Activation of intrinsic afferent pathways in submucosal ganglia of the guinea pig small intestine

The enteric nervous system contains intrinsic primary afferent neurons that allow mucosal stimulation to initiate reflexes without CNS input. We tested the hypothesis that submucosal primary afferent neurons are activated by 5-hydroxytryptamine (5-HT) released from the stimulated mucosa. Fast and/or slow EPSPs were recorded in submucosal neurons after the delivery of exogenous 5-HT, WAY100325 (a 5-HT1P agonist), mechanical, or electrical stimuli to the mucosa of myenteric plexus-free preparations (+/- extrinsic denervation). These events were responses of second-order cells to transmitters released by excited primary afferent neurons. After all stimuli, fast and slow EPSPs were abolished by a 5-HT1P antagonist, N-acetyl-5-hydroxytryptophyl-5-hydroxytryptophanamide, and by 1.0 mu M tropisetron, but not by 5-HT4-selective antagonists (SB204070 and GR113808A) or 5-HT3-selective antagonists (ondansetron and 0.3 mu M tropisetron). Fast EPSPs in second-order neurons were blocked by hexamethonium, and most slow EPSPs were blocked by an antagonist of human calcitonin gene-related peptide (hCGRP(8-37)). hCGRP(8-37) also inhibited the spread of excitation in the submucosal plexus, assessed by measuring the uptake of FM2-10 and induction of c-fos. In summary, data are consistent with the hypothesis that 5-HT from enterochromaffin cells in response to mucosal stimuli initiates reflexes by stimulating 5-HT1P receptors on submucosal primary afferent neurons. Second-order neurons respond to these cholinergic/CGRP-containing cells with nicotinic fast EPSPs and/or CGRP-mediated slow EPSPs. Slow EPSPs are necessary for excitation to spread within the submucosal plexus. Because some second-order neurons contain also CGRP, primary afferent neurons may be multifunctional and also serve as interneurons.