Cerebellar purkinje cell dysfunction contributes to gait impairments in Shank3-mutant mice

Complex motor dysfunction is a common comorbidity of autism spectrum disorder (ASD) and may be attributed to the perturbation of cerebellar function. Purkinje cells (PCs), which serve as the primary output neurons of the cerebellum, are integral to various aspects of cerebellar motor function, such as gait control. However, the specific contribution of PCs to ASD has not been fully characterized. In this study, we utilized adult male homozygous Shank3-knockout (Δ13–16, KO) mice, a well-established autism mouse model, to investigate the relationship between structural and functional deficits in PCs and impaired motor performance. Digital gait analysis was used to examine gait abnormalities. Changes in dendritic arborization and cell body area were measured using sparse labeling technology. In vitro electrophysiology was used to evaluate alterations in PC firing patterns in Shank3-KO mice. Shank3-KO mice displayed autistic-like behaviors (stereotyped grooming behavior and social disorders) and motor impairments. Gait impairment was obvious in Shank3-KO mice and was mainly spatiotemporal and anatomical in nature. The stand duration and swing duration decreased, the body speed and swing speed clearly increased, and the print position parameters changed. PCs, the sole output neurons of the cerebellar cortex, exhibited reduced line density in different lobules and decreased cell body area in Shank3-KO mice, accompanied by cellular hypoactivity. Notably, the optogenetic inhibition of PCs in wild-type (WT) mice induced gait impairments, whereas the excitation of PCs in Shank3-KO mice rescued gait abnormalities. Although cerebellar dysfunction, particularly in PCs, is implicated in gait abnormalities, the contributions of other regions, such as the basal ganglia and cerebral cortex, remain to be fully elucidated. Similarly, within the cerebellar circuitry, other cell types, including granule cells and interneurons, are also likely involved. An important limitation of our study is its restriction to male mice, leaving potential sex-dependent differences unexplored. Future studies are therefore necessary to delineate the roles of these various brain regions and cell types and to examine their interactions in both sexes. In this study, we found that Shank3-KO mice exhibit deficits in motor performance and gait, which are also observed in ASD patients. These motor dysfunctions were strongly linked to cerebellar pathology, and we established that PC dysfunction underlies the disrupted gait patterns in this ASD model.