Role of GluN2A NMDA receptor in homocysteine‐induced prostaglandin E2 release from neurons

Abstract

Hyperhomocysteinemia or systemic elevation of homocysteine is a metabolic condition that has been linked to multiple neurological disorders where inflammation plays an important role in the progression of the disease. However, it is unclear whether hyperhomocysteinemia contributes to disease pathology by inducing an inflammatory response. The current study investigates whether exposure of primary cultures from rat and mice cortical neurons to high levels of homocysteine induces the expression and release of the proinflammatory prostanoid, Prostaglandin E2 (PGE2). Using enzymatic assays and immunoblot analysis we show concurrent increase in the activity of cytosolic phospholipase A2 (cPLA2) and level of cyclooxygenase‐2 (COX2), two enzymes involved in PGE2 biosynthesis. The findings also show an increase in PGE2 release from neurons. Pharmacological inhibition of GluN2A‐containing NMDAR (GluN2A‐NMDAR) with NVP‐AAM077 significantly reduces homocysteine‐induced cPLA2 activity, COX2 expression, and subsequent PGE2 release. Whereas, inhibition of GluN2B‐containing NMDAR (GluN2A‐NMDAR) with Ro 25‐6981 has no effect. Complementary studies in neuron cultures obtained from wild type and GluN2A knockout mice show that genetic deletion of GluN2A subunit of NMDAR attenuates homocysteine‐induced neuronal increase in cPLA2 activity, COX2 expression, and PGE2 release. Pharmacological studies further establish the role of both extracellular‐regulated kinase/mitogen‐activated protein kinase and p38 MAPK in homocysteine‐GluN2A NMDAR‐dependent activation of cPLA2‐COX2‐PGE2 pathway. Collectively, these findings reveal a novel role of GluN2A‐NMDAR in facilitating homocysteine‐induced proinflammatory response in neurons.

Calcium Export from Neurons and Multi-Kinase Signaling Cascades Contribute to Ouabain Neuroprotection in Hyperhomocysteinemia

“…Whereas NMDARs containing GluN2A/C or D subunits did not reveal desensitization during the HCY activation [12,13], currents through GluN1/GluN2B NMDARs briefly declined to about 15% of the peak amplitude because of the desensitization [12]. This observation exhibits why NMDARs containing GluN2A subunits, but not those of GluN2B, mostly contribute to HCY-induced excitotoxicity in cortical neurons [12][13][14][15][16]. Cerebellar neurons expressing NMDARs containing GluN2C or 2D are also sensitive to the excitotoxic action of HCY since these receptors do not reveal desensitization during the HCY activation [13].…”

“…The inhibition of PKA or PKC blocked ouabain-induced neuroprotection against HCY but not against Glu neurotoxicity. This observation can be related to the NMDAR subtype selectivity of HCY because GluN2A, but not GluN2B NMDAR subunits, mostly contribute to HCY toxicity in cortical neurons [12,[14][15][16] and reflect extensive recruitment of different glutamate ionotropic and metabotropic receptors and transporters in glutamate neurotoxicity.…”

“…For example, during long-term action, HCY, but not Glu, induces GluN2A-dependent sustained activation of ERK2 MAPK [8,59], internalization of the Ca 2+ -impermeable GluA2-subunit of AMPA receptors, and increase of intracellular [Ca 2+ ] [60], which both cause permanent activation of p38 MAPK [14], downstream phosphorylation of caspase-9, and apoptosis (for a review see [61]). Conversely, GluN2B-containing extrasynaptic NMDARs over-activated by Glu induce only transient [59] ERK MAPK activation [17][18][19], while HCY causes sustained p38 MAPK activity [14]. As a result, p38 MAPK inhibition can protect neurons from short-term but not long-term activation of NMDARs [62].…”

Calcium export from neurons and multi-kinase signaling cascades contribute to ouabain neuroprotection in hyperhomocysteinemia

“…Whereas NMDARs containing GluN2A/C or D subunits did not reveal desensitization during the HCY activation [12,13], currents through GluN1/GluN2B NMDARs brie y declined to about 15 % of the peak amplitude because of the desensitization [12]. This observation exhibits why NMDARs containing GluN2A subunits, but not the GluN2B ones, mostly contribute to HCY-induced excitotoxicity in cortical neurons [12][13][14][15][16]. Cerebellar neurons expressing NMDARs containing GluN2C or 2D are also sensitive to the excitotoxic action of HCY since these receptors do not reveal desensitization during the HCY activation [13].…”

“…The inhibition of PKA or PKC blocked ouabain induced neuroprotection against HCY, but not against Glu neurotoxicity. This observation can be related to the NMDAR subtype selectivity of HCY because GluN2A, but not GluN2B NMDAR subunits, mostly contribute to HCY toxicity in cortical neurons [12,[14][15][16] [17][18][19], while HCY causes sustained p38 MAPK activity [14]. As a result, p38 MAPK inhibition can protect neurons from short-term, but not long-term activation of NMDARs [62].…”