Figure 1: Retrograde tracing and electrophysiological analysis of single- or dual-virus labelled GCs.

Horizontal sections through (a) area CA3, (b) DG and (c) MEC and LEC, derived from a mouse injected with LVV followed by EnvA-ΔG-MCh rabies virus (red) infection Scale bar, 200 μm. (a) MCh⁺-labelling of area CA3 pyramidal cells. (b) hGFP⁺ GC nuclei labelled with LVV (green). Rabies virus injected in the same location resulted in dual-labelled 'starter' cells (co-localization of green and red). (c) Afferent input from the LEC and MEC to the 'starter' cells in DG. Nuclei were labelled with 4′-6-diaminodino-2-phenylindole (DAPI) (blue). (d) Traces representative of membrane potentials in response to current injections, from newborn GCs (RTG) at different time points (15, 25, 30 and 60 d.p.i., as indicated at the top), mature unlabelled GCs (CON) and cells labelled with LVV. Recordings were made from cells with single (green trace) and double labelling (orange trace). Depolarizing steps were delivered at threshold and twice threshold current. Scale: 10 mV, 50 ms. (e) Traces representative of spontaneous postsynaptic currents recorded under voltage clamp (Vh=−80 mV; scale 10 pA, 100 ms). (f) Photomicrographs showing newborn GCs labelled with retrovirus (RTG) expressing nuclear hGFP, and MCh at 30 d.p.i. Scale bar, 25 μm. (g) Summary histograms of passive and active membrane properties. Measurements did not show significant differences between single and double labelling (P>0.05). Differences between mature and newborn GCs were analysed with Kruskal–Wallis test followed by Dunn's test, ***P<0.001, **P<0.01, *P<0.05; CON (n=20), LVV (n=8), LVV+MCh (n=5), 15 d.p.i. RTG (n=7), 21–25 d.p.i. RTG (n=3), 21–25 d.p.i. RTG+MCh (n=4), 28–35 d.p.i. RTG (n=7), 28–35 d.p.i. RTG+MCh (n=3), 60–65 d.p.i. RTG (n=3), 60–65 d.p.i. RTG+MCh (n=3). Data are means±s.e.m. DG, Dentate Gyrus; MEC, medial entorhinal cortex; LEC, lateral entorhinal cortex.