TOPOGRAPHY OF ALPHAXALONE AND DELTA(16)-ALPHAXALONE IN MEMBRANE BILAYERS CONTAINING CHOLESTEROL

We have used small-angle X-ray diffraction and differential scanning c alorimetry (DSC) to study the topographies of alphaxalone and its biol ogically inactive analog Delta(16)-alphaxalone in dimyristoylphosphati dylcholine (DMPC) and DMPC/cholesterol model membranes. Diffraction pa tterns were obtained and analyzed for preparations of bilayers without and with the steroids. Temperature dependence of the total period rep eat distance (d-spacing) allowed us to identify equivalent temperature s at which the preparations had similar d-spacing and were in the same mesomorphic state. The combination of X-ray and DSC data showed that the anesthetic steroid alphaxalone broadens the membrane phase transit ion and increases the ratio of gauche:trans conformers in the membrane s in contrast to the inactive steroid Delta(16)-alphaxalone which affe cts the membranes only marginally. In model DMPC membranes alphaxalone and Delta(16)-alphaxalone are located near the bilayer interface. Thi s location is maintained by alphaxalone when cholesterol is incorporat ed in the bilayer as evidenced by the X-ray measurements. However, whe n Delta(16)-alphaxalone is incorporated in cholesterol containing bila yers, a decrease in the electron density profile of the preparation is observed. This can be explained by invoking the formation of a Delta( 16)-alphaxalone -cholesterol complex The Delta(16)-alphaxalone complex shows no periodicity and is therefore, not detected in the X-ray diff raction experiment. Presumably, this complex forms aggregates either o n the surface or inside the bilayer. This explanation corroborates DSC results which show that Delta(16)-alphaxalone sharpens the phase tran sition of DMPC/cholesterol preparations, an indication that some chole sterol is excluded from the bilayer preparation after the addition of the biologically inactive steroid.