A new technique for evaluating volume sensitivity of ion channels

Abstract

 A new technique is described for evaluating the volume sensitivity of both endogenous and expressed ion channels in Xenopus oocytes. The technique utilizes vesicles derived from inside-out, excised patches and permits evaluation of volume sensitivity at the single-channel level, which is not possible with whole oocytes since removal of the vitelline membrane renders oocytes too fragile to withstand osmotic stress. Using this method, the volume sensitivity of the normally occurring oocyte mechanosensitive (SA-cat) channel was assessed. In 12 experiments, osmotic swelling increased vesicle volume by an average of 60 ± 8% and increased the surface area of the inner side of the vesicle by 47 ± 10%. This was associated with an increase in mean number of open channels (NP _o) from zero (unswollen) to 0.1 ±0.02 (after 5 min). Hence, oocyte SA-cat channels are volume as well as stretch sensitive, where the volume sensitivity may involve cytoskeletal elements adhering to the membrane but probably does not involve freely diffusible components of the original oocyte.