MACT (Mosaicism with AAV Mediated Conditional Transgenesis) for Single Neuron Analysis of Neurodegeneration in Vivo, a Proof of Principle in Focal Cerebral Ischemia

Abstract

The World Health Organization (WHO) warns neurodegenerative diseases (ND) in our aging population will sharply increase over the coming decades. Developing biological tools is paramount in the search for novel therapies to stop the onset or halt the progression of ND. We have developed a novel genetic method MACT (Mosaicism with AAV mediated Conditional Transgenesis) for single neuron analysis. MACT integrates the retrograde labelling capability of AAV and a conditional genetic reporter mouse model for sparse genetic labelling to reveal detailed morphology of different cellular types. As proof of principle, we subsequently subjected MACT mice to a middle cerebral artery occlusion (MCAO) leading to extensive neurodegeneration. Using two-photon imaging and 3-D reconstruction, we have illustrated the robust neurodegeneration of cortex pyramidal neuron, striatal medium spiny neuron and caught in action the engulfment of neurons and blood vessels by microglia. Moreover, we've documented dramatic axon degradation consistent with Wallerian pathology while the cell bodies remain intact ("dying back"), opening the possibility of therapeutic intervention. We also observed unexpected Tyrosine Hydroxylase (TH) positive cell bodies in the striatum, suggesting the brain responds to neurodegeneration by expressing these TH cells in an attempt to compensate for dopaminergic denervation. Most notably, we've developed a novel genetic method to visualize neuron morphology and gain a more accurate understanding of the cellular mechanisms underlying neurodegeneration. MACT represents a powerful genetics method to explore therapeutic interventions, including opto-and chemo-genetics in neurodegenerative diseases in vivo.