A Novel Subtype of Prostacyclin Receptor in the Central Nervous System *²Yumiko Watanabe, *²Kiyoshi Matsumura, *²Hajime Takechi, ‡Koichi Kato, *²Hiroshi Morii, *¶Margareta Bjo ¨rkman, *¶Bengt Långstro ¨m, ‡Ryoji Noyori, *§Masaaki Suzuki, and *²Yasuyoshi Watanabe *Subfemtomole Biorecognition Project, ICORP, Japan Science and Technology Corporation, and ² Department of Neuroscience, Osaka Bioscience Institute, Osaka, ‡Department of Chemistry and Molecular Chirality Research Unit, Nagoya University Faculty of Science, Nagoya, and §Department of Biomolecular Science, Faculty of Engineering, Gifu University, Gifu, Japan; and Uppsala University PET Centre and ¶Department of Organic Chemistry, Institute of Chemistry, Uppsala, Sweden Abstract: Recently, in the course of our search for the prostacyclin receptor in the brain, we found a novel subtype, designated as IP 2 , which was finely discrim- inated by use of the specific ligand (15R)-16-m-tolyl- 17,18,19,20-tetranorisocarbacyclin (15R-TIC) and spe- cifically localized in the rostral part of the brain. In the present study, the tritiated compound 15R-[15- 3 H]TIC was synthesized and utilized for more specific research on IP 2 . The specificity of binding to rat brain regions was confirmed by use of several prostacyclin deriva- tives including 15S-TIC. Mapping of 15R- and 15S- [ 3 H]TIC binding in adjacent pairs of frozen sections of rat brain demonstrated a quite similar pattern of distri- bution in almost all rostral brain regions, indicating that the regions may contain only the IP 2 subtype. On the other hand, 15R-[ 3 H]TIC binding was very faint as com- pared with 15S-[ 3 H]TIC binding in the caudal medullary region. High densities of 15R-[ 3 H]TIC binding sites were shown in the dorsal part of the lateral septal nucleus, thalamic nuclei, limbic structures, and some of the cortical regions. Scatchard plot analysis showed two components of high-affinity 15R-[ 3 H]TIC binding in the rostral regions, one with a K D value at 1nM and the other with 30 nM. These results strengthen our previous finding that a different subtype of prostacyclin receptor is expressed in the CNS, and the map with 15R-[ 3 H]TIC obtained here could guide further studies on the molecular and functional properties of the IP 2 . Key Words: Prostacyclin receptor—Central nervous system—Tolyl-isocarbacyclin. J. Neurochem. 72, 2583–2592 (1999). Prostaglandin (PG) I 2 (prostacyclin) showed a protec- tive effect against neuronal cell death and damage after experimental ischemia (Cazevieille et al., 1993, 1994). However, the existence of a prostacyclin receptor in the neuronal cells had not been known until we (Matsumura et al., 1995b; Takechi et al., 1996) reported the prosta- cyclin receptor in the brain. Our recent study (Takechi et al., 1996) revealed that two distinct types of prostacyclin receptors exist in the rat brain in terms of their ligand specificity. In the caudal medulla such as the nucleus of the solitary tract (NTS), [ 3 H]iloprost and [ 3 H]isocarba- cyclin, both of which are stable agonists for the known and cloned prostacyclin receptor in platelets and various peripheral tissues (Boie et al., 1994; Namba et al., 1994; Sasaki et al., 1994), showed high affinity with K D values in a range of several nanomolar. On the other hand, in the rostral regions of the brain, [ 3 H]iloprost bound with very low affinity (K D = 159 nM in the thalamus), whereas [ 3 H]isocarbacyclin bound with comparably high affinity (K D = 7.8 nM ) there. From ganglionectomy and nerve ligation experiments, the receptor for [ 3 H]iloprost bind- ing in the caudal medulla was shown to originate in the sensory ganglia and be transported to the central termi- nus of the primary sensory afferents as well as to their peripheral terminals (Matsumura et al., 1995b). In situ hybridization for a cloned peripheral-type prostacyclin receptor showed no significant signals in the rostral re- gions of the brain, other than in the smooth muscle cells of arteries, but a high density of signals in the neurons of the sensory ganglia (Oida et al., 1995). Another impor- tant finding (Takechi et al., 1996) was obtained through displacement studies on [ 3 H]isocarbacyclin binding in both thalamus and NTS: (15R)-16-m-tolyl-17,18,19,20- tetranorisocarbacyclin (15R-TIC) was shown to be highly selective for binding to the rostral regions of the brain. These results clearly indicated the presence of one Received October 15, 1998; revised manuscript received January 25, 1999; accepted January 25, 1999. Address correspondence and reprint requests to Dr. Y. Watanabe at Department of Neuroscience, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita-shi, Osaka 565-0874, Japan. Abbreviations used: IP 1 and IP 2 , peripheral and central type of prostacyclin receptor, respectively; NTS, nucleus of the solitary tract; PG, prostaglandin; TIC, 16-m-tolyl-17,18,19,20-tetranorisocarbacy- clin. 2583 Journal of Neurochemistry Lippincott Williams & Wilkins, Inc., Philadelphia © 1999 International Society for Neurochemistry