Signaling via A2A adenosine receptor in four PC12 cell clones

Abstract

PC12 cells are genetically labile and so-called wild-type cells comprise multiple subclones. We have examined the A2A adenosine receptor signal transduction pathways in four such clones (denoted clones 1, 19, 21 and 27) of PC12 cells. Adenosine A2A, A2B and A1 receptor mRNAs were detected in all four clones by RT-PCR, whereas no A3 receptor mRNA was found. A2A receptors were quantitated by radioligand binding using the antagonist radioligand [3H]SCH 58261 ([3H]-5-amino-7(2-phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4 triazolo [1,5-c] pyrimidine). The Bmax was highest in clone 1 followed by clones 21, 19 and 27. Whereas the amount of G(i) protein appeared similar in all four clones, the amount of G(s) protein was higher in clones 21 and 27 than in the other two clones. Maximal responses to the non-selective adenosine analogue NECA (5'-N-ethylcarboxamidoadenosine) were similar to those observed with the selective adenosine A2A receptor agonist CGS 21680 (2-[p-(2-carbonylethyl) phenylethylamino]-5'-N-ethylcarboxamidoadenosine), and were approximately equal in clones I and 21, but lower in clone 19 and very low in clone 27. For both compounds EC50 was significantly higher in clone 27 than in clone 1. In both clones the response to NECA could be competitively antagonized by a selective adenosine A2A antagonist, SCH 58261. The present results show that different clones of PC 12 cells differ widely in the cAMP increase induced by adenosine analogues and that this is due to differences in the amount of adenosine A2A receptor, G protein and effector. A large difference in receptor number resulted in differences in potency of an agonist.

Formation of a Ternary Complex among NHERF1, β-Arrestin, and Parathyroid Hormone Receptor

“…It is becoming increasingly clear that the cellular context plays an important role for the integration of many signal transduction processes. Several studies indicate that the extent and also the quality of a receptor response are highly dependent on the cellular composition and stoichiometry of intracellular interaction partners (35)(36)(37)(38). Also, the functional implications of NHERF interaction with the PTHR are complex and probably highly cell type-dependent.…”

Distinct Protein Kinase C Isoforms Mediate Regulation of Vascular Endothelial Growth Factor Expression by A2A Adenosine Receptor Activation and Phorbol Esters in Pheochromocytoma PC12 Cells

“…Research examining the signal transduction cascade linking A 2A AR activation to physiologic responses has typically demonstrated a critical role for the AC-PKA pathway (31,37,38). Therefore, the initial focus of this study was the role of AC and PKA in the A 2A AR-mediated down-regulation of VEGF mRNA.…”

“…stimulation of the A 2A AR is linked to AC activation (31,37,38), the signaling pathway mediating the down-regulation of VEGF mRNA induced by CGS21680, a selective agonist for the A 2A AR, was initially explored using activators and inhibitors of PKA (Fig. 1A).…”

“…Additionally, PC12 cells express A 2A and A 2B adenosine receptors (AR) (31) and have been employed to study AR signal transduction and physiologic activity. This laboratory has shown previously that activation of the A 2A AR in PC12 cells results in a substantial reduction of VEGF, which is observed at both the mRNA and protein levels (32).…”

“…The A 2A AR is typically coupled via the G s protein to the stimulation of adenylyl cyclase (AC) and activation of protein kinase A (PKA) (31,37,38). However, certain effects mediated by the A 2A AR have been linked to protein kinase C (PKC) activation (39 -42).…”