Intestinal type 2 proteinase-activated receptors: Expression in opioid-sensitive secretomotor neural circuits that mediate epithelial ion transport

Trypsin and mast cell tryptase cleave within the extracellular N terminus o f proteinase-activated receptor-2 (PAR-2), exposing a tethered ligand (SLIG RL) that binds and activates the cleaved receptor. We examined the neuronal expression of PAR-2 and its role in intestinal ion transport. Short-circui t current elevations in response to trypsin or the receptor-activating pept ide SLIGRL-NH2 were measured in sheets of mucosa-submucosa from porcine ile um. SLIGRL-NH2 or trypsin rapidly elevated short-circuit current after thei r contraluminal application with respective 50% effective concentrations of 184 and 769 nM. Their actions were attenuated after contraluminal administ ration of the neuronal conduction blocker saxitoxin (0.1 mu M); the cycloox ygenase inhibitor indomethacin (10 mu M); or the Na+/K+ /Cl- cotransport in hibitor furosemide (10 mu M), but not by atropine (0.1 mu M), a muscarinic cholinergic antagonist. In addition, soybean trypsin inhibitor (5 mu g/ml) reduced mucosal responses to trypsin. The delta-opioid agonist [D-Pen (2,5) ]-enkephalin (0.1 mu M) inhibited trypsin action, an effect that was preven ted by naltrindole (0.1 mu M), a delta-opioid antagonist. PAR-2 immunofluor escence was localized in the mucosa using a receptor-specific antibody. PAR -2-like immunoreactivity was detected in myenteric and submucosal neurons, nerve fibers innervating ileal smooth muscle and mucosa, and in enteroendoc rine cells. Some neurons coexpressed PAR-2- and choline acetyltransferase-l ike immunoreactivity. These results indicate that PAR-2 is expressed on cho linergic and noncholinergic submucosal neurons in porcine ileum. PAR-2 agon ists stimulate active anion secretion by a neurogenic mechanism that is mod ulated by prostanoids and opioids. These receptors may have a potentially i mportant role in intestinal neuroimmunomodulation.