Spinal cord injury causes a wide-spread, persistent loss of Kir4.1 and glutamate transporter 1: benefit of 17β-oestradiol treatment

During neuronal activity astrocytes function to remove extracellular increases in potassium, which are largely mediated by the inwardly-rectifying potassium channel Kir4.1, and to take up excess glutamate via glutamate transporter 1, a glial-specific glutamate transporter. Here we demonstrate that expression of both of these proteins is reduced by nearly 80% following a crush spinal cord injury in adult male rats, 7 days post-injury. This loss extended to spinal segments several millimetres rostral and caudal to the lesion epicentre, and persisted at 4 weeks post-injury. Importantly, we demonstrate that loss of these two proteins is not a direct result of astrocyte loss, as immunohistochemistry at 7 days and western blots at 4 weeks demonstrate a marked up-regulation in glial fibrillary acidic protein expression. Kir4.1 and glutamate transporter 1 expression were partially rescued by post-spinal cord injury administration of physiological levels of 17 beta-oestradiol (0.08 mg/kg/day) in vivo. Utilizing an in vitro culture system we demonstrate that 17 beta-oestradiol treatment (50 nM) is sufficient to increase glutamate transporter 1 protein expression in spinal cord astrocytes. This increase in glutamate transporter 1 protein expression was reversed and Kir4.1 expression reduced in the presence of an oestrogen receptor antagonist, Fulvestrant 182 780 suggesting a direct translational regulation of Kir4.1 and glutamate transporter 1 via genomic oestrogen receptors. Using whole-cell patch-clamp recordings in cultured spinal cord astrocytes, we show that changes in protein expression following oestrogen application led to functional changes in Kir4.1 mediated currents. These findings suggest that the neuroprotective benefits previously seen with 17b-oestradiol after spinal cord injury may be in part due to increased Kir4.1 and glutamate transporter 1 expression in astrocytes leading to improved potassium and glutamate homeostasis.