ɑO‐Conotoxin GeXIVA isomers modulate N‐type calcium (Ca_V2.2) channels and inwardly‐rectifying potassium (GIRK) channels via GABA_B receptor activation

Abstract

αO‐Conotoxin GeXIVA is a 28 amino acid peptide derived from the venom of the marine snail Conus generalis. The presence of four cysteine residues in the structure of GeXIVA allows it to have three different disulfide isomers, that is, the globular, ribbon or bead isomer. All three isomers are active at α9α10 nicotinic acetylcholine receptors, with the bead isomer, GeXIVA[1,2], being the most potent and exhibiting analgesic activity in animal models of neuropathic pain. The original report of GeXIVA activity failed to observe any effect of the isomers on high voltage‐activated (HVA) calcium channel currents in rat dorsal root ganglion (DRG) neurons. In this study, we report, for the first time, the activity of globular GeXIVA[1,3] at G protein‐coupled GABAB receptors (GABABR) inhibiting HVA N‐type calcium (Cav2.2) channels and reducing membrane excitability in mouse DRG neurons. The inhibition of HVA Ba2+ currents and neuroexcitability by GeXIVA[1,3] was partially reversed by the selective GABABR antagonist CGP 55845. In transfected HEK293T cells co‐expressing human GABABR1 and R2 subunits and Cav2.2 channels, both GeXIVA[1,3] and GeXIVA[1,4] inhibited depolarization‐activated Ba2+ currents mediated by Cav2.2 channels, whereas GeXIVA[1,2] had no effect. The effects of three cyclized GeXIVA[1,4] ribbon isomers were also tested, with cGeXIVA GAG being the most potent at human GABABR‐coupled Cav2.2 channels. Interestingly, globular GeXIVA[1,3] also reversibly potentiated inwardly‐rectifying K+ currents mediated by human GIRK1/2 channels co‐expressed with GABABR in HEK293T cells. This study highlights GABABR as a potentially important receptor target for the activity of αO‐conotoxin GeXIVA to mediate analgesia.

Dual Antagonism of α9α10 nAChR and GABAB Receptor-Coupled CaV2.2 Channels by an Analgesic αO-Conotoxin Analogue

“…40 However, the binding site of conotoxins on the GABA B R has not been precisely determined. 15,21 Until recently, the binding site for several α-conotoxins was determined based on docking and mutagenesis studies. 41 Docking calculations predicted that α-conotoxins form close contacts with VFT residues in both B1 and B2 subunits, which comprise a novel GABA B R ligand-binding site.…”

“…20 In addition, the globular and ribbon disulfide isomers of GeXIVA potently inhibit Ca V 2.2 channels via activation of GABA B receptors with a halfmaximal inhibition concentration (IC 50 ) of 3−22 nM. 21 Another αO-conotoxin, GeXXVIIA (Figure 1C,D), is a relatively potent dimer peptide antagonist of α9α10 nAChRs and its monomer is 13 amino acids longer at the N-terminus than GeXIVA. 22 A common characteristic of GeXIVA and GeXXVIIA is that they are difficult to produce in mass quantity due to their long sequences and high number of disulfide bonds, thus limiting their development as a drug.…”

α7- and α9-Containing Nicotinic Acetylcholine Receptors in the Functioning of Immune System and in Pain

“…Several studies have proposed that α-conotoxins Vc1.1, RgIA and αO-conotoxin GeXIVA can exert analgesic effects through the modulation of the N-type VGCC Ca V 2.2 via the stimulation of G protein-coupled γ-aminobutyric acid type B (GABA B ) receptors instead of the inhibition of α9-containing nAChR [ 217 , 218 , 219 , 220 , 221 , 222 , 223 , 224 ]. This is consistent with the observed inhibition of N-type VGCC CaV2.2 by α-conotoxins Vc1.1 and RgIA in DRG neurons of α9 KO mice [ 222 ].…”

Preparation of uniform-sized GeXIVA[1,2]-loaded PLGA microspheres as long-effective release system with high encapsulation efficiency

“…Conotoxin αO-GeXIVA, a 28-amino acid polypeptide found in the marine animal conus in the South country Sea, can achieve an analgesic effect by specifically blocking the α9α10 nAChR receptor, which has attracted widespread attention (Yu et al., 2018 ; Xu et al., 2020 ; Wang et al., 2019 ). Its isomer GeXIVA[1,2] is by far the most active conotoxin (Yousuf et al., 2022 ), showing excellent analgesic effects in many disease models (Vincler & Mcintosh, 2007 ; Alsharari et al., 2020 ; Khan et al., 2002 ). Notably, no addictive or motor side effects were observed during the treatment period.…”