The aim of this in vivo microdialysis study was to characterise the regulation of prostaglandin D2 (PgD2) synthesis by NMDA receptors in the rabbit hippocampus in relation to changes in extracellular Ca2+ concentration ([Ca2+]e) and nitric oxide (NO) levels. Samples of dialysate were analysed for changes in PgD2 concentration, in [Ca2+]e and in the level of NO. The results demonstrated that a 20-min pulse application of 0.1 - 2.5 mM NMDA via a microdialysis probe induced a prolonged stimulation of PgD2 release that was sensitive to competitive NMDA receptor antagonists. An inhibitor of voltage-sensitive Na+ channels, tetrodotoxin, did not influence this effect but significantly suppressed basal PgD2 production, whereas a NOS inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME), prevented NMDA-evoked NO release and inhibited NMDA-induced PgD2 release in an L-arginine-sensitive manner. NO donors, S-nitroso-N-acetylpenicillamine and sodium nitroprusside, stimulated PgD2 release. NMDA-evoked decrease in [Ca2+]e was insensitive to TTX and L-NAME. These results demonstrate an in vivo NMDA receptor-mediated modulation of PgD2 synthesis in the brain, in which NO participates.