Figure 1.

Transient coexpression of two developmentally relevant transcription factors stably reprograms fibroblasts to attain properties of functionally mature neurons. (a) Transient coexpression of Brn3a with either Ngn1 or Ngn2 for 8 d in fibroblasts induced cells with neural morphology that stained for the pan-neural markers Map2 (red) and Tuj1 (green) Cells were immunostained on day 14. Scale bars represent 100 μm. (b) Coexpression of Brn3a with Ngn1 or Ngn2 was required for induction of Map2 and Tuj1 double-positive cells. Transcription factors were induced for 8 d and immunostained on day 14. Data are presented as means ± s.e.m. from three independent replicates. n = 50,000 cells per genotype. (c) Synapsin expression (red) in BN2 neural cells. Cells were counterstained for Tuj1 (green). (d) Vamp (red) expression in BN1 neural cells. Scale bars in c and d represent 25 μm. (e) The majority of neurons induced with BN1 and BN2 expressed synaptic markers indicative of mature neurons (Vamp, (BN1 = 100%, n = 52, BN2 = 100%, n = 44; Synapsin, BN1 = 93.4%, n = 47, BN2 = 97.9%, n = 44). Data are presented as means ± s.e.m. from two independent experiments. (f) Whole-cell currents recorded in voltage-clamp mode. Inward fast-inactivating Na⁺ and outward currents were observed. (g) Representative subthreshold voltage responses and action potentials evoked from neural cells after 14 d in culture. Current pulses of 350-ms starting at −145 pA. (h) Representative single voltage spike isolated from trace in g measured at +95-pA current level. (i) Representative train of spontaneous action potentials observed in 14-d culture.