Study on multimolecular modulations of ATP-sensitive K^+ channels in pancreatic β-cells and their disharmony in diabetes.

ATP敏感性K^+（KATP）通道由磺酰脲受体单元和内向整流K^+通道的孔单元的异多聚体组成，研究了它们对细胞内Ca^2+活性的影响。研究发现，细胞内Ca^2+升高可抑制磺酰脲类药物与其受体结合和通道关闭之间的功能性转导。暴露于PIP_2可增加KATP通道的活性，并降低通道的ATP敏感性。当用GFP（绿色荧光蛋白）标记的含有PH结构域的探针测量膜PIP_2浓度时，当β细胞暴露于乙酰胆碱时，GFP强度的膜分数降低。在乙酰胆碱的存在下，在移液管中记录在由内向外模式的通道的ATP敏感性增加相比，在控制移液管溶液。在细胞贴附膜片钳实验中，乙酰胆碱作用于β细胞，可诱发动作电流。在用制霉菌素穿孔模式记录的全细胞电流中，乙酰胆碱在葡萄糖浓度为2.8mM时使KATP通道电流降低。从这些结果可以看出，受体刺激胰腺β细胞导致KATP通道活性降低，与膜去极化有关，而在葡萄糖阈值浓度附近胰岛素分泌的输出增加。需要进一步探索来自糖尿病动物的β-细胞在副交感神经刺激期间是否具有乙酰胆碱去膜的能力以及由此产生的胰岛素分泌的扰动。

ATP-sensitive K^+ (KATP) channels, composed of heteromultimers of sulfonylurea-receptor unit and a pore unit of inwardly rectifying K^+ channels, were studied with respect to effects of intracellular Ca^<2+> activity of the channels. It was found that elevation of intracellular Ca^<2+> conferred inhibition of functional transduction between binding of sulfonylureas to their receptors and closure of channels. Exposure of membrane patches to PIP_2 increased activity of KATP channels in association with reduction of ATP sensitivity of the channels. When measured membrane PIP_2 concentration by means of GFP (green fluorescent protein) labeled PH-domain contained probe, membrane fraction of GFP intensity decreased on exposure of β-cells to acetylcholine. The ATP sensitivity of the channel recorded in the presence of acetylcholine in the pipette in inside-out mode was increased as compared to that in control pipette solution. Acetylcholine, when it was applied to β-cell during cell-attached patch experiments, induced action currents. In whole-cell current recorded by nystatin-perforated mode, acetylcholine decreased the KATP channel currents at the glucose concentration of 2.8 mM. From these results, it is suggested that receptor-stimulation of pancreatic β-cells produces reduction of activity of the KATP channels associated with depolarization of membrane and an increase in output of insulin secretion around threshold concentrations of glucose. It is needed to further explore whether β-cells from diabetic animals have the inability of acetylcholine to depolarize the membrane and resultant perturbation of insulin secretion during parasympathetic neural stimulation.