A new procedure for amyloid beta oligomers preparation enables the unambiguous testing of their effects on cytosolic and mitochondrial Ca2+ entry and cell death in primary neurons

作者：Caballero Erica; Calvo Rodriguez Maria; Gonzalo Ruiz Alicia; Villalobos Carlos; Nunez Lucia

来源：Neuroscience Letters, 2016, 612: 66-73.

DOI：10.1016/j.neulet.2015.11.041

• Calcium
• Mitochondria
• Alzheimer's disease
• Amyloid beta peptide
• Oligomers

Oligomers of the amyloid beta peptide (A beta(o)) are becoming the most likely neurotoxin in Alzheimer's disease. Controversy remains on the mechanisms involved in neurotoxicity induced by A beta(o) and the targets involved. We have reported that A beta(o) promote Ca2+ entry, mitochondrial Ca2+ overload and apoptosis in cultured cerebellar neurons. However, recent evidence suggests that some of these effects could be induced by glutamate receptor agonists solved in F12, the media in which A beta(o) are prepared. Here we have tested the effects of different media on A beta(o) formation and on cytosolic Ca2+ concentration ([Ca2+](cyt)) in rat cerebellar and hippocampal cell cultures. We found that A beta(o) prepared according to previous protocols but solved in alternative media including saline, MEM and DMEM do not allow oligomer formation and fail to increase [Ca2+](cyt). Changes in the oligomerization protocol and supplementation of media with selected salts reported to favor oligomer formation enable A beta(o) formation. A beta(o) prepared by the new procedure and containing small molecular weight oligomers increased [Ca2+](cyt), promoted mitochondrial Ca2+ overload and cell death in cerebellar granule cells and hippocampal neurons. These results foster a role for Ca2+ entry in neurotoxicity induced by A beta(o) and provide a reliable procedure for investigating the Ca2+ entry pathway promoted by A beta(o).

• 出版日期2016-1-26