The alpha2delta auxiliary subunit reduces affinity of omega-conotoxins for recombinant N-type (Cav2.2) calcium channels.
The omega-conotoxins from fish-hunting cone snails are potent inhibitors of voltage-gated calcium channels. The omega-conotoxins MVIIA and CVID are selective N-type calcium channel inhibitors with potential in the treatment of chronic pain. The beta and alpha(2)delta-1 auxiliary subunits influence the expression and characteristics of the alpha(1B) subunit of N-type channels and are differentially regulated in disease states, including pain. In this study, we examined the influence of these auxiliary subunits on the ability of the omega-conotoxins GVIA, MVIIA, CVID and analogues to inhibit peripheral and central forms of the rat N-type channels. Although the beta3 subunit had little influence on the on- and off-rates of omega-conotoxins, coexpression of alpha(2)delta with alpha(1B) significantly reduced on-rates and equilibrium inhibition at both the central and peripheral isoforms of the N-type channels. The alpha(2)delta also enhanced the selectivity of MVIIA, but not CVID, for the central isoform. Similar but less pronounced trends were also observed for N-type channels expressed in human embryonic kidney cells. The influence of alpha(2)delta was not affected by oocyte deglycosylation. The extent of recovery from the omega-conotoxin block was least for GVIA, intermediate for MVIIA, and almost complete for CVID. Application of a hyperpolarizing holding potential (-120 mV) did not significantly enhance the extent of CVID recovery. Interestingly, [R10K]MVIIA and [O10K]GVIA had greater recovery from the block, whereas [K10R]CVID had reduced recovery from the block, indicating that position 10 had an important influence on the extent of omega-conotoxin reversibility. Recovery from CVID block was reduced in the presence of alpha(2)delta in human embryonic kidney cells and in oocytes expressing alpha(1B-b). These results may have implications for the antinociceptive properties of omega-conotoxins, given that the alpha(2)delta subunit is up-regulated in certain pain states.
Mould
Jorgen
J
Institute for Molecular Bioscience, University of Queensland, Queensland 4072, Australia.
Yasuda
Takahiro
T
Schroeder
Christina I
CI
Beedle
Aaron M
AM
Doering
Clinton J
CJ
Zamponi
Gerald W
GW
Adams
David J
DJ
Lewis
Richard J
RJ
D000595:Amino Acid Sequence; D000818:Animals; D015220:Calcium Channels / Q000737:chemistry*; D020864:Calcium Channels, N-Type / Q000737:chemistry* / Q000378:metabolism; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D005091:Exons; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism; D010146:Pain; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D018075:RNA, Complementary / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D015854:Up-Regulation; D014982:Xenopus laevis; D020960:omega-Conotoxins / Q000737:chemistry* / Q000378:metabolism
D015220:Calcium Channels; D020864:Calcium Channels, N-Type; D010455:Peptides; D018075:RNA, Complementary; D011994:Recombinant Proteins; D020960:omega-Conotoxins