Spatial mapping of RNA turnover kinetics in the mouse brain

Gene regulation requires coordinated control of RNA synthesis and degradation, yet measuring RNA turnover across intact tissues remains challenging. Here we present spatial NT-seq, a method that combines transgenesis-free metabolic RNA labeling with in situ chemical recoding on spatial transcriptomics platforms to co-map newly synthesized and pre-existing RNAs. Applying spatial NT-seq to the mouse brain reveals pronounced regional heterogeneity in RNA turnover and identifies the dentate gyrus as a spatial hotspot marked by coordinated upregulation of basal RNA synthesis and decay. Moreover, spatial NT-seq uncovers rapid, brain region-specific transcriptional and post-transcriptional responses to electroconvulsive stimulation, a clinically relevant treatment for refractory depression. Finally, we leverage computational modeling to identify sequence features and post-transcriptional regulators that shape transcriptome-wide mRNA stability across spatial and cellular contexts in the mouse brain. Together, this integrated 'in vivo timescope' framework provides a spatially resolved view of RNA turnover kinetics and reveals the regulatory architecture of RNA stability in vivo.