Enhanced Neuronal Damage After Ischemic Insults in Mice Lacking Kir6.2-Containing ATP-Sensitive K⁺Channels

Abstract

. Enhanced neuronal damage after ischemic insults in mice lacking Kir6.2-containing ATP-sensitive K ϩ channels. J Neurophysiol 95: 2590 -2601, 2006. First published December 14, 2005 doi:10.1152/jn.00970.2005. Adenosine triphosphate (ATP)-sensitive potassium (K ATP ) channels, incorporating Kir6.x and sulfonylurea receptor subunits, are weak inward rectifiers that are thought to play a role in neuronal protection from ischemic insults. However, the involvement of Kir6.2-containing K ATP channel in hippocampus and neocortex has not been tested directly. To delineate the physiological roles of Kir6.2 channels in the CNS, we used knockout (KO) mice that do not express Kir6.2. Immunocytochemical staining demonstrated that Kir6.2 protein was expressed robustly in hippocampal neurons of the wild-type (WT) mice and absent in the KO. To examine neuronal sensitivity to metabolic stress in vitro, and to ischemia in vivo, we 1) exposed hippocampal slices to transient oxygen and glucose deprivation (OGD) and 2) produced focal cerebral ischemia by middle cerebral artery occlusion (MCAO). Both slice and whole animal studies showed that neurons from the KO mice were severely damaged after anoxia or ischemia, whereas few injured neurons were observed in the WT, suggesting that Kir6.2 channels are necessary to protect neurons from ischemic insults. Membrane potential recordings from the WT CA1 pyramidal neurons showed a biphasic response to OGD; a brief hyperpolarization was followed by a small depolarization during OGD, with complete recovery within 30 min after returning to normoxic conditions. By contrast, CA1 pyramidal neurons from the KO mice were irreversibly depolarized by OGD exposure, without any preceding hyperpolarization. These data suggest that expression of Kir6.2 channels prevents prolonged depolarization of neurons resulting from acute hypoxic or ischemic insults, and thus protects these central neurons from the injury.

Neuroprotective role of ATP-sensitive potassium channels in cerebral ischemia

“…During the initial phase of hypoxia and ischemia, energy failure reduces the ATP/ADP ratio, which activates neuronal K ATP channels, causing hyperpolarization of the neuronal cell membrane and suppression of neuronal activity (Figure 2). Similar to previous reports [88,95,129,130] , we reported that membrane hyperpolarization was induced during the first 5 min of ischemia in both hippocampal and cortical neurons [92,93] . Using Kir6.2 knockout mice, we further reported that hippocampal CA1 neurons that were deficient for Kir6.2 revealed rapid depolarization and increased neuronal activity in response to hypoxia.…”

“…Using Kir6.2 knockout mice, we further reported that hippocampal CA1 neurons that were deficient for Kir6.2 revealed rapid depolarization and increased neuronal activity in response to hypoxia. In contrast, neuronal suppression occurred in wild-type neurons under similar conditions [92] . Hypoxic hyperpolarization in wild-type hippocampal CA1 neurons could be prevented by the K ATP channel blocker tolbutamide.…”

“…Single cell RT-PCR analysis showed that Kir6.2 mRNA is predominantly expressed in interneurons and pyramidal, granule and neuroglial cells of the hippocampus in the brains from young rats aged 10-13 d [84] . In the adult brain, Kir6.2 subunits have been found in hippocampal [87,92] , cortical [93] , and hypothalamic neurons [94] , as well as in the SNr pars reticulata [95] . Immunohistochemical studies showed that Kir6.2 subunits are mainly located in the somata and dendrites of the central neurons [92,93,96] .…”

“…In the adult brain, Kir6.2 subunits have been found in hippocampal [87,92] , cortical [93] , and hypothalamic neurons [94] , as well as in the SNr pars reticulata [95] . Immunohistochemical studies showed that Kir6.2 subunits are mainly located in the somata and dendrites of the central neurons [92,93,96] . Mitochondrial K ATP channels are also found in the rat brain, and the expression level of Kir6.1 (per milligram of mitochondrial protein) is six to seven times higher than that in the heart and liver [12] .…”

Inhibition of Caspase-Mediated Apoptosis by Peroxynitrite in Traumatic Brain Injury

“…They were sectioned into 25 m free-floating slices in PBS using a Kryostat 1720 (Leica, Wetzler, Germany). Immunostaining was performed as described previously (Sun et al, 2006). In brief, sections were blocked using a 1% BSA, 3% goat serum, 0.3% Triton X-100 in PBS solution for 90 min and doubled labeled with rabbit anti-3-nitrotyrosine (1:50; Upstate Biotechnology) and mouse anti-neuronal-specific nuclear protein (NeuN; 1:100; Chemicon, Temecula, CA) overnight at 4°C.…”