Protease-activated receptors (PARs) belong to the family of membrane receptors coupled to G-proteins; their presence is reported in a wide variety of cells. The object of this study was to demonstrate the presence of PAR-1 and PAR-2 in myenteric glia of the guinea pig, and to elucidate the cellular mechanisms that are triggered upon receptor activation. Thrombin and PAR-1 agonist peptide (PARP-1) activate PAR-1 with a maximum mean +/- SEM change in intracellular calcium concentration with respect to basal level (Delta[Ca2+]i) of 183 +/- 18 nm and 169 +/- 6 nm, respectively. Trypsin and PAR-2 agonist peptide (PARP-2) activate PAR-2 with a maximum Delta[Ca2+]i of 364 +/- 28 nm and 239 +/- 19 nm, respectively. Inhibition of phospholipase C by U73312 (1 microm) decreased the Delta[Ca2+]i due to PAR-1 activation from 167 +/- 10 nm to 87 +/- 6 nm. The PAR-2-mediated Delta[Ca2+]i decreased from 193 +/- 10 nm to 124 +/- 8 nm when phospholipase C activity was inhibited. Blockade of sphingosine kinase with dimethylsphingosine (1 microm) decreased the Delta[Ca2+]i due to PAR-2 activation from 149 +/- 19 nm to 67 +/- 1 nm, but did not influence the PAR-1-mediated Delta[Ca2+]i. PAR-1 and PAR-2 were localized in myenteric glia by immunolabeling. Our results indicate that PAR-1 and PAR-2 are present in myenteric glia of the guinea pig, and their activation leads to increases in intracellular calcium via different signal transduction mechanisms that involve activation of phospholipase C and sphingosine kinase.