Histamine protects against NMDA-induced necrosis in cultured cortical neurons through H 2 receptor/cyclic AMP/protein kinase A and H 3 receptor/GABA release pathways Haibin Dai,* ,  Zhongmiao Zhang, Yongpin Zhu,* Yao Shen,* Weiwei Hu,* Yuwen Huang,* ,   Jianhong Luo,* Henk Timmerman,à Rob Leursà and Zhong Chen* *Department of Pharmacology, School of Medicine, Zhejiang University, Hangzhou, China 310031  Department of Pharmacy, Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China 310009 àLeiden/Amsterdam Center for Drug Research, Division of Medicinal Chemistry, Faculty of Chemistry, De Boelelaan 1083, 1081 HV, Amsterdam, the Netherlands Abstract Using histamine and the H 3 receptor antagonist thioperamide, the roles of histamine receptors in NMDA-induced necrosis were investigated in rat cultured cortical neurons. Within 3 h of intense NMDA insult, most neurons died by necrosis. Hista- mine reversed the neurotoxicity in a concentration-dependent manner and showed peak protection at a concentration of 10 )7 M. This protection was antagonized by the H 2 receptor antagonists cimetidine and zolantidine but not by the H 1 receptor antagonists pyrilamine and diphenhydramine. In addition, the selective H 2 receptor agonist amthamine mim- icked the protection by histamine. This action was prevented by cimetidine but not by pyrilamine. 8-Bromo-cAMP also mimicked the effect of histamine. In contrast, both the adenylyl cyclase inhibitor 9-(tetrahydro-2-furanyl)-9H-purine-6-amine and the cAMP-dependent protein kinase inhibitor N-[2-(p-bromocinna- mylamino) ethyl]-5-isoquinolinesulfonamide reversed the pro- tection by histamine. Thioperamide also attenuated NMDA- induced excitotoxicity, which was reversed by the H 3 receptor agonist (R)-a-methylhistamine but not by pyrilamine and ci- metidine. In addition, the protection by thioperamide was inhibited by the GABA A receptor antagonists picrotoxin and bicuculline. Further study demonstrated that the protection by thioperamide was due to increased GABA release in NMDA- stimulated samples. These results indicate that not only the H 2 receptor/cAMP/cAMP-dependent protein kinase pathway but also the H 3 receptor/GABA release pathway can attenuate NMDA-induced neurotoxicity. Keywords: amthamine, GABA, histamine, neurotoxicity, NMDA, thioperamide. J. Neurochem. (2006) 96, 1390–1400. It is well known that histamine acts as a neurotransmitter and neuromodulator in the peripheral and CNS (Schwartz et al. 1991; Haas and Panula 2003). The histaminergic system is thought to be involved in various physiological and behavioral functions including sleep/wake cycles, emotion, appetite control, locomotor activity, stress behavior, neuro- endocrine control, and learning and memory through hista- mine H 1 ,H 2 and H 3 receptors (Schwartz et al. 1991; Passani et al. 2000). The NMDA receptor has also been implicated in a number of important brain functions and pathologies, including learning, neuronal development and stroke (McBain and Mayer 1994). Recently, the interaction between histamine and the NMDA receptor has attracted considerable attention in view of the effects on synaptic plasticity, learning and ischemic neuronal damage (Brown and Haas 1999). Histamine can affect long-term potentiation, which may involve NMDA receptors, in a number of ways (Behnisch and Reymann 1993; Brown et al. 1995). Histamine selectively facilitates currents evoked by NMDA in neurons in the hippocampal slice and in culture (Bekkers 1993; Vorobjev et al. 1993; Brown et al. 1995). On the other hand, histaminergic neurotransmission Received August 18, 2005; revised manuscript received September 27, 2005; accepted November 4, 2005. Address correspondence and reprint requests to Prof. Zhong Chen, Department of Pharmacology, School of Medicine, Zhejiang University, Hangzhou, China, 310031. E-mail: chenzhong@zju.edu.cn Abbreviations used: MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl- tetrazolium bromide; PKA, cAMP-dependent protein kinase. Journal of Neurochemistry , 2006, 96, 1390–1400 doi:10.1111/j.1471-4159.2005.03633.x 1390 Journal Compilation Ó 2006 International Society for Neurochemistry, J. Neurochem. (2006) 96, 1390–1400 Ó 2006 The Authors