Exploring the Zinc Metallome of Cultured Cortical Neurons Using Synchroton Radiation X-Ray Fluorescence Microscopy

Abstract

Zinc is abundant in the brain, and dysregulation of zinc ion homeostasis has been implicated in Alzheimer’s disease pathology and neuronal death after brain ischemia. Many studies have established that cytosolic free zinc ion concentrations are much lower than measured total average cellular concentrations, suggesting a large capacity for intracellular zinc buffering. It is generally believed that protein binding constitutes a major source of intracellular zinc ion buffering. In this study, we explored the subcellular distribution of zinc-binding sites in neurons using synchrotron radiation X-ray fluorescence microscopy. We observed that nearly all the intracellular zinc in resting neurons is bound and that zinc-binding sites were uniformly distributed in neurons resulting in similar average zinc concentrations (≈300 μM) throughout the cell body including the nucleus, peri-nuclear region, and processes. However, sites of elevated zinc concentration were observed in resting neurons, and were prevalent throughout the neuron (including the cell body and processes) when neurons were loaded with zinc. These sites of elevated zinc could represent either zinc bound to proteins or zinc sequestered in subcellular organelles. Additional studies are needed to completely describe the neuronal zinc metallome and the function of zinc-binding proteins in buffering zinc loads.