(A–B) schemes illustrating the method used to study the coupling properties of mGlu heterodimers composed of mGlu2 (group-II) and a group-III subunit with the wild-type C-terminal tails. In (A), activating specifically the mGlu2 subunit unable to activate G protein (F756S, red cross) can generate a signal only if associated with a functional group-III subunit. (B) Same as in (A) with the inactive group-III subunit (F781S, F773S, F784S, F777S in mGlu4-6-7-8, respectively) and a specific group-III agonist. (C–F) functional coupling of the indicated subunits under the condition indicated on the top (black, control; green, group-II agonist (DCG-IV, 30 μM); blue, group-II agonist with mGlu2 NAM (DCG-IV, 30 μM and MNI137 10 μM); red, group-III agonist (L-AP4, 30 μM for mGlu4-6-8, LSP4-2022, 300 μM for mGlu7). (C) Data obtained with cells expressing both mGlu2 and mGlu4, with either the inactive mGlu2 (2X) or the inactive mGlu4 (4X). (D, E and F), same as in C using mGlu6, mGlu7 or mGlu8 constructs, respectively. Data are means ±SEM (n ≥ 3). **p<0.01, ***p<0.001 (unpaired t test).