Pharmacological characterization of adenosine receptors in PGT-β mouse pineal gland tumour cells

1 The adenosine receptor in mouse pinealocytes was identified and characterized using pharmacological and physiological approaches, 2 Expression of the two adenosine receptor subtypes A(2B) and A(3) was detected in mouse pineal glands and PGT-beta cells by polymerase chain reaction and nucleotide sequencing. 3 Adenosine and 5'-N-ethylcarboxamidoadenosine (NECA) evoked cyclic AMP generation but the A(2A)-selective agonist 2-(4-(2-carboxyethyl)phenylethylamino)adenosine-5'-N-ethylcarboxamideadenosine (CGS 21680) and the A(1)-specific agonists R-N-6-(2-phenylisopropyl)adenosine (R-PIA) and N-6-cyclopentyladenosine (CPA) had little effect on intracellular cyclic AMP levels. The A(2B) receptor selective antagonists alloxazine and enprofylline completely blocked NECA-mediated cyclic AMP accumulation. 4 Treatment of cells with the A(3)-selective agonist N-6-(3-iodobenzyl)-5'-(N-methylcarbamo,I)adenosine (IB-MECA) inhibited the elevation of the cyclic AMP level induced by NECA or isoproterenol in a concentration-dependent manner with maximal inhibition of 40-50%. These responses were blocked by the specific A3 adenosine receptor antagonist MRS 1191. Pretreatment of the cells with pertussis toxin attenuated the IB-MECA-induced responses, suggesting that this effect occurred ria the pertussis toxin-sensitive inhibitory G proteins. 5 IB-MECA also caused a concentration-dependent elevation in [Ca2+](i) and IP3 content. Both the responses induced by IB-MECA were attenuated by treatment with U73122 or phorbol 12-myristate 13-acetate. 6 These data suggest the presence of both A(2B) and A(3) adenosine receptors in mouse pineal tumour cells and that the A(2B) receptor is positively coupled to adenylyl cyclase whereas the A(3) receptor is negatively coupled to adenylyl cyclase and also coupled to phospholipase C.