Specificities of Gβγ subunits for the SNARE complex before and after stimulation of α_2a-adrenergic receptors

Ligand binding to G protein-coupled receptors (GPCRs), such as the α_2a-adrenergic receptor (α_2aAR), results in the activation of heterotrimeric G proteins, which consist of functionally distinct Gα subunits and Gβγ dimers. α_2aAR-dependent inhibition of synaptic transmission regulates functions such as spontaneous locomotor activity, anesthetic sparing, and working memory enhancement and requires the soluble NSF attachment protein receptor (SNARE) complex, a Gβγ effector. To understand how the Gβγ-SNARE complex underlies the α_2aAR-dependent inhibition of synaptic transmission, we examined the specificity of Gβγ subunits for the SNARE complex in adrenergic neurons, in which auto-α_2aARs respond to epinephrine released from these neurons, and nonadrenergic neurons, in which hetero-α_2aARs respond to epinephrine released from other neurons. We performed a quantitative, targeted multiple reaction monitoring proteomic analysis of Gβ and Gγ subunits bound to the SNARE complex in synaptosomes from mouse brains. In the absence of stimulation of auto-α_2aARs, Gβ₁ and Gγ₃ interacted with the SNARE complex. However, Gβ₁, Gβ₂, and Gγ₃ were found in the complex when auto-α_2aARs were activated by epinephrine. Further understanding of the specific usage of distinct Gβγ subunits in vivo may provide insights into the homeostatic regulation of synaptic transmission and the mechanisms of dysfunction that occur in neurological diseases.