Quantitative histofluorescence of hippocampal mossy fiber zinc

N-(6-methoxy-8-quinolyl)-p-toluenesulfonamide (TS-Q) is a fluorescent probe for histochemically detecting a chelatable pool of zinc in the hippocampal formation and other brain regions. Utilization of zinc:TS-Q histofluorescence as a quantitative tool required further characterization of zinc:TS-Q fluorescence and the development of appropriate zinc standards. Studies of zinc:TS-Q fluorescence in aqueous solutions were conducted to optimize the reaction conditions for TS-Q chelation with zinc. Possible interference of zinc:TS-Q fluorescence by chelation of other biologically significant cations with zinc:TS-Q was shown to be negligible. Zinc standards were developed using a water soluble plastic embedding medium. Varying the concentration of zinc in the plastic sections yielded a standard curve exhibiting a log-linear relationship between 0.15 and 11 fg zinc/100 μm². Zinc:TS-Q histofluorescence was characterized in horizontal sections of rat ventral hippocampal formation using a computerized image analysis system. Section thickness and TS-Q concentration were optimized. The concentration of zinc varied between 1.5 and 17 fg/100 μm² in different regions of the mossy fiber projection of the ventral hippocampal formation. The development of this quantitative histofluorescence technique should facilitate our understanding of the functional role of zinc in the mossy fiber projection and can be utilized to investigate changes in mossy fiber zinc in a variety of pathological states.