Phospholipids and bile acids as diffusional carriers of Na+ across nonpolar media

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Abstract
Phospholipids and bile acids, by virtue of their amphiphilic properties, can interact in nonpolar media forming “inverted” structures (micelles) which presumably have an hydrophilic core and might act as diffusional carriers (ionophores) of electrolytes across low dielectric constant media or lipid membranes. The Na+ ionophoretic capability of various purified phospholipids and the modulating effects of bile acids and phospatidylcholine was examined by: (a) measurement of 22Na+ partition into the organic phase (chloroform) of a two-phase system and (b) direct measurement of the translocation of 22Na+ across a bulk chloroform phase separating two aqueous phases in a Pressman cell. All phospholipids tested, except for phosphatidylcholine, showed ionophoretic capability for Na+ at micromolar concentrations. Cardiolipin and phosphatidylserine were the most efficient Na+ carriers, comparable with monensin, an established Na+ ionophore. In contrast, cholic acid as well as other bile acids demonstrated only marginal or no Na+ ionophoretic capability. However, hydroxylated bile acids (particularly cholic acid), sodium dodecyl sulfate and Triton X-100, which can induce and stabilize inverted structures in lipid membranes, were able to increase 5- to 8-fold the phospholipid-mediated Na+ transport. Interaction of cardiolipin with Na+ in the chloroform phase followed a rectangular hyperbolic function with an apparent Kd within the physiological Na+ concentration range (16.9 ± 5.1 mM). Addition of cholic acid to the cardiolipin-containing organic phase resulted in a 10-fold increase of maximal Na+ uptake and no change in apparent Kd. The effect of cholic acid on both cardiolipin-mediated Na+ partition and Na+ translocation across the chloroform phase showed a marked dependence on pH, being greater at pH 7.4. On the other hand, phosphatidylcholine, which is reported to stabilize phospholipid bilayers and to inhibit formation of inverted structures, inhibited Na+ cardiolipin interactions. Cholic acid addition completely prevented the inhibitory effect of phosphatidylcholine. This study permits us to establish that the acidic phospholipids cardiolipin and phosphatidylserine can act as efficient Na+ ionophores in this in vitro system, at physiological Na+ concentrations and with kinetics comparable to those of systems involved in Na+ transport (e.g. Na+/H+ antiports) in liver surface membrane and other biomembranes. Modulation by bile acids (stimulation) and phosphatidylcholine (inhibition) of cardiolipin-mediated Na+ transport suggests that the underlying mechanism probably involves inverted-mixed micelle formation in the organic phase. The possibility exists that the Na+ ionophoretic properties of acidic phospholipids are relevant to the bile secretory process.