ARACHIDONATE AND OTHER FATTY-ACIDS MOBILIZE CA2+ IONS AND STIMULATE BETA-GLUCURONIDASE RELEASE IN A CA2+-DEPENDENT FASHION FROM UNDIFFERENTIATED HL-60 CELLS

Arachidonate (1-300 mu M) mobilized Ca2+ ions from an intracellular store and stimulated the entry of Ca2+ ions from the extracellular fluid in undifferentiated HL-60 cells that had been loaded with Fura-2. The integrated response was biphasic in farm: arachidonate liberated Ca2+ ions from the intracellular store first, resulting in a transient increase in cytosolic free Ca2+ concentration ([Ca2+](i)). Ca2+ entry from the extracellular fluid was not evident for a further 1-2 min, At baseline, [Ca2+](i) was 48.1 +/- 14.0 nM (SEM, n = 5), Upon addition of arachidonate (100 mu M), [Ca2+](i) rose to a transient peak level of 217 +/- 38.6 nM (SEM, n = 5) and a later plateau level of 427 +/- 118 nM (SEM, n = 5). Removal of added Ca2+ ions from the extracellular fluid in the presence of EGTA (1.0 mM) had no effect on the initial transient response but abolished the second phase of the response, In HL-60 cells that had been loaded with BAPTA/AM, the initial transient phase of the response was abolished but the elevation in [Ca2+](i) due to Ca2+ entry from the extracellular fluid was unaffected. Undifferentiated HL-60 cells also responded to arachidonate (100 mu M) with an increase in the release of the lysosomal enzyme beta-glucuronidase. Arachidonate-induced beta-glucuronidase release from BAPTA-loaded cells or in control cells exposed to Ca2+-free solutions was inhibited by about 50%. In BAPTA-loaded cells that were incubated with Ca2+-free solutions, arachidonate-induced beta-glucuronidase release was inhibited by about 90%. Leukotriene B-4 failed to elevate [Ca2+](i) in the concentration range 0.01-1 mu M and failed to activate beta-glucuronidase release in concentrations up to 10 mu M. Furthermore, the cyclo-oxygenase/lipoxygenase inhibitor ETYA (100 mu M) was without effect on secretion. Consistent with this finding, we found that a large number- of unsaturated fatty acids could reproduce the effect of arachidonate on [Ca2+](i) and beta-glucuronidase release. Fatty acids belonging to the omega-3, omega-6 and omega-7 unsaturated fatty acid families were effective in elevating [Ca2+](i) and stimulating beta-glucuronidase release. However, three unsaturated fatty acids, all belonging to the omega-9 fatty acid family, were ineffective.