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The role of insulin in the regulation of adipose tissue lipoprotein lipase activity in humans was investigated in 11 normal subjects and compared with the effects of 0.9% saline infusions in five control subjects. After a basal adipose tissue biopsy for lipoprotein lipase activity, insulin was rapidly infused to achieve and maintain serum levels of approximately 70 microunits/ml while plasma glucose was kept at basal concentrations. Free fatty acids in serum fell to 27 +/- 3% of basal by 20 min (t = 5.19, P less than 0.001) and triglycerides decreased to 77 +/- 3% of basal by 80 min (t = 3.76, P less than 0.01). Adipose tissue lipoprotein lipase activity failed to increase significantly above that measured in controls by the first 3 h of the study. By 6 h of the infusion a stimulatory effect of insulin on adipose tissue lipoprotein lipase was found (t = 3.94, P less than 0.01). There was no relationship between the amount of glucose infused and the insulin effect on the enzyme. The increase in adipose tissue lipoprotein lipase activity at 6 h, however, was inversely related to the basal lipase activity (r = -0.690, P less than 0.02). Thus, insulin appears to stimulate adipose tissue lipoprotein lipase activity in humans. This effect of insulin is delayed when compared with antilipolysis and the fall in plasma triglyceride. The inverse relationship between insulin-stimulated adipose tissue lipoprotein lipase activity and basal enzyme activity suggests that adipose tissue itself is the main regulator of the lipase response to insulin.