Continuous inhibition of secretase regulates synapse formation

Abstract

Alzheimer's disease (AD) is a progressive neurodegenerative brain disorder and the most common cause of dementia in the elderly. An excessive accumulation of amyloid β peptide (Aβ), derived from amyloid precursor protein (APP). APP is sequentially cleaved by membrane bound protease called α-, β-and γ-secretase, which is being explored as a target for therapy and prevention of AD. Although previous reports indicated that these secretases have a significant role in development, these functions in developed neurons and synapses are still largely unknown.We investigated the roll of these secretase in synaptic formation and maturation using organotypic hippocampal slice cultures because they have advantages over both in vivo and in vitro platforms. The hippocampal slices were obtained from anaesthetized rats at postnatal day 7-8 and placed on a polytetrafluoroethylene membrane filter, and culture medium (50% MEM, 25% HBSS, 25% horse serum) was added up to the bottom surface of the filter. For the analysis of synapse formation and maturation, we evaluated the expression level of synaptic proteins and synaptic activity. In this study, we utilized hippocampal slice cultures as an ex vivo model that enabled continuous and long-term analysis for secretases. Furthermore, we suggested that 3-20 days in vitro were a critical period of synapse formation and maturation in hippocampal slice cultures. During the period of synapse formation and maturation, pharmacological inhibition of secretases changed the expression levels of synaptic proteins. After synaptic maturation, secretase inhibition did not affect the synaptic protein, except for APP. Our results and previous reports indicate that secretase activity is important for synaptogenesis and maturation. Furthermore, we suggest that cultured hippocampal slices are useful tools for understanding physiological functions of membrane associated protease. This report is the first step toward ex vivo continuous analysis of APP and secretases.