Long-term exposure to fatty acids impairs beta cell function in type 2 diabetes while little is known about the chronic effects of fatty acids on alpha cell. We therefore studied the prolonged impact of palmitate on alpha cell function and on the expression of genes related to fuel metabolism. We also investigated if the antihyperglycaemic agent, stevioside, was able to counteract these effects of palmitate. Clonal alpha TC1-6 cells were cultured with palmitate in the presence or absence of stevioside. After 72h we evaluated glucagon secretion, glucagon content, triglyceride content and changes in the gene expression. Glucagon secretion was dose-dependently increased after 72h culture with palmitate (0.25 mM 1.0mM) (P<0.05). Palmitate (0.5mM) enhanced triglyceride content of alpha cells by 73% (P<0.01). Interestingly, stevioside (10^-8 and 10^-6 M) reduced palmitate-stimulated glucagon release by 22% and 45%, respectively (P<0.01). There was no significant change in glucagon content after 72h culture with palmitate and/or stevioside. Palmitate increased CPT-1 mRNA level while stevioside enhanced CPT-1, PPARr and SCD gene expressions in the presence of palmitate (P<0.05). In conclusion, long-term exposure to elevated fatty acids leads to a hypersecretion of glucagon and an accumulation of triglyceride content in clonal alpha TC1-6 cells. Stevioside was able to counteract the alpha cell hypersecretion caused by palmitate and enhanced the expression of genes involved in fatty acid oxidation and disposal. This indicates that stevioside may be a promising antidiabetic agent in treatment of type 2 diabetes.