Pakkanen Jukka S, Stenfors Jan, Jokitalo Eija, Tuominen Raimo K
Division of Pharmacology and Toxicology, Faculty of Pharmacy, University of Helsinki, Finland.
Synapse. 2006 Jun 1;59(7):383-93. doi: 10.1002/syn.20249.
Chronic nicotine treatment increases the number of neuronal nicotinic acetylcholine receptors (nAChRs). Localization of nAChRs at a cellular level determines their functional role. However, changes in the localization of nAChRs caused by chronic nicotine treatment are not well known. In this study, we have examined the effects of chronic nicotine treatment on alpha7 and beta2 nAChR subunits in vitro in cell lines and in vivo in mouse striatum. In vitro, two different cell lines were used, SH-SY5Y cells endogenously expressing several nAChR subtypes and SH-EP1-halpha7 cells, transfected with the human alpha7 nAChR subunit gene. Effects of chronic nicotine treatment (10 microM, 3 days) were studied in vitro by using confocal and electron microscopy and calcium fluorometry. In vitro in SH-SY5Y cells, alpha7 and beta2 subunits formed groups, unlike alpha7 subunits in SH-EP1-halpha7 cells, which were partially localized on endoplastic reticulum. Chronic nicotine treatment did not change the localization of nAChRs in endosomes, but caused clustering of alpha7 subunits in SH-EP1-halpha7 cells. In vivo, nicotine was given to mice in their drinking water for 7 weeks. Results showed that alpha7 and beta2 subunits formed groups, and that chronic nicotine treatment increased the size of the clusters. As a conclusion, our data show that there are large intracellular pools of nAChR subunits, which are partially localized on endoplastic reticulum. Chronic nicotine treatment does not change endocytotic trafficking of nAChRs. Chronic nicotine treatment increased clustering of nAChRs, which could have a role in the release of dopamine (DA) evoked by nicotine.
慢性尼古丁处理会增加神经元烟碱型乙酰胆碱受体（nAChRs）的数量。nAChRs在细胞水平上的定位决定了它们的功能作用。然而，慢性尼古丁处理引起的nAChRs定位变化尚不清楚。在本研究中，我们在体外细胞系和体内小鼠纹状体中研究了慢性尼古丁处理对α7和β2 nAChR亚基的影响。在体外，使用了两种不同的细胞系，内源性表达几种nAChR亚型的SH-SY5Y细胞和转染了人α7 nAChR亚基基因的SH-EP1-hα7细胞。通过共聚焦显微镜、电子显微镜和钙荧光测定法研究了慢性尼古丁处理（10 microM，3天）在体外的作用。在体外的SH-SY5Y细胞中，α7和β2亚基形成簇，这与SH-EP1-hα7细胞中的α7亚基不同，后者部分定位于内质网。慢性尼古丁处理并未改变nAChRs在内体中的定位，但导致SH-EP1-hα7细胞中α7亚基聚集。在体内，给小鼠饮用含尼古丁的水7周。结果表明，α7和β2亚基形成簇，并且慢性尼古丁处理增加了簇的大小。总之，我们的数据表明存在大量nAChR亚基的细胞内池，它们部分定位于内质网。慢性尼古丁处理不会改变nAChRs的内吞运输。慢性尼古丁处理增加了nAChRs的聚集，这可能在尼古丁诱发的多巴胺（DA）释放中起作用。
