Unknown

COS-7 cells were transiently transfected with the human GPR119 receptor. Transfection of the COS-7 cells was performed by the calcium phosphate precipitation method . The transiently transfected cells were seeded at 2.5 x 10^5 cells/well in 12-well plates and incubated for 24 h with 2mkCi/ml of 3H-adenine in 0.5 ml growth medium per well. Cells were washed twice in buffer of 25 mM HEPES (pH 7.2), supplemented with 0.75 mM NaH2PO4, 140 mM NaCl, and 0.05% (wt/vol) BSA, and 0.5 ml buffer of 25 mM HEPES (pH 7.2), supplemented with 0.75 mM NaH2PO4, 140 mM NaCl, and 0.05% (wt/vol) BSA supplemented with 1mMof the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine was added together with increasing concentrations of the different ligands. After 25 min incubation at 37C (310 K), cells were placed on ice, the medium was removed, and the cells were lysed in 1 ml of 5% (wt/vol) trichloroacetic acid, supplemented with 0.1 mM cAMP and 0.1 mM ATP for 30 min