Figure 5. Partial loss of Homer1b/c from distal dendrites of Snx6^-/- neurons.

(A) Neurons were co-transfected with pLL3.7.1 and EGFP or EGFP-SNX6 construct on DIV13, fixed on DIV18 and immunostained with antibodies to Homer1b/c. Shown are representative confocal images of dendritic segments. (B) Quantification of puncta number per 100 μm dendrite length and mean intensity in spines for Homer1b/c (mean ± SEM, n = 30, N = 3). (C) Neurons were transfected with pLL3.7.1 on DIV13, fixed on DIV18 and immunostained with antibodies to PSD95. (D) Quantification of PSD95 distribution in dendrites (mean ± SEM, n = 30, N = 3). (E) DIV14 neurons were co-transfected with constructs overexpressing EGFP and mCherry or mCherry-SNX6, fixed on DIV16 and immunostained with antibodies to Homer1b/c. shown are representative confocal images of transfected neurons. Dashed lines outline the cell bodies. (F) Quantification of Homer1b/c distribution in the cell body and dendrites, and its mean intensity in spines in (E) (mean ± SEM, n = 30, N = 3). (G) DIV13 neurons were co-transfected with constructs expressing DsRed and EGFP, EGFP-SNX6, mEmerald-Homer1c-FL or EGFP-Homer1c-C and fixed on DIV18. (H) Quantification of spine density in (G) (mean ± SEM, n = 30, N = 3). Bars: 20 μm in (E), 2 μm in other panels.

DOI: http://dx.doi.org/10.7554/eLife.20991.011

Figure 5—figure supplement 1. Immunoblotting analysis of Homer1b/c and PSD95 in hippocampi from Snx6^fl/fl and Nestin-Cre; Snx6^fl/fl mice.

β-actin serves as loading control. Shown are samples from three pairs of littermates. Data represent mean ± SEM.

DOI: http://dx.doi.org/10.7554/eLife.20991.012

Figure 5—figure supplement 2. Ablation of SNX6 causes decrease in spine density of CA1 but not CA3 neurons.

(A) Hippocampal neurons in dissociated culture were transfected with pLL3.7.1 on DIV14, fixed on DIV18 and immunostained with antibodies to CTIP2, a transcription factor specifically expressed in DG and CA1, but not CA3 neurons. Compared with DG neurons, CA1/CA3 neurons have larger cell bodies and more primary dendrites. Shown are representative confocal images of CA1 and CA3 neurons. (B) Quantification of spine density in (A) (mean ± SEM, n = 30 cells, N = 3). Bar: 2 μm.

DOI: http://dx.doi.org/10.7554/eLife.20991.013