Kaiser S A, Soliakov L, Harvey S C, Luetje C W, Wonnacott S
Department of Biology and Biochemistry, University of Bath, England, UK.
J Neurochem. 1998 Mar;70(3):1069-76. doi: 10.1046/j.1471-4159.1998.70031069.x.
The presynaptic nicotinic modulation of dopamine release from striatal nerve terminals is well established, but the subtype(s) of neuronal nicotinic acetylcholine receptor (nAChR) underlying this response has not been identified. Recently, alpha-conotoxin-MII has been reported to inhibit potently and selectively the rat alpha3beta2 combination of nAChR subunits. Here we have synthesised the peptide, confirmed its specificity, and examined its effect on the (+/-)-anatoxin-a-evoked release of [3H]dopamine from rat striatal synaptosomes and slices. Alpha-conotoxin-MII (112 nM) completely blocked acetylcholine-evoked currents of alpha3beta2 nAChRs expressed in Xenopus oocytes (IC50 = 8.0 +/- 1.1 nM). Pairwise combinations of other nicotinic subunits were not blocked by 112 nM alpha-conotoxin-MII. On perfused striatal synaptosomes and slices, alpha-conotoxin-MII dose-dependently inhibited [3H]dopamine release evoked by 1 microM (+/-)-anatoxin-a with IC50 values of 24.3 +/- 2.9 and 17.3 +/- 0.1 nM, respectively. The dose-response curve was shifted to the right with increasing agonist concentrations. However, the maximal inhibition of responses achieved by alpha-conotoxin-MII (112 nM) was 44.9 +/- 5.4% for synaptosomes and 25.0 +/- 4.1% for slices, compared with an inhibition by 10 microM mecamylamine of 77.9 +/- 3.7 and 88.0 +/- 2.1%, respectively. These results suggest the presence of presynaptic alpha3beta2-like nAChRs on striatal dopaminergic terminals, but the incomplete block of (+/-)-anatoxin-a-evoked [3H]dopamine release by alpha-conotoxin-MII also supports the participation of nAChRs composed of other subunits. The lower inhibition found in slices is consistent with an additional indirect nicotinic stimulation of dopamine release via an alpha-conotoxin-MII-insensitive nAChR.
纹状体神经末梢多巴胺释放的突触前烟碱调节作用已得到充分证实，但介导该反应的神经元烟碱型乙酰胆碱受体（nAChR）亚型尚未明确。最近，据报道α-芋螺毒素-MII能有效且选择性地抑制大鼠nAChR亚基的α3β2组合。在此，我们合成了该肽段，证实了其特异性，并研究了其对（±）-anatoxin-a诱发的大鼠纹状体突触体和脑片中[3H]多巴胺释放的影响。α-芋螺毒素-MII（112 nM）完全阻断了非洲爪蟾卵母细胞中表达的α3β2 nAChRs的乙酰胆碱诱发电流（IC50 = 8.0 ± 1.1 nM）。112 nM的α-芋螺毒素-MII不会阻断其他烟碱亚基的两两组合。在灌注的纹状体突触体和脑片上，α-芋螺毒素-MII剂量依赖性地抑制1 μM（±）-anatoxin-a诱发的[3H]多巴胺释放，IC50值分别为24.3 ± 2.9 nM和17.3 ± 0.1 nM。随着激动剂浓度增加，剂量反应曲线向右移动。然而，与10 μM美加明分别抑制77.9 ± 3.7%和88.0 ± 2.1%相比，α-芋螺毒素-MII（112 nM）对突触体反应的最大抑制率为44.9 ± 5.4%，对脑片的最大抑制率为25.0 ± 4.1%。这些结果表明纹状体多巴胺能终末存在突触前α3β2样nAChRs，但α-芋螺毒素-MII对（±）-anatoxin-a诱发的[3H]多巴胺释放的不完全阻断也支持由其他亚基组成的nAChRs的参与。在脑片中发现的较低抑制率与通过α-芋螺毒素-MII不敏感的nAChR对多巴胺释放的额外间接烟碱刺激一致。
