Correlation between behavioral deficits in HD and WT mice and disease progression.

<p>(A) Body weights of HD mice are significantly lower than those of their WT littermates at 22 and 30 weeks. (B) HD mice had lower whole brain weights at 22 and 30 weeks than their WT littermates. (C) At 22 and 30 weeks HD mice have poorer motor ability and fall off the accelerating rota-rod at lower speeds than WT littermates. (D) The percentage of HD mice expressing the rear-paw clasping (RPC) motor phenotype increases to 100% by 30 weeks. Data are expressed as mean ± s.e.m, <i>n</i>≥3 animals, *p≤0.05.</p

Representative images from the immunstaining of 33-week-old WT and HD hippocampus.

<p>Representative images of BrdU (A) and DCX (B) staining in the WT hippocampus. Composite images of BrdU (red) and DCX (green) double-staining in the WT (C) and HD (D) hippocampus. Composite images of BrdU (red) and GFAP (green) double-staining in the WT (E) and HD (F) hippocampus. All images were taken using the 20x objective.</p

Quantification of <i>in vivo</i> neurogenesis in the hippocampus of symptomatic HD and WT mice.

<p>Significantly fewer BrdU-positive (A, <i>n</i> = 8 HD and <i>n</i> = 9 WT animals), DCX-positive (B, <i>n</i> = 5 HD and <i>n</i> = 6 WT animals), BrdU/DCX double-positive (C, <i>n</i> = 5 HD and <i>n</i> = 6 WT animals) and BrdU/GFAP double-positive (D, <i>n</i> = 8 HD and <i>n</i> = 9 WT animals) cells were observed in the hippocampus of 33-week old HD mice compared to WT littermates. Data are expressed as mean ± s.e.m, *p≤0.05, **p≤0.01.</p

Increased precursor activity in the hippocampus but not the SVZ of symptomatic HD mice.

<p>(A) Similar numbers of SVZ-derived neurospheres were generated from HD and WT littermates at all time points examined. (B) A significant increase in the number of hippocampal-derived neurospheres in HD mice compared to WT littermates was observed at 30 (<i>n</i> = 5) and 33 weeks of age (<i>n</i> = 4). (C) Younger (16-week old) HD mice generated a similar number of hippocampal neurospheres to their WT littermates and the number of neurospheres could be increased by the addition of depolarizing levels of KCl (<i>n</i> = 3). More neurospheres were generated from older symptomatic HD mice (33 weeks; <i>n</i> = 4) than the corresponding WT mice. However, unlike the WT mice, which show a significant increase in neurosphere number following <i>in vitro</i> depolarization, no further increase was observed in the HD mice in the presence of additional KCl. All data are expressed as mean ± s.e.m, *p≤0.05, **p≤0.01, ***p≤0.001.</p

In vivo assessment of 5-HT_1A and 5-HT₂ post-synaptic receptor function using 8-OH-DPAT-induced change of corticosterone levels and DOI-induced head-twitches.

<p>Compared to paired-saline injected animals, administration of the 5-HT_1A agonist 8-OH-DPAT (0.3 mg/kg, s.c.) significantly increased corticosterone levels in both (A) male and (B) female mice, regardless of the genotype. In the DOI-induced head-twitches experiment (bottom panel), there was a significant effect of genotype in both sexes. Indeed, the number of head-twitches following DOI (1 mg/kg) were decreased in (C) HD male and (D) HD female mice when compared to WT animals. Values represent means (± SEM) of n = 5–10 mice per group. Saline vs. DPAT0.3: (+) p<0.05, (++) p<0.01, (+++) p<0.001 WT vs. HD: (*) p<0.05, (**) p<0.01, (***) p<0.001.</p

Concentrations of 5-HT and 5-HIAA in brain tissue.

<p>Using the HPLC system, we measured the tissue levels of serotonin (5-HT) and its metabolite 5-hydroxyindolacetic acid (5-HIAA) in several brain areas. Both 5-HT and 5-HIAA concentrations were decreased in the hippocampus (A/B), the cortex (C/D) and the striatum (E/F) in female HD mice when compared to WT animals. Levels of 5-HIAA were also reduced in male HD. Finally, male mice exhibited higher hippocampal tissue levels of both 5-HT and 5-HIAA when compared to female animals. 5-HT and 5-HIAA are expressed in ng/g tissue. Values represent means (± SEM) of n = 6–8 mice per group. WT vs. HD: (*) p<0.05, (**) p<0.01, (***) p<0.001. Male vs. female: (##) p<0.01.</p

Effect of sex and HD mutation on saccharin-preference test and total fluid intake.

<p>(A) We found a significant interaction between sex and genotype (F_(1,44) = 5.84, p<0.05) on saccharin preference (expressed as % of total fluid intake). Indeed compared to WT animals, only HD female mice exhibited reduced saccharin preference. (B) Interestingly looking at total fluid intake (expressed in mL), we revealed an overall effect of genotype (F_(1,44) = 12.3, p<0.01) but no significant effect of sex or interactions. Values represent means (± SEM) of n = 8–14 mice per group. WT vs. HD: (*) p<0.05, (**) p<0.01.</p

Effect of sex and HD mutation on the novelty suppressed feeding test (NSFT).

<p>(A) Analyzing the time (expressed in sec) to complete the task in the NSFT, we found a significant interaction between sex and genotype (F_(1,55) = 4.2, p<0.05). Indeed compared to WT animals, only HD female mice exhibited greater delay to complete the NSFT. (B) Interestingly measuring the amount of food consumed after NSFT, we did not find any effect of genotype (F_(1,55) = 1.7, p = 0.19) or interaction with the sex (F_(1,55) = 0.39, p = 0.54). Values represent means (± SEM) of n = 12–17 mice per group. WT vs. HD: (***) p<0.001.</p

Gene expression of 5-HT transporter (5-HTT), 5-HT_1A receptor (5-HT_1AR) and tryptophan hydroxylase-2 (TPH2) in the raphe.

<p>Measuring mRNA levels of several genes encoding proteins that regulate 5-HT homeostasis, we found significant genotype-sex interactions for both mRNA levels of (A) 5-HTT and (B) TPH2 but not for (C) 5-HT_1AR gene expression. Both 5-HTT and TPH2 mRNA levels were decreased in female WT when compared to male WT. In addition, HD mutation decreased both 5-HTT and TPH2 gene expressions in male only. Finally 5-HT_1AR mRNA levels were not affected by either the sex or the genotype. Values represent means (± SEM) of n = 5–6 mice per group. WT vs. HD: (**) p<0.01. Male vs. female: (##) p<0.01.</p