Characterization of adenosine A 1 receptor in cultured myoblast C 2C 12 cells of mice

Abstract

In an attempt to investigate the presence of adenosine A 1 receptor in cell line, we used N 6-cyclopentyladenosine (CPA), an agonist of adenosine A 1 receptor, to incubate with C 2C 12 cells in vitro. CPA increased the uptake of radioactive glucose into C 2C 12 cells in a concentration-dependent manner and this action was abolished by the antagonists, both 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) (1,3-dipropyl-8-cyclopentylxanthine) and 8-( p-sulfophenyl)theophylline (8-SPT), at concentrations sufficient to block adenosine A 1 receptor. Northern blot analysis showed the expression of adenosine A 1 receptor mRNA by C 2C 12 cells. Western blotting also indicated a positive correlation ( r=0.99) of antibody recognized adenosine A 1 receptor with membrane protein. The presence of adenosine A 1 receptor in C 2C 12 cells can thus be considered. In the presence of U73312 (1-[6[[(17β)-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione), the specific inhibitor of phospholipase C, glucose uptake stimulated by CPA into C 2C 12 cells was reduced concentration-dependently while it was not modified by U73343 (1-[6[[(17β)-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-2,5-pyrrolidinedione), the negative control of U73312. Moreover, chelerythrine and GF 109203X (3-[1-[3-(dimethylamino)propyl]-1H-indol-3-yl]-4-(1H-indol-3-yl)-1H-pyrrole-2,5-dione) also diminished the CPA-stimulated glucose uptake at concentrations sufficient to inhibit protein kinase C. The obtained data suggest that activation of adenosine A 1 receptor in C 2C 12 cells may increase the glucose uptake via phospholipase C-protein kinase C pathway.