Inhibition or attenuation of mucous hypersecretion in middle ear epithelium is a key step toward resolution of mucoid otitis media. Mucous hypersecretion induced by platelet-activating factor (PAF) in cultured Chinchilla middle ear epithelial cells is dependent on arachidonic acid metabolites via PAF receptors, suggestive of the role of phospholipase A2 (PLA2) in mucous glycoprotein (MGP) secretion. In this study, dexamethasone added to cultured Chinchilla middle ear epithelial cells inhibited baseline and PAF-induced MGP secretion in a concentration-dependent manner. A definite time lag (16 h) was observed between administration of dexamethasone and MGP inhibition. This inhibition was reversed by the addition of exogenous PLA2 (the rate-limiting enzyme of arachidonic acid metabolism) and actinomycin D (an inhibitor of mRNA synthesis). This suggests that dexamethasone inhibits baseline and PAF-induced MGP secretion via a PLA2-dependent mechanism.