ROLE OF EXTRACELLULAR CALCIUM IN IN-VITRO UPTAKE AND INTRAPHAGOCYTIC LOCATION OF MACROLIDES

We compared the uptakes and intracellular locations of four 14-membere d-ring macrolides (roxithromycin, dirithromycin, erythromycin, and ery thromycylamine) in human polymorphonuclear neutrophils (PMNs) in vitro , Intracellular location was assessed by cell fractionation and uptake kinetics in cytoplasts (granule-poor PMNs), Trapping of dirithromycin within PMN granules (up to 80% at 30 min) was significantly more mark ed than the intracellular trapping of the other drugs (erythromycylami ne, 45% +/- 5.1%; erythromycin, 42% +/- 3.7%; roxithromycin, 35% +/- 3 .0%), A new finding was that, in the absence of extracellular calcium, the uptakes of all of the macrolides by PMNs and cytoplasts were sign ificantly impaired, by about 50% (PMN) and 90% (cytoplasts), Furthermo re, inorganic Ca2+ channel blockers inhibited macrolide uptake in a co ncentration-dependent manner, with 50% inhibitory concentrations of 1. 6 to 2.0 mM and 29 to 35 mu M, respectively, for Ni2+ and La3+. The in tracellular distributions of the drugs were unchanged in the presence of Ni2+ and La3+ and in Ca2+-free medium supplemented with ethylene gl ycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. The organi c Ca2+ channel blocker nifedipine had no effect on macrolide uptake, w hereas verapamil inhibited it in a time- and concentration-dependent m anner. These data show the importance of extracellular Ca2+ in macroli de uptake by phagocytes and suggest a link with Ca2+ channels or a Ca2 + channel-operated mechanism.