3D optical reconstruction of whole-brain neuropeptidergic connectome in Caenorhabditis elegans

Understanding how distinct, spatially organized cell types in the brain interact to generate specific functions is the central challenge in neuroscience. While efforts to map synaptic “wiring” networks have advanced our understanding of neural circuits, synapses represent only one type of neuronal communication. Many critical interactions occur via volume transmission in which chemical massagers – particularly neuropeptides - are released extrasynaptically, diffuse to target cells, and activate receptors that trigger downstream gene expression changes. This form of “wireless” signaling plays essential roles in all nervous systems. Despite their importance, neuropeptide signaling remain poorly characterized, largely due to the slow dynamics of neuropeptide signaling and the lack of direct physical contact between communicating cells. Here, we propose a highly interdisciplinary approach – which we refer to as Peptide-Clamp - that integrates patterned illumination, optogenetics, spatial transcriptomics, and machine learning to enable high-throughput, functional mapping of neuropeptidergic connectivity and transcriptome at single-cell resolution. This approach consists of three components: (1) high-throughput optical control of neuropeptide release in individually 'sender' neurons; (2) followed by volumetric transcriptome imaging of the same brain to measure gene expression in situ; (3) machine learning to identify significant transcriptional changes to casually link the responding ‘receiver’ cells to the ‘sender’ cells. We will apply this approach to generate the whole-body transcriptome and neuropeptidergic connectome maps in C. elegans. The resulting data will allow us to uncover the organizational principles of neuromodulatory communication and systematically reveal how neuropeptide signaling drives cell-type-specific transcriptional responses at the organism level.