Fluorophilia: Fluorophore-containing compounds adhere non-specifically to injured neurons

Ionic (free) zinc (Zn2 +) is implicated in apoptotic neuronal degeneration and death. In our attempt to examine the effects of Zn2 + in neurodegeneration following brain injury, we serendipitously discovered that injured neurons bind fluorescein moieties, either alone or as part of an indicator dye, in histologic sections. This phenomenon, that we have termed “fluorophilia”, is analogous to the ability of degenerating neuronal somata and axons to bind silver ions (argyrophilia — the basis of silver degeneration stains). To provide evidence that fluorophilia occurs in sections of brain tissue, we used a wide variety of indicators such as Fluoro-Jade (FJ), a slightly modified fluorescein sold as a marker for degenerating neurons; Newport Green, a fluorescein-containing Zn2 + probe; Rhod-5N, a rhodamine-containing Ca2 + probe; and plain fluorescein. All yielded remarkably similar staining of degenerating neurons in the traumatic brain-injured tissue with the absence of staining in our sham-injured brains. Staining of presumptive injured neurons by these agents was not modified when Zn2 + in the brain section was removed by prior chelation with EDTA or TPEN, whereas staining by a non-fluorescein containing Zn2 + probe, N-(6-methoxy-8-quinolyl)-p-toluenesulfonamide (TSQ), was suppressed by prior chelation. Thus, certain fluorophore-containing compounds nonspecifically stain degenerating neuronal tissue in histologic sections and may not reflect the presence of Zn2 +. This may be of concern to researchers using indicator dyes to detect metals in brain tissue sections. Further experiments may be advised to clarify whether Zn2 +-binding dyes bind more specifically in intact neurons in culture or organotypic slices.

► We examined non-specific staining of injured hippocampal neurons.
► Multiple ion indicator dyes and neurodegeneration stains were used.
► We used two models of injury: trauma and chemically-induced seizures.