PROJECT SUMMARY Prevention, treatment, and cure of Type 2 diabetes requires understanding of the underlying causes, at the genetic and/or pathophysiological level. In our laboratory we have long been interested in identifying underlying causes leading to the disease. We have identified several underlying causes; in particular a reduction in insulin compensation for insulin resistance (lowered Disposition Index) and reduced insulin clearance. We have identified and additional potential factor – reduced Glucose Effectiveness (GE). The latter represents the ability of glucose itself to self-normalize (after oral or intravenous administration) independent of a dynamic insulin response. Yet the mechanisms underlying GE are not clearly understood. In this proposal we describe a series of studies to identify underlying mechanisms. We propose that GE is reflective of the ability of the liver to phosphorylate absorbed glucose via glucokinase (GCK), after which triose phosphates can be stored, utilized for energy or exported as lactate. Studies are don in the dog model -- We will use glucose clamps at basal insulin to measure GE; we will be able to quantitate liver glucose uptake and lactate export by arteriovenous difference across the organ. Also, we will assess GCK and a panoply of the regulatory proteins directly by biopsy of liver tissue, and other pathway components by biopsy of muscle and adipose. We compare GE with insulin sensitivity using clamps. To test the importance of GCK activity (and glucokinase regulatory protein (GKRP) we will either activate GCK with FGF1 or suppress GKRP with antisense oligonucleotide (ASO) designed for our experiments. These experiments will test the hypothesis that GE measured in vivo is determined by the activity of glucokinase in liver. We will examine the putative importance of GE itself in experimental models of metabolic dysfunction, including obesity (overfeeding) and a model of prediabetes (insulin resistance and mild β-cell dysfunction). We have proposed a computer model which allows for estimate of hepatic glucokinase from the intravenous glucose tolerance test, by modeling the relationship between plasma glucose and lactate. The validity of this model will be tested by determining its ability to detect increases in GCK and GE due to FGF1 administration into the CNS (intranasal) and to assess increase in GE due to an exercise regimen. These studies will, for the first time, elucidate the factors underlying glucose effectiveness, and test the extent to which hepatic glucokinase activity can account for GE measured in vivo. We will provide a model-based method to measure GE in vivo which can be applied to studies of the role of GE in pathogenesis of metabolic disease including impaired glucose tolerance, diabetes, PCOS, and others.