Fig. 1: Gs-GPCR mobilization of intracellular Ca²⁺ fully depends on active Gq.

In all HEK293 lines, calcium signals were recorded following a two-step addition protocol. This is exemplified in a for the β₂AR. At t = 20 s, either solvent (a_i) or Gq stimulus ATP 100 µM (a_ii–a_iv) was added, followed by a second addition at t = 140 s of either Iso or Calcium ionophore A23187. a_iv Cells were pretreated with 1 µM of the Gq inhibitor FR. b–d Concentration-effect curves derived from the maximum calcium response of the second addition of b Iso on β₂AR, c PGE₁ on prostanoid EP₂ and EP₄, d NECA on A_2A and A_2B receptors, or A23187 (5 µM) after prior addition of solvent (no priming), ATP (100 µM) or CCh (100 µM). To exclude the contribution of endogenous Gi/o-coupled prostanoid and adenosine receptors to Gs-Ca²⁺, cells were pretreated overnight (16 h) with 100 ng/ml of the Gi/o inhibitor pertussis toxin (PTX). Representative traces are means + SEM, averaged data are mean ± SEM of n biologically independent experiments (b: CCh and solvent n = 3, ATP n = 7; c: n = 3; d: n = 3), each performed in duplicate. Source data are provided as a Source Data file.