Figure 6.

Rbp4-Cre neurons form active circuits already at E14.5

(A) Left: schematic of NMDA+AMPA injection during in vivo embryonic two-photon imaging. Right: Rbp4-Cre neurons; color: normalized calcium activity).

(B) Change in fluorescence before (Pre) and after (Post) application of either cortex buffer (blue) or NMDA+AMPA (red). Wilcoxon signed rank test. n = number of Rbp4-Cre neuron ROIs from 3 E14.5 and 3 E18.5 embryos (Figure S7).

(C) Pairwise correlations of Rbp4-Cre neurons’ calcium activity, that are significantly greater than random (Figure S7). Shuffled data is on the left. Filled circles: correlations; gray shading: distribution; red line: median. Bars: percent of neuron pairs with correlations significantly greater than random (red). Wilcoxon rank-sum test. n = pairs of Rbp4-Cre neurons recorded from 3 (E13.5), 9 (E14.5), 5 (E15.5), 4 (E16.5), 5 (E17.5), and 6 (E18.5) embryos.

(D) Immunostaining of Rbp4-Cre neurons (green), Map2 (dendrites, red), NF (axons, white), Hoechst (blue).

(E) Left: schematic in vivo para-uterine imaging using 3D acousto-optic two-photon microscope. Top middle: mean projections around each soma. Bottom middle: Three zoomed examples (red outline, top middle). Right: Δf/f activity from examples. Cells 1 and 3 have high correlation.

(F) Pairwise correlations of E14.5 Rbp4-Cre neurons’ activity that are significantly greater than random, within and across layers. Dots: pairwise correlations; box (25–75 percentile) and whisker (5–95 percentile); line: median.

Scale bars: 10 μm (A), 30 μm (top, D), 100 μm (bottom, D), 20s and 5 %ΔF/F (E).

See also Figure S7.