Analgesic -Conotoxins CVIE and CVIF Selectively and
Voltage-Dependently Block Recombinant and Native N-Type
Calcium Channels
G. Berecki,
1
L. Motin, A. Haythornthwaite, S. Vink, P. Bansal, R. Drinkwater, C. I. Wang,
M. Moretta, R. J. Lewis, P. F. Alewood, M. J. Christie, and D. J. Adams
2
The Queensland Brain Institute (G.B., L.M., D.J.A.), the School of Biomedical Sciences (G.B., A.H., D.J.A.), and Institute for
Molecular Bioscience (S.V., P.B., C.I.W., R.J.L., P.F.A.), the University of Queensland, Queensland, Australia; Xenome Ltd.,
Indooroopilly, Queensland, Australia (R.D.); and Brain and Mind Research Institute, University of Sydney, New South Wales,
Australia (M.M., M.J.C.)
Received June 24, 2009; accepted October 23, 2009
ABSTRACT
Neuronal (N)-type Ca
2+
channel-selective -conotoxins
have emerged as potential new drugs for the treatment of
chronic pain. In this study, two new -conotoxins, CVIE and
CVIF, were discovered from a Conus catus cDNA library.
Both conopeptides potently displaced
125
I-GVIA binding to
rat brain membranes. In Xenopus laevis oocytes, CVIE and
CVIF potently and selectively inhibited depolarization-
activated Ba
2+
currents through recombinant N-type (1
B-b
/
2
1/
3
) Ca
2+
channels. Recovery from block increased with
membrane hyperpolarization, indicating that CVIE and CVIF
have a higher affinity for channels in the inactivated state.
The link between inactivation and the reversibility of -cono-
toxin action was investigated by creating molecular diversity
in subunits: N-type channels with
2a
subunits almost
completely recovered from CVIE or CVIF block, whereas
those with
3
subunits exhibited weak recovery, suggesting
that reversibility of the -conotoxin block may depend on the
type of -subunit isoform. In rat dorsal root ganglion sensory
neurons, neither peptide had an effect on low-voltage-
activated T-type channels but potently and selectively inhib-
ited high voltage-activated N-type Ca
2+
channels in a volt-
age-dependent manner. In rat spinal cord slices, both
peptides reversibly inhibited excitatory monosynaptic trans-
mission between primary afferents and dorsal horn superfi-
cial lamina neurons. Homology models of CVIE and CVIF
suggest that -conotoxin/voltage-gated Ca
2+
channel inter-
action is dominated by ionic/electrostatic interactions. In the
rat partial sciatic nerve ligation model of neuropathic pain,
CVIE and CVIF (1 nM) significantly reduced allodynic behav-
ior. These N-type Ca
2+
channel-selective -conotoxins are
therefore useful as neurophysiological tools and as potential
therapeutic agents to inhibit nociceptive pain pathways.
Neuronal (N)-type voltage-gated calcium channels (VGCCs)
play important roles in regulating neuronal excitability and
nociceptive transmission and are prominently involved in the
transduction of acute and chronic pain perception (Snutch,
2005; Yasuda and Adams, 2007). These channels represent
important drug targets for the management of chronic and
neuropathic pain and have been investigated in the develop-
ment of new analgesic agents (McGivern, 2006; Schroeder et
al., 2006). A number of structurally related -conopeptides of
the genus Conus (cone snails) selectively inhibit N-type
VGCCs of pain-sensing primary nociceptors (Olivera et al.,
1994). Among these, the -conotoxin MVIIA (ziconotide) still
maintains its orphan drug status as a valuable alternative
intrathecal analgesic for the management of chronic intrac-
table pain, especially in patients refractory to opioids (Klotz,
2006). Other conopeptides, such as CVID, are currently in
clinical trials (McGivern, 2006) and hold promise for the
This work was supported by the National Health and Medical Research
Council Program [Grant 351446]; and a University of Queensland Postdoctoral
Research Fellowship.
1
Current affiliation: Department of Physiology and Pharmacology, Hotch-
kiss Brain Institute, University of Calgary, Calgary, Alberta, Canada.
2
Current affiliation: Health Innovations Research Institute, Royal Mel-
bourne Institute of Technology University, Bundoora, Victoria, Australia.
Article, publication date, and citation information can be found at
http://molpharm.aspetjournals.org.
doi:10.1124/mol.109.058834.
ABBREVIATIONS: VGCC, voltage-gated calcium channel; CVID, -conotoxin CVID; CVIE, -conotoxin CVIE; CVIF, -conotoxin CVIF; DRG,
dorsal root ganglion; EPSC, excitatory postsynaptic current; HP, holding potential; PNL, partial nerve ligation; HPLC, high-performance liquid
chromatography; RP-HPLC, reversed-phase high-performance liquid chromatography; PCR, polymerase chain reaction; ANOVA, analysis of
variance; BAPTA, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; TFA, trifluoroacetic acid; N, neuronal; SNX-331, Y13W derivative of
-conotoxin MVIIC.
0026-895X/10/7702-139–148$25.00
MOLECULAR PHARMACOLOGY Vol. 77, No. 2
Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics 58834/3551986
Mol Pharmacol 77:139–148, 2010 Printed in U.S.A.
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