The objective is elucidation of the role of long-chain polyenoic acids (LCPA) in spermatogenesis.
Specific aims are directed by the hypothesis that function of LCPA as 1) precursors to prostaglandin-like messenger or regulatory molecules or 2) necessary components of specific membranes, will be reflected in properties and distribution of key enzyme activities, in lipid composition of LCPA-rich membranes and in response of enzyme activities and membrane compositions to regulators and other modifiers of spermatogenesis. Key enzymes, including acyl-CoA synthetases, transacylases, desaturases, phospholipases, triacylglycerol lipases and cholesteryl ester hydrolases, which specify distribution of LCPA among cell types and lipid classes of rat testis, will be measured and characterized in whole testis, Sertoli cells, spermatocytes, spermatids and spermatozoa from testis and epididymus. Polar metabolites of 20 and 22 carbon polyenoic acids will be isolated from incubations of the cell types by HPLC and characterized by GC-Mass Spectrometry. Biosynthesis of polar metabolites from radioactive LCPA will be measured in the presence of polyenoic acids, prostaglandins, hormones and other effectors of spermatogenesis. Concentrations and fatty acid compositions of phospholipid classes will be measured in cellular total lipid and in microsomal, mitochondrial and plasma membranes from each of the cell types. Membrane lipid composition will be perturbed by treatments known to modify spermatogenesis, such as essential fatty acid deficiency, feeding of omega-3 fatty acids, hypophysectomy and hormonal replacement. Enzyme activities of specific cell types and membrane classes will be correlated with cell and membrane lipid compositions at various stages of differentiation and with interventions affecting spermatogenesis. Lipid composition and enzyme activities of cell types and membranes will be correlated with morphology by light and electron microscopy with the various interventions. These studies will increase understanding of spermatogenesis on a molecular level and yield insight into its regulation, which will facilitate clinical intervention. These studies will also address regulation of membrane properties, function and metabolism of prostaglandins and functional significance of essential fatty acids, which are relevant to normal functioning of every organ system, many related pathologies and proposed increases in dietary omega-3 fatty acids, which are LCPA.
