CHLOROTETRACYCLINE‐ASSOCIATED FLUORESCENCE CHANGES DURING CALCIUM UPTAKE AND RELEASE BY RAT BRAIN SYNAPTOSOMES¹

Abstract

Abstract– The fluorescent divalent metal chelate‐probe, chlorotetracycline (CTC), was used as a dynamic monitor of calcium association with rat brain snynaptosomes. The determined fluorescence excitation and emission maxima, 412 nm and 522 nm respectively, were used to monitor membrane‐calcium interactions as a function of various parameters. Positive correlations were observed between increased or decreased fluorescence quantum yield and the uptake of both CTC and 45 Ca by synpatosomes. The divalent metal ionophore A23187 enhanced fluorescence as well as probe and 45 Ca uptake. Whereas, the polar chelator, EGTA, markedly reduced fluorescence, and the synaptosomal bound CTC and 45 Ca. The CTC fluorescence changes also demonstrated the saturable manner in which 45 Ca bound synaptosomes. At concentrations greater than 100μg/ml, CTC bound to the synaptosomes in a manner which quenched fluorescence at 522 nm. Also, CTC, at concentrations above 15 μg/ml, enhanced the uptake of 45 Ca. At CTC concentrations between 10 and 15 μg/ml the quenching and iono‐phoretic properties of the probe were minimized without affecting the capability of using the probe to visualize calcium interactions with synaptosomal membranes. Also, at a low CTC concentration (12.5 μg/ml) the inhibition of calcium uptake by increasing monovalent ion concentrations was clearly demonstrated.