Electrophysiology, shape, and chemistry of neurons that project from guinea pig colon to inferior mesenteric ganglia  Keith A. Sharkey, Alan E.G. Lomax, Paul P. Bertrand, John B. Furness  Gastroenterology  Volume 115, Issue 4, Pages 909-918 (October 1998) DOI: 10.1016/S0016-5085(98)70263-X Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 1 A composite map showing the positions of the cells from which recordings were made. Records were taken from retrogradely labeled nerve cells both close to the mesenteric attachment and toward the antimesenteric region. In some cases, the immunoreactivities of neurons were determined as indicated. ●, Fast Blue; ■, Fast Blue/NOS; ×, Fast Blue/calbindin; ▴, Fast Blue/ChAT. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 2 Intracellular electrophysiological recordings from retrogradely labeled cells. (A and B) Large-amplitude fast EPSPs in response to interganglionic nerve strand stimulation, typical of the responses elicited in the retrogradely labeled neurons, recorded from two different neurons. (C and D) Examples of spontaneous fast EPSPs from different neurons. In C (record from the same neuron as in A), the effects of several fast EPSPs seem to be summated, whereas single events are seen in D (record from the same neuron as in B). (E) Reduction in EPSP amplitude after hexamethonium was added to the bathing solution. (F) An example of an EPSP that led to an action potential. The action potential does not have a hump on its falling phase. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 3 Action potentials of retrogradely labeled neurons. (A and B) Responses to 500-millisecond intracellular depolarizing pulses in two different neurons. The neurons fire phasically, i.e., only at the beginnings of the depolarizing pulses. (C) The shape of an action potential on an expanded time base. (D) Plot of the differentiated record from C. Zero slope is marked by the horizontal line. Neither the voltage record nor the differentiated record reveals a hump on the falling phase of the action potential. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 4 The same nerve cell, revealed by retrogradely transported Fast Blue, by streptavidin–Texas Red complexed to biocytin that had been injected through the recording electrode, and in a camera lucida drawing made after the biocytin had been converted to a permanent diaminobenzidine deposit. This is a Dogiel type I neuron with short and long dendrites, which in places abut the surface of the ganglion. It responded to interganglionic strand stimulation with a fast EPSP. The outline of the ganglion is marked with dashed lines. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 5 Examples of the shapes of retrogradely labeled cells, revealed by conversion of injected biocytin to a permanent deposit, that were drawn by camera lucida. Each cell had a single axon and an irregular cell body with lamellar dendrites, typical of Dogiel type I neurons. Some cells had only short lamellar dendrites, whereas others had a mixture of short and long dendrites. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 6 Immunoreactivities of retrogradely labeled nerve cells. All images were taken after fixation and processing for immunohistochemistry. Each row of micrographs shows one nerve cell that was filled through the recording electrode with biocytin (that was reacted with Texas Red to reveal the cell; panels marked Biocytin-TR), had been labeled by retrograde transport of Fast Blue (middle panel of each series), and then reacted for immunohistochemistry. (A) This nerve cell was immunoreactive for ChAT. (B) Retrogradely labeled neuron that was calbindin immunoreactive. (C) This neuron was immunoreactive for NOS; it contains a cytoplasmic vacuole caused by the recording electrode. (D) Neuron in a preparation processed for calretinin immunoreactivity. The retrogradely labeled nerve cell is not calretinin immunoreactive. Arrows in C and D indicate positions of the same cells. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 7 Diagram to illustrate possible relationships between intrinsic neurons of the colon and the cell bodies of intestinofugal neurons. The intestinofugal neurons are uniaxonal and project to the IMG. They receive synaptic inputs from other neurons, including intrinsic primary afferent neurons, that have been identified in other studies to have Dogiel type II (DII) morphology. The inputs from intrinsic primary afferent neurons may be direct or indirect, via interneurons; this has not been determined. Gastroenterology 1998 115, 909-918DOI: (10.1016/S0016-5085(98)70263-X) Copyright © 1998 American Gastroenterological Association Terms and Conditions