Functional protease-activated receptors in the dorsal motor nucleus of the vagus.
BACKGROUND
Protease-activated receptors (PARs), a family member of G-protein coupled receptors, are present and functionally active in a wide variety of cells. The object of this study was to demonstrate the presence and function of PAR-1 and PAR-2 in the dorsal motor nucleus of the vagus (DMV).


METHODS
DMNV neurons were isolated from neonatal rat brainstems using micro-dissection and enzymatic digestion. Neurons were cultured in Neurobasal medium A containing 2% B27 supplement. Intracellular calcium concentration ([Ca(2+)](i)) was measured using fura-2 based microspectrometry. Expression of PARs was detected by RT-PCR and immunofluorescent staining. KEY RESULT: Thrombin and PAR-1 agonist peptide activate PAR-1 with a maximum change in [Ca(2+)](i) expressed as DeltaF/F0 of 229 +/- 14% and 137 +/- 7%, respectively. Trypsin and PAR-2 agonist peptide activate PAR-2 with a maximum DeltaF/F0 change of 258 +/- 12% and 242 +/- 10%, respectively. Inhibition of phospholipase C (PLC) by U73312 (1 microm) decreased the maximal change in DeltaF/F0 induced by PAR-1 activation from 140 +/- 17% to 21 +/- 3%, while the PAR-2-mediated maximal change in DeltaF/F0 decreased from 185 +/- 21% to 19 +/- 6%. Blockade of IP3 receptor with 2APB inhibited the maximal change in DeltaF/F0 due to PAR-1 and PAR-2 activation by 72 +/- 13% and 71 +/- 20% respectively. PAR-1 immnuoreactivity was present in DMV neurons. Increase in transcripts for PAR-1 and PAR-2 were detected in DMV tissues derived from IBD rats relative to control animals.


CONCLUSIONS
Our results indicate that PAR-1 and PAR-2 are present in the DMV neurons, and their activation leads to increases in intracellular calcium via signal transduction mechanism that involves activation of PLC and the production of IP3.
Wang
H
H
Department of Surgery, The Sixth Affiliated Hospital, Sun Yat-sen University, Guangzhou, China.
Wu
X
X
Li
J-Y
JY
Chai
B-X
BX
Wang
J
J
Mulholland
M W
MW
Zhang
W
W
D000704:Analysis of Variance; D000818:Animals; D001933:Brain Stem / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D044463:Receptor, PAR-1 / Q000235:genetics / Q000378:metabolism*; D044464:Receptor, PAR-2 / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology; D014630:Vagus Nerve / Q000378:metabolism*
D012333:RNA, Messenger; D044463:Receptor, PAR-1; D044464:Receptor, PAR-2; D002118:Calcium