A Critical Period for Development of Cerebellar-Mediated Autism-Relevant Social Behavior.

The cerebellum has been increasingly implicated in autism spectrum disorder (ASD) with many ASD-linked genes impacting both cerebellar function and development. However, the precise timing and critical periods of when abnormal cerebellar neurodevelopment contributes to ASD-relevant behaviors remains poorly understood. In this study, we identify a critical period for the development of ASD-relevant behaviors in a cerebellar male mouse model of tuberous sclerosis complex (TSC), by using the mechanistic target of rapamycin (mTOR) inhibitor, rapamycin, to pharmacologically inhibit dysregulated downstream signaling. We find independent critical periods during which abnormal ASD-relevant behaviors develop for the two core ASD diagnostic criteria, social impairments and behavioral flexibility, and delineate an anatomic, physiological, and behavioral framework. These findings not only further our understanding of the genetic mechanisms underlying the timing of ASD-relevant behaviors but also have the capacity to inform potential therapies to optimize treatment interventions.SIGNIFICANCE STATEMENT No targeted treatments currently exist for autism spectrum disorder (ASD). This complex developmental disorder has established links to genetic and circuit aberrations, yet the precise timing and coordination of these underlying mechanisms that contribute to the spectrum of physiological and behavioral abnormalities remains unclear. Cerebellar pathology is consistently seen in ASD individuals; therefore, we sought to identify the specific windows for cerebellar involvement in the development of ASD-relevant behaviors. Using pharmacologic treatment paradigms, we outline distinct critical periods of developmental vulnerability for ASD-relevant social and inflexible behaviors. From this study, we posit a refined window of time during which ASD symptoms develop that will inform therapeutic timing.

Hydrophobic and Hydrogel-Based Methods for Passive Tissue Clearing.

Optical tissue clearing enables the precise imaging of cellular and subcellular structures in whole organs and tissues without the need for physical tissue sectioning. By combining tissue clearing with confocal or lightsheet microscopy, 3D images can be generated of entire specimens for visualization and large-scale data analysis. Here we demonstrate two different passive tissue clearing techniques that are compatible with immunofluorescent staining and lightsheet microscopy: PACT, an aqueous hydrogel-based clearing method, and iDISCO+, an organic solvent-based clearing method.