Evidence for the presence of functional protease activated receptor 4 (PAR4) in the rat colon.
OBJECTIVE
Protease activated receptors (PARs) have been postulated to play a role during intestinal inflammation. The presence and role played by PAR(4) in gastrointestinal functions have not been fully clarified. The aims of this study were: (i) to examine expression of PAR(4) in rat proximal colon; (ii) to determine the mechanical effects induced by PAR(4) activation in longitudinal muscle; and (iii) to characterise the underlying mechanisms.


METHODS
PAR(4) expression was determined by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. Mechanical activity was recorded as changes in isometric tension.


RESULTS
A PCR product corresponding to the predicted size of the PAR(4) signal was amplified from RNA prepared from the colon of rats, showing the presence of PAR(4) in those tissues. Immunohistochemistry revealed that PAR(4) protein was expressed on epithelial surfaces and submucosa. PAR(4) activating peptides, GYPGKF-NH(2) and AYPGKG-NH(2), produced concentration dependent contractile effects on longitudinal muscle. Tetrodotoxin (TTX) or atropine significantly reduced the contractile responses to AYPGKG-NH(2), and atropine after TTX did not cause any further reduction. NK(1) receptor antagonist, SR140333, or NK(2) receptor antagonist, SR48968, alone or in combination, produced a reduction in PAR(4) induced contractile effect, and when coadministered with TTX abolished it. Capsaicin markedly reduced the contractions evoked by AYPGKG-NH(2).


CONCLUSIONS
The present results suggest that PAR(4) is functionally expressed in rat colon and its activation induces contraction of the longitudinal muscle both through TTX sensitive release of acetylcholine and release of tachykinins, probably from sensory nerves. These actions may contribute to motility disturbances during intestinal trauma and inflammation.
Mulè
F
F
Dipartimento di Biologia cellulare e dello Sviluppo, Laboratorio di Fisiologia Generale, Università di Palermo, Viale delle Scienze, 90128 Palermo, Italy. fmule@unipa.it
Pizzuti
R
R
Capparelli
A
A
Vergnolle
N
N
D000818:Animals; D001285:Atropine / Q000494:pharmacology; D001549:Benzamides / Q000494:pharmacology; D002211:Capsaicin / Q000494:pharmacology; D003092:Colitis / Q000378:metabolism* / Q000503:physiopathology; D003106:Colon / Q000737:chemistry* / Q000503:physiopathology; D004305:Dose-Response Relationship, Drug; D005769:Gastrointestinal Motility; D007150:Immunohistochemistry / Q000379:methods; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000503:physiopathology; D064729:Neurokinin-1 Receptor Antagonists; D009842:Oligopeptides / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D011812:Quinuclidines / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D018041:Receptors, Neurokinin-2 / Q000037:antagonists & inhibitors; D018179:Receptors, Thrombin / Q000032:analysis* / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013779:Tetrodotoxin / Q000494:pharmacology
C482003:AYPGKG-NH(2); D001549:Benzamides; C482002:GYPGKF-NH(2); D064729:Neurokinin-1 Receptor Antagonists; D009842:Oligopeptides; D010880:Piperidines; D011812:Quinuclidines; D012333:RNA, Messenger; D018041:Receptors, Neurokinin-2; D018179:Receptors, Thrombin; C085215:SR 140333; D013779:Tetrodotoxin; C073839:SR 48968; D001285:Atropine; C112908:protease-activated receptor 4; D002211:Capsaicin