title: Exendin-4 improved rat cortical neuron survival under oxygen/glucose deprivation through PKA pathway.
abstract: Previous studies demonstrated that exendin-4 (Ex-4) may possess neurotrophic and neuroprotective functions in ischemia insults, but its mechanism remained unknown. Here, by using real-time PCR and ELISA, we identified the distribution of active GLP-1Rs in the rat primary cortical neurons. After establishment of an in vitro ischemia model by oxygen/glucose deprivation (OGD), neurons were treated with various dosages of Ex-4. The MTT assay showed that the relative survival rate increased with the dosage of Ex-4 ranging from 0.2 to 0.8 μg/ml (P<0.001, vs. OGD group). The apoptosis rate was reduced from (49.47±2.70)% to (14.61±0.81)% after Ex-4 treatment (0.4 μg/ml) 12h after OGD (P<0.001). Moreover, immunofluorescence staining indicated that Ex-4 increased glucose-regulated proteins 78 (GRP78) and reduced C/EBP-homologous protein (CHOP). Western blot analysis demonstrated that, after neurons were treated with Ex-4, GRP78 was up-regulated over time (P<0.01, vs. OGD group), while CHOP levels rose to a peak 8h after OGD and then decreased (P<0.05, vs. OGD group). This effect was changed by both the protein kinase A (PKA) inhibitor H89 (P<0.01, P<0.05, respectively, vs. Ex-4 group) and the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 (P<0.01, P<0.01, respectively, vs. Ex-4 group) but not by the mitogen-activated protein kinase (MAPK) inhibitor U0126. Our study also revealed that, compared with the Ex-4 group, inhibition of the PKA signaling pathway significantly decreased the survival rate of neurons, down-regulated the expression of B-cell lymphoma 2 (Bcl-2) and up-regulated the Bax expression 3h after ODG (P<0.05, P<0.01, respectively), while neither PI3K nor MAPK inhibition exerted such effects. Furthermore, Western blotting exhibited that PKA expression was elevated in the presence or absence of OGD insults (P<0.05). This study indicated that Ex-4 protected neurons against OGD by modulating the unfolded protein response (UPR) through the PKA pathway and may serve as a novel therapeutic agent for stroke.
authors.affiliation: Department of Neurology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Jiefang Avenue, Wuhan 430022, PR China.
authors.forename: M-D
authors.initials: MD
authors.lastname: Wang
authors.forename: Y
authors.initials: Y
authors.lastname: Huang
authors.forename: G-P
authors.initials: GP
authors.lastname: Zhang
authors.forename: L
authors.initials: L
authors.lastname: Mao
authors.forename: Y-P
authors.initials: YP
authors.lastname: Xia
authors.forename: Y-W
authors.initials: YW
authors.lastname: Mei
authors.forename: B
authors.initials: B
authors.lastname: Hu
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002545:Brain Ischemia / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects*; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004396:Coloring Agents; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology*; D004721:Endoplasmic Reticulum / Q000187:drug effects; D000077270:Exenatide; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D052216:Glucagon-Like Peptide 1 / Q000378:metabolism; D005947:Glucose / Q000172:deficiency*; D000860:Hypoxia / Q000473:pathology*; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D018696:Neuroprotective Agents; D016716:PC12 Cells; D010455:Peptides / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D013778:Tetrazolium Salts; D013844:Thiazoles; D014688:Venoms / Q000494:pharmacology*
chemical_list: D004396:Coloring Agents; D018696:Neuroprotective Agents; D010455:Peptides; D013778:Tetrazolium Salts; D013844:Thiazoles; D014688:Venoms; D052216:Glucagon-Like Peptide 1; D000077270:Exenatide; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; C022616:thiazolyl blue; D005947:Glucose