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Tzu-Ying Lee College of Medicine, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Department of Physiology, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Medical College, Graduate Institutes of Medical Sciences and Neuroscience, Taipei Medical University, 250 Wu-Hsing Street, Taipei 110, Taiwan, ROC

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Ke-Li Tsai College of Medicine, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Department of Physiology, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Medical College, Graduate Institutes of Medical Sciences and Neuroscience, Taipei Medical University, 250 Wu-Hsing Street, Taipei 110, Taiwan, ROC

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Wen-Sen Lee College of Medicine, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Department of Physiology, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Medical College, Graduate Institutes of Medical Sciences and Neuroscience, Taipei Medical University, 250 Wu-Hsing Street, Taipei 110, Taiwan, ROC

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Chin Hsu College of Medicine, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Department of Physiology, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC
Medical College, Graduate Institutes of Medical Sciences and Neuroscience, Taipei Medical University, 250 Wu-Hsing Street, Taipei 110, Taiwan, ROC

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Previously, we showed that predominant expression of the N-methyl-d-aspartate (NMDA) receptor in the neurons of the sexually dimorphic nucleus of the preoptic area of male rats plays an important role in preventing neurons from apoptosis during sexual development. Blocking of the NMDA receptor by dizocilpine ((+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d] cyclohepten-5,10-iminemaleate (MK-801) causes down-regulation of some survival-related genes including cytochrome oxidase subunit II (COII), a mitochondria-encoded complex IV subunit, which in turn induces ATP depletion and the occurrence of apoptosis. The aim of this study is to investigate the molecular events during down-regulation of the COII gene expression induced by blocking of the NMDA receptor. Treatment of the GnRH cell line (GT1-7) with MK-801 caused 1) a decrease of intracellular calcium concentration ([Ca2+]i) after 20 h; 2) significant decreases of the levels of peroxisome proliferator-activated receptor γcoactivator-1 (PGC-1) mRNA and protein after 24 h; 3) down-regulation of COII mRNA after 36 h; and 4) the occurrence of neuronal apoptosis after 48 h. Accordingly, we hypothesize that blocking of the NMDA receptor may cause a decrease of the [Ca2+]i, which in turn inhibits the expressions of PGC-1 and COII and then leads to subsequent neuronal apoptosis.

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Hseng-Kuang Hsu Department of Physiology, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
School of Nursing, Mei-Ho Institute of Technology, Pingtung, Taiwan

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Pei-Lin Shao Department of Physiology, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
School of Nursing, Mei-Ho Institute of Technology, Pingtung, Taiwan

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Ke-Li Tsai Department of Physiology, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
School of Nursing, Mei-Ho Institute of Technology, Pingtung, Taiwan

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Huei-Chuan Shih Department of Physiology, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
School of Nursing, Mei-Ho Institute of Technology, Pingtung, Taiwan

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Tzu-Ying Lee Department of Physiology, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
School of Nursing, Mei-Ho Institute of Technology, Pingtung, Taiwan

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Chin Hsu Department of Physiology, Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
School of Nursing, Mei-Ho Institute of Technology, Pingtung, Taiwan

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The present study was designed to identify possible signaling pathways, which may play a role in prevention of neuronal apoptosis in the sexually dimorphic nucleus of the preoptic area (SDN-POA) after physiological activation of the N-methyl-D-aspartate (NMDA) receptor. Gene response to the blockage of the NMDA receptor by an antagonist (dizocilpine hydrogen maleate; MK-801) was screened after suppression subtractive hybridization (SSH). The results showed that dfferential screening after SSH detected the presence of some neurotrophic genes (RNA binding motif protein 3 (RBM3), α-tubulin) as well as apoptosis-related genes (Bcl-2, cytochrome oxidase subunit II, cytochrome oxidase subunit III) in the SDN-POA of male rats, which were down-regulated by blocking the NMDA receptor. The RT-PCR products of the aforementioned genes in MK-801-treated males were significantly less than that in untreated males. In particular, the expression of Bcl-2 mRNA, including Bcl-2 protein, in male rats were significantly suppressed by MK-801 treatment. Moreover, the binding activity of nuclear factor κB (NFκB) was significantly higher in male rats than in females, but significantly diminished by blocking the NMDA receptor with MK-801 in male rats. No significant difference in cAMP response element-binding protein (CREB) binding activity was observed among untreated male, MK-801-treated male, untreated female and MK-801-treated female groups. These results suggest that genes regulated by NMDA receptor activation might participate in neuronal growth and/or anti-apoptosis, and support an important signaling pathway of NFκB activation and its target gene, Bcl-2, in preventing neuronal apoptosis in the SDN-POA of male rats during sexual development.

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