Mould Jorgen, Yasuda Takahiro, Schroeder Christina I, Beedle Aaron M, Doering Clinton J, Zamponi Gerald W, Adams David J, Lewis Richard J
Institute for Molecular Bioscience, University of Queensland, Queensland 4072, Australia.
J Biol Chem. 2004 Aug 13;279(33):34705-14. doi: 10.1074/jbc.M310848200. Epub 2004 May 27.
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.
来自食鱼芋螺的ω-芋螺毒素是电压门控钙通道的强效抑制剂。ω-芋螺毒素MVIIA和CVID是选择性N型钙通道抑制剂，具有治疗慢性疼痛的潜力。β和α(2)δ-1辅助亚基影响N型通道α(1B)亚基的表达和特性，并且在包括疼痛在内的疾病状态中受到不同程度的调节。在本研究中，我们研究了这些辅助亚基对ω-芋螺毒素GVIA、MVIIA、CVID及其类似物抑制大鼠N型通道外周和中枢形式能力的影响。虽然β3亚基对ω-芋螺毒素的结合和解离速率影响很小，但α(2)δ与α(1B)共表达显著降低了N型通道中枢和外周亚型的结合速率和平衡抑制。α(2)δ还增强了MVIIA对中枢亚型的选择性，但未增强CVID的选择性。在人胚肾细胞中表达的N型通道也观察到了类似但不太明显的趋势。α(2)δ的影响不受卵母细胞去糖基化的影响。从ω-芋螺毒素阻断中恢复的程度，GVIA最小，MVIIA中等，CVID几乎完全恢复。施加超极化钳制电位（-120 mV）并没有显著提高CVID的恢复程度。有趣的是，[R10K]MVIIA和[O10K]GVIA从阻断中恢复得更好，而[K10R]CVID从阻断中恢复得减少，这表明第10位对ω-芋螺毒素的可逆程度有重要影响。在人胚肾细胞和表达α(1B-b)的卵母细胞中，α(2)δ的存在会降低CVID阻断后的恢复。鉴于α(2)δ亚基在某些疼痛状态下上调，这些结果可能对ω-芋螺毒素的抗伤害感受特性有影响。
