Neuropeptide Y suppresses epileptiform discharges by regulating AMPA receptor GluR2 subunit in rat hippocampal
Wei Bu1, Wen-Qing Zhao2, Wen-Ling Li3
1Department of Neurosurgery, The Third Hospital of Hebei Medical University, Shijiazhuang, Hebei 050051, P.R. China.
Abstract:
The present study aimed to investigate the effects of neuropeptide Y (NPY) on the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor glutamate receptor 2 (GluR2) subunit in epileptiform discharge hippocampal neurons. Hippocampal neurons were harvested from neonatal Sprague‑Dawley rats aged <24 h and primarily cultured in vitro. At day 12 following culture, hippocampal neurons were divided into the following groups: Control, Mg2+‑free, NPY+Mg2+‑free and BIBP3226+NPY+Mg2+‑free. The action potential of neurons was measured using the whole cell patch clamp technique in the control, Mg2+‑free and NPY+Mg2+‑free groups. AMPA current (IAMPA) was detected and peak current density was calculated in each group. Alterations in total protein and phosphorylation of the GluR2 subunit were detected by western blot analysis, and GluR2 mRNA expression levels were detected by reverse transcription‑quantitative polymerase chain reaction, in each group. The whole cell patch clamp technique demonstrated an abnormal action potential in the Mg2+‑free group. The frequency and amplitude of the action potential were significantly greater in the Mg2+‑free group compared with the control group, and significantly reduced in the NPY+Mg2+‑free group compared with the Mg2+‑free group (P<0.05). In the Mg2+‑free group, compared with the control group, peak current density was significantly reduced (P<0.05), GluR2 subunit protein content was slightly reduced (P>0.05), phosphorylation levels of GluR2 subunit were significantly greater (P<0.05) and GluR2 mRNA was significantly reduced (P<0.05). In the NPY+Mg2+‑free group, compared with the Mg2+‑free group, peak current density was significantly greater (P<0.05), phosphorylation levels of GluR2 subunit were significantly reduced (P<0.05) and GluR2 mRNA expression was significantly greater (P<0.05). In the BIBP3226+NPY+Mg2+‑free group, compared with the NPY+Mg2+‑free group, peak current density was significantly reduced (P<0.05), phosphorylation levels of GluR2 subunit were significantly greater (P<0.05) and GluR2 mRNA expression was significantly reduced (P<0.05). After 3 h of treatment with Mg2+‑free extracellular fluid, epileptiform discharge was detected in the cells. NPY inhibited the discharge and its underlying mechanism may be that epileptiform discharge suppressed the function of the AMPA receptor GluR2 subunit. NPY relieved the inhibition of the GluR2 subunit via the Y1 receptor. This may provide a novel direction for future studies on the pathogenesis and treatment of epilepsy.
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