Impaired basal and running-induced hippocampal neurogenesis coincides with reduced Akt signaling in adult R6/1 HD mice

Abstract

Huntington's disease (HD) is a fatal neurodegenerative disorder affecting a range of cellular and molecular functions in the brain. Deficits in adult hippocampal neurogenesis (AHN) have been documented in the R6/1 mouse model of HD. Here we examined basal and running-induced neuronal precursor proliferation in adult female and male R6/1 HD mice. We further tested whether sequential delivery of voluntary running followed by environmental enrichment could synergistically enhance functional AHN in female R6/1 HD mice. R6/1 HD mice engaged in significantly reduced levels of voluntary running, with males showing a more severe deficit. Basal neural precursor proliferation in the hippocampal sub-granular zone remained unchanged between female and male R6/1 HD mice and neither sex significantly responded to running-induced proliferation. While discrete provision of running wheels and enriched environments doubled AHN in adult female R6/1 HD mice it did not reflect the significant 3-fold increase in female wildtypes. Nevertheless, triple-label c-Fos/BrdU/NeuN immunofluorescence and confocal microscopy provided evidence that the doubling of AHN in female R6/1 HD mice was functional. Intrinsic cellular dysfunction mediated by protein aggregates containing mutant huntingtin (mHtt) did not appear to coincide with AHN deficits. In the hippocampus of female R6/1 HD mice, proliferating precursors and 6 week old adult-generated neurons were devoid of mHtt immuno-reactive aggregates, as were endothelial, microglial and astroglial cells populating the neurogenic niche. Serum transforming growth factor-β concentrations remained unaltered in female R6/1 HD mice as did the hippocampal levels of proliferating microglia and glial fibrillarly acidic protein expression. Examining the growth hormone/insulin-like growth factor 1 (GH/IGF-1) axis showed no change in base-line serum GH between genotypes. However, despite a reduced distance, acute running increases serum GH in both female wildtype and R6/1 HD mice. Serum IGF-1 levels were increased in female R6/1 HD mice compared to wildtypes during daytime inactive period, while hippocampal levels of the IGF-1 receptor remained unchanged. Running induced Akt phosphorylation in the hippocampus of female wildtype mice, which was not reflected in R6/1 HD mice. Total Akt levels were decreased in the hippocampus of both control and running R6/1 HD mice. Our results show adult-generated hippocampal neurons in female R6/1 HD mice express c-Fos and that running and Akt signaling deficits may mediate reduced basal and running-induced AHN levels.

Introduction

Huntington's disease (HD) is an autosomal dominantly inherited neurodegenerative disorder. The pathophysiology of HD manifests primarily in the brain with progressive symptomology that includes cognitive, psychiatric and motor dysfunction (Borrell-Pages et al., 2006, Cattaneo et al., 2005, HDCRG, 1993, Reiner et al., 1988). The onset of HD typically occurs in the adult with a disease outcome that is invariably fatal. At the genetic level HD results from an expanded cytosine–adenine–guanine (CAG) trinucleotide repeat that encodes a poly-glutamine (polyQ) tract in the N-terminus of the huntingtin protein (Hatters, 2008, HDCRG, 1993). The expanded polyQ stretch imparts a propensity for the mutant huntingtin protein (mHtt) to form intracellular aggregates and proteolytic cleavage allows mutant fragments to enter the nucleus and form protein aggregates (Wellington et al., 1998). The presence of polyQ-containing aggregates in the brain is a pathological hallmark of HD and is postulated to contribute to neuronal dysfunction and death (Davies et al., 1997). The R6/1 transgenic mouse line closely models human HD, including the development of aggregates, and provides an important platform to study underlying disease mechanisms and therapeutic strategies (Gil and Rego, 2009).

Cell-based approaches remain a focus of therapeutic intervention for HD, which hitherto remains without effective therapeutic intervention (Clelland et al., 2008). However, cell-based therapy for HD has had equivocal outcomes and preclinical studies are paramount in developing effective neural regeneration approaches. The emergence of adult neurogenesis as an endogenous neuronal repair mechanism has provided therapeutic appeal to a variety of neurodegenerative disorders including HD (Vandenbosch et al., 2009). Adult neurogenesis ensues throughout the life of mammals in discreet niches, including the sub-granular zone (SGZ) of the hippocampus. Adult hippocampal neurogenesis (AHN) is postulated to be an important neuronal substrate mediating the positive effects of running and enrichment on learning and memory (Lafenetre et al., 2011). Deficits in AHN have been demonstrated in several rodent models of HD (Gil et al., 2004, Gil et al., 2005, Lazic et al., 2004, Simpson et al., 2011) and reviewed in Gil-Mohapel et al. (2011). AHN deficits in the R6/1 HD transgenic mouse model manifest progressively and could underlie the impaired cognitive and depressive-like behaviors that develop in this mouse model (Grote et al., 2005, Lazic et al., 2004, Lazic et al., 2006, Ransome et al., 2012).

A diversity of environmental, physiological and genetic factors contributes to the regulation of AHN levels. Exercise in the form of voluntary running stimulates neuronal precursor proliferation to increase AHN levels and improve cognition (Ransome and Turnley, 2008, van Praag et al., 1999a, van Praag et al., 1999b). R6/1 HD mice given access to voluntary running have sex-dependent responses to exercise-induce neurotrophin expression (Zajac et al., 2010). However, whether sex is a determinant of exercise-induced AHN in R6/1 HD mice remains unknown. Cognitive stimulation through environmental enrichment also increases AHN levels but does so by promoting neuronal survival (Kempermann et al., 1997). Although R6/1 HD mice remain responsive to environmental enrichment strategies aimed at improving AHN and cognition (Lazic et al., 2004, Lazic et al., 2006, Nithianantharajah et al., 2008, Pang et al., 2006), there remains a paucity of evidence demonstrating functional AHN. Our current study was performed to examine whether sex affects basal or running-induced AHN in R6/1 HD and wildtype mice and determine whether adult-generated hippocampal neurons express c-Fos in R6/1 HD mice.