Aunis D, García A G
Br J Pharmacol. 1981 Jan;72(1):31-40. doi: 10.1111/j.1476-5381.1981.tb09101.x.
1 Secretion of catecholamines (CA) evoked by ouabain, chlormadinone acetate (CMA), phenoxybenzamine (Pbz) and vanadate, four agents known to inhibit Na(+), K(+)-dependent Mg(2+)-activated adenosine triphosphatase (ATPase) activity has been studied in suspensions of bovine isolated adrenal medullary cells.2 Acetylcholine (ACh) evoked a 5 fold increase of the basal CA secretion from isolated cells suspended in oxygenated Krebs-bicarbonate solution kept at 27 degrees C. Secretion was antagonized by Ca(2+)-deprivation or hexamethonium, indicating good functional viability of the cells.3 Ouabain (10(-7) to 10(-4) M) evoked a progressive, dose-dependent release of CA from cell suspensions. Study of the time course of the secretory response for 2 h allowed the separation of two components in the secretory response at all doses studied: a slow initial component (0.011 pg/min CA) and a second faster component (0.032 pg/min CA).4 CMA evoked a clear-cut CA secretory response. The ED(50) for CMA was 10(-4) M, as compared to 3 x 10(-6) M for ouabain. Pbz and vanadate did not induce CA release.5 [(3)H]-ouabain was taken up and bound to intact isolated cells by a non-saturable binding process. However, in semi-purified plasma membranes from bovine adrenal medulla a saturable specific [(3)H]-ouabain binding process was observed with a K(D) of 8.1 nM. Binding to the membranes was ATP-dependent and antagonized by K(+).6 [(3)H]-ouabain specific binding to membranes was antagonized by ouabain and CMA, but not by Pbz or vanadate; the ID(50) for ouabain and CMA were 10(-6) and 10(-5) M respectively.7 Ouabain partially inhibited, in a dose-dependent manner, Na(+), K(+)-Mg(2+) ATPase activity of the semi-purified plasma membranes.8 The results demonstrate a good correlation between the ability of different drugs, known to inhibit ATPase activity, to displace [(3)H]-ouabain binding to adreno-medullary plasma membranes and their capacity to evoke a CA secretory response from isolated chromaffin cells. The data also suggest that the CA secretory effects of ouabain may not be due simply to inhibition of the Na(+) pump and the subsequent ionic redistribution across the plasma membrane; a second mechanism may also be involved.
哇巴因、醋酸氯地孕酮（CMA）、酚苄明（Pbz）和钒酸盐这四种已知可抑制钠钾依赖的镁激活三磷酸腺苷酶（ATP酶）活性的物质所诱发的儿茶酚胺（CA）分泌，已在牛离体肾上腺髓质细胞悬液中进行了研究。
乙酰胆碱（ACh）使悬浮于27℃含氧 Krebs - 碳酸氢盐溶液中的离体细胞的基础CA分泌增加了5倍。钙缺乏或六甲铵可拮抗分泌，表明细胞具有良好的功能活性。
哇巴因（10⁻⁷至10⁻⁴M）可引起细胞悬液中CA的渐进性、剂量依赖性释放。对2小时分泌反应的时间进程研究表明，在所有研究剂量下，分泌反应可分为两个成分：一个缓慢的初始成分（0.011 pg/分钟CA）和第二个较快的成分（0.032 pg/分钟CA）。
CMA诱发了明显的CA分泌反应。CMA的半数有效剂量（ED₅₀）为10⁻⁴M，而哇巴因的ED₅₀为3×10⁻⁶M。Pbz和钒酸盐未诱导CA释放。
[³H] - 哇巴因通过非饱和结合过程被完整的离体细胞摄取并结合。然而，在牛肾上腺髓质的半纯化质膜中，观察到了一个可饱和的特异性[³H] - 哇巴因结合过程，解离常数（K(D)）为8.1 nM。与膜的结合依赖于ATP，并被钾离子拮抗。
[³H] - 哇巴因与膜的特异性结合被哇巴因和CMA拮抗，但不被Pbz或钒酸盐拮抗；哇巴因和CMA的半数抑制剂量（ID₅₀）分别为10⁻⁶和10⁻⁵M。
哇巴因以剂量依赖的方式部分抑制了半纯化质膜的钠钾 - 镁ATP酶活性。
结果表明，已知抑制ATP酶活性不同药物取代[³H] - 哇巴因与肾上腺髓质质膜结合的能力与其从离体嗜铬细胞诱发CA分泌反应的能力之间存在良好的相关性。数据还表明，哇巴因对CA的分泌作用可能不仅仅是由于抑制钠泵以及随后跨质膜的离子重新分布；可能还涉及第二种机制。
