Metabolically active rat brain slices as a model to study the regulation of protein phosphorylation in mammalian brain

Abstract

The reversible protein phosphorylation is the most important cellular regulation of the biological functions of many proteins. Disregulation of protein phosphorylation is involved in pathogeneses of several human diseases. The abnormal hyperphosphorylation of microtubule-associated protein tau and its aggregation into neurofibrillary tangles in selective neurons is one of the major brain pathologies of Alzheimer's disease and several other related neurodegenerative diseases. Here we present metabolically competent rat brain slices as a model to study the regulation of protein phosphorylation in brain. Employing this model we have been able to study the abnormal hyperphosphorylation of tau and other microtubule-associated proteins. We have evaluated the activity and intactness of the rat brain slices both biochemically and morphologically. Selective inhibition of protein phosphatase 2A in these rat brain slices by the treatment with okadaic acid induced hyperphosphorylation of tau at many abnormal sites seen in Alzheimer's disease brain and the accumulation of hyperphosphorylated tau in pyramidal neurons of the cortex and hippocampus. The regulation of the phosphorylation of high-molecular-weight microtubule-associated protein, MAP1b, was also studied with this model. This model enables studies on the regulation of protein phosphorylation not only biochemically, but also histochemically and immunocytochemically. Furthermore, unlike cultured cells, the neurons in the brain slices reside in the physiological environment of the brain consisting of natural extracellular matrix, neuronal connectivity, and neuronal-glial interactions.