Rohlicek V, Schmid A
Czech Academy of Science, Prague.
Pflugers Arch. 1994 Aug;428(1):30-8. doi: 10.1007/BF00374749.
A dual-frequency method was developed to monitor changes of membrane capacitance, membrane conductance and serial resistance between patch pipette and cytoplasm of the cell in the whole-cell configuration. Measurement of real and imaginary components of cell admittance during excitation with two superimposed sinusoidal voltage of different frequencies provides mathematical solutions for all three variables. The validity of the method was verified with experiments on mast cells and exocrine pancreatic acinar cells. During degranulation of mast cells, induced by GTP gamma S in the pipette solution, a stepwise increase in membrane capacitance could be observed, indicating that the resolution of the method is below 10 fF. Precalibration of the setup allows all calculated data to be expressed as absolute values. The capacitance measurement proved to be rather independent of changes in the access resistance and in the cell membrane resistance over a wide range. The huge changes in membrane conductance of mouse pancreatic acinar cells during hormonal stimulation with acetylcholine produced a relative error of less than 0.3% in the capacitance trace. This allows a clear distinction between changes of membrane conductance and cell capacitance. The method therefore offers great advantages in the study of exocytosis as well as endocytosis in cell types, such as exocrine gland cells, with major changes in membrane conductance during hormonal stimulation.
开发了一种双频方法，用于监测全细胞配置下细胞膜电容、膜电导以及膜片钳与细胞质之间串联电阻的变化。在施加两个不同频率的叠加正弦电压进行激发时，测量细胞导纳的实部和虚部，可得出这三个变量的数学解。通过对肥大细胞和胰腺外分泌腺泡细胞进行实验，验证了该方法的有效性。在移液管溶液中用GTPγS诱导肥大细胞脱颗粒过程中，可观察到膜电容呈逐步增加，这表明该方法的分辨率低于10 fF。对仪器进行预校准可使所有计算数据以绝对值表示。电容测量结果在很大程度上不受接入电阻和细胞膜电阻变化的影响。在用乙酰胆碱进行激素刺激时，小鼠胰腺腺泡细胞膜电导发生巨大变化，但电容记录中的相对误差小于0.3%。这使得能够清晰区分膜电导变化和细胞电容变化。因此，该方法在研究诸如外分泌腺细胞等在激素刺激期间膜电导发生重大变化的细胞类型的胞吐作用和胞吞作用方面具有很大优势。
