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b-Amyloid protein is the major component of senile plaques found in the brains of Alzheimer's patients. Previously, a new biochemical property of amyloid, its ability to disrupt ester and peptide bonds, was described [Elbaum, D., Brzyska, M., Bacia, A. & Alkon, D. (2000) Biochem. Biophys. Res. Commun. 267, 733±738]. In the present work we compare the ability of b-amyloid to hydrolyse and oxidize model fluorescent derivatives of dichlorofluorescein [dichlorodihydrofluorescein (H 2 DCF) or dichlorofluorescein diacetate (DCF-DA), respectively] to the same final product (dichlorofluorescein). Although there is accumulating evidence of oxidative properties of b-amyloid, little is known about its hydrolytic abilities. Chemical modification studies revealed that hydrolytic properties are related to a His, Ser and Asp/Glu triad, while residues of His, Tyr and Met are involved in the oxidative activity of amyloid. Studies with the rat homologue of human b-amyloid (1±40), containing three amino-acid substitutions (Arg53Gly, Tyr103Phe and His133Arg) confirmed a role of His in the studied processes. Reduction of the hydrolysis product caused by inhibitors of Ser esterases (phenylmethylsulphonyl fluoride and eserine) suggests that b-amyloid-mediated hydrolysis is Ser sensitive. Antioxidants and metal chelators that reduced H 2 DCF oxidation did not change or increase DCF-DA hydrolysis. Solvent isotope effects suggest the involvement of hydrogen bonds in the hydrolysis reaction. Hydrolysis was inhibited by redox-active metal ions and was practically oxygen independent while the oxidation process was redox-active-metal enhanced [Cu(II) and Fe(II) primarily], and oxygen dependent. Product formation was significantly inhibited by catalase and superoxide dismutase as well as benzoquinone, a specific superoxide anion radical scavenger. Increase of fluorescence by oxidation was strongly inhibited by azide and His and enhanced in samples prepared with deuterated phosphate buffer, suggesting singlet oxygen intermediacy. Our data are consistent with superoxide-mediated singlet oxygen intermediate in this Fenton mechanism-driven reaction. These results indicate that hydrolytic and oxidative properties of b-amyloid are distinct features of this peptide and probably require different mechanisms to occur, but both of them may contribute to b-amyloid toxicity.Keywords: amino acid; b-amyloid; hydrolysis; metal; oxidation.Alzheimer's disease is a progressive neurodegenerative disorder affecting a large proportion of the ageing population. It is characterized by gradual loss of cognitive function and synaptic integrity, selective neuronal death and the abnormal formation of neuritic and core plaques in the cerebral cortex. The major proteinaceous component of the extracellular amyloid deposits is the b-amyloid peptide, consisting of 39±43 amino acids, which is produced by proteolytic processing of its precursor, the b-amyloid precursor protein (APP). This is a 70-kDa transmembrane protein, expressed in various cell types, including neural cel...

The adult CNS has the capacity to remyelinate following metabolic, toxic and autoimmune demyelinating insults. In cuprizone-induced demyelination, spontaneous remyelination occurs after the cessation of cuprizone diet. We used the cuprizone model to investigate the role of glial K(+) channels in oligodendroglial (OLG) regeneration and remyelination in vivo. We found that treatment with 4-aminopyridine (4-AP), a broad-spectrum K(+) channel antagonist, results in: (1) decreased number of oligodendroglial progenitors (OP) and OLGs; (2) diminished astrogliosis; and (3) decreased remyelination in the corpus callosum based on the immunoreactivity to myelin basic protein (MBP), Rip monoclonal antibody, and by electron microscopy. Our findings support the concept that glial K(+) channels play an important role during OLG regeneration and remyelination, a crucial factor to be considered during the development of therapeutic strategies to facilitate recovery in demyelinating diseases and spinal cord injury.

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