A Role for Neuromedin U in Stress Response Reiko Hanada,* Masamitsu Nakazato,† ,1 Noboru Murakami,‡ Satoru Sakihara,§ Hironobu Yoshimatsu,* Koji Toshinai,† Takeshi Hanada, ¶ Toshihiro Suda,§ Kenji Kangawa, Shigeru Matsukura,† and Toshiie Sakata* *First Department of Internal Medicine, Oita Medical University, Oita 879-5593, Japan; †Department of Internal Medicine, Miyazaki Medical College, Miyazaki 889-1692, Japan; ‡Department of Veterinary Physiology, Faculty of Agriculture, Miyazaki University, Miyazaki 889-2192, Japan; §Third Department of Internal Medicine, Hirosaki University School of Medicine, Hirosaki 036-8562, Japan; ¶ Suntory Institute for Medicinal Research & Development, Gunma 370-0503, Japan; and Department of Biochemistry, National Cardiovascular Center Research Institute, Osaka 565-8565, Japan Received October 2, 2001 Neuromedin U (NMU) is a hypothalamic peptide that has been recently found to reduce food intake, but few is known about its other functions in the central ner- vous system. We here studied behavioral activities in- duced by an intracerebroventricular (ICV) adminis- tration of NMU in rats and mice. NMU increased gross locomotor activity, face washing behavior, and groom- ing. NMU-induced stress response was significantly abolished by pretreatment with an antagonist of cor- ticotropin-releasing hormone (CRH), -helical CRH (9 – 41) (-hCRH), or anti-CRH IgG. NMU did not induce locomotor activity in CRH knockout mice. NMU that interacts anatomically and/or functionally with the CRH system is a novel physiological regulator of stress response. © 2001 Academic Press Key Words: neuromedin U; hypothalamus; CRH; stress; CRH KO mice. NMU, a 23-amino-acid peptide, was first isolated as a smooth-muscle-contracting peptide from the porcine spinal cord and later from the brains of other species (1). Two receptors for NMU, NMU1R, and NMU2R, have recently been identified by using an intracellular calcium influx assay in a cell line expressing NMU1R (2–7). Rat NMU1R is expressed at a low level in the brain, but NMU2R is abundantly expressed in the hy- pothalamic paraventricular nucleus (PVN), along the wall of the third ventricle in the hypothalamus, and CA1 region of the hippocampus (2). We and another group have shown that an ICV administration of NMU suppressed feeding in rats (2, 3, 8). The PVN is a major part that produces CRH, a 41-amino-acid peptide, which functions in stress response, anorectic behavior, autonomic regulation, and the hypothalamo-pituitary- adrenal axis (9, 10). An ICV injection of CRH to rodents increased gross locomotor activity, especially groom- ing, and face washing behavior (11–13). We here stud- ied behavioral activities of NMU given to rats cen- trally. We also investigated the functional relationship between NMU and CRH in the stress response. Fur- thermore, we examined the effect of NMU on behav- ioral responses to stress by the use of CRH-deficient (knockout) mice. MATERIALS AND METHODS Rat experiment. Ten-week-old male Wistar rats weighing 300 – 325 g (Charles River Japan, Inc., Shiga, Japan) were maintained in individual cages under controlled temperature (21–23°C) and light (light on 0700 –1900 h) with ad libitum access to chow and water. ICV cannulae were implanted into the lateral cerebral ventricle. Proper placement of the cannulae was verified at the end of the experiment by the administration of dye. Rat NMU ( M r = 2641.3) was synthesized by the solid phase technique in our laboratory. All experiments were performed twice. All procedures were done in accordance with the Japanese Physiological Society’s guidelines for animal care. First, movement of rats ( n = 8 per group) that had received an ICV administration of NMU (1 nmol), CRH (1 nmol), or vehicle was measured using a Rat Locomotor Activity Recording Systems Device (Muromachi Co. Ltd., Tokyo, Japan) as described previously (14). After materials were injected to free-moving rats at 0900 h, they were immediately returned to the individual sound- and light-proof cages equipped with infrared light-beam detectors. Locomotor activ- ity counts were made every 15 min and summed up for 60 min after administration. Second, rats ( n = 10 per group) were administered an ICV injec- tion at 0900 h with the following reagents: NMU 1 nmol, NMU 1 nmol + -hCRH (50 g, Sigma Chemical Co., St. Louis, MO), NMU 1 nmol + anti-CRH IgG (1 g, Peptide Institute, Inc., Osaka, Japan), NMU 1 nmol + control serum IgG (1 g), CRH 1 nmol (Peptide Institute, Inc.), CRH 1 nmol + anti-CRH IgG, CRH 1 nmol + control serum IgG. IgG was injected 2 h before peptide administration. Locomotor activity counts were made as above. Third, rats ( n = 6 per group) were administered an ICV injection at 0900 h with following reagents: 5 nmol NMU, 5 nmol NMU + 100 1 To whom correspondence should be addressed. E-mail: nakazato@ post.miyazaki-med.ac.jp. Biochemical and Biophysical Research Communications 289, 225–228 (2001) doi:10.1006/bbrc.2001.5945, available online at http://www.idealibrary.com on 225 0006-291X/01 $35.00 Copyright © 2001 by Academic Press All rights of reproduction in any form reserved.