Nonspecific Interaction of Prefibrillar Amyloid Aggregates with Glutamatergic Receptors Results in Ca2+ Increase in Primary Neuronal Cells

Francesca Pellistri,Monica Bucciantini,Annalisa Relini,Daniele Nosi,Alessandra Gliozzi,Mauro Robello,Massimo Stefani

doi:10.1074/jbc.m803992200

Abstract

It is widely reported that the Ca(2+) increase following nonspecific cell membrane permeabilization is among the earliest biochemical modifications in cells exposed to toxic amyloid aggregates. However, more recently receptors with Ca(2+) channel activity such as alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA), N-methyl D-aspartate (NMDA), ryanodine, and inositol 1,4,5-trisphosphate receptors have been proposed as mediators of the Ca(2+) increase in neuronal cells challenged with beta-amyloid peptides. We previously showed that prefibrillar aggregates of proteins not associated with amyloid diseases are toxic to exposed cells similarly to comparable aggregates of disease-associated proteins. In particular, prefibrillar aggregates of the prokaryotic HypF-N were shown to be toxic to different cultured cell lines by eliciting Ca(2+) and reactive oxygen species increases. This study was aimed at assessing whether NMDA and AMPA receptor activations could be considered a generic feature of cell interaction with amyloid aggregates rather than a specific effect of some aggregated protein. Therefore, we investigated whether NMDA and AMPA receptors were involved in the Ca(2+) increase following exposure of rat cerebellar granule cells to HypF-N prefibrillar aggregates. We found that the intracellular Ca(2+) increase was associated with the early activation of NMDA and AMPA receptors, although some nonspecific membrane permeabilization was also observed at longer times of exposure. This result matched a significant co-localization of the aggregates with both receptors on the plasma membrane. Our data support the possibility that glutamatergic channels are generic sites of interaction with the cell membrane of prefibrillar aggregates of different peptides and proteins as well as the key structures responsible for the resulting early membrane permeabilization to Ca(2+).

Highlights

Many human degenerative diseases are associated with the intracellular and/or extracellular deposition, in the affected tissues, of fibrillar aggregates arising from the ordered polymerization of specific proteins or peptides [1]
A␤ can drive the loss from the cell surface of both N-methyl D-aspartate (NMDA) receptors (NMDA-Rs) and amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors (AMPA-Rs) [23], whereas Ca2ϩ-permeable AMPA-Rs are involved in the cell death associated with neurological disorders such as amyotrophic lateral sclerosis and Alzheimer disease [24]
We investigated whether cultured rat cerebellar granule cells are damaged by HypF-N amyloid aggregates, whether cell damage is accomplished via a nonspecific pathway or through aggregate interaction with specific cell surface receptors, and whether NMDA-Rs and AMPA-Rs are involved in the biochemical modifications resulting from cell exposure to HypF-N or muscle acylphosphatase (AcP, a related protein) aggregates

Summary

Introduction

Many human degenerative diseases are associated with the intracellular and/or extracellular deposition, in the affected tissues, of fibrillar aggregates arising from the ordered polymerization of specific proteins or peptides [1]. We investigated whether cultured rat cerebellar granule cells are damaged by HypF-N amyloid aggregates, whether cell damage is accomplished via a nonspecific pathway or through aggregate interaction with specific cell surface receptors, and whether NMDA-Rs and AMPA-Rs are involved in the biochemical modifications resulting from cell exposure to HypF-N or muscle acylphosphatase (AcP, a related protein) aggregates. To this purpose, we measured the Ca2ϩ influx following aggregate interaction with our experimental cell model by blocking selectively the possible permeation pathways. We investigated whether the immunoreactivity of the AMPA-Rs and/or NMDA-Rs at the cell membrane was reduced in cells exposed to the aggregates for 1 h

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Conclusion