PROJECT SUMMARY/ABSTRACT The brainstem is a complex and early-developing brain region that is responsible for sensory, motor, autonomic, and critical-for-life functions. The first biology-based hypothesis of autism suggested that the brainstem's reticular formation was responsible for the behavioral features of autism. However, technological barriers have prevented the field from reliably characterizing brainstem substructures in vivo. Excitingly, recent technological advances now allow us to examine the microstructural properties of the brainstem's individual nuclei and white matter tracts using Magnetic Resonance Imaging. In our original R01, our group contributed to these technological advances through innovative development and refinement of acquisition and post-processing techniques that optimize brainstem imaging. We applied these techniques in autistic children to show how brainstem substructures relate to sensorimotor and core diagnostic features. Yet, the brainstem is highly connected, and we still do not know how the brainstem contributes to autistic whole-brain networks. Leveraging the findings of the original R01, the overall scientific premise of this renewal is that brainstem substructures, as key hubs in whole-brain networks, may hold significant insights into the neurobiological underpinnings of primary and secondary features of autism. Given the functions of the brainstem, we hypothesize that the whole-brain connections of key brainstem nuclei will explain variation in sensorimotor, autonomic, feeding, and core autism features. This hypothesis will be tested through two specific aims: 1) Determine brainstem-inclusive whole-brain network differences and markers of social and repetitive behavior features in autistic children compared to non-autistic children. 2) Identify brainstem- inclusive network associations with sensorimotor and autonomic features in autistic and non-autistic children. We will perform brainstem-inclusive whole-brain tractography across three datasets: 1) our original R01's data of brainstem-optimized imaging and measures of sensorimotor and autistic features in 74 autistic and 74 non-autistic children; 2) a new dataset with further brainstem-imaging improvements, feeding, and respiratory sinus arrhythmia measures in 74 autistic and 74 non-autistic children/youth; and 3) publicly available core diagnostic and neuroimaging data to which we will apply brainstem-optimized processing. An innovative Mahalanobis Distance measure and graph theory measures will quantify group and individual differences in brainstem-inclusive networks. Successful completion of this research will provide a quantitative characterization of brainstem-inclusive whole-brain networks that will advance the understanding of the neurobiological basis for core and co-occurring autism features. These contributions will be significant by paving the way for determining age-appropriate and biology-informed interventions for the prevalent sensorimotor...