Modulation of GH4 cell cycle via A1 adenosine receptors

Identification and characterization of A1 adenosine receptors (A1Rs) in a tumor cell line derived from rat pituitary (GH4 cells) was performed by ligand binding and immunocytochemistry. Subsequently, the involvement of A1Rs in the regulation of cell proliferation was studied in these cells. The agonist N6-(R)-phenylisopropyladenosine (R-PIA) did not modify the number of cultured cells, but it regulated the kinetics of the cell cycle. By means of experiments of pulse and of pulse and chase with bromodeoxyuridine and further labeling with Hoechst 33258, propidium iodide, and/or fluorescein-conjugated antibodies against bromodeoxyuridine, it was demonstrated that R-PIA, via A1Rs, accelerated progression from G0/G1 to S phase and from S to G2/M phase of the cell cycle, whereas the initiation of a new cycle occurred at the same time in treated and untreated cells. As a consequence, R-PIA did not change the total length of the cycle. This is the first description of cell cycle regulation without modification of cell proliferation. Although pertussis toxin blocked the R-PIA-induced inhibition of cyclic AMP production in these cells, it did not affect the R-PIA action on the cell cycle. In contrast, cholera toxin mimicked the action of R-PIA. Thus, it is likely that regulation of the cell cycle via A1Rs is mediated by heterotrimeric G proteins different from those that mediate inhibition of adenylate cyclase. Due to the fact that cells in G0/G1 phase were less susceptible to secretory signals, adenosine, in an autocrine manner and by regulating the cell cycle kinetics, may contribute to the modulation of the secretory capacity of pituitary cells.