In vivo visualization of stochastic expression of clustered protocadherin β

Neuronal individuality is fundamental to the brain’s remarkable capacity for learning and memory, and complex information processing. A major molecular mechanism underlying this individuality is the stochastic expression of clustered protocadherin (cPcdh), which confer a unique cell surface identity to each neuron and mediate self/non-self-recognition. However, in vivo analysis of the spatiotemporal dynamics of cPcdh expression has been hampered by the lack of isoform-specific detection tools. To address this, we developed knock-in reporter mouse lines expressing enhanced fluorescent proteins (tdTomato or GFP) under the control of individual cPcdh isoforms (cPcdh-β3 or -β19). These models enable robust, isoform-specific visualization of cPcdh expression throughout the brain, from development to aging. Our findings reveal that stochastic cPcdh-β expression is cell type-specific, spatially biased, left-right asymmetry, varies across individuals, and dynamically regulated across the lifespan, including in adulthood. Live imaging further reveals that cPcdh-β3 expression changes over time within individual neurons, suggesting a potential role in long-term plasticity. We also provide in vivo evidence for monoallelic cPcdh gene expression. These reporter lines constitute a powerful resource for dissecting the physiological and pathological roles of the cPcdh-based neuronal identity codes, offering novel insights into the molecular basis of neural individuality and circuit function.