Lectins from Agaricus bisporus and Agaricus campestris stimulate insulin and glucagon release from isolated rat islets in the presence of 2 mM glucose. In the case of insulin release, maximal stimulation was observed at lectin concentrations above 58 μg. per milliliter (approximately 1 μM). A. bisporus PHA B stimulated insulin release was independent of a source of metabolic energy but was abolished by deuterium oxide. The lectin did not alter islet glucose oxidation to CO2 or incorporation of [3H] leucine into trichloracetic acid precipitable material nor did it modify rates of insulin secretion induced by 20 mM glucose. None of nine other lectins tested stimulated insulin release, whereas stimulation of fat cell glucose was a general property of the lectins. Binding of 125I labeled A. bisporus PHA B to islets increased with time up to one hour and after attainment of equilibrium was very slowly reversible. Binding was directly proportional to islet number and the estimated K(diss) of the binding reaction was 17μg per milliliter. The total number of A. bisporus PHA B binding sites per islet was approximately 2 x 1010. Binding of A. bisporus PHA B to the islets and A. bisporus PHA B stimulated insulin release were inhibited in parallel by a glycopeptide containing the oligosaccharide receptor for the lectin, suggesting that lectin binding is essential for the expression of insulin releasing activity. It is proposed that the specific interaction between mushroom lectin and its receptors may lead to conformational changes in the structure of the membranes of the islet A2 and B cells that facilitate exocytosis.