A molecular architecture of the vagal interoceptive system

Interoception, the ability to timely and precisely sense changes inside the body, is critical for survival. Vagal sensory neurons (VSNs) form an important body-to-brain axis, navigating along body's rostral-caudal axis to their target organs and dive across organ's surface-lumen axis into appropriate tissue layers to sense stimuli with various physical properties. The brain can discriminate numerous body signals through VSNs, yet the underlying molecular coding strategy remains poorly understood. Here we show that VSNs code visceral organ, tissue layer, and stimulus modality, three key features of an interoceptive signal, in different dimensions. Large-scale single-cell profiling of VSNs from seven major organs, including the lung, heart, stomach, esophagus, duodenum, pancreas, and transverse colon, using multiplexed projection-barcodes reveals a 'visceral organ' dimension composed of differentially expressed gene modules coding VSN target organs along body's rostral-caudal axis. Surprisingly, we discover another 'tissue layer' dimension with gene modules coding VSN ending locations along organ's surface-lumen axis. The three independent feature-coding dimensions together specify many parallel VSN pathways in a combinatorial fashion and facilitate complex VSN projection in the brainstem. Our study thus highlights a novel multidimensional coding architecture of the mammalian vagal interoceptive system for effective signal communication. Overall design: For each sample of Projection-seq, 10 age and gender-matched wildtype mice were first injected with different SNAP-seq AAVs into seven thoracic and abdominal organs, including the lung, heart, stomach, esophagus, duodenum, pancreas, and transverse colon. Vagal ganglia were harvested and acutely isolated 7 days later. Single cell libraries were prepared using 10x Genomics Chromium Single Cell 3' Reagent Kit v3 chemistry and sequenced on an Illumina NovaSeq6000 S4 sequencer.