Specific role of N-methyl-D-aspartate (NMDA) receptor in elastin-derived VGVAPG peptide-dependent calcium homeostasis in mouse cortical astrocytes in vitro

Abstract

Under physiological and pathological conditions, elastin is degraded to produce elastin-derived peptides (EDPs). EDPs are detected in the healthy human brain, and its concentration significantly increases after ischemic stroke. Both elastin and EDPs contains replications of the soluble VGVAPG hexapeptide, which has a broad range of biological activities. Effects of VGVAPG action are mainly mediated by elastin-binding protein (EBP), which is alternatively spliced, enzymatically inactive form of the GLB1 gene. This study was conducted to elucidate the activation and role of the N-methyl-D-aspartate receptor (NMDAR) in elastin-derived VGVAPG peptide-dependent calcium homeostasis in mouse cortical astrocytes in vitro. Cells were exposed to 10 nM VGVAPG peptide and co-treated with MK-801, nifedipine, verapamil, or Src kinase inhibitor I. After cell stimulation, we measured Ca2+ level, ROS production, and mRNA expression. Moreover, the Glb1 and NMDAR subunits (GluN1, GluN2A, and GluN2B) siRNA gene knockdown were applied. We found the VGVAPG peptide causes Ca2+ influx through the NMDA receptor in mouse astrocytes in vitro. Silencing of the Glb1, GluN1, GluN2A, and GluN2B gene prevented VGVAPG peptide-induced increase in Ca2+. Nifedipine does not completely reduce VGVAPG peptide-activated ROS production, whereas MK-801, verapamil, and Src inhibitor reduce VGVAPG peptide-activated Ca2+ influx and ROS production. These data suggest the role of Src kinase signal transduction from EBP to NMDAR. Moreover, the VGVAPG peptide affects the expression of NMDA receptor subunits.

Postnatal prebiotic supplementation in rats affects adult anxious behaviour, hippocampus, electrophysiology, metabolomics, and gut microbiota

“…While GluN2A receptors have mostly been associated with neuronal NMDARs located in the synapse, GluN2A receptors have also been detected in astrocytes in vitro ( Lee et al., 2010 ; Zhou et al., 2010 ; Szychowski and Gminski, 2019 ; Conti et al., 1996 ). It is therefore conceivable that the elevated protein levels of hippocampal GluN2A we observed previously ( Williams et al., 2016 ) were driven by non-neuronal cells.…”

Elastin-Derived Peptides in the Central Nervous System: Friend or Foe

“…It should be noted that the di-heteromeric GluN1/GluN2B and GluN1/GluN2A receptors are the most important receptors in the developing nervous system of mice (Ritter et al 2002). Besides classic calcium channels, such as L-type calcium channels (LTCC) and N-type calcium channels (NTCC), EDP-VGVAPG also activates NMDAR in mouse astrocytes cultured in vitro, which was demonstrated for the first time by Szychowski and Gmiński (Szychowski and Gmiński 2019c). In addition, silencing of the Glb1, GluN1, GluN2A, and GluN2B genes prevented the increase in Ca 2+ levels induced by the VGVAPG peptide.…”

“…So far, only two articles have reported that EDPs can affect Ca 2+ influx into nervous system-derived cells such as normal mouse astrocytes and in human glioblastoma cell lines (C6, CB74, CB109, and CB191) (Coquerel et al 2009;Szychowski and Gmiński 2019c). The most important excitatory receptor permeable to Ca 2+ , sodium (Na + ), and potassium (K + ) ions in the cells of the nervous system is the N-methyl-d-aspartate receptor (NMDAR) (Paoletti and Neyton 2007).…”

Review of the Relationship between Reactive Oxygen Species (ROS) and Elastin-Derived Peptides (EDPs)

“…The increase in the level of ROS in the cell is believed to be caused by at least two main processes, i.e., Ca 2+ influx and disruption of the expression and/or activity of antioxidant enzymes mediated through the peroxisome proliferator-activated receptor gamma (PPARγ) pathway (Figure 2) [9]. A number of studies have reported that tropoelastin, κ-elastin, EDPs, and the VGVAPG peptide increase Ca 2+ influx in human monocytes, fibroblasts, human umbilical venous endothelial cells (HUVEC), different glioma cell lines (C6, CB74, CB109, and CB191), and smooth muscle cells from pig aorta or mouse astrocytes [10,[46][47][48][49]. It is well known that different Ca 2+ signaling pathways can increase the level of cell ROS [50].…”

“…However, during various physiological and pathological processes, elastin is degraded to elastin-derived peptides (EDPs) [7,8]. To date, a number of papers have reported that κ-elastin, EDPs, or peptide VGVAPG (signaling sequences from elastin) affect the ROS level in cell culture models or in vitro in organisms [9][10][11][12]. It has been described that, similar to the ROS level, the level of EDPs increases during aging and in various pathological processes [13,14].…”