The electrostatic properties of membranes are known to play an important role in the modulation of ion channel activity. One of the best characterized of these is the effect of transmembrane potentials on the gating of the voltage-dependent channels in nerve. Membranes also possess an intrinsic internal dipole potential that is much larger than the transmembrane potential, and this dipole potential likely plays an important role in determining membrane protein structure and activity. Because of the lipid solubility and structure of many anesthetics, these compounds are expected to modulate this internal dipole potential. The work proposed here is directed at quantitatively examining the effect of anesthetics on the internal dipole potential of membranes. These electrical measurements will be carried out in vesicle systems using a number of novel electron paramagnetic resonance (EPR) techniques. Some of these EPR measurements will be corroborated by comparable experiments in planar bilayer systems. To provide a structural basis for understanding the electrical changes, nuclear magnetic resonance (NMR) methods will be used to determine the location and orientation of anesthetics as well as their effects on the structure of the lipid interface. These NMR measurements will include novel solid state deuterium experiments along with rotating frame nuclear Overhauser effect (NOE) measurements. A simple electrostatic model will be used to interpret the electrical and structural data from these experiments. The goal of this work is to determine whether changes in the magnitude of the membrane dipole potential can provide a mechanism for the anesthetic induced modulation of membrane protein activity. While this work may ultimately provide information that will allow for a better understanding of anesthesia, the immediate objectives are directed at examining extremely important yet poorly characterized membrane forces that could function in the control of protein structure and dynamics.
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