Rivera-Aponte D E, Méndez-González M P, Rivera-Pagán A F, Kucheryavykh Y V, Kucheryavykh L Y, Skatchkov S N, Eaton M J
Department of Biochemistry, Universidad Central del Caribe, Bayamón, PR, USA.
Department of Biochemistry, Universidad Central del Caribe, Bayamón, PR, USA; Department of Physiology, Universidad Central del Caribe, Bayamón, PR, USA.
Neuroscience. 2015 Dec 3;310:216-23. doi: 10.1016/j.neuroscience.2015.09.044. Epub 2015 Sep 25.
Diabetics are at risk for a number of serious health complications including an increased incidence of epilepsy and poorer recovery after ischemic stroke. Astrocytes play a critical role in protecting neurons by maintaining extracellular homeostasis and preventing neurotoxicity through glutamate uptake and potassium buffering. These functions are aided by the presence of potassium channels, such as Kir4.1 inwardly rectifying potassium channels, in the membranes of astrocytic glial cells. The purpose of the present study was to determine if hyperglycemia alters Kir4.1 potassium channel expression and homeostatic functions of astrocytes. We used q-PCR, Western blot, patch-clamp electrophysiology studying voltage and potassium step responses and a colorimetric glutamate clearance assay to assess Kir4.1 channel levels and homeostatic functions of rat astrocytes grown in normal and high glucose conditions. We found that astrocytes grown in high glucose (25 mM) had an approximately 50% reduction in Kir4.1 mRNA and protein expression as compared with those grown in normal glucose (5mM). These reductions occurred within 4-7 days of exposure to hyperglycemia, whereas reversal occurred between 7 and 14 days after return to normal glucose. The decrease in functional Kir channels in the astrocytic membrane was confirmed using barium to block Kir channels. In the presence of 100-μM barium, the currents recorded from astrocytes in response to voltage steps were reduced by 45%. Furthermore, inward currents induced by stepping extracellular [K(+)]o from 3 to 10mM (reflecting potassium uptake) were 50% reduced in astrocytes grown in high glucose. In addition, glutamate clearance by astrocytes grown in high glucose was significantly impaired. Taken together, our results suggest that down-regulation of astrocytic Kir4.1 channels by elevated glucose may contribute to the underlying pathophysiology of diabetes-induced CNS disorders and contribute to the poor prognosis after stroke.
糖尿病患者面临多种严重健康并发症的风险，包括癫痫发病率增加以及缺血性中风后恢复较差。星形胶质细胞通过维持细胞外稳态以及通过摄取谷氨酸和缓冲钾来预防神经毒性，在保护神经元方面发挥着关键作用。星形胶质细胞的膜中存在钾通道，如内向整流钾通道Kir4.1，有助于这些功能的实现。本研究的目的是确定高血糖是否会改变Kir4.1钾通道的表达以及星形胶质细胞的稳态功能。我们使用q-PCR、蛋白质免疫印迹法、研究电压和钾阶跃反应的膜片钳电生理学以及比色谷氨酸清除试验，来评估在正常和高糖条件下培养的大鼠星形胶质细胞的Kir4.1通道水平和稳态功能。我们发现，与在正常葡萄糖（5mM）条件下培养的星形胶质细胞相比，在高糖（25mM）条件下培养的星形胶质细胞的Kir4.1 mRNA和蛋白质表达降低了约50%。这些降低在暴露于高血糖4 - 7天内出现，而在恢复到正常葡萄糖水平7 - 14天后恢复。使用钡阻断Kir通道证实了星形胶质细胞膜中功能性Kir通道的减少。在存在100μM钡的情况下，星形胶质细胞对电压阶跃反应所记录的电流减少了45%。此外，在高糖条件下培养的星形胶质细胞中，将细胞外[K(+)]o从3mM升至10mM（反映钾摄取）所诱导的内向电流减少了50%。此外，在高糖条件下培养的星形胶质细胞的谷氨酸清除能力明显受损。综上所述，我们的结果表明，葡萄糖升高导致星形胶质细胞Kir4.1通道下调可能促成糖尿病诱导的中枢神经系统疾病的潜在病理生理学，并导致中风后预后不良。
