Abstract
Hypoxia ischemia (HI)-related brain injury is the major cause of long-term morbidity in neonates. One characteristic hallmark of neonatal HI is the development of reactive astrogliosis in the hippocampus. However, the impact of reactive astrogliosis in hippocampal damage after neonatal HI is not fully understood. In the current study, we investigated the role of Na+/H+ exchanger isoform 1 (NHE1) protein in mouse reactive hippocampal astrocyte function in an in vitro ischemia model (oxygen/glucose deprivation and reoxygenation, OGD/REOX). 2 h OGD significantly increased NHE1 protein expression and NHE1-mediated H+ efflux in hippocampal astrocytes. NHE1 activity remained stimulated during 1–5 h REOX and returned to the basal level at 24 h REOX. NHE1 activation in hippocampal astrocytes resulted in intracellular Na+ and Ca2+ overload. The latter was mediated by reversal of Na+/Ca2+ exchange. Hippocampal astrocytes also exhibited a robust release of gliotransmitters (glutamate and pro-inflammatory cytokines IL-6 and TNFα) during 1–24 h REOX. Interestingly, inhibition of NHE1 activity with its potent inhibitor HOE 642 not only reduced Na+ overload but also gliotransmitter release from hippocampal astrocytes. The noncompetitive excitatory amino acid transporter inhibitor TBOA showed a similar effect on blocking the glutamate release. Taken together, we concluded that NHE1 plays an essential role in maintaining H+ homeostasis in hippocampal astrocytes. Over-stimulation of NHE1 activity following in vitro ischemia disrupts Na+ and Ca2+ homeostasis, which reduces Na+-dependent glutamate uptake and promotes release of glutamate and cytokines from reactive astrocytes. Therefore, blocking sustained NHE1 activation in reactive astrocytes may provide neuroprotection following HI.
Highlights
Hippocampal astrocytes are in intimate physical relationship with neurons
Hippocampal astrocytes respond to neonatal hypoxia ischemia (HI) by developing reactive astrogliosis, which is characterized by up-regulation of glial fibrillary acid protein (GFAP) expression, astrocyte hypertrophy, and astrocyte proliferation [3,4]
These results demonstrate that OGD/REOX triggered transformation of hippocampal astrocytes to reactive astrocytes
Summary
Hippocampal astrocytes are in intimate physical relationship with neurons. Each hippocampal astrocyte is in contact with several hundred dendrites from multiple neurons and envelope hundreds of thousands of synapses [1]. Hippocampal astrocytes respond to neonatal hypoxia ischemia (HI) by developing reactive astrogliosis, which is characterized by up-regulation of glial fibrillary acid protein (GFAP) expression, astrocyte hypertrophy, and astrocyte proliferation [3,4]. We recently reported that expression of NHE1 protein was dramatically increased in hippocampal GFAP-positive reactive astrocytes at 72 h following HI in a neonatal mouse model [6]. NHE1 protein expression was up-regulated in hippocampal reactive astrocytes after in vitro ischemia (oxygen and glucose deprivation and reoxygenation, OGD/REOX). We detected concurrent elevation of NHE1 activity, increased intracellular sodium concentration ([Na+]i) and intracellular calcium concentration ([Ca2+]i), and release of glutamate and cytokines from hippocampal astrocytes following OGD/REOX. We concluded that over-stimulation of NHE1 promotes gliotransmitter and cytokine release from reactive astrocytes, which can subsequently contribute to hippocampal neuronal damage under hypoxic and ischemic conditions
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