Insulin resistance is a hallmark of the metabolic syndrome. Muscle comprises the vast majority of insulin sensitive tissue and is a site of dysregulation in the insulin resistant state. Insulin resistance cannot be understood without defining underlying defects in insulin resistant muscle. Despite the central role of muscle metabolism to overall metabolic health, the mechanism(s) for its effectiveness in healthy physically active states, the factors responsible for dysfunction, and the means to correct dysfunction are poorly understood. The objective of the proposed studies is to bridge the biochemical and histological characteristics of the insulin resistant muscle with their functional consequences. The proposed studies will expand on three major findings from the last funding cycle. These are that (a) extramyocellular barriers to muscle glucose uptake are an important component of insulin resistance; (b) the myocellular extracellular matrix in the insulin resistant state is characterized by increased collagen deposition and reduced matrix metalloprotease-9 activity; and (c) unique and diverse mechanisms for enhancing insulin sensitivity in insulin resistant muscle also reduce collagen deposition and increase MMP9 activity. The aims of the proposed studies are to study in the whole organism: (i) the magnitude and mechanism whereby selective phosphodiesterae-5A inhibition prevents and reverses high fat (HF) diet-induced muscle insulin resistance; (ii) the means by which genetic expression of mitochondrial-targeted catalase prevents insulin resistance in HF-fed mice; (iii) the means by which a physiological intervention, regular physical exercise, protects against HF-fed insulin resistance; and (iv) the relative importance of endothelial dysfunction and extracellular matrix modifications to the extramyocellular defects associated with insulin resistance and interventions that enhance insulin action. These aims will be addressed using novel experimental models (chronically-catheterized, unstressed mice), genetically modified mouse models, isotopic techniques, and high-speed slit-confocal imaging. Results from these studies will lead to a greater understanding of the sites involved in the dysregulation of muscle glucose uptake associated with insulin resistance and will identify potential treatment strategies.