Figure 9. Responses of mitotracker +ve neurons to local stimulation.

High power imaging (×60) was used to record responses in neurons to local stimulation. A, upper panel shows average calcium in neurons and location of selected neurons. Lower panel shows location of neurons stained with mitotracker (filled arrows) in the same ganglia. Following local mucosal stimulation (a puff of nitrogen administered to the mucosa under the recording site) two of the 4 mitotracker +ve neurons (left hand panel; e.g. neurons 1 and 2) responded to local stimulation with a brief Ca²⁺ transient; whereas, two other mitotracker +ve neurons did not respond (see left hand panel, neurons 3 and 4). However, 3 of these 4 neurons (1–3) exhibited a sustained response following anal and oral mucosal stimulation. B, upper panel: shows average calcium in neurons before stimulation, and location of selected neurons; middle panel shows average calcium immediately following stimulation, and lower panel shows location of neurons stained with mitotracker (filled arrows). A single brush stroke applied to the mucosa over the recording site evoked Ca²⁺ transients in both mitotracker +ve neurons (1–4) and in mitotracker −ve neurons (5–8), although neuron 5 failed to respond. Ondansetron (3 μm) significantly reduced the responses in these neurons (1–8) to mucosal stimulation (3 strokes). Apparent changes in activity in CM and displacement are stimulus artifacts. C, left hand panel: average calcium in ganglia and location of selected neurons; right hand panel: location of mitotracker +ve (filled arrows) and mitotracker −ve (open arrows) neurons. In the presence of hexamethonium (100 μm), brushing the mucosa (5 strokes) over the stimulation site evoked a sustained Ca²⁺ response in both mitotracker +ve neurons (1–3) and in mitotracker −ve neurons (4 and 5). Ondansetron (3 μm) significantly reduced the responses to stroking the mucosa in these neurons, leaving mostly the stimulus artifact. In these experiments, probenecid (0.5 mm; anion transport pump inhibitor) was added to the Krebs solution bathing the tissue.