Ž . Brain Research 763 1997 63–68 Research report Morphine- and anandamide-stimulated nitric oxide production inhibits presynaptic dopamine release George B. Stefano a,c, ) , Beatrice Salzet c , Christos M. Rialas a , McElvin Pope a , Adam Kustka b , Kevin Neenan a , Stephen Pryor a , Michel Salzet a,c a Multidisciplinary Center for the Study of Aging, Neuroscience Research Institute, State UniÕersity of New York, College at Old Westbury, Old Westbury, NY 11568, USA b Marine Sciences Center, State UniÕersity of New York at Stony Brook, Stony Brook, NY 11794, USA c Centre de Biologie Cellulaire, Laboratoire de Phylogenie Moleculaire des Annelides EA DRED 1027, Groupe de Neuroendocrinologie des Hirudinees, ´ ´ ´ ´ UniÕersite des Sciences et Technologies de Lille, 59655 VilleneuÕe d’Ascq, Cedex, France ´ ´ Accepted 25 March 1997 Abstract Ž . Morphine and anandamide stimulate the release of nitric oxide NO in diverse tissues. The present study examines the consequences of this action on neurotransmitter release in ganglia from two invertebrates: ventral chain ganglia from the leech Hirudo medicinalis and Ž . Ž . the pedal ganglion from the mussel Mytilus edulis. In these ganglia, preloaded serotonin 5-HT and dopamine DA can be released by w 3 x Ž . 50 mM KCl. Anandamide, an endogenous cannabinoid substance, suppresses the potassium-stimulated release of H DA 80% , but not 5-HT, in a concentration-dependent manner, from the neural tissues of both. The effect of anandamide can be antagonized by pre-exposing the neural tissues of both animals to SR 141716A, a potent cannabinoid receptor antagonist. Prior treatment of the ganglia Ž . with N-v-nitro-L-arginine methyl ester L-NAME , a nitric oxide synthase inhibitor, significantly diminishes the inhibitory effect of w 3 x anandamide. Morphine also inhibits H DA release in a naloxone- and L-NAME-sensitive manner. Anandamide and morphine act through separate mechanisms since the respective antagonists show no cross-reactivity. The NO donor, SNAP, depressed the w 3 x potassium-stimulated release of preloaded H DA, but not 5-HT, in the neural tissues of both animals. D-Ala -Met enkephalinamide 2 5 Ž . w 3 x DAMA also inhibited the potassium-stimulated release of H DA in a naloxone-sensitive process. However, the effect of DAMA was Ž y4 . seen in the presence of L-NAME 10 M , indicating that the opioid peptide inhibition of the presynaptic release of DA is not coupled to NO. We postulate that cannabinoids and their endogenous effectors play a prominent role in the regulation of catecholamine release in invertebrates via NO release as is the case for opiate alkaloids. q 1997 Elsevier Science B.V. Keywords: Morphine; Invertebrate ganglia; Dopamine; Anandamide; Serotonin; Presynaptic release 1. Introduction Anandamide is a naturally occurring cannabinoid sub- Ž . stance that can induce the release of nitric oxide NO from human monocytes, invertebrate immunocytes and w x microglia 25–27 . We have previously demonstrated that in invertebrates and human monocytes anandamide bind- ing sites are monophasic, high affinity, stereoselective and coupled NO release. In the leech, the ventral chain ganglia respond to anandamide by releasing NO; this process can be antagonized by a cannabinoid antagonist as well as the nitric oxide synthase inhibitor N-v-nitro-L-arginine methyl ) Ž . Corresponding author, at address a. Fax: q1 516 876-2727; E-mail: stefanog@surg.som.sunysb.edu Ž w x. ester L-NAME; 27 . These cannabinoid actions resemble those of opiate alkaloids in that they release NO but use w x separate receptors 21,22,25 . Given the similarity of opiate and cannabinoid pro- cesses we surmised that monoaminergic neurons might be the target for the naturally occurring cannabinoid ligands since opiate signaling molecules inhibit catecholamine re- w x lease in invertebrate neural tissues 21 . In the present study we demonstrate that anandamide selectively inhibits Ž . the presynaptic release of dopamine DA from the pedal ganglia of Mytilus edulis and from the ventral chain ganglia of the leech Hirudo medicinalis. We also demon- strate that the presynaptic release of DA can be inhibited by opioid alkaloids and opioid peptides; however, only the opiate alkaloids act via the release of NO. These data 0006-8993r97r$17.00 q 1997 Elsevier Science B.V. All rights reserved. Ž . PII S0006-8993 97 00403-4