DISC1-mediated dysregulation of adult hippocampal neurogenesis in rats

Abstract

Adult hippocampal neurogenesis, the constitutive generation of new granule cells in the dentate gyrus of the mature brain, is a robust model of neural development and its dysregulation has been implicated in the pathogenesis of psychiatric and neurological disorders. Previous studies in mice have shown that altered expression of Disrupted-In-Schizophrenia 1 (Disc1), the mouse homolog of a risk gene for major psychiatric disorders, results in several distinct morphological phenotypes during neuronal development. Although there are advantages to using rats over mice for neurophysiological studies, genetic manipulations have not been widely utilized in rat models. Here, we used a retroviral-mediated approach to knockdown DISC1 expression in dividing cells in the rat dentate gyrus and characterized the morphological development of adult-born granule neurons. Consistent with earlier findings in mice, we show that DISC1 knockdown in adult-born dentate granule cells in rats resulted in accelerated dendritic growth, soma hypertrophy, ectopic dendrites, and mispositioning of new granule cells due to overextended migration. Our study thus demonstrates that the Disc1 genetic manipulation approach used in prior mouse studies is feasible in rats and that there is a conserved biological function of this gene across species. Extending gene-based studies of adult hippocampal neurogenesis from mice to rats will allow for the development of additional models that may be more amenable to behavioral and in vivo electrophysiological investigations. These models, in turn, can generate additional insight into the systems-level mechanisms of how risk genes for complex psychiatric disorders may impact adult neurogenesis and hippocampal function.

Keywords: DISC1; adult neurogenesis; dentate gyrus; hippocampus; psychiatric disorders; schizophrenia.

Figure 1

Stereotaxic injection of retrovirus in the dentate gyrus of the adult rat hippocampus. (A) Schematic diagram of the retroviral vector used for in vivo birth-dating and genetic manipulation. (B) Sample projections of Z-series confocal images at 4 wpi of retrovirus either expressing GFP (control) or co-expressing shRNA against Disc1 and GFP (sh-D1) in the dentate gyrus of the adult rat hippocampus. Scale bar: 200 μm.

Figure 2

DISC1 regulates morphogenesis of adult born neurons in the rat hippocampus. (A) Sample projections of Z-series confocal images of GFP⁺ neurons at 2 and 4 wpi of retrovirus expressing GFP (control) or co-expressing shRNA against Disc1 and GFP (sh-D1). Scale bar: 10 μm. Note that the newborn neurons with DISC1 knockdown show increased soma size and ectopic primary dendrites. (B) Quantification of soma size of GFP⁺ neurons at 2 and 4 wpi. Numbers indicated in each bar graph represent the total number of neurons analyzed from 3–8 animals under each condition. Values represent mean ± SEM (*p < 0.05; Two- way ANOVA; 2 wpi: control, n = 87, sh-D1, n = 88; 4 wpi: control 105, sh-D1, n = 98). (C) Quantification of number of primary dendrites of GFP⁺ neurons at 2 and 4 wpi. Numbers indicated in each bar graph represent the total number of neurons analyzed from 3–8 animals under each condition. Values represent mean ± SEM (*p < 0.05; Two-way ANOVA; 2 wpi: control, n = 202, sh-D1, n = 346; 4 wpi: control, n = 155, sh-D1, n = 323).

Figure 3

DISC1 regulates the positioning of the adult born neurons in the rat hippocampus. (A) Sample confocal images of GFP and DAPI. Scale bar: 20 μm. (B) Schematic diagram of the adult rat dentate gyrus region divided into four domains. (C) Distribution of GFP⁺ cells within each area as defined in (B). Numbers indicated in each bar graph represent the total number of neurons analyzed from 3–8 animals under each condition (2 wpi: control, n = 404, sh-D1, n = 295; 4 wpi: control, n = 358, sh-D1, n = 353).

Figure 4

DISC1 regulates the dendritic development of adult born neurons in the rat hippocampus. (A) Sample projections of Z-series confocal images of GFP⁺ neurons at 2 and 4 wpi. Scale bar: 10 μm. (B) Total dendritic length of GFP⁺ neurons at 2 and 4 wpi. Numbers indicated in each bar graph represent the total number of neurons analyzed from 3–8 animals under each condition. Values represent mean ± SEM (*p < 0.05; Two-way ANOVA; 2 wpi: control, n = 87, sh-D1, n = 89; 4 wpi: control, n = 105, sh-D1, n = 99). (C) Number of branches of GFP⁺ neurons at 2 and 4 wpi. Values represent mean ± SEM (*p < 0.05; Two-way ANOVA; 2 wpi: control, n = 87, sh-D1, n = 89; 4 wpi: control, n = 105, sh-D1, n = 99).