Specific effects which may contribute to ethanol-induced teratogenesis have been documented extensively, but no unifying hypothesis has been accepted for the mechanism of action. In whole rat embryo cultures, ethanol produces dose-dependent decreases in protein content and glucose utilization, both of which are ameliorated by glucose supplementation, and protein content correlates closely with anabolic glucose utilization. Ethanol also reduces glucose transporter gene expression and inhibits glucose oxidation via the pentose phosphate pathway (PPP), which supplies ribose precursors for nucleic acid synthesis and NADPH for lipid synthesis. Ribose supplementation also enhances the growth of embryos in the presence of ethanol. Reduced PPP activity has been implicated in fuel-mediated teratogenesis, which produces growth retardation and many of the same abnormalities that characterize the effects of ethanol. It is suggested that ethanol may impair embryonic and fetal growth and differentiation via inhibition of glucose transport and reduction of PPP activity, which limits the supply of substrate for nucleic acid synthesis and of coenzyme for lipid synthesis. Studies are proposed using two culture systems, intact rat embryos and brain astrocytes, which display dose-related effects of ethanol on growth and differentiation and in which the PPP has particular functional significance. The effects of ethanol on glycolytic, PPP, and tricarboxylic acid cycle activity will be differentiated developmentally in embryo cultures, which are characterized by high glycolytic and PPP activity between days 9.5 and 10.5 of gestation and by the appearance of the TCA cycle between days 10.5 and 11.5. The effects of ethanol on the activities of these pathways, on the incorporation of glucose carbon into DNA, and on growth will be compared in media containing normal, low, and supplemented glucose levels. The activities of key enzymes nd the levels of NADPH, NADH, and high energy phosphates also will be determined. Studies in astrocytes will be conducted during exponential growth, when PPP activity is expected to be high, and at confluence, and will parallel those done with embryo cultures. To determine whether effects on glucose transport can account for reduced PPP activity, the transporters and glucose transport activity will be quantitated and characterized under the same conditions, and the mechanism of any effects will be investigated.
