The endocannabinoid anandamide (Arachidonylethanolamide, AEA) is known to induce apoptosis in a wide variety of nucleated cells. The present study explored whether anandamide induces suicidal death of erythrocytes or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine exposure at the erythrocyte surface. Eryptotic cells are phagocytosed and thus cleared from circulating blood. Triggers of eryptosis include increase of cytosolic Ca2+ activity, formation of PGE(2), oxidative stress and excessive cell shrinkage. Erythrocyte Ca2+ activity was estimated from Fluo3 fluorescence, phosphatidylserine exposure from annexin V binding, and erythrocyte volume from forward scatter in FACS analysis. Exposure of erythrocytes to anandamide (= 2.5 microM) increased cytosolic Ca2+ activity, enhanced the percentage of annexin V binding erythrocytes and decreased erythrocyte forward scatter, effects significantly blunted in the presence of cycloxygenase inhibitors acetylsalicylic acid (50 microM) or ibuprofen (100 microM) and in the nominal absence of extracellular Ca2+. Anandamide further enhanced the stimulating effects of hypertonic (addition of 550 mM sucrose) or isotonic (isosmotic replacement of Cl- with gluconate) cell shrinkage on annexin V binding. The present observations demonstrate that anandamide increases cytosolic Ca2+ activity, thus leading to cell shrinkage and cell membrane scrambling of mature erythrocytes.