ACTIVATION OF MAST-CELL K+ CHANNELS THROUGH MULTIPLE G-PROTEIN-LINKED RECEPTORS

The rat basophilic leukemia (RBL) mast cell line possesses cell surface receptors for adenosine whose ligation markedly potentiates antigen-driven Ca2+ influx and secretion. Here we show that engagement of these receptors and of separate P2 purinergic receptors rapidly activates an outwardly rectifying K+ conductance [G(K(OR))] in RBL cells. Activation of G(K(OR)) by the ligands 5'-(N-ethylcarboxamido)adenosine (NECA), ADP, and ATP was prevented by cytoplasmic guanosine 5'-[beta-thio]diphosphate as well as by pretreatment of the cells with pertussis toxin, implicating mediation by a G protein. Multiple cycles of induction and decay of G(K(OR)) were produced upon application and removal of ligand. Induction of G(K(OR)) by either ligand was much faster than the induction caused by guanosine 5'-[gamma-thio]triphosphate (t1/2 < 10 sec vs. 210 sec). In control cells the maximal whole-cell conductance elicited by ADP (2.25 +/- 0.30 nS) or ATP (2.50 +/- 0.33 nS) was about twice as large as that induced by NECA (1.03 +/- 0. 11 nS), and similar to that previously reported for the guanosine 5'-[gamma-thio]triphosphate-elicited GK(OR) in RBL cells (2.58 +/- 1.59 nS). Treatment of RBL cells with dexamethasone upregulated Ca2+ responses to NECA, and it also nearly doubled the maximal conductance elicited by NECA without appreciable effect on responses to ADP or ATP. The failure of water-soluble second messengers to activate G(K(OR)) and the inability of 11 mM EGTA (< 10 nM Ca2+) to prevent activation by ADP suggest that the relevant pathway is membrane-delimited. Two ion-channel blockers inhibited antigen-stimulated secretion with IC50 values similar to those at which they blocked G(K(OR)), suggesting that activity of the outwardly rectifying K+ channel may be important for stimulus-response coupling in these cells. Potentiation of the secretory response by NECA may reflect, in part, the activation of G(K(OR)), which serves to repolarize the membrane more effectively than does the constitutive mechanism, thereby enhancing antigen-driven Ca2+ influx. This channel and its functionally associated receptors may allow neighboring cells of the host to modulate the response of mast cells to exogenous antigen.