Abstract Using α-linolenic acid (ALA), one of several polyunsaturated fatty acids (PUFAs) that have previously been shown to both mobilize intracellular Ca 2+ from the inositol 1,4,5-trisphosphate (IP 3)-sensitive Ca 2+ pool independently of IP 3 production  and inhibit Ca 2+ influx , the relationship between Ca 2+ mobilization from intracellular stores and Ca 2+ influx in T cells (JURKAT) was studied. JURKAT cells were treated with 30 μM ALA to deplete the IP 3-sensitive Ca 2+ pool. When the intracellular free Ca 2+ concentration ([Ca 2+] i) returned to basal level, fatty acid free bovine serum albumin (BSA) was added to remove extracellular and membrane bound ALA. This resulted in a sustained increase in [Ca 2+] i in the absence of inositol phosphates' formation. This sustained increase in [Ca 2+] i was insensitive to protein kinase C activation but was inhibited by Ni 2+ ions. The extent of Ca 2+ inflex was found to be correlated to the amount of Ca 2+ initially discharged from the IP 3-sensitive Ca 2+ pool by sub-optimal concentrations of ALA. Ligation of the CD3 complex or the T cell antigen receptor with an anti-CD3 antibody (OKT3) during the sustained [Ca 2+] i increase (induced by a sub-optimal concentration of ALA), produced a greater response. No increase in the sustained response was observed when the CD3 complex was activated in cells pretreated with an optimal concentration of ALA. In summary, Ca 2+ entry in T cells is activated by emptying of the IP 3-sensitive Ca 2+ pool which can be dissociated from inositol phosphate production. The rate of Ca 2+ influx appears to be closely correlated to the initial discharge of Ca 2+ from the IP 3-sensitive Ca 2+ pool, suggesting that Ca 2+ may first enter the depleted pool and then is released into the cytosol.