Evidence against adenosine analogues being agonists at the growth hormone secretagogue receptor

Adenosine and adenosine analogues have been reported to act as agonists or partial agonists at the growth hormone secretagogue receptor 1a (GHSR1a). We have re-examined this question. A concentration-dependent increase in intracellular calcium concentration ([Ca2+]i) was observed in GHSR1a transfected HEK 293-EBNA cells stimulated with adenosine (EC50: 0.2 μM) or 2-chloroadenosine (EC50: 1.1 μM) but also in untransfected HEK 293-EBNA cells stimulated with 2-chloroadenosine (EC50: 0.67 μM) or 5′-N-ethylcarboxamidoadenosine (NECA) (EC50: 0.045 μM). These findings support endogenous expression of adenosine receptors, presumably A2B receptors in HEK 293-EBNA cells. In GHSR1a transfected CHO cells, lacking adenosine receptors, the GHSR1a agonist hGhrelin (EC50: 2.4 nM) increased [Ca2+]i, but no effects of adenosine, 2-chloroadenosine or NECA were detected. An inverse agonist of GHSR1a, [d-Arg-1, d-Phe-5, d-Trp-7, 9, Leu-11] substance P, reduced hGhrelin effects but adenosine, 2-chloroadenosine or 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) did not. NECA increased the [Ca2+]i in co-transfected (GHSR1a and A2B receptor) CHO cells (EC50: 0.053 μM), but no additive or synergistic effects on [Ca2+]i or cAMP formation were observed after stimulation with NECA in the absence or in the presence of hGhrelin. In binding studies on GHSR1a transfected CHO cell membranes, [125I]-hGhrelin binding could be displaced by the GHSR1a agonist MK-0677 (IC50: 0.34 nM), hGhrelin (IC50: 1.5 nM), and the substance P analogue (IC50: 0.64 μM) but not by adenosine or 2-chloroadenosine. We conclude that adenosine and analogues do not act as agonists or partial agonists at the GHSR1a and that cross-talk between the GHSR1a and A2B receptors is limited.