SARS-CoV-2 Viroporin E Induces Ca Release and Neuron Cell Death in Primary Cultures of Rat Hippocampal Cells Aged In Vitro

The COVID-19 pandemic was caused by infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), which may lead to serious respiratory, vascular and neurological dysfunctions. The SARS-CoV-2 envelope protein (E protein) is a structural viroporin able to form ion channels in cell membranes, which is critical for viral replication. However, its effects in primary neurons have not been addressed. Here we used fluorescence microscopy and calcium imaging to study SARS-CoV-2 viroporin E localization and the effects on neuron damage and intracellular Ca homeostasis in a model of rat hippocampal neurons aged in vitro. We found that the E protein quickly enters hippocampal neurons and colocalizes with the endoplasmic reticulum (ER) in both short-term (6-8 days in vitro, DIV) and long-term (20-22 DIV) cultures resembling young and aged neurons, respectively. Strikingly, E protein treatment induces apoptosis in aged neurons but not in young neurons. The E protein induces variable increases in cytosolic Ca concentration in hippocampal neurons. Ca responses to the E protein are due to Ca release from intracellular stores at the ER. Moreover, E protein-induced Ca release is very small in young neurons and increases dramatically in aged neurons, consistent with the enhanced Ca store content in aged neurons. We conclude that the SARS-CoV-2 E protein quickly translocates to ER endomembranes of rat hippocampal neurons where it releases Ca, probably acting like a viroporin, thus producing Ca store depletion and neuron apoptosis in aged neurons and likely contributing to neurological damage in COVID-19 patients.