SUPPLEMENTARY INFORMATION
Cellular and Molecular Basis for Stress-Induced Depression
Ji-Seon Seo1, Jing Wei2, Luye Qin2, Yong Kim1, Zhen Yan2, Paul Greengard1,*
1Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, NY 10065 and 2Department of Physiology and Biophysics, School of Medicine and Biomedical Sciences, State University of New York, Buffalo, New York 14214
* Correspondence to Paul Greengard ().
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SUPPLEMENTARY FIGURE LEGENDS
SUPPLEMENTARY FIGURES S1-S4
SUPPLEMENTARY TABLES S1-S3
SUPPLEMENTARY FIGURE LEGENDS
Supplementary Figure S1. Knockdown of p11 in PrL induces depression-like behavior.(a) Immunofluorescence image illustrating the expression of viral transgene (GFP) after injecting the p11 shRNA lentivirus (p11-shRNA) into PrL. Scale bar, 20 μm. (b) p11 expression in PrL with or without the injection of GFP-shRNA or p11-shRNA lentivirus (n = 6 per group). **P < 0.01, one-way ANOVA. (c,d) Depression-like behaviors in mice with or without the injection of GFP-shRNA or p11-shRNA lentivirus, as measured by TST (c) and FST (d) (n = 8, CON; n = 10, GFP-shRNA; n = 10, p11-shRNA). * p < 0.05, ** p < 0.01, one-way ANOVA. Data are presented as the means ± s.e.m.
Supplementary Figure S2. Alteration of p11 expression is correlated with depression-like behavior. (a,b) Western blot analysis and quantification showing the expression of p11 in control versus stressed mice at 14, 30, 60, and 90 days after the cessation of stress (n = 6 per group). * p < 0.05, two-way ANOVA. (c,d) Depression-like behaviors in control versus stressed mice at 14, 30, 60, and 90 days after the cessation of stress, as measured by TST (c) and FST (d) (n = 16 for each group). Different animals were used at each time point. ** p < 0.01, two-way ANOVA. Data are presented as the means ± s.e.m.
Supplementary Figure S3. Identification of p11-expressing neurons in layer II/III PrL. (a) Images showing the co-localization of p11 (blue) with dopamine D1 receptor (D1; D1-Cre x tdTomato; red) and dopamine D2 receptor (D2; D2-eGFP; green) in layer II/III PrL neurons from D1-tdT-D2-eGFP (D1-Cre x tdTomato x D2-eGFP) mice. Scale bar, 15 μm. The percentage of neurons expressing p11, D2, or D1 is indicated in Venn diagrams. See Supplementary Table 1 for data. (b) Images showing the co-localization of p11 (blue) with D2 (D2; D2-tdT; red) and CaMKII (green, a marker of glutamatergic neurons) in layer II/III PrL neurons from D2-tdT (D2-Cre x tdTomato) mice. Scale bar, 10 μm. The percentage of neurons expressing p11, D2, or CaMKII is indicated in Venn diagrams. See Supplementary Table 2 for data. (c) Images showing the co-localization of p11 (blue) with D2 (D2; D2-tdT; red) and GAD67 (green, a marker of GABAergic neurons) in layer II/III PrL neurons from D2-tdT mice. Scale bar, 10 μm. The percentage of neurons expressing p11, D2, or GAD67 is indicated in Venn diagrams. See Supplementary Table S3 for data. In all the Venn diagrams, arrows point to the percentages of co-localized neurons.
Supplementary Figure S4. Chronic stress or p11 overexpression does not affect presynaptic functions. (a-c) Coefficient of variations (CV) of AMPAR-EPSC (a) and NMDAR-EPSC (b) and pre-paired ratio (PPR) of AMPAR-EPSC (c) in D2+ layer II/III PrL neurons from control mice (D2-tdT) and stressed mice with PrL injection of AAV_eYFP or AAV_p11 (a,b, n = 12 CON(D2), n = 8 RST(D2), n = 8, RST(D2)+AAV_eYFP, n = 9 RST(D2)+AAV_p11; c, n = 9 CON(D2), n = 8, RST(D2)+AAV_eYFP, n = 8 RST(D2)+AAV_p11). Inset: Representative traces of AMPAR-EPSC evoked by paired pulses at different intervals.
Supplementary Figure S1.
Supplementary Figure S2.
Supplementary Figure S3.
Supplementary Figure S4.
SUPPLEMENTARY TABLES
Supplementary TableS1. Quantification of the co-localization of p11, D1 and D2 in layer II/III PrL neurons.
p11+ / D1+ / D2+ / Triple+Total p11+ / 100 ± 12.04 / 5.29 ± 1.32 / 53.33 ± 3.91 / 1.32 ± 0.81
Total D1+ / 8.69 ± 2.17 / 100 ± 4.3 / 5.43 ± 1.71 / 2.17 ± 1.33
Total D2+ / 88.23 ± 12.10 / 1.82 ± 12.10 / 100 ± 11.86 / 1.96 ± 1.20
Red: Percentage of D2+ neurons in total p11-expressing cells.
Green: Percentage of p11+ cells in total D2-expressing neurons.
Supplementary TableS2. Quantification of the co-localization of p11, D2 and CaMKII in layer II/III PrL neurons.
p11+ / D2+ / CaMKII+ / Triple+Total p11+ / 100 ± 8.23 / 47.83 ± 3.01 / 86.61 ± 7.74 / 40.15 ± 1.76
Total D2+ / 90.75 ± 3.98 / 100 ± 4.13 / 94.21 ± 3.51 / 88.43 ± 3.87*
Total CaMKII+ / 76.92 ± 6.87 / 37.76 ± 1.48 / 100 ± 4.51 / 35.66 ± 1.56
Red: Percentage of D2+ neurons in total p11-expressing cells.
Green: Percentage of p11+ cells in total D2-expressing neurons.
Blue: Percentage of CaMKII+ neurons in total p11-expressing cells or D2-expressing neurons.
*: Percentage of CaMKII+ and p11+ neurons in total D2-expressing neurons.
Supplementary TableS3. Quantification of the co-localization of p11, D2 and GAD67 in layer II/III PrL neurons.
p11+ / D2+ / GAD67+ / Triple+Total p11+ / 100 ± 14.86 / 51.25 ± 5.78 / 14.37 ± 2.28 / 1.87 ± 0.88
Total D2+ / 93.82 ± 9.69 / 100 ± 11.29 / 7.40 ± 1.23 / 3.7 ± 1.74
Total GAD67+ / 25.28 ± 13.92 / 5.74 ± 1.48 / 100 ± 12.92 / 3.44 ± 1.62
Red: Percentage of D2+ neurons in total p11-expressing cells.
Green: Percentage of p11+ cells in total D2-expressing neurons.
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