Extended Data Fig. 7: Spatial patterning of NTS responses to gut and upper airway stimuli.
a, Representative two-photon image of H2B-jRGECO1a fluorescence in the NTS (transverse view), with neurons color-coded based on their relative peak response amplitudes above threshold to leg pinch (red) and stomach stretch (green, successive 150, 300, 600, and 900 μl distensions), scale bar: 100 μm. b, Positions of neurons selectively responsive to stimuli indicated are charted as in Fig. 3b, with analysis here involving 9 randomly selected mice from Fig. 3b to equalize sample size, oral/stomach: 925 neurons, larynx/stomach: 1128 neurons, duodenum/stomach: 2347 neurons, jejunum/stomach: 1153 neurons, cecum/stomach: 415 neurons (same data as Fig. 3b), color scale depicts neuron density (see methods), axis length: 150 μm, R: rostral, M: medial. c, Representative two-photon images at various depths of H2B-jRGECO1a fluorescence in the NTS (transverse view), with neurons color-coded based on their relative peak response amplitudes above threshold to stomach stretch (green, successive 150, 300, 600, and 900 μl distensions) and duodenum stretch (red, successive 90, 115, or 140 μl distensions), scale bar: 100 μm. d, Positions of neurons at various depths selectively responsive to stimuli as described in c are charted with axis origin corresponding to the centroid for stomach-stretch responsive neurons, 0 μm: 455 neurons, 17 mice, 80 μm: 2517 neurons, 46 mice, 160 μm: 4909 neurons, 53 mice, 240 μm: 4445 neurons, 52 mice, 320 μm: 2204 neurons, 39 mice, color scale depicts neuron density, axis length: 150 μm, R: rostral, M: medial. e, Quantifying spatial segregation of neurons in d, mean ± sem, see methods for Segregation Index. f, Pairwise distances between neurons responsive to the same stimulus (colored) or different stimuli (black). Data from Fig. 3b (real) were compared with data from a simulation where neuron responses were randomly assigned based on the observed response frequency in each field of view (shuffled) to control for regional variation in neuron density, oral/stomach: 1204 neurons, 11 mice, larynx/stomach: 10610 neurons, 57 mice, duodenum/stomach: 28556 neurons, 107 mice, jejunum/stomach: 13050 neurons, 66 mice, cecum/stomach: 415 neurons, 9 mice, mean ± sem, ####P < 0.0001, significant interaction in two-way analysis of variance between responder types and shuffling, ****P < 0.0001, Šídák multiple comparisons test. g, Cartoon depicting sites of balloon distension in the gastrointestinal tract. h, Positions of neurons responsive to stimuli indicated are charted relative to the centroid (coordinate origin) of neurons responsive to distention of stomach site 2, left: 1155 neurons, 4 mice, right: 927 neurons, 4 mice, axis length: 150 μm, R: rostral, M: medial. i, Quantifying spatial segregation of neurons in h responsive to stimuli indicated, mean ± sem, ****P < 0.0001, two-tailed Mann-Whitney test, see methods for Segregation Index.
