CaV2.2 model system to be tested by NK-1R activation. (A) Flow chart representing the signaling cascade used by SP to modulate N current. (B) Schematic of model to be tested: N channels consist of the pore-forming CaV2.2, which is made up of four homologous domains (I–IV) also referred to as pseudosubunits, α2δ-1, and a CaVβ. Palmitoylated CaVβ2a blocks endogenously liberated free AA from binding to CaV2.2's inhibitory sites after exposure of cells to SP. AA's enhancement site remains available and is shown here in the outer regions of CaV2.2, although the actual location of this site remains uncharacterized. (C) Topological organization of CaV2.2 showing the six transmembrane segments and pore loop (P) of each pseudosubunit. An intracellular linker tethers each pseudosubunit to the subsequent one. CaVβ binds the AID region on the I–II linker at a site (delineated by the dotted box) that overlaps with a binding site for Gβγ. (D) The amino acid deletions in the region proximal to the AID result in Bdel1 and Bdel2 mutant channels. (E) Cross-sectional views from the inner pore region of wt CaV2.2 and the two mutant channels. Sequential amino acid deletions in the IS6-AID segment of Bdel1 and Bdel2 are predicted to reorient CaVβ2a such that the two palmitoyl groups (small white circles) are displaced from their wt positions.
