Influence of Cholesterol Microstructures on Fluctuation Spikes in Nystatin Channel Currents in Phospholipid/Cholesterol Bilayers

Abstract

Nystatin (NYS) is an antifungal agent that preferentially forms ion channels in membranes containing ergosterol (ERG) or cholesterol (CHOL). In phospholipid bilayers ERG or CHOL segregate into ordered (Lo) and disordered (Ld) domains. We prepared POPC bilayers containing CHOL mol fraction 0.18 ≤ χERG ≤ 0.50 separating two chambers containing KCl solutions. The concentration gradient between chambers was 435/150 mM KCl. We allowed the CHOL/POPC bilayer to settle 15 min then added 19 or 38 microM NYS to the reference (Cis) chamber, imposed 50 mV across the bilayer, and stirred the Cis chamber at 4 Hz. The NYS molecules adhered to the bilayer forming channels on the boundaries of the Lo domains. [1] The resultant bilayer current exhibited prominent spikes of very short duration. A plot of the frequency of these spikes against χERG revealed prominent spikes at χERG = 0.19, 0.25 and 0.40. These correspond well to the dips in dehydroergosterol fluorescence observed by Chong [2], which he understood in terms of a superlattice. We conclude that fluctuations in NYS channels on the perimeter of the Lo CHOL domains depend strongly on Lo domain structure.1. Helrich, C. S., J. A. Schmucker, and D. J. Woodbury 2006. Evidence that Nystatin Channels Form at the Boundaries, Not the Interiors of Lipid Domains. Biophys. J. 91: 1116-1127.2. Chong, P.L-G. 1994. Evidence for regular distribution of sterols in liquid crystalline phosphatidylcholine bilayers, Proc. Nati. Acad. Sci. USA 91:10069-10073.