To determine if increased secretion of amylin can be implicated in the pathogenesis of non-insulin-dependent diabetes mellitus (NIDDM) in vitro and in vivo, we studied its relationships to insulin in insulin-resistant rats with and without NIDDM. In obesity-associated and dexamethasone-induced insulin resistance without diabetes, basal and stimulated secretion of amylin and insulin by isolated pancreata were proportionately elevated, leaving the amylin-to-insulin ratio (A/I) unchanged. By contrast, whenever diabetes occurred in dexamethasone-treated rats or in spontaneously diabetic obese insulin-resistant ZDF-drt male rats, a doubling of A/I was invariably observed due to an increase in amylin without a proportional increase in insulin secretion. Correction of dexamethasone-induced hyperglycemia with the glucocorticord receptor antagonist RU-486 was accompanied by a decline in A/I. Longitudinal in vivo studies demonstrated in both spontaneous and dexamethasone-induced models of NIDDM an increase in plasma A/I at the onset of hyperglycemia. In dexamethasone-induced diabetes, the increased A/I was associated with a high proamylin mRNA relative to proinsulin mRNA. We conclude that amylin and insulin expression and secretion rise in concert in compensated insulin-resistant states, but when hyperglycemia is present the increase in amylin exceeds that of insulin. Although a role of an increased A/I in the pathogenesis of NIDDM has not been established directly, these studies indicate that such a role could be possible.