The polyunsaturated fatty acids (PUFA) of the linolenic (n-3) and linoleic (n-3) classes are not synthesized by animals. Mammalian tissues cannot interconvert the different classes of PUFA and must therefore derive both n-3 and n-6 precursors from exogeneous sources. The fatty acid composition of neural tissues is highly enriched in PUFA and contain a relatively large amount of long- chain PUFA of the n-3 class. The importance of plasma membrane PUFA composition on neuronal function remains unknown, although there is evidence to support the hypothesis that membrane lipids can affect ion channel functions. The objective of the proposed research is to test the hypothesis that changes in membrane PUFA composition can affect neuronal excitability by altering passive membrane properties and/or properties of voltage-dependent ion channels. The approach is to grow rat embryonic neocortical neurons in serum-free defined culture medium and to alter the membrane PUFA composition by supplementing the culture with the n-3 or n-6 classes of PUFA. The electrophysiological properties of neurons grown in PUFA supplemented media will be compared with neurons grown in medium without the addition of exogenous fatty acids or in medium supplemented with saturated or monounsaturated fatty acids. The differences in the electrophysiological properties will be correlated with differences in membrane lipid composition. The gigaseal whole cell voltage-clamp recording technique will be used to study the electrophysiological properties, and high pressure liquid chromotography combined with mass spectrometry and gas chromatography will be used analyze plasma membrane lipid composition. The information that will be gained from studying this relatively simple system of dissociated primary cultures will aid in the understanding of the role of essential polyunsaturated fatty acids and their metabolites in neuronal excitability in the more complex mammalian brain in vivo. This will serve as an important baseline study for my longer term research goal, which is to understand the regulation of expression of membrane excitability by epigenetic factors in developing mammalian brain neurons. This study will increase our understanding of the functional defects of CNS neurons induced by prenatal and postnatal essential fatty acid deficiency.
