Preservation of GABA _A Receptor Function by PTEN Inhibition Protects Against Neuronal Death in Ischemic Stroke

Abstract

Background and Purpose-Downregulation of the tumor suppressor, phosphatase and tensin homolog deleted on chromosome 10 (PTEN), is thought to be a novel neuroprotective strategy in ischemic stroke, but the underlying mechanisms remain unclear. In this study, we aimed to validate the use of PTEN regulation of ␥-aminobutyric acid subtype A receptors (GABA A Rs) as a molecular target for the treatment of ischemic stroke. Because suppression of GABA A Rs contributes to ischemic neuron death, describing the intracellular signaling that interacts with GABA A Rs in ischemic neurons would provide a molecular basis for novel stroke therapies. Methods-We measured surface GABA A R expression by immunocytochemical labeling and surface protein biotinylation assay. Knockdown and overexpression approaches were used to test the effects of PTEN on the expression and function of GABA A Rs. Neuronal death was detected in both in vitro and in vivo stroke models. Results-The knockdown and overexpression approaches provided the first evidence that PTEN negatively regulated membrane expression and function of GABA A Rs in rat hippocampal neurons. Importantly, we demonstrated that a PTEN inhibitor prevented the reduction of surface GABA A Rs in injured hippocampal neurons subjected to oxygen-glucose deprivation, an in vitro insult that mimics ischemic injury, whereas a GABA A R antagonist significantly reduced this PTEN inhibitor-induced neuroprotection in both the in vitro and in vivo ischemic stroke models. Conclusions-Our study provides direct evidence that downregulation of PTEN protects against ischemic neuron death by preserving GABA A R function. Targeting this pathway may be an effective strategy for development of selective, potent stroke treatments.

Stabilization of GABAAReceptors at Endocytic Zones Is Mediated by an AP2 Binding Motif within the GABAAReceptor β3 Subunit

“…However, hypertonicity has multiple cellular effects (Rosenmund and Stevens, 1996;Carroll et al, 1999), and dynamin inhibitors may block other (e.g., caveolar) endocytosis pathways (Nabi and Le, 2003;Doherty and McMahon, 2009). By imaging super-ecliptic pHluorin-tagged GABA A Rs, we show a decrease in synaptic GABA A R clusters during OGD treatment of hippocampal neurons for 30 min, which is consistent with other studies (Mielke and Wang, 2005;Liu et al, 2010). Using the ␤3-subunit RRR motif mutant, we then directly tested the pathway of pathological GABA A R endocytosis during OGD and showed that acute loss of synaptic GABA A Rs during OGD is mediated by an AP2 and clathrin-dependent internalization pathway.…”

“…Several studies have reported a decrease in surface and synaptic GABA A R expression during ischemia, suggesting an increase in receptor endocytosis (Schwartz-Bloom and Sah, 2001;Mielke and Wang, 2005;Zhan et al, 2006;Kelley et al, 2008;Liu et al, 2010), but whether this is via an AP2-dependent pathway remains unclear.…”

Pro-Brain-Derived Neurotrophic Factor Inhibits GABAergic Neurotransmission by Activating Endocytosis and Repression of GABAAReceptors

“…All together, the observations obtained here may reflect the functional antagonism between p75 NTR and TrkB receptor signaling (Lu, 2003;Yang et al, 2014). BDNF, acting via the TrkB receptor, has different modulatory effects on the efficacy of GABAergic synapses and transmission.…”

“…BDNF is expressed mainly by glutamatergic neurons and secreted through a regulated and constitutive pathway (Haubensak et al, 1998;Wu et al, 2004;Kuczewski et al, 2008a;Lessmann and Brigadski, 2009). BDNF is stored as proneurotrophin (proBDNF) within secretion vesicles (Lu, 2003). To produce mature BDNF (mBDNF), proBDNF is cleaved by furin in the trans-Golgi network (Mowla et al, 2001), or by extracellular plasmin or matrixmetalloprotease-7 once secreted (Lee et al, 2001).…”

Therapeutic Targets for Cerebral Ischemia Based on the Signaling Pathways of the GluN2B C Terminus

“…61 Therefore, PTEN may be activated by the GluN2B signal through nNOS. PTEN can induce neuronal damage after ischemic insults through several pathways (Figure 3): antagonizing the phosphatidylinositol-3 kinase (PI3K) signaling pathway, 62 negatively regulating the membrane expression and function of GABAA receptors (GABA A Rs), 63 positively regulating extrasynaptic NMdARs 64 and interfering with nuclear signaling. 65 However, PTEN also induces a neuroprotective effect through activating the GluN2A cell prosurvival pathway.…”