Leukotriene D4-induced increases in cytosolic calcium in THP-1 cells: dependence on extracellular calcium and inhibition with selective leukotriene D4 receptor antagonists.
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Agonist-induced changes in intracellular calcium ion concentration ([Ca++]i) were examined in human monocytic leukemia THP-1 cells loaded with fura 2/acetoxymethyl ester (fura 2/AM). Leukotriene (LT)D4 induced a concentration-dependent biphasic response consisting of a transient phase (up to 5-fold peak increase) followed by a sustained phase, showing characteristics of a receptor-operated calcium channel. Homologous desensitization to LTD4 was observed. The responses to LTD4 were reduced by 80 to 90% in calcium-free buffer. The responses to LTD4 in a calcium-free buffer were dependent upon the duration of prior exposure of the cells to a calcium-free environment. The response at 30 or 60 min after exposure to calcium-free buffer was greater than that at earlier time points (time-dependent sensitization). Similar responses were obtained with THP-1 cells exposed to EDTA-containing buffer. It is speculated that such time-dependent sensitization is a result of changes at the receptor level. The responses to LTD4 were blocked by two specific LTD4 antagonists, MK-0571 and ICI-204,219, in a concentration-dependent manner. When given after addition of LTD4, MK-0571 or ICI-204,219 reversed the sustained phase of the LTD4-induced response, suggesting that maintenance of the response requires persistent activation of the LTD4 receptor. ICI-204,219 was 5 to 10 times more potent than MK-0571 (IC50 values of 1.1 and 9.3 nM, respectively), in agreement with results from radioligand binding studies reported separately.
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