THAPSIGARGIN ACTIVATES UNIVALENT-CATION AND BIVALENT-CATION ENTRY IN HUMAN NEUTROPHILS BY A SK-AND-F-96365-SENSITIVE AND GD3-SENSITIVE PATHWAY AND IS A PARTIAL SECRETAGOGUE - INVOLVEMENT OF PERTUSSIS-TOXIN-SENSITIVE G-PROTEINS AND PROTEIN PHOSPHATASE-1()PHOSPHATASE-2A AND PHOSPHATASE-2B IN THE SIGNAL-TRANSDUCTION PATHWAY/

The Ca2+-ATPase inhibitor thapsigargin (TG) activates bivalent-cation entry in human neutrophils via depletion of intracellular Ca2+ stores, bur little is known about the underlying mechanism and the functional role of TG-induced cation entry. We studied the effects of TG on univ alent- and bivalent-cation entry, lysozyme release and superoxide-anio n (O-2(-)) formation in human neutrophils. TG, like the chemotactic pe ptide, N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), stimulate d entry of Ca2+, Mn2+, Ba2+, Sr2+ and Na+ in a xyphenyl)propoxy]-4-met hoxyphenethyl}-1H-imidazole hydrochloride (SK&F 96365)- and Gd3+-sensi tive manner. The inhibitors of protein phosphatases 1/2A, calyculin A and okadaic acid, diminished TG-induced cation influxes, whereas the i nhibitors of protein phosphatase 2B, cyclosporin A and FK-506, were po tentiators. Pertussis toxin (PTX) partially inhibited the effects of T G on Ca2+ and Mn2+ entry. TG and fMLP activated inward currents with a linear current-voltage relationship and a reversal potential at about 0 mV. TG activated lysozyme release and potentiated fMLP-induced O-2( -) formation. TG-induced lysozyme release was inhibited by SK&F 96365, PTX and the removal of extracellular Ca2+ or Na+. Our data show that TG activates a non-selective and SK&F 96365- and Gd3+-sensitive cation entry pathway and is a partial secretagogue. TG-stimulated cation ent ry involves PTX-sensitive G-proteins and protein phosphatases, with pr otein phosphatases 1/2A and 2B playing opposite roles.