Department of Anesthesiology, Far-Eastern Memorial Hospital, New Taipei City 22060, Taiwan.
Department of Mechanical Engineering, Yuan Ze University, Taoyuan 32003, Taiwan.
Int J Mol Sci. 2022 Feb 27;23(5):2641. doi: 10.3390/ijms23052641.
The neurotransmitter glutamate plays an essential role in excitatory neurotransmission; however, excessive amounts of glutamate lead to excitotoxicity, which is the most common pathogenic feature of numerous brain disorders. This study aimed to investigate the role of butyl 2-[2-(2-fluorophenyl)acetamido]benzoate (HFP034), a synthesized anthranilate derivative, in the central glutamatergic system. We used rat cerebro-cortical synaptosomes to examine the effect of HFP034 on glutamate release. In addition, we used a rat model of kainic acid (KA)-induced glutamate excitotoxicity to evaluate the neuroprotective potential of HFP034. We showed that HFP034 inhibits 4-aminopyridine (4-AP)-induced glutamate release from synaptosomes, and this inhibition was absent in the absence of extracellular calcium. HFP034-mediated inhibition of glutamate release was associated with decreased 4-AP-evoked Ca level elevation and had no effect on synaptosomal membrane potential. The inhibitory effect of HFP034 on evoked glutamate release was suppressed by blocking P/Q-type Ca channels and protein kinase C (PKC). Furthermore, HFP034 inhibited the phosphorylation of PKC and its substrate, myristoylated alanine-rich C kinase substrate (MARCKS) in synaptosomes. We also observed that HFP034 pretreatment reduced neuronal death, glutamate concentration, glial activation, and the levels of endoplasmic reticulum stress-related proteins, calpains, glucose-regulated protein 78 (GRP 78), C/EBP homologous protein (CHOP), and caspase-12 in the hippocampus of KA-injected rats. We conclude that HFP034 is a neuroprotective agent that prevents glutamate excitotoxicity, and we suggest that this effect involves inhibition of presynaptic glutamate release through the suppression of P/Q-type Ca channels and PKC/MARCKS pathways.
神经递质谷氨酸在兴奋性神经传递中发挥着重要作用；然而，过量的谷氨酸会导致兴奋性毒性，这是许多脑部疾病最常见的致病特征。本研究旨在探讨 2-[2-(2-氟苯基)乙酰胺基]苯甲酸丁酯（HFP034）在中枢谷氨酸能系统中的作用。我们使用大鼠脑皮质突触体来研究 HFP034 对谷氨酸释放的影响。此外，我们使用红藻氨酸（KA）诱导的谷氨酸兴奋性毒性大鼠模型来评估 HFP034 的神经保护潜力。结果表明，HFP034 抑制 4-氨基吡啶（4-AP）诱导的突触体谷氨酸释放，而在不存在细胞外钙的情况下，这种抑制作用不存在。HFP034 介导的谷氨酸释放抑制与 4-AP 诱导的 Ca 水平升高减少有关，并且对突触体膜电位没有影响。HFP034 对诱发谷氨酸释放的抑制作用被阻断 P/Q 型钙通道和蛋白激酶 C（PKC）所抑制。此外，HFP034 抑制了突触体中 PKC 及其底物豆蔻酰化丙氨酸丰富 C 激酶底物（MARCKS）的磷酸化。我们还观察到，HFP034 预处理可减少 KA 注射大鼠海马神经元死亡、谷氨酸浓度、神经胶质激活以及内质网应激相关蛋白钙蛋白酶、葡萄糖调节蛋白 78（GRP 78）、C/EBP 同源蛋白（CHOP）和半胱天冬酶-12 的水平。综上所述，HFP034 是一种神经保护剂，可防止谷氨酸兴奋性毒性，我们认为这种作用涉及通过抑制 P/Q 型钙通道和 PKC/MARCKS 途径抑制突触前谷氨酸释放。
