Characterization of HTT Inclusion Size, Location, and Timing in the zQ175 Mouse Model of Huntington´s Disease: An In Vivo High-Content Imaging Study

Abstract

Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the huntingtin gene. Major pathological hallmarks of HD include inclusions of mutant huntingtin (mHTT) protein, loss of neurons predominantly in the caudate nucleus, and atrophy of multiple brain regions. However, the early sequence of histological events that manifest in region- and cell-specific manner has not been well characterized. Here we use a high-content histological approach to precisely monitor changes in HTT expression and characterize deposition dynamics of mHTT protein inclusion bodies in the recently characterized zQ175 knock-in mouse line. We carried out an automated multi-parameter quantitative analysis of individual cortical and striatal cells in tissue slices from mice aged 2–12 months and confirmed biochemical reports of an age-associated increase in mHTT inclusions in this model. We also found distinct regional and subregional dynamics for inclusion number, size and distribution with subcellular resolution. We used viral-mediated suppression of total HTT in the striatum of zQ175 mice as an example of a therapeutically-relevant but heterogeneously transducing strategy to demonstrate successful application of this platform to quantitatively assess target engagement and outcome on a cellular basis.

Frequency of nuclear mutant huntingtin inclusion formation in neurons and glia is cell‐type‐specific

“…Interestingly, only neuronal nuclear inclusions changed in size with disease progression, increasing from 0.6 µm to a maximum of 2 µm at the end stage. This is in agreement with earlier findings in R6/2 mice, zQ175 mice, and human patients (Carty et al, ; Gutekunst et al, ; Li et al, ). This suggests that nuclear inclusions persist for a long period of time and increase in size, most likely due to sequestration of newly synthesized mHTT but also the sequestering of other proteins.…”

Polyglutamine-Expanded Huntingtin Exacerbates Age-Related Disruption of Nuclear Integrity and Nucleocytoplasmic Transport

“…Htt Q175 mice closely mimic the human genetic context, express mutant huntingtin containing expanded polyglutamine (polyQ Htt), and gradually develop behavioral and pathological abnormalities similar to those observed in HD patients (Heikkinen et al, 2012, Menalled et al, 2012, Smith et al, 2014, Carty et al, 2015). Strikingly, expression of Htt Q175 from both alleles markedly increased the number of cells with altered nuclear envelopes to 89% in cortex and 62% in striatum of 24 month-old Htt Q175/Q175 mice (Figures 1A, C and S2A,B).…”

“…We investigated nuclear envelope alterations in mice expressing physiological levels of a ~175 CAG trinucleotide repeat expansion within one ( Htt Q7/Q175 ) or both ( Htt Q175/Q175 ) endogenous huntingtin ( Htt ) alleles (Heikkinen et al, 2012, Menalled et al, 2012, Smith et al, 2014, Carty et al, 2015). We first tested whether wild-type mice with 7 glutamines (Q) in both endogenous Htt alleles ( Htt Q7/Q7 ) develop age-dependent nuclear envelope defects in cortex and striatum, two HD-vulnerable brain regions in which >80% of the cells are neurons detectable with the neuron-specific marker Neuronal Nuclear Antigen (NeuN) (Gusel’nikova and Korzhevskiy, 2015) (Figure S1A).…”

Overview of Huntington's Disease Models: Neuropathological, Molecular, and Behavioral Differences

“…Cortical aggregate density matched striatum only when the mice reached 14 to 16 months of age (Bayram‐Weston, Torres, Jones, Dunnett, & Brooks, ; Yu et al., ). In heterozygous Q175s, the accumulation of mutant HTT inclusions is age‐dependent; aggregates are detected as early as 3 months, with an age‐dependent progressive increase in both cortex and striatum (Carty et al., ; Peng et al., ; Smith et al., ). However, the size of the nuclear inclusions is significantly larger in the striatum at 6 months, but larger in the cortex at 8 months (Carty et al., ).…”

“…In heterozygous Q175s, the accumulation of mutant HTT inclusions is age‐dependent; aggregates are detected as early as 3 months, with an age‐dependent progressive increase in both cortex and striatum (Carty et al., ; Peng et al., ; Smith et al., ). However, the size of the nuclear inclusions is significantly larger in the striatum at 6 months, but larger in the cortex at 8 months (Carty et al., ). Accordingly, evidence suggests that mutant HTT nuclear inclusions are responsible for altered gene expression in striatum (Kuhn et al., ; Luthi‐Carter et al., ).…”