Signals generated by free fatty acid receptor 2 (FFA2R) can activate the neutrophil NADPH-oxidase without involvement of any orthosteric FFA2R agonist. The initiating signals may be generated by P2Y 2 R, the receptor for ATP. An FFA2R specific allosteric modulator (PAM; Cmp58) was required for this response and used to investigate the mechanism by which signals generated by ATP/P2Y 2 R activate an FFA2R dependent process. The P2Y 2 R induced signal that together with the modulated FFA2R activates neutrophils, was generated downstream of the Gα q containing G protein coupled to P2Y 2 R. A rise in the cytosolic concentration of ionized calcium ([Ca 2+ ] i ) was hypothesized to be the important signal. The hypothesis gained support from the finding that the modulator transferred the neutrophils to a Ca 2+ sensitive state. The rise in [Ca 2+ ] i induced by the Ca 2+ specific ionophore ionomycin, activated the neutrophils provided that an allosteric modulator was bound to FFA2R. The activity of the superoxide generating NADPH-oxidase induced by ionomycin was rapidly terminated and the FFA2Rs could then no longer be activated by the FFA2R agonist propionate or by the signal generated by ATP/P2Y 2 R. The non-responding state of FFA2R was, however, revoked by a cross-activating allosteric FFA2R modulator. The [Ca 2+ ] i mediated activation of neutrophils with their FFA2Rs allosterically modulated, represent a unique regulatory receptor crosstalk mechanism by which the activation potency of a G protein coupled receptor is controlled by a receptor-crosstalk signaling system operating from the cytosolic side of the plasma membrane.