THE TEMPORAL AND SPATIAL APPEARANCE OF HUNTINGTIN AGGREGATES IN THE BRAINS OF THE ZQ175 MOUSE MODEL OF HUNTINGTON'S DISEASE

Background Huntington’s disease (HD) is caused by a CAG repeat expansion in exon 1 of the HTT gene. The zQ175 knock in mouse model displays extensive behavioural, histopathological, and molecular phenotypes. In the preparation for testing HTT-lowering approaches, it is essential that temporal and spatial appearance of HTT aggregates has been defined in the brains of zQ175 mice. Aim To map the appearance of aggregate species in the brains of zQ175 mice at one monthly intervals from one to six months of age. Methods DAB based immunohistochemistry (IHC) with the S830 antibody was optimized. In order to demonstrate that a given staining pattern represented an aggregated form of HTT, the anti-polyglutamine antibody, 4H7H7, was used in conjunction with formic acid treatment to retrieve polyglutamine epitopes, that had been masked through the aggregation process. These techniques were applied to coronal brain sections from heterozygote zQ175 and wild type mice. Results HTT aggregates could be detected in brains of two month old zQ175 mice, but not in brains from mice at one month of age. At two months, S830 detected diffuse nuclear staining and small puncta in striatum, CA1 region of hippocampus and neuronal layer IV of the cortex. Immunostaining with 4H7H7, only detected these HTT species after pretreatment with formic acid, demonstrated that they represented an aggregated form of HTT. The aggregate distribution and formation of inclusions increases with age over the six month period. Conclusion Aggregated forms of HTT can be detected in the brains of zQ175 mice at two months of age by IHC methods. We were able to demonstrate that the diffuse HTT immunostaining pattern in neuronal nuclei represents an aggregated form of HTT. We shall use sensitive seeding assays to determine the youngest postnatal age, or embryonic stage, at which the aggregation process can first be detected. This detailed analysis of the appearance of HTT aggregates will be important to assess the effects of HTT-lowering strategies on HTT aggregation when administered to zQ175 mice at different ages. Funding This work was funded by the CHDI Foundation.