AJP - Cell Physiology, Vol 248, Issue 3 372-C378, Copyright © 1985 by American Physiological Society
Ion movements in membrane vesicles: a new fluorescence method and application to smooth muscle
A. K. Grover, A. P. Singh, P. K. Rangachari and P. Nicholls
A method is described for studying ion permeabilities of membrane vesicles
based on the principle that when membrane permeability to H+ is very high,
the H+ movement is determined by the membrane potential generated by the H+
movement. The rate of H+ movement under these conditions thus gives a
measure of the rate of dissipation of this membrane potential by comovement
of anions or countermovement of cations present. Thus, by studying the H+
efflux using an impermeant cation and different anions, the membrane
permeability to the anions can be assessed. Similarly, the use of an
impermeant anion allows the study of the permeation of various cations. H+
movement was followed across the membranes by monitoring a change in the
fluorescence intensity of the pH-sensitive dye pyranine trapped inside the
membranes. This method when tested using phosphatidylcholine liposomes
yielded the expected results, i.e., permeability of the liposomal membrane
was: Cl- greater than SO2-4 and K+ greater than Na+. A plasma
membrane-enriched fraction loaded with pyranine was isolated from
estrogen-dominant rat myometrium. The anion permeability characteristics of
this membrane were studied using tetramethylammonium (TMA+) as the poorly
permeant cation, and the cation permeability was studied using L-glutamate-
as the poorly permeant anion. The anion permeabilities were D-glutamate-
less than L-glutamate- less than glutarate2- less than Cl- less than or
equal to SO2-4, and the cation permeabilities were TMA+ less than K+ less
than Na+. It is hypothesized that the observed anomalously higher Na+ and
SO2-4 movements may involve special mechanisms.
