The in vivo specificity of synaptic Gβ and Gγ subunits to the α(2a) adrenergic receptor at CNS synapses

G proteins are major transducers of signals from G-protein coupled receptors (GPCRs). They are made up of α, β, and γ subunits, with 16 Gα, 5 Gβ and 12 Gγ subunits. Though much is known about the specificity of Gα subunits, the specificity of Gβγs activated by a given GPCR and that activate each effector in vivo is not known. Here, we examined the in vivo Gβγ specificity of presynaptic α(2a)-adrenergic receptors (α(2a)ARs) in both adrenergic (auto-α(2a)ARs) and non-adrenergic neurons (hetero-α(2a)ARs) for the first time. With a quantitative MRM proteomic analysis of neuronal Gβ and Gγ subunits, and co-immunoprecipitation of tagged α(2a)ARs from mouse models including transgenic FLAG-α(2a)ARs and knock-in HA-α(2a)ARs, we investigated the in vivo specificity of Gβ and Gγ subunits to auto-α(2a)ARs and hetero-α(2a)ARs activated with epinephrine to understand the role of Gβγ specificity in diverse physiological functions such as anesthetic sparing, and working memory enhancement. We detected Gβ(2), Gγ(2), Gγ(3), and Gγ(4) with activated auto α(2a)ARs, whereas we found Gβ(4) and Gγ(12) preferentially interacted with activated hetero-α(2a)ARs. Further understanding of in vivo Gβγ specificity to various GPCRs offers new insights into the multiplicity of genes for Gβ and Gγ, and the mechanisms underlying GPCR signaling through Gβγ subunits.