TY - JOUR
T1 - Expression of protease-activated receptors (PARs) in OLN-93 oligodendroglial cells and mechanism of PAR-1-induced calcium signaling
AU - Wang, Yingfei
AU - Richter-Landsberg, C.
AU - Reiser, G.
N1 - Funding Information:
The authors thank Dr. Theo Hanck for the help with DNA sequencing; Dr. Gregor Zündorf, Dr. Rolf Stricker and Mohan Tulapurkar for the help with confocal laser scanning microscopy. This work was supported by grants from the Deutsche Forschungsgemeinschaft (Graduiertenkolleg für ‘Biologische Grundlagen von Erkrankungen des Nervensystems’). Land Sachsen-Anhalt (grant 2923A/0028H), Bundesministerium für Bildung und Forschung (01-ZZ 9505) and Fonds der chemischen Industrie.
PY - 2004
Y1 - 2004
N2 - Protease-activated receptors (PARs) are a group of four members of the superfamily of G protein-coupled receptors that transduce cell signaling by proteolytic activity of extracellular serine proteases, such as thrombin. Possible expression and functions of PARs in oligodendrocytes, the myelin forming cells of the CNS, are still unclear. Here, the oligodendrocyte cell line OLN-93 was used to investigate the signaling of PARs. By reverse transcription-polymerase chain reaction (RT-PCR), immunostaining and Ca 2+ imaging studies, we demonstrate that OLN-93 cells functionally express PAR-1. PAR-3 seems to be expressed without apparent activity, and PAR-2 and PAR-4 cannot be detected. Short-term stimulation of the OLN-93 cells with PAR-1 agonists, such as thrombin, trypsin and PAR-1 activating peptide, dose-dependently induced a transient rise of [Ca2+]i. Concentration-effect curves display a sigmoidal concentration dependence. Elevation of [Ca2+]i induced by PAR-1 mainly resulted from Ca2+ release from intracellular stores. Studies on the effects of pertussis toxin (PTX), phospholipase C antagonist and 2-APB, showed that in OLN-93 cells (i) the calcium signaling cascade from PAR-1 was mediated through PTX-insensitive G proteins, (ii) activation of phospholipase C and liberation of InsP3 were events upstream of the Ca2+ release from the stores. In addition, the present study analyzed PAR-1 desensitization caused by exposure to thrombin, trypsin, and PAR-1 activating peptide, elucidated the influence of the protease cathepsin G on PAR-1 activation, and also characterized PAR-1 desensitization. This is the first study, which shows that OLN-93 oligodendrocytes functionally express PAR-1, and identifies the receptor coupling to mobilization of intracellular calcium. Moreover, the expression of PAR-1 was demonstrated by RT-PCR in primary oligodendrocytes from rat brain.
AB - Protease-activated receptors (PARs) are a group of four members of the superfamily of G protein-coupled receptors that transduce cell signaling by proteolytic activity of extracellular serine proteases, such as thrombin. Possible expression and functions of PARs in oligodendrocytes, the myelin forming cells of the CNS, are still unclear. Here, the oligodendrocyte cell line OLN-93 was used to investigate the signaling of PARs. By reverse transcription-polymerase chain reaction (RT-PCR), immunostaining and Ca 2+ imaging studies, we demonstrate that OLN-93 cells functionally express PAR-1. PAR-3 seems to be expressed without apparent activity, and PAR-2 and PAR-4 cannot be detected. Short-term stimulation of the OLN-93 cells with PAR-1 agonists, such as thrombin, trypsin and PAR-1 activating peptide, dose-dependently induced a transient rise of [Ca2+]i. Concentration-effect curves display a sigmoidal concentration dependence. Elevation of [Ca2+]i induced by PAR-1 mainly resulted from Ca2+ release from intracellular stores. Studies on the effects of pertussis toxin (PTX), phospholipase C antagonist and 2-APB, showed that in OLN-93 cells (i) the calcium signaling cascade from PAR-1 was mediated through PTX-insensitive G proteins, (ii) activation of phospholipase C and liberation of InsP3 were events upstream of the Ca2+ release from the stores. In addition, the present study analyzed PAR-1 desensitization caused by exposure to thrombin, trypsin, and PAR-1 activating peptide, elucidated the influence of the protease cathepsin G on PAR-1 activation, and also characterized PAR-1 desensitization. This is the first study, which shows that OLN-93 oligodendrocytes functionally express PAR-1, and identifies the receptor coupling to mobilization of intracellular calcium. Moreover, the expression of PAR-1 was demonstrated by RT-PCR in primary oligodendrocytes from rat brain.
KW - 2′,3′-cyclic nucleotide 3′-phosphodiesterase
KW - AP
KW - CNP
KW - DMEM
KW - Dulbecco's Modified Eagle medium
KW - GAPDH
KW - activating peptide
KW - base pair
KW - bp
KW - calcium signaling
KW - cathepsin G
KW - protease-activated receptors
KW - thrombin
KW - trypsin
UR - http://www.scopus.com/inward/record.url?scp=2342450576&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=2342450576&partnerID=8YFLogxK
U2 - 10.1016/j.neuroscience.2004.03.024
DO - 10.1016/j.neuroscience.2004.03.024
M3 - Article
C2 - 15145074
AN - SCOPUS:2342450576
SN - 0306-4522
VL - 126
SP - 69
EP - 82
JO - Neuroscience
JF - Neuroscience
IS - 1
ER -