Voltage-dependent activation of slow vacuolar (SV) channels has been studied on isolated patches from red beet (Beta vulgaris L.) vacuoles. Isoosmotic variation of vacuolar K+ from 10 to 400 mM in Ca2+-free solutions at the vacuolar side shifted the SV channel activation threshold to more positive voltages. The effect of K+ could be mimicked by additions of choline or N-methyl D-glucamine and could be explained by unspecific screening of the negative surface charge. Fitting the dependence of voltage shift on K+ concentration to the Gouy-Chapman model yields a surface charge density of 0.36 ± 0.05 e−/nm2. Negative surface potential also tended to increase the local concentration of permeable ions (K+), resulting in anomalously high single-channel conductance, ∼200 pS in 10 mM KCl. An increase of ionic strength due to addition of impermeable cations greatly reduced the unitary conductance. Large positive shift of the SV channel voltage dependence, caused by physiological (0.5 mM) free vacuolar Ca2+, was partly ameliorated by increasing luminal K+. We interpreted these results as follows: K+ induced a reduction of surface potential, hence i) causing a positive shift of the voltage dependence and ii) a dilution of Ca2+ in the membrane vicinity, thus reducing the inhibitory effect of vacuolar Ca2+ and causing a negative shift of the SV channel voltage dependence, with a sum of the two shifts being negative.
The Journal of Membrane Biology – Springer Journals
Published: Jan 1, 2005
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