Selective modulation of neuronal nicotinic acetylcholine receptor channel subunits by Go-protein subunits
Selective modulation of neuronal nicotinic acetylcholine receptor channel subunits by Go-protein subunits
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DOI：
10.1523/jneurosci.4971-04.2005
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发表时间：
2005-04-06
影响因子：
5.3
通讯作者：
Adams, DJ
中科院分区：
文献类型：
--
作者：
Fischer, H;Liu, DM;Adams, DJ
采用全细胞透析和切除膜膜片记录的方法，研究了大鼠心脏内源性神经节神经元烟碱乙酰胆碱受体（nAChR）通道的蛋白调节。GTP γ S细胞透析增加了nachr的激动剂亲和力，导致低浓度尼古丁诱发的全细胞电流增强。在GTP γ S存在的情况下，乙酰胆碱和尼古丁诱发的电流振幅增加了约两倍。在内外膜斑块中，相对于对照组，浸泡0.2mM GTP γ S后，nachr介导的单一电流的打开概率（NPo）可逆地增加了四倍，但在GTP γ S存在的情况下没有变化。胆碱能激动剂乙酰胆碱、烟碱和1,1-二甲基-4-苯基哌嗪引起的电流在GTP γ s存在下增强，但不包括胱氨酸或胆碱。G蛋白亚基与nachr之间的直接相互作用通过G(o) α或G β γ亚基在内外膜斑块和谷胱甘肽s转移酶拉下和共免疫沉淀实验进行了研究。50 nM的G β γ使ACh激活的单通道电流的打开概率增加了5倍，而G(o) α (50 nM)没有显著增加NPo。神经元nAChR亚基α 3- α 5和α 2与G(o) α和G β γ表现出积极的相互作用，而β 4和α 7与G蛋白亚基中的任何一个都没有相互作用。这些结果为nAChR和g蛋白亚基之间的直接相互作用提供了证据，揭示了大鼠心内神经节副交感神经元中乙酰胆碱激活的单通道电流的打开概率增加和nAChR介导的全细胞电流的增强。
protein modulation of neuronal nicotinic acetylcholine receptor ( nAChR) channels in rat intrinsic cardiac ganglia was examined using dialyzed whole-cell and excised membrane patch-recording configurations. Cell dialysis with GTP gamma S increased the agonist affinity of nAChRs, resulting in a potentiation of nicotine-evoked whole-cell currents at low concentrations. ACh- and nicotine-evoked current amplitudes were increased approximately twofold in the presence of GTP gamma S. In inside-out membrane patches, the open probability (NPo) of nAChR-mediated unitary currents was reversibly increased fourfold after bath application of 0.2mM GTP gamma S relative to control but was unchanged in the presence of GDP gamma S. The modulation of nAChR-mediated whole- cell currents was agonist specific; currents evoked by the cholinergic agonists ACh, nicotine, and 1,1-dimethyl-4-phenylpiperazinium iodide, but not cytisine or choline, were potentiated in the presence of GTP gamma S. The direct interaction between G-protein subunits and nAChRs was examined by bath application of either G(o)alpha or G beta gamma subunits to inside-out membrane patches and in glutathione S-transferase pull-down and coimmunoprecipitation experiments. Bath application of 50 nM G beta gamma increased the open probability of ACh- activated single-channel currents fivefold, whereas G(o)alpha( 50 nM) produced no significant increase in NPo. Neuronal nAChR subunits alpha 3-alpha 5 and alpha 2 exhibited a positive interaction with G(o)alpha and G beta gamma, whereas beta 4 and alpha 7 failed to interact with either of the G-protein subunits. These results provide evidence for a direct interaction between nAChR and G-protein subunits, underlying the increased open probability of ACh-activated single-channel currents and potentiation of nAChR-mediated whole-cell currents in parasympathetic neurons of rat intrinsic cardiac ganglia.