Unbiased Multidimensional Analysis Reveals Novel Principles of Cortical Interneuron Synaptic Organization

• Main Author: Dummer, Patrick Daniel
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-2024
• Subjects: Artificial intelligence; Cellular biology; Developmental biology; Neurosciences; Systematic biology
• ISBN: 9798383570593

Neurons display exquisite specificity in synaptic connectivity, but we lack a complete understanding of neuronal connectivity and the rules that govern it. A major impediment to addressing this question lies in the vast diversity of neurons, the small size and large number of synapses formed by any given neuron over a wide territory, and the need to study these connections in intact tissue. We therefore developed an image-based tool to assess synaptic specificity in tissue sections and dissociated culture. We focused on three interneuron subpopulations that target distinct sub-cellular regions of the post-synaptic cell: soma-targeting basket cells (BCs), axon initial segment (AlS)-targeting chandelier cells (ChCs), and distal dendrite-targeting somatostatin cells (SstCs). Using mouse dissociated cortical culture as a starting point, we built a machine learning (ML) based image processing and analysis pipeline to classify individual presynaptic boutons at scale. Supervised ML classification revealed similar subcellular targeting profiles for these interneuron populations in slice and culture, indicating that targeting is primarily regulated by cell intrinsic programs. We also observed a remarkable target-dependent laminar organization in vivo. An unsupervised ML analysis using the same input data not only identified the same three canonical targeting classes, but also revealed that these classes are comprised of multiple subpopulations. In slice, these synaptic subpopulations displayed distinct laminar organization. In dissociated culture, two soma-targeting synaptic subpopulations mapped to target cells with different cellular profiles. The six dendrite-targeting synaptic subpopulations were found at increasing distances from target soma, suggesting molecularly distinct proximal, medial, and distal dendritic compartments in culture. Tracking subtype targeting across axonal branches of individual neurons indicated that SstCs and BCs utilize distinct targeting strategies in culture that accord with established findings in vivo. In sum, our synaptic analysis pipeline revealed novel synaptic subpopulations in interneurons. Further analysis uncovered novel aspects of interneuron synaptic biology that, remarkably, are retained in culture.