Ullrich Nina D, Voets Thomas, Prenen Jean, Vennekens Rudi, Talavera Karel, Droogmans Guy, Nilius Bernd
Laboratorium voor Fysiologie, Department of Physiology, Campus Gasthuisberg, KU Leuven, Herestraat 49, B-3000 Leuven, Belgium.
Cell Calcium. 2005 Mar;37(3):267-78. doi: 10.1016/j.ceca.2004.11.001.
Non-selective cation (NSC) channels activated by intracellular Ca2+ ([Ca2+]i) play an important role in Ca2+ signaling and membrane excitability in many cell types. TRPM4 and TRPM5, two Ca2+-activated cation channels of the TRP superfamily, are potential molecular correlates of NSC channels. We compared the functional properties of mouse TRPM4 and TRPM5 heterologously expressed in HEK 293 cells. Dialyzing cells with different Ca2+ concentrations revealed a difference in Ca2+ sensitivity between TRPM4 and TRPM5, with EC50 values of 20.2+/-4.0 microM and 0.70+/-0.1 microM, respectively. Similarly, TRPM5 activated at lower Ca2+ concentration than TRPM4 when [Ca2+]i was raised by UV uncaging of the Ca2+-cage DMNP-EDTA. Current amplitudes of TRPM4 and TRPM5 were not correlated to the rate of changes in [Ca2+]i. The Ca2+ sensitivity of both channels was strongly reduced in inside-out patches, resulting in approximately 10-30 times higher EC50 values than under whole-cell conditions. Currents through TRPM4 and TRPM5 deactivated at negative and activated at positive potentials with similar kinetics. Both channels were equally sensitive to block by intracellular spermine. TRPM4 displayed a 10-fold higher affinity for block by flufenamic acid. Importantly, ATP4- blocked TRPM4 with high affinity (IC50 of 0.8+/-0.1 microM), whereas TRPM5 is insensitive to ATP4- at concentrations up to 1 mM.
由细胞内钙离子（[Ca2+]i）激活的非选择性阳离子（NSC）通道在许多细胞类型的Ca2+信号传导和膜兴奋性中起着重要作用。TRPM4和TRPM5是瞬时受体电位（TRP）超家族的两个Ca2+激活阳离子通道，是NSC通道的潜在分子关联物。我们比较了在HEK 293细胞中异源表达的小鼠TRPM4和TRPM5的功能特性。用不同Ca2+浓度透析细胞揭示了TRPM4和TRPM5之间Ca2+敏感性的差异，其半数有效浓度（EC50）值分别为20.2±4.0微摩尔和0.70±0.1微摩尔。同样，当通过Ca2+笼合物DMNP-EDTA的紫外线解笼提高[Ca2+]i时，TRPM5在比TRPM4更低的Ca2+浓度下被激活。TRPM4和TRPM5的电流幅度与[Ca2+]i的变化速率无关。在内外向外膜片钳记录中，两个通道的Ca2+敏感性都大大降低，导致EC50值比全细胞条件下高约10 - 30倍。通过TRPM4和TRPM5的电流在负电位时失活，在正电位时以相似的动力学激活。两个通道对细胞内精胺的阻断同样敏感。TRPM4对氟芬那酸阻断的亲和力高10倍。重要的是，ATP4-以高亲和力阻断TRPM4（IC50为0.8±0.1微摩尔），而TRPM5在浓度高达1毫摩尔时对ATP4-不敏感。
