Figure 1. Correlating calcium activity in identified neurons.

A, colonic preparation used for stimulating the mucosa at the oral and anal cut ends. The colonic wall was opened and pinned in the middle with the longitudinal muscle (LM) uppermost. Strips of LM were peeled away to reveal the myenteric plexus. The colon was also opened at the oral and anal ends and pinned with the mucosa uppermost to allow stimulation of the mucosa with a brush (see symbol). The stimulus was registered by activating a light emitting diode (LED), which was located under the organ bath, at the beginning of the stimulus regime. Recordings were made in the middle of the preparation ∼2.5 cm from the sites of stimulation. B, to locally activate the mucosa, nitrogen was spritzed onto the mucosa via a polyethylene tube with a small hole under the recording site. C, to brush the mucosa directly over the imaging site, the preparation was pinned with the mucosa uppermost over a coverslip and imaged using an inverted microscope. D, following calcium imaging experiments the preparation was stained with an antibody to nitric oxide synthase (NOS) and the ganglia on which imaging was performed located (NOS +ve neurons, closed arrows; NOS −ve neurons, open arrows). The square in the circle locates the ganglion that was imaged in E. E, maximum Ca²⁺ induced fluorescence in myenteric neurons within the ganglion, and later NOS staining of the same ganglion. F, the calcium fluorescence in an individual neuron was converted into a spatio-temporal map (ST map; lower panel). Note that the ST map corresponds to the activity (see line trace) in the neuron. The width of the ST map corresponds to the length of the long axis of the neuron.