Glutamate as a potential “survival factor” in an in vitro model of neuronal hypoxia/reoxygenation injury: leading role of the Na+/Ca2+ exchanger

Abstract

In brain ischemia, reduction in oxygen and substrates affects mitochondrial respiratory chain and aerobic metabolism, culminating in ATP production impairment, ionic imbalance, and cell death. The restoration of blood flow and reoxygenation are frequently associated with exacerbation of tissue injury, giving rise to ischemia/reperfusion (I/R) injury. In this setting, the imbalance of brain bioenergetics induces important metabolic adaptations, including utilization of alternative energy sources, such as glutamate. Although glutamate has long been considered as a neurotoxin, it can also be used as intermediary metabolite for ATP synthesis, and both the Na+/Ca2+ exchanger (NCX) and the Na+-dependent excitatory amino-acid transporters (EAATs) are essential in this pathway. Here we analyzed the role of NCX in the potential of glutamate to improve metabolism and survival of neuronal cells subjected to hypoxia/reoxygenation (H/R). In SH-SY5Y neuroblastoma cells differentiated into a neuron-like state, H/R produced a significant cell damage, a decrease in ATP cellular content, and intracellular Ca2+ alterations. Exposure to glutamate at the onset of the reoxygenation phase attenuated H/R-induced cell damage and evoked a significant raise in intracellular ATP levels. Furthermore, we found that in H/R cells NCX reverse-mode activity was reduced, and that glutamate limited such reduction. All the effects induced by glutamate supplementation were lost when cells were transfected with small interfering RNA against NCX1 and EAAT3, suggesting the need of a specific functional interplay between these proteins for glutamate-induced protection. Collectively, our results revealed the potential beneficial effect of glutamate in an in vitro model of H/R injury and focused on the essential role exerted by NCX1. Although preliminary, these findings could be a starting point to further investigate in in vivo systems such protective effect in ischemic settings, shedding a new light on the classical view of glutamate as detrimental factor.

Neuroprotective effects of human amniotic fluid stem cells-derived secretome in an ischemia/reperfusion model

“…14,15 Particularly, this cell line is used to develop the in vitro OGD/IR model. [15][16][17][18][19][20][21] Current findings in regenerative medicine have strengthened the search for new stem cell sources with beneficial properties; in particular, amniotic fluid has been identified as a valid resource of stem cells in this field. Adult stem cells, even after reprogramming, could preserve epigenetic alterations, thus representing a limitation in their application.…”

Peripheral blood-derived monocytes show neuronal properties and integration in immune-deficient rd1 mouse model upon phenotypic differentiation and induction with retinal growth factors

“…Upon induction for a period of 8 days, RNLCs exhibited morphological alteration showing elongated structure, neuron-like axonal spread out and cell-cell contacts. Additionally, we found a significant upregulation in neuro-signalling molecules like l -glutamate, Ach and NO that also act as paracrine factors in retina and facilitate cell interactions [ 39 – 41 ]. qPCR results showed that RNLCs also exhibited an upregulation in a few neuronal and retinal markers (IRBP and RHO mRNA).…”

“…We also find that RNLCs have the potential to secrete neurotransmitters like nitric oxide, acetylcholine and l -glutamate in vitro, which also have a broad range of paracrine signalling function in neurons besides synaptic functions [ 39 – 41 ] (Sup Fig. 4 ).…”

Gateways for Glutamate Neuroprotection in Parkinson’s Disease (PD): Essential Role of EAAT3 and NCX1 Revealed in an In Vitro Model of PD

“…We have previously demonstrated that in different cell models and experimental settings, glutamate supplementation stimulates ATP synthesis under pathophysiological conditions [38][39][40][41]. Remarkably, this glutamate-related response strictly relies on the activity of two main players: Na + -dependent excitatory amino acid transporters (EAATs) and the Na + /Ca 2+ exchanger (NCX).…”

“…In this regard, we have previously demonstrated a physical and functional interaction between a specific EAATs subtype, EAAT3 (excitatory amino acid carrier 1, EAAC1 in rodents) and NCX1 at both the plasma membrane and the mitochondrial levels [38,41,50]. In both cardiac and neuronal models, their interaction modulates the glutamate-driven ATP synthesis under pathophysiological conditions [38][39][40][41]51].…”