Adult neurogenesis and neurite outgrowth are impaired in LRRK2 G2019S mice - PubMed (original) (raw)

Comparative Study

doi: 10.1016/j.nbd.2010.12.008. Epub 2010 Dec 16.

H L Melrose, C Zhao, K M Hinkle, M Yue, C Kent, A T Braithwaite, S Ogholikhan, R Aigner, J Winkler, M J Farrer, F H Gage

Affiliations

Comparative Study

Adult neurogenesis and neurite outgrowth are impaired in LRRK2 G2019S mice

B Winner et al. Neurobiol Dis. 2011 Mar.

Abstract

The generation and maturation of adult neural stem/progenitor cells are impaired in many neurodegenerative diseases, among them is Parkinson's disease (PD). In mammals, including humans, adult neurogenesis is a lifelong feature of cellular brain plasticity in the hippocampal dentate gyrus (DG) and in the subventricular zone (SVZ)/olfactory bulb system. Hyposmia, depression, and anxiety are early non-motor symptoms in PD. There are parallels between brain regions associated with non-motor symptoms in PD and neurogenic regions. In autosomal dominant PD, mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are frequent. LRRK2 homologs in non-vertebrate systems play an important role in chemotaxis, cell polarity, and neurite arborization. We investigated adult neurogenesis and the neurite development of new neurons in the DG and SVZ/olfactory bulb system in bacterial artificial chromosome (BAC) human Lrrk2 G2019S transgenic mice. We report that mutant human Lrrk2 is highly expressed in the hippocampus in the DG and the SVZ of adult Lrrk2 G2019S mice. Proliferation of newly generated cells is significantly decreased and survival of newly generated neurons in the DG and olfactory bulb is also severely impaired. In addition, after stereotactic injection of a GFP retrovirus, newly generated neurons in the DG of Lrrk2 G2019S mice exhibited reduced dendritic arborization and fewer spines. This loss in mature, developed spines might point towards a decrease in synaptic connectivity. Interestingly, physical activity partially reverses the decrease in neuroblasts observed in Lrrk2 G2010S mice. These data further support a role for Lrrk2 in neuronal morphogenesis and provide new insights into the role of Lrrk2 in adult neurogenesis.

Published by Elsevier Inc.

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Figures

Figure 1

Figure 1. Transgenic human G2019S Lrrk2 is highly expressed in the hippocampus

A: Real time RT-PCR with a human-specific LRRK2 Taqman probe reveals highest transgene mRNA expression in the hippocampus. B: Finer mapping of transgene expression using i_n situ_ hybridization with a human-specific probe confirms high DG (arrow) expression of Lrrk2. C: Immunoblot with a Lrrk2-specific antibody shows robust transgenic human expression.

Figure 2

Figure 2. Proliferation is decreased in G2019S transgenic mice

A: Experimental design for the non-transgenic control (NT) and Lrrk2 G2019S mice. Animals were perfused 24 hours after a single BrdU injection (100 mg/kg). B: BrdU cell numbers indicating mitotic cells within the hippocampal DG of the respective groups are decreased in G2019S Lrrk2 transgenic mice; mean number of BrdU-positive cells ± SD, *p<0.01. C: Representative images of DAB staining of BrdU-positive cells in the DG of the different groups, NT: non-transgenic. Scale bar, 50 µm. D–F: Proliferation, presented as the mean number of BrdU-positive cells ± SD, is decreased in the SVZ of G2019S Lrrk2 transgenic mice; E, F show representative images of an overview of the BrdU-positive cells, that are decreased in the SVZ/ RMS of Lrrk2 G2019S mice (F) compared to NT (E). Scale bar in E, F, 50 µm. G: Schematic drawing of the SVZ/olfactory bulb (OB) system. H, I: A thinner stream of newly generated cells (BrdU-positive) is present in the rostral migratory stream (RMS) of Lrrk2 G2019S mice (I) compared to controls (H), Scale bar, 250 µm. Expression of LRRK2 is observed in the SVZ of NT mice (J, Lrrk2 in green merged with Dapi, blue in K) and more strongly in the SVZ of LRRK2 G2019S mice (L, Lrrk2 in green merged with Dapi, blue in M). Scale bar, 20 µm (E, F, H, I).

Figure 3

Figure 3. Olfactory neurogenesis is decreased in G2010S transgenic mice

A: Experimental design for the non-transgenic control (NT) and Lrrk2 G2019S mice. Animals received BrdU injections (50 mg/kg) on five consecutive days and were perfused one month after the BrdU injections. B: Schematic overview of the two neurogenic regions of the olfactory bulb analyzed, the granule cell layer (GCL) and the glomerular layer (GLOM). C–E: Analysis of neurogenesis in the GCL in NT (C) and Lrrk2 G2010S mice (D) reveals a significant decrease in BrdU/NeuN-positive cells in the transgenic mice (E, *p<0.01). BrdU-positive cells are depicted in green in C, D; NeuN-positive cells are in blue in C, D. (Scale bar in C, D, 25 µm). F–M: Analysis of neurogenesis in the GLOM. Neurogenesis (I) and dopaminergic neurogenesis (M) are both decreased in transgenic mice (*p<0.01). F–H: examples of newly generated neurons (BrdU/NeuN) and J–L: examples of newly generated dopaminergic neurons (BrdU/TH, Scale bars F–H and J–L, 10 µm). BrdU is depicted in green, TH is depicted in red and NeuN is depicted in green.

Figure 4

Figure 4. Fewer new neurons survive in G2019S transgenic mice

A: Experimental design for the non-transgenic control (NT) and Lrrk2 G2019S mice. B: Cell numbers of newly generated cells (BrdU-positive) in the hippocampal DG from the respective groups are presented as the mean number of BrdU-positive cells ± SD, *p<0.01. C: Representative images of BrdU-positive cells (green) colabeled with NeuN (red) for the groups indicated. Scale bar, 50 µm. D: Cell numbers of newly generated neurons (BrdU/ NeuN-positive) in the DG, *p<0.0.01, compared to NT.

Figure 5

Figure 5. Defects in dendrite outgrowth of new neurons in G2019S transgenic mice

A: Experimental design for the non-transgenic control (NT) and Lrrk2 G2019S mice. Animals were stereotactically injected with CAG-GFP retrovirus and were perfused one month later. B: Representative images of GFP-positive cells in the DG of NT and Lrrk2 G2019S mice. Scale bar, 20 µm. Quantitative analysis for dendrite length (C) and branching points (D), ± SD, *p<0.01.

Figure 6

Figure 6. Spine number and maturation are decreased in G2019S transgenic mice

A: Experimental design for the non-transgenic control (NT) and Lrrk2 G2019S mice. Animals were stereotactically injected with CAG-GFP retrovirus and were perfused one month later. B: Representative images of GFP-positive dendrites in the DG of NT and Lrrk2 G2019S mice. Scale bar, 5 µm. Quantitative analysis for C: dendritic spines/µm and D: percentage of mushroom spines, * p<0.001.

Figure 7

Figure 7. Running partly reverses negative impact of G2019S on neurogenesis

A: Experimental design for the non-transgenic control (NT) and Lrrk2 G2019S mice, running (run) or non-running (non-run). B: Representative images of DCX-positive neuroblasts in the DG of NT and Lrrk2 G2019S mice (red: DCX, blue: NeuN). Note the increase in numbers of DCX-positive cells after running. C: Quantitative analysis for the total number of DCX-positive cells in the DG. Scale bar, 50 µm, * p<0.05.

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