THE USE OF RESISTANCE NETWORKS AND IMPROVED ESTIMATES OF SEAL RESISTANCE TO ESTIMATE TONOPLAST CONDUCTIVITY IN WHOLE-VACUOLE PATCH CLAMPING

The stability of the seal resistance is a major assumption in patch-cl amping making its estimation critical especially in the whole-vacuole configuration. A clamp voltage protocol was used that enhanced the cha nce of:closure of channels in the patch, making possible a better esti mate of the seal resistance while in the vacuole-attached configuratio n. In the whole-vacuole configuration the resistance of the tonoplast was often found to be close in magnitude to the seal resistance, makin g direct measurement of the tonoplast resistance problematic. Therefor e, a resistance network model was used to separate by calculation the influence of the seal resistance from the tonoplast resistance. Using unadjusted data the specific conductance of red beet tonoplast in whol e-vacuole configuration was estimated to be 0.507 pS/mu m(2) (S.E. = 0 .176 pS/mu m(2), n = 8), but after correcting for the influence of the seal and tip resistances the tonoplast specific conductivity was esti mated to be 0.365 pS/mu m(2) (S.E. = 0.140 pS/mu m(2), n = 8), a 28% s hift in the estimate. Careful attention to seal resistance estimation, and correction of data for its influence is suggested for all work in volving whole-vacuole patch-clamping. (C) 1997 Elsevier Science Irelan d Ltd.