title: In vivo EPR pharmacokinetic evaluation of the redox status and the blood brain barrier permeability in the SOD1G93AALS rat model.
abstract: Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder affecting the motor pathways of the central nervous system. Although a number of pathophysiological mechanisms have been described in the disease, post mortem and animal model studies indicate blood-brain barrier (BBB) disruption and elevated production of reactive oxygen species as major contributors to disease pathology. In this study, the BBB permeability and the brain tissue redox status of the SOD1G93AALS rat model in the presymptomatic (preALS) and symptomatic (ALS) stages of the disease were investigated by in vivo EPR spectroscopy using three aminoxyl radicals with different cell membrane and BBB permeabilities, Tempol, 3-carbamoyl proxyl (3CP), and 3-carboxy proxyl (3CxP). Additionally, the redox status of the two brain regions previously implicated in disease pathology, brainstem and hippocampus, was investigated by spectrophotometric biochemical assays. The EPR results indicated that among the three spin probes, 3CP is the most suitable for reporting the intracellular redox status changes, as Tempol was reduced in vivo within minutes (t1/2=2.0±0.5min), thus preventing reliable kinetic modeling, whereas 3CxP reduction kinetics gave divergent conclusions, most probably due to its membrane impermeability. It was observed that the reduction kinetics of 3CP in vivo, in the head of preALS and ALS SOD1G93Arats was altered compared to the controls. Pharmacokinetic modeling of 3CP reduction in vivo, revealed elevated tissue distribution and tissue reduction rate constants indicating an altered brain tissue redox status, and possibly BBB disruption in these animals. The preALS and ALS brain tissue homogenates also showed increased nitrilation, superoxide production, lipid peroxidation and manganese superoxide dismutase activity, and a decreased copper-zinc superoxide dismutase activity. The present study highlights in vivo EPR spectroscopy as a reliable tool for the investigation of changes in BBB permeability and for the unprecedented in vivo monitoring of the brain tissue redox status, as early markers of ALS.
authors.affiliation: University of Belgrade - Faculty of Biology, Center for Laser Microscopy, Studentski trg 3, 11158 Belgrade, Serbia.
authors.forename: Stefan
authors.initials: S
authors.lastname: Stamenković
authors.affiliation: University of Belgrade - Faculty of Physical Chemistry, EPR Laboratory, Studentski trg 12-16, 11158 Belgrade, Serbia.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Pavićević
authors.affiliation: University of Belgrade - Faculty of Physical Chemistry, EPR Laboratory, Studentski trg 12-16, 11158 Belgrade, Serbia.
authors.forename: Miloš
authors.initials: M
authors.lastname: Mojović
authors.affiliation: University of Belgrade - Faculty of Physical Chemistry, EPR Laboratory, Studentski trg 12-16, 11158 Belgrade, Serbia.
authors.forename: Ana
authors.initials: A
authors.lastname: Popović-Bijelić
authors.affiliation: Institute for Medical Research, Military Medical Academy, Crnotravska 17, 11000 Belgrade, Serbia.
authors.forename: Vesna
authors.initials: V
authors.lastname: Selaković
authors.affiliation: University of Belgrade - Faculty of Biology, Center for Laser Microscopy, Studentski trg 3, 11158 Belgrade, Serbia. Electronic address: pandjus@bio.bg.ac.rs.
authors.forename: Pavle
authors.initials: P
authors.lastname: Andjus
authors.affiliation: University of Belgrade - Faculty of Physical Chemistry, EPR Laboratory, Studentski trg 12-16, 11158 Belgrade, Serbia.
authors.forename: Goran
authors.initials: G
authors.lastname: Bačić
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D000818:Animals; D001812:Blood-Brain Barrier / Q000473:pathology*; D001921:Brain / Q000502:physiology*; D003497:Cyclic N-Oxides / Q000493:pharmacokinetics; D004195:Disease Models, Animal; D018450:Disease Progression; D004578:Electron Spin Resonance Spectroscopy; D006801:Humans; D009154:Mutation / Q000235:genetics; D010084:Oxidation-Reduction; D051381:Rats; D055647:Rats, Transgenic; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013113:Spin Labels; D000072105:Superoxide Dismutase-1 / Q000235:genetics / Q000378:metabolism*
chemical_list: D003497:Cyclic N-Oxides; D017382:Reactive Oxygen Species; C000606290:SOD1 protein, human; D013113:Spin Labels; D000072105:Superoxide Dismutase-1; C001803:tempol
keywords: ALS; Blood-brain barrier; EPR; Pharmacokinetics of nitroxides; Redox status; SOD1(G93A) rat