a mature hESC-derived or b mouse RPE. Nav subtypes 1.4, 1.6, and 1.8 (green) were immunolabeled together with filamentous actin (phalloidin stain, red). Scale bars 10 μm. Right side panels show a higher magnification of the highlighted regions. Patch clamp recordings were performed on mature hESC-derived RPE using selective blockers for channel subtypes. c Nav subtypes were sequentially blocked by extracellularly applied 4,9-AnhydroTTX (30 nM, Nav1.6 blocker), A-803467 (1 μM, Nav1.8 blocker) and μ-Conotoxin GIIB (600 nM, Nav1.4 blocker). The average normalized peak current–voltage relationship (I/Imax vs Vm) was determined from all recordings (mean ± SEM, n = 7). d Applying the selective blockers in combination with TTX (10 μM) removed most of the Nav currents (n = 11). e Mass spectrometry analysis of Nav channel expression in hESC-derived RPE. Specific peptides were identified for all Nav subtypes, excluding Nav1.2