Haass M, Serf C, Gerber S H, Krüger C, Haunstetter A, Vahl C F, Nobiling R, Kübler W
Department of Cardiology, University of Heidelberg, Germany.
J Mol Cell Cardiol. 1997 Jun;29(6):1615-27. doi: 10.1006/jmcc.1997.0398.
It was the aim of the present study (1) to characterize the influence of Na+/K(+)-ATPase inhibition by the digitalis glycoside ouabain on both spontaneous and nicotine-evoked norepinephrine release from the human heart; and (2) to further investigate the role of glycoside-induced changes in [Na+]i and [Ca2+]i (determined by microfluorimetry) for catecholamine release. The latter experiments were performed in bovine adrenal medullary chromaffin cells (BCC), an established cell culture model for sympathetic nerves. Ouabain (1-1000 mumol/l) exerted a dual effect on norepinephrine release (determined by HPLC) from incubated human atrial tissue: (I) Ouabain induced a concentration-dependent increase in norepinephrine release, that was calcium-independent and almost completely prevented by blockade of the uptake1-carrier by desipramine (1 mumol/l). The characteristics of this release process are consistent with a non-exocytotic mechanism. (II) In addition, ouabain augmented the nicotine-evoked (1-100 mumol/l) calcium-dependent norepinephrine release, which can be considered to be exocytotic. Na+/K(+)-ATPase inhibition also reduced the threshold concentration of nicotine from 10 to 1 mumol/l and it delayed the rapid tachyphylaxis of its norepinephrine releasing effect in human atrial tissue. In BCC, ouabain increased [Na+]i, [Ca2+]i and [3H]-norepinephrine release in parallel. Under calcium-free conditions, not only the ouabain-induced increase in [Na+]i, but also [3H]-norepinephrine release were enhanced. The ouabain-induced [3H]-norepinephrine release was always closely related to changes in [Na+]i, indicating a key role of [Na+]i for this calcium-independent non-exocytotic norepinephrine release. In addition, pretreatment with ouabain (1 mmol/l) augmented the nicotine-evoked (0.1-10 mumol/l) increments in [Na+]i, [Ca2+]i and [3H]-norepinephrine release. As nicotine-induced norepinephrine release depends on an increase in both [Na+]i and [Ca2+]i, these findings are indicative of an ouabain-mediated facilitation of exocytosis. In conclusion, increasing [Na+]i and [Ca2+]i inhibition of Na+/K(+)-ATPase by ouabain triggers non-exocytotic norepinephrine release, and facilitates nicotine-evoked exocytotic norepinephrine release.
（1）描述洋地黄苷哇巴因抑制Na+/K(+)-ATP酶对人心脏自发释放和尼古丁诱发去甲肾上腺素释放的影响；（2）进一步研究糖苷诱导的细胞内钠离子浓度（[Na+]i）和钙离子浓度（[Ca2+]i）变化（通过显微荧光测定法测定）在儿茶酚胺释放中的作用。后一项实验在牛肾上腺髓质嗜铬细胞（BCC）中进行，BCC是一种成熟的交感神经细胞培养模型。哇巴因（1 - 1000 μmol/L）对孵育的人心房组织中去甲肾上腺素释放（通过高效液相色谱法测定）产生双重作用：（I）哇巴因诱导去甲肾上腺素释放呈浓度依赖性增加，该增加不依赖钙离子，且几乎完全被地昔帕明（1 μmol/L）阻断摄取1载体所抑制。这种释放过程的特征与非胞吐机制一致。（II）此外，哇巴因增强了尼古丁诱发的（1 - 100 μmol/L）依赖钙离子的去甲肾上腺素释放，这可被认为是胞吐作用。抑制Na+/K(+)-ATP酶还将尼古丁在人心房组织中诱发去甲肾上腺素释放的阈值浓度从10 μmol/L降低到1 μmol/L，并延迟了其去甲肾上腺素释放效应的快速耐受性。在BCC中，哇巴因使细胞内钠离子浓度（[Na+]i）、钙离子浓度（[Ca2+]i）和[3H]-去甲肾上腺素释放同时增加。在无钙条件下，不仅哇巴因诱导的细胞内钠离子浓度（[Na+]i）增加，而且[3H]-去甲肾上腺素释放也增强。哇巴因诱导的[3H]-去甲肾上腺素释放始终与细胞内钠离子浓度（[Na+]i）的变化密切相关，表明细胞内钠离子浓度（[Na+]i）在这种不依赖钙离子的非胞吐性去甲肾上腺素释放中起关键作用。此外，用哇巴因（1 mmol/L）预处理可增强尼古丁诱发的（0.1 - 10 μmol/L）细胞内钠离子浓度（[Na+]i）、钙离子浓度（[Ca2+]i）和[3H]-去甲肾上腺素释放的增加。由于尼古丁诱导的去甲肾上腺素释放依赖于细胞内钠离子浓度（[Na+]i）和钙离子浓度（[Ca2+]i）的增加，这些发现表明哇巴因介导促进了胞吐作用。总之，哇巴因抑制Na+/K(+)-ATP酶增加细胞内钠离子浓度（[Na+]i）和钙离子浓度（[Ca2+]i），触发非胞吐性去甲肾上腺素释放，并促进尼古丁诱发的胞吐性去甲肾上腺素释放。
