Calcium imaging of the nodose ganglion reveals specific cell responses to specific cytokines

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Neural reflex circuits regulate the progression and resolution of inflammation. In the inflammatory reflex, sensory responses to injury or infection are relayed via afferent vagus nerve fibers to the brainstem that in turn sends efferent neural signals to spleen, which inhibits inflammation. Prior vagus nerve recording techniques demonstrated that information linked to specific cytokines is carried through the afferent arm of the vagus nerve. However, these electrical recordings cannot observe sensory signaling at the single neuron level. Here, using calcium imaging we demonstrate that individual nodose ganglion neurons respond to specific cytokines. To explore the role of the vagus sensory neurons in reporting specific cytokine signals, we utilized Vglut2-gCaMP6 mice expressing green fluorescent protein in response to intracellular Ca2+as a proxy for neuronal activity. Nodose ganglion was isolated from the brainstem, and placed under a Miniscope, leaving intact connections to the peripheral organs. Using this preparation, approximately 69±16.3 cells per recording were monitored in a plane of focus for individual neuronal activity, continuously for over 1 hour. After baseline imaging, two proinflammatory cytokines, TNF and IL-1β, were administered directly on the nerve in 5 min intervals with saline washes in between cytokines. The viability of the neuronal preparation was confirmed using KCl as a positive control. Analysis of the calcium traces for individual cells using the computational toolbox CaImAn revealed that individual sensory neurons either selectively respond to only one cytokine at a time (responding cells TNF: 23.6±10.4; IL-1β: 19.75±15.7) or with a subset of neurons that respond to both (responding cells TNF+IL-1β: 13±10.3). Interestingly, the peak relative fluorescence (dF/F) values of TNF responsive neurons are significantly higher than the neurons responding to both TNF and IL1β (TNF vs. TNF+IL-1β, p < 0.01). These findings suggest that individual sensory neurons in the nodose ganglion encode information about a specific cytokine and be recruited by multiple cytokines.