Arachidonic acid has protective effects on oxygen-glucose deprived astrocytes mediated through enhancement of potassium channel TREK-1 activity

Highlights

• AA suppressed glutamate uptake and decreased GLT-1 levels under normal conditions.

• AA alleviated OGD-induced astrocyte death.

• AA rescued the decreased glutamate uptake under OGD conditions.

• AA increased expression levels of GLT-1 and TREK-1 under OGD conditions.

Abstract

Polyunsaturated fatty acids (PUFAs) have neuroprotective effects against ischemic brain diseases. The newly discovered potassium channel “TREK-1” is a promising target for therapies against neurodegeneration. Arachidonic acid (AA) is an n-6 PUFA, as well as a potent TREK-1 activator. We previously showed that TREK-1 is expressed at high levels in astrocytes. However, the effect of AA on astrocytes in ischemia remains unknown. Here, we assessed the effects of 3–30 μM AA on astrocyte apoptosis, glutamate uptake, and expression of the astrocytic glutamate transporter 1 (GLT-1) and TREK-1 under different conditions. Under normal conditions, 3–30 μM AA showed no effect on astrocytic apoptosis or TREK-1 expression, whereas glutamate uptake decreased significantly and its change paralleled the decreased expression of GLT-1. When astrocytes were subjected to 4 h of oxygen-glucose deprivation (OGD), 10 μM AA markedly alleviated OGD-induced cell death, recovering from 63.50 ± 1.90% to 82.96 ± 4.63% of the control value. AA also rescued the decreased glutamate uptake and increased mRNA, as well as protein levels of GLT-1 and TREK-1. Our results provide new evidence of a protective effect of AA on astrocytes under OGD conditions, suggesting that a low concentration of AA may protect against brain ischemic diseases.

Introduction

There is now a considerable literature describing the beneficial effects of polyunsaturated fatty acids (PUFAs) in the prevention of central nervous disease, such as global or focal brain ischemia [1]. The beneficial effects of PUFAs contribute to a decrease in synaptic glutamate transmission. A newly discovered two-pore-domain potassium channel, TREK-1, has been shown to be another important factor in the neuroprotective action of PUFAs [2], [3], [4].

The TREK-1 channel is involved in background leak K⁺ currents and plays an important role in regulating neuronal excitability [5]. TREK-1, which is widely distributed in the central nervous system (CNS), especially in the cerebral cortex, hippocampus, and hypothalamus, is activated by pathological factors, including membrane stretching (as in cell swelling), intracellular acidification, and a high body temperature, which occur commonly in ischemia and other pathological conditions. After acute and chronic cerebral ischemia, enhanced expression of TREK-1 has been observed in the cortex and hippocampus of rats, suggesting a correlation between TREK channels and acute cerebral ischemia [6]. Reduced resistance to cerebral ischemia was also observed in TREK-1^−/− mice [7]. Taken together, these data suggest that TREK-1 may be a mediator of neuroprotection. This hypothesis is reinforced by the effects of PUFAs, which are potent TREK-1 activators. Indeed, PUFAs induce dilation in the cerebral circulation, mediated by TREK-1, because the dilation response is abolished in isolated basilar arteries of TREK-1^−/− mice, and the corresponding increase in blood flow in vivo is also abolished [1], [4]. Thus, TREK-1 channels are required for the protective properties of PUFAs against global ischemia [8].

Arachidonic acid (AA), a PUFA, is a potent activator of TREK-1. AA has been shown to inhibit several sodium-coupled amino acid transporters, including the uptake systems for glutamate, in astrocytes under physiological conditions [9]. Astrocytes are the most abundant glial cell type and outnumber neurons by more than five‐fold. Astrocytes provide metabolic and trophic support to neurons, participate in synaptic function and plasticity, and maintain the extracellular balance of ions, fluid, and transmitters [10]. Additionally, astrocytes respond to various CNS insults, such as ischemia, neurodegenerative disease, and infection. In previous work, we demonstrated that TREK-1 is abundantly distributed in astrocytes and is involved in neurotrophin (such as BDNF) secretion [11].

Although it is now clear that PUFAs play a role in neuroprotection, the role of AA in brain ischemia remains controversial. In this study, we were interested in whether AA has a protective role in ischemic astrocytes and, if so, what the underlying cellular mechanism(s) are. Thus, an in vitro ischemia model, under oxygen-glucose deprivation (OGD) conditions, was used to mimic cerebral ischemia in vivo. We compared the effects of AA on astrocytes under normal and OGD conditions. Our results showed that while 3–30 μM AA had no cytotoxicity and no effect on TREK-1 expression by astrocytes, it could inhibit glutamate uptake by decreasing mRNA and protein levels of the glutamate transporter GLT-1 under normal conditions. However, AA could rescue OGD-induced astrocyte apoptosis, and enhance glutamate clearance, through increasing the expression of GLT-1 and TREK-1 under OGD conditions. We believe this study sheds light on AA-mediated neuroprotection in brain ischemia.