Knopp A, Thierfelder S, Koopmann R, Biskup C, Böhle T, Benndorf K
Institut für Physiologie, Abt. Herz-Kreislauf-Physiologie, Friedrich-Schiller-Universität Jena, Germany.
Cardiovasc Res. 1999 Mar;41(3):629-40. doi: 10.1016/s0008-6363(98)00238-7.
The aim was to improve the measurement of both the time course and amplitude of anoxia-induced KATP-channel current (IKATP) in isolated heart cells to specify the role of these channels in the time course of K+ accumulation in the ischemic myocardium.
Ionic currents in isolated ventricular heart cells of the mouse were measured with a patch clamp technique under normoxic conditions (atmospheric pO2), during wash-out of oxygen, and under anoxic conditions (pO2 < 0.2 mmHg). During the measurement, the actual pO2 in the close proximity of the cell was determined with an optical technique by exciting Pd-meso-tetra(4-carboxyphenyl)porphin with light flashes of 508-570 nm and evaluating the quenching kinetics of the emitted phosphorescence signal at 630-700 nm. These quenching kinetics steeply depend on pO2 and can be evaluated best at pO2 values near 0 mmHg.
Out of 28 cells, 23 cells started to develop IKATP at pO2 values between 0 and 0.4 mmHg, i.e. in the range of the level of half maximum activity of the cytochrome oxidase. The remaining five cells developed IKATP between 0.4 and 1.8 mmHg. With respect to the time course, 18 out of 27 cells started to develop IKATP within the first minute after pO2 had decreased to values below 0.2 mmHg. The amplitude of IKATP induced by anoxia and various metabolic inhibitors was large, 29 +/- 12 and 48 +/- 21 nA (+40 mV), respectively. The anoxia-induced IKATP was significantly smaller than IKATP induced by metabolic inhibitors. During the pulses of 50 ms duration to +40 mV, the amplitude of IKATP decayed and, after clamping back to -80 mV, IKATP generated large tail currents. This suggests a notable change in the concentration gradient of K+ ions in the time range of tens of milliseconds.
The results in isolated myocytes indicate that KATP channels open sufficiently rapidly after starting anoxia and generate sufficiently large conductance at maintained anoxia to explain both the time course and magnitude of the ischemic K+ accumulation if an appropriate counter-ion flux is available.
旨在改进对分离的心肌细胞中缺氧诱导的ATP敏感性钾通道电流（IKATP）的时间进程和幅度的测量，以明确这些通道在缺血心肌中钾离子蓄积的时间进程中的作用。
采用膜片钳技术，在常氧条件下（大气氧分压）、氧洗脱期间以及缺氧条件下（氧分压<0.2 mmHg），测量小鼠分离的心室肌细胞中的离子电流。在测量过程中，通过用508 - 570 nm的光脉冲激发钯-中-四（4-羧基苯基）卟啉并评估630 - 700 nm处发射的磷光信号的猝灭动力学，采用光学技术测定细胞附近的实际氧分压。这些猝灭动力学强烈依赖于氧分压，并且在氧分压值接近0 mmHg时能得到最佳评估。
在28个细胞中，23个细胞在氧分压值介于0和0.4 mmHg之间时开始出现IKATP，即处于细胞色素氧化酶最大活性一半的水平范围内。其余5个细胞在0.4至1.8 mmHg之间出现IKATP。就时间进程而言，27个细胞中有18个在氧分压降至低于0.2 mmHg后的第一分钟内开始出现IKATP。缺氧和各种代谢抑制剂诱导的IKATP幅度较大，分别为29±12和48±21 nA（+40 mV）。缺氧诱导的IKATP明显小于代谢抑制剂诱导的IKATP。在持续50 ms至+40 mV的脉冲期间，IKATP的幅度衰减，并且在钳制回-80 mV后，IKATP产生大的尾电流。这表明在几十毫秒的时间范围内钾离子浓度梯度发生了显著变化。
分离的心肌细胞中的结果表明，如果有合适的抗衡离子通量，ATP敏感性钾通道在开始缺氧后足够迅速地开放，并在持续缺氧时产生足够大的电导，以解释缺血性钾离子蓄积的时间进程和幅度。
