A Concerted Role of Na+—K+—Cl− Cotransporter and Na+/Ca2+ Exchanger in Ischemic Damage

Luo, Jing; Wang, Yanping; Chen, Hai; Kintner, Douglas B.; Cramer, Samuel W.; Gerdts, Josiah; Chen, Xinzhi; Shull, Gary E.; Philipson, Kenneth D.; Sun, Dandan

doi:10.1038/sj.jcbfm.9600561

Cited by 30 publications

(24 citation statements)

References 35 publications

(64 reference statements)

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“…Focal cerebral ischemia was induced by 60-min middle cerebral artery (MCA) occlusion, as previously described 26 and detailed description is provided in the online-only Data Supplement.…”

Section: Methodsmentioning

confidence: 99%

Inhibition of WNK3 Kinase Signaling Reduces Brain Damage and Accelerates Neurological Recovery After Stroke

Begum

Yuan

Kahle

et al. 2015

Stroke

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107

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BACKGROUND AND PURPOSE WNK kinases, including WNK3, and the associated downstream SPAK and OSR1 kinases, comprise an important signaling cascade that regulates the cation-chloride cotransporters. Ischemia-induced stimulation of the bumetanide-sensitive Na+-K+-Cl- cotransporter (NKCC1) plays an important role in the pathophysiology of experimental stroke, but the mechanism of its regulation in this context is unknown. Here, we investigated the WNK3-SPAK/OSR1 pathway as a regulator of NKCC1 stimulation and their collective role in ischemic brain damage. METHOD Wild-type WNK3 (WT) and WNK3 knockout (KO) mice were subjected to ischemic stroke via transient middle cerebral artery (MCA) occlusion. Infarct volume, brain edema, blood brain barrier (BBB) damage, white matter demyelination, and neurological deficits were assessed. Total and phosphorylated forms of WNK3 and SPAK/OSR1 were assayed by immunobloting and immunostaining. In vitro ischemia studies in cultured neurons and immature oligodendrocytes were conducted using the oxygen-glucose deprivation/reoxygenation method. RESULTS WNK3 KO mice exhibited significantly decreased infarct volume and axonal demyelination, less cerebral edema, and accelerated neurobehavioral recovery compared to WNK3 WT mice subjected to MCA occlusion. The neuroprotective phenotypes conferred by WNK3 KO were associated with a decrease in stimulatory hyper-phosphorylations of the SPAK/OSR1 catalytic T-loop and of NKCC1 stimulatory sites Thr203/Thr207/Thr212, as well as with decreased cell surface expression of NKCC1. Genetic inhibition of WNK3 or siRNA knockdown of SPAK/OSR1 increased the tolerance of cultured primary neurons and oligodendrocytes to in vitro ischemia. CONCLUSION These data identify a novel role for the WNK3-SPAK/OSR1-NKCC1 signaling pathway in ischemic neuroglial injury, and suggest the WNK3-SPAK/OSR1 kinase pathway as a therapeutic target for neuroprotection following ischemic stroke.

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“…Focal cerebral ischemia was induced by 60-min middle cerebral artery (MCA) occlusion, as previously described 26 and detailed description is provided in the online-only Data Supplement.…”

Section: Methodsmentioning

confidence: 99%

Inhibition of WNK3 Kinase Signaling Reduces Brain Damage and Accelerates Neurological Recovery After Stroke

Begum

Yuan

Kahle

et al. 2015

Stroke

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107

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“…4,26,27 Either pharmacological inhibition or genetic ablation of NKCC1 significantly reduced ischemic cell death in vivo and in in vitro ischemic models. 4,26,27 Although the reduction in ischemic brain injury in WNK3 KO and SPAK KO mice may be largely attributable to inhibition of the WNK3-SPAK-NKCC1 cascade, increased Cl À efflux via K-Cl cotransporter (KCC) proteins may also play a role. Yang et al 28 have detected increased basal KCC2 protein expression and decreased ratios of NKCC1/KCC2 in SPAK KO brains.…”

Section: Discussionmentioning

confidence: 99%

Deletion of the WNK3-SPAK kinase complex in mice improves radiographic and clinical outcomes in malignant cerebral edema after ischemic stroke

Zhao

Nepomuceno

Gào

et al. 2016

J Cereb Blood Flow Metab

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The WNK-SPAK kinase signaling pathway controls renal NaCl reabsorption and systemic blood pressure by regulating ion transporters and channels. A WNK3-SPAK complex is highly expressed in brain, but its function in this organ remains unclear. Here, we investigated the role of this kinase complex in brain edema and white matter injury after ischemic stroke. Wild-type, WNK3 knockout, and SPAK heterozygous or knockout mice underwent transient middle cerebral artery occlusion. One cohort of mice underwent magnetic resonance imaging. Ex-vivo brains three days postischemia were imaged by slice-selective spin-echo diffusion tensor imaging magnetic resonance imaging, after which the same brain tissues were subjected to immunofluorescence staining. A second cohort of mice underwent neurological deficit analysis up to 14 days post-transient middle cerebral artery occlusion. Relative to wild-type mice, WNK3 knockout, SPAK heterozygous, and SPAK knockout mice each exhibited a >50% reduction in infarct size and associated cerebral edema, significantly less demyelination, and improved neurological outcomes. We conclude that WNK3-SPAK signaling regulates brain swelling, gray matter injury, and demyelination after ischemic stroke, and that WNK3-SPAK inhibition has therapeutic potential for treating malignant cerebral edema in the setting of middle cerebral artery stroke.

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“…In addition, bumetanide may protect OLs by promoting ionic homeostasis and preventing osmotic overload, cytotoxic edema, and massive calcium influx. Specifically, NKCC1's role as an Na + -importer, in conjunction with the activity of Na + /Ca 2+ exchanger proteins, has been shown in animals to cause pathological Ca 2+ increases in OLs, cortical neurons, and astrocytes (31,32). NKCC1 has also been linked to AMPAR and NMDAR function and activity.…”

Section: Discussionmentioning

confidence: 99%

Chloride cotransporter NKCC1 inhibitor bumetanide protects against white matter injury in a rodent model of periventricular leukomalacia

Jantzie

Park

et al. 2015

Pediatr Res

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Background: Periventricular leukomalacia (PVL) is a major form of preterm brain injury. Na + -K + -Cl − 1 cotransporter (NKCC1) expression on neurons and astrocytes is developmentally regulated and mediates Cl − reversal potential. We hypothesized that NKCC1 is highly expressed on oligodendrocytes (OLs) and increases vulnerability to hypoxia-ischemia (HI) mediated white matter injury, and that the NKCC1 inhibitor bumetanide would be protective in a rodent PVL model. Methods: Immunohistochemistry in Long-Evans rats and PLP-EGFP transgenic mice was used to establish cell-specific expression of NKCC1 in the immature rodent brain. HI was induced on postnatal day 6 (P6) in rats and the protective efficacy of bumetanide (0.3 mg/kg/i.p. q12h × 60 h) established. results: NKCC1 was expressed on OLs and subplate neurons through the first 2 postnatal weeks, peaking in white matter and the subplate between P3-7. Following HI, NKCC1 is expressed on OLs and neurons. Bumetanide treatment significantly attenuates myelin basic protein loss and neuronal degeneration 7 d post-HI. conclusion: Presence and relative overexpression of NKCC1 in rodent cerebral cortex coincides with a period of developmental vulnerability to HI white matter injury in the immature prenatal brain. The protective efficacy of bumetanide in this model of preterm brain injury suggests that Cl − transport is a factor in PVL and that its inhibition may have clinical application in premature human infants.

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A Concerted Role of Na⁺—K⁺—Cl⁻ Cotransporter and Na⁺/Ca²⁺ Exchanger in Ischemic Damage

Cited by 30 publications

References 35 publications

Inhibition of WNK3 Kinase Signaling Reduces Brain Damage and Accelerates Neurological Recovery After Stroke

Inhibition of WNK3 Kinase Signaling Reduces Brain Damage and Accelerates Neurological Recovery After Stroke

Deletion of the WNK3-SPAK kinase complex in mice improves radiographic and clinical outcomes in malignant cerebral edema after ischemic stroke

Chloride cotransporter NKCC1 inhibitor bumetanide protects against white matter injury in a rodent model of periventricular leukomalacia

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