title: On the nature of the Cu-rich aggregates in brain astrocytes.
abstract: Fulfilling a bevy of biological roles, copper is an essential metal for healthy brain function. Cu dyshomeostasis has been demonstrated to be involved in some neurological conditions including Menkes and Alzheimer's diseases. We have previously reported localized Cu-rich aggregates in astrocytes of the subventricular zone (SVZ) in rodent brains with Cu concentrations in the hundreds of millimolar. Metallothionein, a cysteine-rich protein critical to metal homeostasis and known to participate in a variety of neuroprotective and neuroregenerative processes, was proposed as a binding protein. Here, we present an analysis of metallothionein(1,2) knockout (MTKO) mice and age-matched controls using X-ray fluorescence microscopy. In large structures such as the corpus callosum, cortex, and striatum, there is no significant difference in Cu, Fe, or Zn concentrations in MTKO mice compared to age-matched controls. In the astrocyte-rich subventricular zone where Cu-rich aggregates reside, approximately 1/3 as many Cu-rich aggregates persist in MTKO mice resulting in a decrease in periventricular Cu concentration. Aggregates in both wild-type and MTKO mice show XANES spectra characteristic of CuxSymultimetallic clusters and have similar [S]/[Cu] ratios. Consistent with assignment as a CuxSymultimetallic cluster, the astrocyte-rich SVZ of both MTKO and wild-type mice exhibit autofluorescent bodies, though MTKO mice exhibit fewer. Furthermore, XRF imaging of Au-labeled lysosomes and ubiquitin demonstrates a lack of co-localization with Cu-rich aggregates suggesting they are not involved in a degradation pathway. Overall, these data suggest that Cu in aggregates is bound by either metallothionein-3 or a yet unknown protein similar to metallothionein.
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Brendan
authors.initials: B
authors.lastname: Sullivan
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Gregory
authors.initials: G
authors.lastname: Robison
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Jenna
authors.initials: J
authors.lastname: Osborn
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Martin
authors.initials: M
authors.lastname: Kay
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Peter
authors.initials: P
authors.lastname: Thompson
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Katherine
authors.initials: K
authors.lastname: Davis
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States.
authors.forename: Taisiya
authors.initials: T
authors.lastname: Zakharova
authors.affiliation: BioCAT, Advanced Photon Source, Argonne National Laboratory, 9700 S. Cass Ave., Argonne, IL 60439, United States; XSD, Argonne National Laboratory, 9700 S. Cass Ave., Argonne, IL 60439, United States.
authors.forename: Olga
authors.initials: O
authors.lastname: Antipova
authors.affiliation: Department of Physics and Astronomy, Purdue University, 525 Northwestern Ave., West Lafayette, IN 47907, United States. Electronic address: ypushkar@purdue.edu.
authors.forename: Yulia
authors.initials: Y
authors.lastname: Pushkar
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000378:metabolism* / Q000648:ultrastructure; D015415:Biomarkers / Q000378:metabolism; D002413:Cations, Divalent; D002540:Cerebral Cortex / Q000378:metabolism / Q000648:ultrastructure; D011232:Chemical Precipitation; D003300:Copper / Q000737:chemistry*; D003337:Corpus Callosum / Q000378:metabolism / Q000648:ultrastructure; D003342:Corpus Striatum / Q000378:metabolism / Q000648:ultrastructure; D017353:Gene Deletion; D015870:Gene Expression; D007501:Iron / Q000737:chemistry; D020547:Lateral Ventricles / Q000378:metabolism* / Q000648:ultrastructure; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism / Q000648:ultrastructure; D008668:Metallothionein / Q000172:deficiency* / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D061848:Optical Imaging; D051381:Rats; D017207:Rats, Sprague-Dawley; D013052:Spectrometry, X-Ray Emission; D025801:Ubiquitin / Q000235:genetics / Q000378:metabolism; D015032:Zinc / Q000737:chemistry; D052118:Lysosomal-Associated Membrane Protein 1
chemical_list: D015415:Biomarkers; D002413:Cations, Divalent; D003300:Copper; D007501:Iron; D051907:Lysosomal Membrane Proteins; D008668:Metallothionein; D025801:Ubiquitin; D015032:Zinc; C052255:Lamp1 protein, mouse; D052118:Lysosomal-Associated Membrane Protein 1
keywords: Cu; Metallothionein; Subventricular zone; X-ray fluorescence microscopy