The Molecular Regulation of Wiring Specificity in the Serotonin System Across Key Postnatal Stages
The serotonin system in the brain is implicated in a myriad of functions and pathologies, ranging from fundamental processes such as sleep and breathing to complex cognitive functions like learning and memory. Notably, it is the primary target for treating depression and anxiety. Reflecting its broad functional spectrum, serotonin neuron projections influence nearly every brain region. Despite their extensive reach, most forebrain-projecting serotonin neurons originate from discrete nuclei in the dorsal and median raphe (DR, MR) and are surprisingly few in number compared to other neuronal types. This raises the question of how such a limited population can innervate nearly all central nervous system circuitry during development. Recent research has highlighted the diversity within midbrain serotonin neurons, revealing heterogeneity in connectivity, electrophysiology, and gene expression. Prior studies have identified distinct cortical and subcortical subsystems with varying transcriptomic profiles. These genetically distinct, projection specific subsystems require appropriate signals during axon innervation. However, the molecular mechanisms behind how serotonin neurons specifically innervate the correct target region during development is un-known. To address this question, we profiled the single-cell transcriptomic trajectories of midbrain serotonin neurons across eight developmental timepoints. We further integrated snRNA-seq datasets from projection-defined subpopulations targeting five distinct brain regions in adult mice. This comprehensive approach enabled us to establish developmental transcriptomic profiles for projection-defined serotonin neurons, uncovering dynamic gene expression changes over time. Through this analysis, we identified candidate genes with specificity for both temporal and connectivity attributes, particularly those implicated in axon guidance. Concurrently, we are generating comprehensive maps of whole-brain serotonin axon projections across the same developmental time window using tissue clearing and light-sheet imaging. This approach provides a 3D view of serotonergic axon innervation at the whole-brain level, facilitating direct comparisons between anatomical and molecular progression. Furthermore, we are laying the groundwork for CRISPR-Cas9 knockdown experiments targeting selected genes, followed by light-sheet imaging of axon projections in target regions to validate their functional roles in vivo. Integrating these strategies will enable us to elucidate the developmental projection dynamics of serotonin neuron subpopulations and identify the molecular components that regulate these processes across critical developmental stages.
Neuroscience, Development, Serotonin, Transcriptomics
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