Fig. 3.
Temporal development of the presynaptic connectome of newborn DG granule neurons. (A) Injection schemes (1–3) used in hGFAP-TVA mice. (B) Example of a presynaptic GFP-only positive neuron contacting a double-transduced newborn neuron (yellow arrowhead, enlarged in Insets). An individual stack of the boxed area (i) showing single and merged channels is depicted on the right. Arrowheads point to the axon of the presynaptic neuron. (Scale bars, 20 μm.) (C) Quantification of RABV-traced cells obtained following injection schemes (1–3) based on their morphology and location (n = 3–5). (D) Example of a RABV-traced mossy cell (white arrowhead, enlarged in the inset) located in the hilus of hGFAP-TVA mice following injection scheme 1. Enlargements of single and merged channels of a nearby double-transduced newborn neuron (yellow arrowhead, i) are shown on the right. (Scale bars, 30 μm.) (E) Example of RABV-traced neurons in the MS and NDB. (Scale bar, 70 μm.) (F) Ratio of RABV-traced neurons versus double-transduced newborn neurons following injection paradigms 1–3. (n = 3–5 mice per experimental condition; *P < 0.05). (G) Injection schemes (4 and 5) used in C57BL/6 mice. (H) Example of presynaptic tracing, 7 wk after retrovirus injection; double-transduced adult-generated granule neurons (yellow arrowheads) and putative presynaptic neurons (white arrowheads) are indicated. (Insets) Single channel images of the boxed cell. (Scale bars, 30 µM.) (I) Phenotypic characterization of RABV-traced local interneurons in the DG. (J) Example of a reconstructed RABV-traced basket cell profusely innervating the GCL. (Scale bars, 20 μm) (K) Visual and electrophysiological identification of presynaptic neurons in the DG and subiculum. GFP-only positive neurons (RABV-traced) were classified based on the location of their cell body (see micrographs) and the voltage responses following current injections. Several examples of presynaptic neurons differing in their IV traces and firing pattern are shown. (Scale bar, 20 μm.)