Sandle G I, Butterfield I, Higgs N B, Warhurst G
Molecular Medicine Unit, St. James's University Hospital, Leeds LS9
Pflugers Arch. 1999 Mar;437(4):596-602. doi: 10.1007/s004240050822.
The most abundant basolateral K+ channels in human colonic crypt cells have a low conductance (23 pS), respond to increases in intracellular Ca2+ and cAMP, and have been implicated in intestinal electrogenic Cl- secretion. The effect of nicardipine on the activity of these K+ channels was examined by patch-clamp recording in the cell-attached and excised inside-out configurations from the basolateral membrane of single crypts isolated from biopsied samples of human distal colon. During cell-attached recordings, addition of 2 micromol/l nicardipine to crypts pretreated with 200 micromol/l dibutyryl cAMP decreased single-channel open probability by 87%, but in parallel studies nicardipine had no effect on the intracellular Ca2+ concentration. Using inside-out patches from crypts pretreated with dibutyryl cAMP (bathed in 1.2 mmol/l Ca2+), the addition of increasing concentrations of nicardipine (200 nmol/l, 2 micromol/l and 20 micromol/l) decreased single-channel open probability in a concentration-dependent manner (IC50 0.47 micromol/l). In additional experiments using stripped rat distal colonic mucosa mounted in conventional Ussing chambers, serosal addition of nicardipine at increasing concentrations (ranging from 200 nmol/l to 20 micromol/l) produced a concentration-dependent inhibition of dibutyryl-cAMP-stimulated electrogenic Cl- secretion (IC50 2 micromol/l). Taken together, these results indicate that nicardipine has a direct inhibitory action on 23-pS basolateral K+ channels in human intestinal crypt cells, which is likely to decrease cAMP-stimulated electrogenic Cl- secretion. These basolateral K+ channels may provide a focal point for the development of new strategies in the treatment of secretory diarrhoeal diseases.
人类结肠隐窝细胞中最丰富的基底外侧钾通道具有低电导（23 pS），对细胞内Ca2+和cAMP的增加有反应，并与肠道电生性氯分泌有关。通过膜片钳记录，在从人远端结肠活检样本中分离的单个隐窝的基底外侧膜上，采用细胞贴附式和内面向外式记录配置，研究了尼卡地平对这些钾通道活性的影响。在细胞贴附式记录期间，向用200 μmol/l二丁酰cAMP预处理的隐窝中加入2 μmol/l尼卡地平，单通道开放概率降低了87%，但在平行研究中，尼卡地平对细胞内Ca2+浓度没有影响。使用用二丁酰cAMP预处理（浸泡在1.2 mmol/l Ca2+中）的隐窝的内面向外膜片，加入浓度逐渐增加的尼卡地平（200 nmol/l、2 μmol/l和20 μmol/l）以浓度依赖性方式降低单通道开放概率（IC50为0.47 μmol/l）。在另外的实验中，使用安装在传统Ussing室中的剥离大鼠远端结肠黏膜，向浆膜侧加入浓度逐渐增加的尼卡地平（范围从200 nmol/l到20 μmol/l），对二丁酰cAMP刺激的电生性氯分泌产生浓度依赖性抑制（IC50为2 μmol/l）。综上所述，这些结果表明，尼卡地平对人肠道隐窝细胞中的23-pS基底外侧钾通道具有直接抑制作用，这可能会减少cAMP刺激的电生性氯分泌。这些基底外侧钾通道可能为分泌性腹泻疾病治疗新策略的开发提供一个重点。
