Cellular and Molecular Neurobiology, Vol. 23, No. 6, December 2003 ( C  2003) Long-Term Imipramine Treatment Increases Nitrate Levels in the Rat Hypothalamus Eiji Suzuki, 1,5 Toshio Nakaki, 2 Shigenobu Kanba, 3 Futoshi Shintani, 4 and Hitoshi Miyaoka 1 Received December 16, 2002; accepted March 10, 2003 SUMMARY 1. Animal experiments have shown nitric oxide synthase inhibitors to have antidepressant-like properties. However, the effects of clinically available antidepressants on nitric oxide production in the brain remain unclear. In the present study, we examined whether imipramine, a conventional antidepressant, changes the levels of type-II nitric ox- ide synthase mRNA and nitrate, a final nitric-oxide-oxidation product measurable in vivo, in the rat brain. 2. Type-II nitric oxide synthase mRNA was detected using a reverse transcription- polymerase chain reaction method and nitrate was measured with a combination of high- performance liquid chromatography and the Griess reaction. 3. In untreated rats, type-II nitric oxide synthase mRNA was not detected in the hy- pothalamus, hippocampus, cerebral cortex, brain stem, or cerebellum. However, after 28-day oral administration of imipramine, it was detected in every brain region tested. Nitrate levels in the hypothalamus and cerebral cortex increased after 28-day treatment. In the hypothala- mus, nitrate levels increased dose-dependently. These dose-dependent nitrate level changes were prevented by pretreatment with a nitric oxide synthase inhibitor. Moreover, the pre- ventive effect of N G -nitro-L-arginine methyl ester was reversed by coadministration of L-arginine, a nitric oxide substrate. 4. These results suggest that chronic imipramine treatment induces nitric oxide synthase gene expression in the brain, followed by augmented NO production. KEY WORDS: imipramine; nitric oxide; hypothalamus; nitric oxide synthase; nitrate. INTRODUCTION Nitric oxide (NO), which is biosynthesized from L-arginine by nitric oxide synthase (NOS), was first identified as an endothelium-derived relaxing factor, but is currently recognized as an important bioregulatory molecule in the nervous, immune, and cardiovascular systems (Garthwaite, 1991; Sessa, 1994). 1 Department of Psychiatry, Kitasato University School of Medicine, Kanagawa, Japan. 2 Department of Pharmacology, Teikyo University School of Medicine, Tokyo, Japan. 3 Department of Psychiatry, Yamanashi University School of Medicine, Yamanashi, Japan. 4 Department of Psychiatry, Saitama Medical University, Saitama, Japan. 5 To whom correspondence should be addressed at Department of Psychiatry, Kitasato University School of Medicine, 2-1-1 Asamizodai, Sagamihara, Kanagawa 228-8520, Japan; e-mail: e-suzuki@ kitasato-u.ac.jp. 953 0272-4340/03/1200-0953/0 C  2003 Plenum Publishing Corporation