Hundreds of G-protein coupled receptors (GPCR) are encoded in the human genome. All GPCRs react to a vast variety of ligands and initiate the G-protein activation cycle, by catalyzing the exchange of GDP by GTP on the Gα subunit. Classically, this mode of activation has been proposed to be of catalytic collision coupling nature, where a single receptor sequentially activates several G-proteins. However, recent biophysical and imaging studies challenged this concept and suggested that some GPCR and G-proteins form stable non-dissociating complexes prior to and after activation. We were interested in determining the mode of coupling between the human muscarinic 2 receptor (m2R) and Gαi3βγ. We used the G-protein activated K+ channel (GIRK) as a reporter for receptor activation and systematically quantified receptor's and channel's plasma membrane concentrations using fluorescent methods and radioligand assays. We found a decrease in activation time at high receptor density, with no change in channel concentration. However, maximal amplitude was attained at lower receptor density, suggesting an amplification process. No change in Gβγ concentration was observed, as judged by the unchanged Gβγ-dependent basal activity of GIRK. Additionally, increasing amounts of m2R did not increase Gαi concentration. Together, these results suggest a catalytic collision coupling mechanism. We constructed a model describing m2R's activation scheme and predicted that excessive Gα subunits should slow the activation process by occupying the activated receptor in “dead-end” interactions, not leading to channel activation. Increasing amounts of two fluorescent Gαi3 subunits were used to test the prediction. Indeed, both subunits slowed the evoked-current, without change in current amplitude. These results, together with our previous observations, suggest that the m2R activates Gαi3 via a catalytic collision coupling mechanism, where one receptor diffuses and activates several Gαβγ subunits, leading to the activation of GIRK.