Peptides, Vol. 7, pp. 815-820, 1986. © Ankho International Inc. Printed in the U.S.A. 0196-9781/86 $3.00 + .00 Neuromedin B-Like Peptides in Rat Brain: Biochemical Characterization, Mechanism of Release and Localization in Synaptosomes TERRY W. MOODY,*' LOUIS Y. KORMANt AND THOMAS L. O'DONOHUE:~ *Department of Biochemistry, The George Washington University School of Medicine and Health Sciences Washington, DC 20037 tGastroenterology Section, VA Medical Center, Washington, DC 20901 and SExperimental Therapeutics Branch National Institute of Neurological and Communicative Disorders and Stroke National Institutes of Health, Bethesda, MD 20205 Received 31 March 1986 MOODY, T. W., L. Y. KORMAN AND T. L. O'DONOHUE. Neuromedin B-like peptides in rat brain: Biochemical characterization, mechanism of release and localization in synapto.~omes. PEPTIDES 7(5) 815-820, 1986.--Neuromedin B-like peptides were characterized in the rat brain. A rabbit antisera was utilized which recognized neuromedin B but not bombesin or GRP. Using gel filtration and HPLC techniques, a major and minor peak of immunoreactivity was present in rat brain extracts. In both cases the main peak of immunoreactivity coeluted with synthetic neuromedin B. The density of neuromedin B-like peptides ranged 50-fold being greatest in the olfactory bulb and hypothalamus, intermediate in the hippocampus, spinal cord, medulla/pons, pituitary, midbrain, thalamus, striatum and cortex and lowest in the cerebellum. Release studies indicated that neuromedin B-like peptides were secreted from bypothalamic, olfactory bulb and thalamic slices in a Ca++-dependent manner when KCI (75 mM) was present. Also, the neuromedin B-like peptides in the rat brain were localized to synaptosomes. These data indicate that neuromedin B-like peptides may function as regulatory peptides in the CNS distinct from bombesin/GRP. Neuromedin B-like peptides Synaptosomes lmmunoreactivity Localization Characterization ONE class of peptides biologically active in the CNS is rep- resented by bombesin/gastrin releasing peptide (BN/GRP). In the CNS, BN/GRP is a satiety agent after direct injection into the hypothalamus [28,30] and inhibits the increase in gastric acid secretion caused by various chemical and surgi- cal procedures in rats [29]. Also, intraventricular injection of BN/GRP causes intense grooming in rodents [6, 8, 10, 12] and intraspinal injection elicits a vigorous and long lasting bite scratch response [22]. These actions may be mediated by the endogenous BN/GRP-Iike peptides which are dis- cretely distributed in the rat brain and spinal cord [20, 24, 26]. These BN/GRP-like peptides are localized to synapto- somes and are released by depolarizing stimuli in a Ca + ~- dependent manner [18,19]. When released from CNS neurons, these BN/GRP-like peptides likely interact with re- ceptors which have been detected in discrete brain regions [31,32]. Thus, BN/GRP-like peptides may function as regula- tory agents in the CNS. The structural similarities between members of the BN family of peptides is shown in Table 1. BN, initially isolated from frog skin [1], GRP, isolated from porcine nonantral stomach tissue [11] and neuromedin C, which was purified from porcine spinal cord [14] have a common C-terminal heptapeptide. In contrast, neuromedin B, which was se- quenced from porcine spinal cord [13], has 7 of the same 10 C-terminal amino acid residues as does BN. Recently, Minamino et al. [15] used an antiserum which reacted with neuromedin B and ranatensin but not bombesin/GRP and determined that high levels of endogenous neuromedin B were present in the rat brain. Here we investigated the dis- tribution of neuromedin B in the rat CNS using an antiserum which recognized neuromedin B but not ranatensin or 'Requests for reprints should be addressed to Dr. Terry W. Moody. Department of Biochemistry, GWU Medical Center, 2300 Eye St., N.W.. Washington, DC 20037. 815