Spatial Restriction of Apoptotic Machinery during Neuronal Apoptosis and Pruning

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Project Summary/Abstract Neurons are capable of activating pathways that induce either the degeneration of the entire cell by apoptosis or to selectively degenerate only the axons by pruning. While the main components of the caspase activating machinery during apoptosis and pruning have been identified, a fundamental question of whether the apoptotic machinery is activated throughout the neuronal soma and axons, or is spatially restricted, during these events remains unknown. This question is most relevant in the context of pruning where one predicts the apoptotic machinery to be localized to the targeted axons undergoing degeneration. However, we were surprised to find an unexpected spatial restriction of caspase activation even during apoptosis, where we found these to be restricted primarily to the soma, even though both soma and axons degenerate. In this proposal, we will mechanistically examine the spatial localization of the apoptotic machinery in neurons during apoptosis and pruning. We will utilize neurons cultured in microfluidic chamber devices to allow for the spatial segregation and manipulation of neuronal somas and axons. Our hypothesis is that during apoptosis and pruning, the restricted caspase activity causes the “physiological axotomy” of axons, activating the Sarm1-mediated axotomy pathway of axon degeneration. The concept that the developmental pathways of apoptosis and pruning can cause axotomy is novel because these pathways were considered to be distinct from the injury-induced axotomy pathway. In Aim 1, we will define the spatial restriction of the apoptotic machinery in neurons during apoptosis and axon pruning. In Aim 2, we focus on examining the function of Sarm1 during apoptosis and axon pruning. In Aim 3, we will investigate the Sarm1-deficient mice for pruning defects in vivo and evaluate if these mice exhibit behavioral deficits. This project opens exciting areas of research not only because of its new concepts for apoptosis and pruning, but also because it brings into focus a developmental function of Sarm1 that is beyond its recognized role in axon injury.