LEUKOTRIENE D-4-INDUCED CA2-CELLS( MOBILIZATION IN EHRLICH ASCITES TUMOR)

Stimulation of Ehrlich ascites tumor cells with leukotriene D-4 (LTD(4 )) within the concentration range 1-100 nM leads to a concentration-de pendent, transient increase in the intracellular, free Ca2+ concentrat ion, [Ca2+](i). The Ca2+ peak time, i.e., the time between addition of LTD, and the highest measured [Ca2+](i) value, is in the range 0.20 t o 0.21 min in ten out of fourteen independent experiments. After addit ion of a saturating concentration of LTD(4) (100 nM), the highest meas ured increase in [Ca2+](i) in Ehrlich cells suspended in Ca2+-containi ng medium is 260+/-14 nM and the EC(50) value for LTD(4)-induced Ca2mobilization is estimated at 10 nM. Neither the peptido-leukotrienes L TC(4) and LTE(4) nor LTB(4) are able to mimic or block the LTD(4)-indu ced Ca2+ mobilization, hence the receptor is specific for LTD(4). Remo val of Ca2+ from the experimental buffer significantly reduces the siz e of the LTD(4)-induced increase in [Ca2+](i). Furthermore, depletion of the intracellular Ins(1,4,5)P-3-sensitive Ca2+ stores by addition o f the ER-Ca2+-ATPase inhibitor thapsigargin also reduces the size of t he LTD(4)-induced increase in [Ca2+](i) in Ehrlich cells suspended in Ca2+-containing medium, and completely abolishes the LTD(4)-induced in crease in [Ca2+](i) in Ehrlich cells suspended in Ca2+-free medium con taining EGTA. Thus, the LTD(4)-induced increase in [Ca2+](i) in Ehrlic h cells involves an influx of Ca2+ from the extracellular compartment as well as a release of Ca2+ from intracellular Ins(1,4,5)P-3-sensitiv e stores. The Ca2+ peak times for the LTD(4)-induced Ca2+ influx and f or the LTD(4)-induced Ca2+ release are recorded in the time range 0.20 to 0.21 min in four out of five experiments and in the time range 0.3 4 to 0.35 min in six out of eight experiments, respectively. Stimulati on with LTD(4) also induces a transient increase in Ins(1,4,5)P-3 gene ration in the Ehrlich cells, and the Ins(1,4,5)P-3 peak time is record ed in the time range 0.27 to 0.30 min. Thus, the Ins(1,4,5)P-3 content seems to increase before the LTD(4)-induced Ca2+ release from the int racellular stores but after the LTD(4)-induced Ca2+ influx. Inhibition of phospholipase C by preincubation with U73122 abolishes the LTD(4)- induced increase in Ins(1,4,5)P-3 as well as the LTD(4)-induced increa se in [Ca2+](i), indicating that a U73122-sensitive phospholipase C is involved in the LTD(4)-induced Ca2+ mobilization in Ehrlich cells. Th e LTD(4)-induced Ca2+ influx is insensitive to verapamil, gadolinium a nd SK&F 96365, suggesting that the LTD(4)-activated Ca2+ channel in Eh rlich cells is neither voltage gated nor stretch activated and most pr obably not receptor operated. In conclusion, LTD(4) acts in the Ehrlic h cells via a specific receptor for LTD(4), which upon stimulation ini tiates an influx of Ca2+, through yet unidentified Ca2+ channels, and an activation of a U73122-sensitive phospholipase C, Ins(1,4,5)P-3 for mation and finally release of Ca2+ from the intracellular Ins(1,4,5)P- 3-sensitive stores.