全細胞パッチクランプ法での流動電位測定法の開発

The streaming potential (V_stream) is an outcome of the electroosmosis that arises from coupled flux of ion and water in the pore. V_stream gives quantitative information on the coupling ratio of ion-water flux and, thus, it is prerequisite for investigating ion permeation mechanisms. V_stream is evaluated as a shift of the reversal potential (V_rev) upon changes in osmotic gradient. Measurements of V_stream are, however, technically challenging. High osmotic gradient must be imposed across the membrane and planar bilayer method has been used exclusively. Moreover, the value of V_stream is so small that precise evaluation of V_rev is necessary. Here, we developed a novel method for measuring V_stream under the whole-cell patch-clamp configuration. Two features are essential in this method. Firstly, exposure time to high osmotic gradient was defined for short period of time by the ultra-fast solution-exchange system, thus circumvented cellular damages and subsequent responses. Secondly, V_rev was evaluated with great accuracy from macroscopic current measurements upon a series of ramp command. Whole time courses of current traces during a ramp voltage were used for optimizing the parameters of an equivalent membrane circuit. We applied this method to HERG potassium channel expressed in HEK293 cells and revealed that the ion-water coupling ratio changed with potassium concentration. [Jpn J Physiol 55 Suppl:S136 (2005)]