Figure 7.

Loss of CGRP and TH after organ culture. A, B, An example of CGRP (A; green) and TH (B; cyan) immunofluorescence in the same biotinamide labeled (red) control preparation. Numerous varicosities containing TH or CGRP can be seen within the myenteric plexus, with some co-labeling of biotinamide-labeled axons and varicosities apparent with CGRP (yellow; A), but not TH (n = 3). This is consistent with the presence of spinal afferent axons in control preparations. In organ cultured preparations, immunohistochemically detectable CGRP (C; green) and TH (D; cyan) were dramatically reduced, showing degeneration of extrinsic nerve fibers, while viscerofugal nerve cell bodies persisted (biotinamide; red). Additionally, no co-labeling of these markers occurred with biotinamide-labeled axons. Expectedly, faintly CGRP-immunoreactive varicosities and nerve fibers persisted in organ cultured preparations. This is consistent with a population of intrinsic enteric neurons. Together, these data support a viscerofugal origin of the activity recorded from rectal nerve trunks in organ cultured preparations.