Inhibitory action of linoleamide and oleamide toward sarco/endoplasmic reticulum Ca(2+)-ATPase.

Abstract Background SERCA maintains intracellular Ca 2+ homeostasis by sequestering cytosolic Ca 2+ into SR/ER stores. Two primary fatty acid amides (PFAAs), oleamide (18:1 9- cis ) and linoleamide (18:2 9,12- cis ), induce an increase in intracellular Ca 2+ levels, which might be caused by their inhibition of SERCA. Methods Three major SERCA isoforms, rSERCA1a, hSERCA2b, and hSERCA3a, were individually overexpressed in COS-1 cells, and the inhibitory action of PFAAs on Ca 2+ -ATPase activity of SERCA was examined. Results The Ca 2+ -ATPase activity of each SERCA was inhibited in a concentration-dependent manner strongly by linoleamide (IC 50 15–53 μM) and partially by oleamide (IC 50 8.3–34 μM). Inhibition by other PFAAs, such as stearamide (18:0) and elaidamide (18:1 9- trans ), was hardly or slightly observed. With increasing dose, linoleamide decreased the apparent affinity for Ca 2+ and the apparent maximum velocity of Ca 2+ -ATPase activity of all SERCAs tested. Oleamide also lowered these values for hSERCA3a. Meanwhile, oleamide uniquely reduced the apparent Ca 2+ affinity of rSERCA1a and hSERCA2b: the reduction was considerably attenuated above certain concentrations of oleamide. The dissociation constants for SERCA interaction varied from 6 to 45 μM in linoleamide and from 1.6 to 55 μM in oleamide depending on the isoform. Conclusions Linoleamide and oleamide inhibit SERCA activity in the micromolar concentration range, and in a different manner. Both amides mainly suppress SERCA activity by lowering the Ca 2+ affinity of the enzyme. General significance Our findings imply a novel role of these PFAAs as modulators of intracellular Ca 2+ homeostasis via regulation of SERCA activity.