Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCA272

Poster Communications

Store-operated calcium entry is profoundly impaired in a mouse model of Alzheimer's disease.

M. P. Weir1, A. Kaar1, M. G. Rae1

1. Physiology, University College Cork, Cork, Cork, Ireland.

Although Alzheimer's disease (AD) is the most common form of dementia, its cause is still unclear and a cure has yet to be developed. We have previously shown however that young (P2-P5) neurons derived from transgenic murine models of AD1,2 exhibit significantly altered endoplasmic reticulum (ER)-mediated Ca2+ handling compared to controls, suggesting that intracellular calcium homeostasis may be a primary initiator of AD pathogenesis. In addition to excitation-mediated Ca2+ entry, one of the primary means of loading the neuronal ER with Ca2+ is via store-operated calcium entry (SOCE). This process is initiated by the depletion of ER Ca2+ stores and involves the movement of Ca2+ directly through plasma membrane-located channels into the ER. However, to date, little is known about how this Ca2+ entry pathway is affected by mutations which predispose to the development of AD. The aims of this study were therefore to, firstly, determine if SOCE was present in mouse primary cultured hippocampal neurons and, if so, to investigate if it was altered in primary cultured hippocamapal neurons from the 3xTg mouse model of AD relative to controls. Hippocampal neurons were cultured from 3-5 day, wild type (non-Tg) and transgenic (3xTg) mice. Neurons were loaded with the Ca2+-sensitive dye, fluo-2 AM (4μM) and then superfused with standard physiological saline (HBSS) containing TTX (1mM). Intracellular somatic Ca2+ levels were monitored using Winfluor software. SOCE was evoked by depleting the ER using the SERCA pump inhibitor thapsigargin (TPN) in conjunction with Ca2+-free HBSS. The SOCE antagonist gadolinium (Gd3+) was also utilised to determine the relative susceptibility of SOCE in the two different hippocampal genotypes. Data are expressed as mean ± SEM and statistical significance was determined using Student's unpaired t-test. Our results demonstrated that SOCE in 3xTg neurons was significantly reduced compared to that evoked in non-Tg neurons (by 46.3±9.8%, P<0.0001; non-Tg, n=65; 3xTg, n=57). Additionally, the intracellular Ca2+ elevation evoked by TPN (1µM) was significantly reduced in 3xTg neurons relative to non-Tg neurons (by 84.7±9.7%, P<0.001; non-Tg, n=25; 3xTg, n=11). Gd3+ (50mM) significantly inhibited SOCE in both non-Tg and 3xTg hippocampal neurons but the degree of SOCE inhibition was significantly greater in non-Tg neurons (by 21±4.1%, P<0.001; non-Tg, n=35; 3xTg, n=29). The significantly smaller SOCE displayed by transgenic AD neurons appears to be yet another example of dysregulated calcium handling processes within neurons expressing AD-linked genetic mutations. The reason(s) underlying the disrupted SOCE in the 3xTg neurons remain(s) unclear but may be linked to incomplete emptying of the ER stores due to the particular PS1 mutation expressed within these transgenic mice, which is proposed to inhibit the normal leak of calcium from the ER3.

Where applicable, experiments conform with Society ethical requirements