Fluorescence response in chlortetracycline-loaded neutrophils measures release of Ca2+ from intracellular membrane enclosed storage sites.

Chlortetracycline complexes with di- and trivalent cations resulting in an enhancement of its fluorescence emission intensity. Rabbit peritoneal neutrophils loaded with chlortetracycline gave a fluorescence response, even in the absence of extracellular Ca2+ and Mg2+, by a decrease in fluorescence intensity. The shift in the fluorescence emission maximum to lower wavelengths after the response suggested the response to be due to Ca2+ and not Mg2+ flux. The response was elicited by three mechanisms--a receptor-mediated mechanism by the chemotactic peptide, an ionophore-mediated one by lasalocid, and a detergent-mediated response by digitonin. These observations indicated that the response was due to transport of calcium across membranes in the intracellular compartments and may be physiologically significant. Whereas extracellular Ca2+ did not significantly affect the chemotactic peptide and lasalocid-mediated responses, Ca2+ inhibited the digitonin-mediated responses in a dose-dependent manner possibly due to extracellular Ca2+ flooding the cytosol through the digitonin-permeabilized plasma membrane and equilibrating the Ca2+ gradient across the intracellular membranes. The data collectively indicate that the fluorescence response is due to release of Ca2+ across intracellular membranes from a Ca2+ storage site into the cytosol.