Fig. 3: In vivo calcium imaging of the gastric myenteric plexus in response to VNS.
a Steps for the gastric emptying rate test in C57BL/6J mice. b VNS significantly increases the rate of gastric emptying. Control 1 refers to the group of mice without implanted electrodes and Control 2 refers to the group of mice implanted with electrodes but no VNS (n = 5 mice per group). Control 1 vs Control 2, p = 0.6262; Control 1 vs 20 Hz, p = 0.0009; Control 1 vs 30 Hz, p < 0.0001; Control 2 vs 20 Hz, p = 0.0008; Control 2 vs 30 Hz, p < 0.0001; 20 Hz vs 30 Hz, p = 0.0299. ns: p > 0.05, *p < 0.05, ***p < 0.001, ****p < 0.0001, unpaired student t-test (two-tailed). c Cartoon depiction of a strategy for tracing the vagus nerves from the stomach. d Native fluorescence whole-map analysis of left- and right- ganglion and vagus nerves (white arrow) after gastric infection with CTB-555 in C57BL/6J mice. All scale bars are 200 μm. e Schematic of in vivo imaging of mouse stomach. The blue box shows a detail of electrode implantation for left VNS and the purple box shows a detailed view of the gastric window during imaging. f Frames were captured from in vivo confocal imaging of myenteric neurons expressing GCaMP6s during 20 Hz VNS. All scale bars are 50 μm. g Changes in calcium intensity from (f) ganglia and cells (white arrow) in response to 20 Hz VNS (red arrow, 3 times). The longitudinal coordinates of the curves for ganglia and cells 1, 2, and 3 are ΔF/F0 = 100% and cells 4, 5, and 6 are ΔF/F0 = 300%. h Myenteric ganglia displacement (blue, longitudinal muscle axis; purple, circular muscle axis) to 20 Hz VNS (3 times). i Tissue movement to 20 Hz VNS of the stomach. j Calcium intensity traces of responsive myenteric ganglia (dark red) and resulting tissue movement (dark blue) to 20 Hz VNS (red arrow) of the stomach. All data are mean \(\pm\) \({\mbox{s.e.m}}\).
