Abstract
Neuronal Store-Operated Ca2+ Entry (nSOCE) plays an essential role in refilling endoplasmic reticulum Ca2+ stores and is critical for Ca2+-dependent neuronal processes. SOCE sensors, STIM1 and STIM2, can activate Orai, TRP channels and AMPA receptors, and inhibit voltage-gated channels in the plasma membrane. However, the link between STIM, SOCE, and NMDA receptors, another key cellular entry point for Ca2+ contributing to synaptic plasticity and excitotoxicity, remains unclear. Using Ca2+ imaging, we demonstrated that thapsigargin-induced nSOCE was inhibited in rat cortical neurons following NMDAR inhibitors. Blocking nSOCE by its inhibitor SKF96365 enhanced NMDA-driven [Ca2+]i. Modulating STIM protein level through overexpression or shRNA inhibited or activated NMDA-evoked [Ca2+]i, respectively. Using proximity ligation assays, immunofluorescence, and co-immunoprecipitation methods, we discovered that thapsigargin-dependent effects required interactions between STIMs and the NMDAR2 subunits. Since STIMs modulate NMDAR-mediated Ca2+ levels, we propose targeting this mechanism as a novel therapeutic strategy against neuropathological conditions that feature NMDA-induced Ca2+ overload as a diagnostic criterion.
Highlights
As a critical intracellular signaling ion, calcium (Ca2+ ) coordinates numerous cellular processes, such as fertilization, proliferation, development, learning, and memory [1]
We explored if N-methyl-d-aspartate receptor (NMDAR) participate in the mechanisms underlying TG-induced Neuronal Store-Operated Ca2+ Entry (nSOCE) using the addback assay
To eliminate the possibility that inhibitory effect of the NMDAR antagonists on nSOCE occurs through direct inhibition of STIM1, STIM2 or Orai proteins, we examined these responses in HeLa cells, since they do not express endogenous NMDARs [48], in the presence of one NMDAR antagonist
Summary
As a critical intracellular signaling ion, calcium (Ca2+ ) coordinates numerous cellular processes, such as fertilization, proliferation, development, learning, and memory [1]. The main store of intracellular Ca2+ ions is the endoplasmic reticulum (ER). Maintaining intracellular Ca2+ homeostasis is vital for cell survival [2], which underscores the need to elucidate the mechanisms underlying cellular Ca2+ dynamics. In non-excitable cells, such as lymphocytes, extracellular Ca2+ influx through the tightly regulated store-operated Ca2+ channels (SOCCs) in the plasma membrane (PM) drives store-operated Ca2+ entry (SOCE) [3] that regulate Ca2+ influx. Cells 2020, 9, 160 and STIM2 [4,5,6]. STIM proteins were initially discovered as potential tumor suppressor proteins [7]
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