ELSEVIER Journal of ChemicalNeuroanatomy 10 (1996) 287-293 Autoradiographic localizaton of [3H]-cyclic GMP binding sites in the rat brain Christopher Bladen, David Loewen, Steven R. Vincent* Division of Neurological Sciences, Department of Psychiatry, The University of British Columbia, Vancouver, B. C, V6T I Z3, Canada Received 4 September 1995;revised 1 February 1996;accepted I February 1996 Alma.act Both the atriopeptides and nitric oxide act in the nervous system by activating guanylyl cyclases to stimulate the production of cyclicGMP. Thus a key to understanding the roles of these messengers is to understand the functions of cyclic GMP in the nervous symm. Three potential targets for cyclic GMP have been identified, phosphodiesterases, protein kinases and ion channels. In this study we _~_a,~i_ "be a method using autoradiography to localize specific [3H]-cGMPbinding sites in the brain. The specificbinding of [)H]-cGMP to rat brain sections was saturable (Bnm= 1.5 pmoi/mg protein) and of high atYmity (Kv = 164 nM). The phar- macological characteristics were consistent with binding to the cGMP-dependent protein kinase. Highest densities of binding were seen in the medial habenula, basal ganglia, locus ceruleus and nucleus of the solitary tract. The CA1 pyramidal cells of the hip- pocampus, the neocortex, thalamus and cerebellum were also labelled. This method should prove useful in studies of potential tar- gets for cyclic GMP in the brain. Keywords: cGMP-dependent protein kinase; Nitric oxide; Guanylyl cyclase 1. lltrodu~on Nitric oxide and the atriopeptides appear to affect their target cells by causing the synthesis of cGMP. The atriopeptides act on cell surface receptor molecules which have a guanylyl cyclase moiety in their intracellu- lar domain. In contrast, nitric oxide interacts with a heine group associated with a soluble heterodimeric type of guanylyl cyclase to stimulate cGMP synthesis (Koesl- ing et al., 1995). There is a complementary distribution of NO synthase and NO stimulated cGMP accumula- tion in the brain (Southam and Garthwaite, 1993). How- ever, a major question concerning signal transduction by both the atriopeptides and NO is the role of cGMP. Three main targets for cGMP have been described, cGMP-dependent protein kinases, cGMP-gated ion channels and cGMP.regnlated phosphodiesterases (Lin- coin and Cornwell, 1993). Two types of cGMP- dependent serine/threonine protein kinases have been * Corrmponding author. Tel.: (604) 822-7038; fax: (604) 822-7981. 0891-0618/96/$15.00 © 1996 ElsevierScienceB.V. All rights leserved 1"11:S0891-06 ! 8(96)00146-9 identified in brain, the type I, which is primarily express- ed in the Puridnje cells of the cerebellar cortex (Lohmann et al., 1981), and the more widely distributed type II (EI-Husseini et al., 1995a). Both kinases are ac- tivated by low concentrations of cGMP. Cyclic nucleo- tide gated ion channels have been identified in sensory cells in the retina and the olfactory epithelium (Kaup, 1991). Recently, the olfactory type channel, which can be gated by low concentrations of either cAMP or cGMP, has been shown to be expressed in restricted regions of the brain (E1-Husseini et al., 1995b). It may thus serve as a target for cGMP in some neurons. A large family of phosphodiesterases has been described, some of which specifically hydrolyze cGMP, while others can be activated by or inhibited by cGMP (Beavo and Reifsnyder, 1990). The phosphodiesterases are widely expressed, and these latter types may thus be important physiological targets for cGMP in neurons. Autoradiographic binding studies have recently been used to localize components of a variety of signal trans- duction pathways in the brain. Thus [3H]-forskolin has