Identification and characterization of the leech CNS cannabinoid receptor: coupling to nitric oxide release.
The present study demonstrates that stereoselective binding sites for anandamide, a naturally occurring cannabinoid substance, can be found in leech (Theromyzon tessulatum and Hirudo medicinalis) central nervous system. The anandamide binding site is monophasic and of high affinity exhibiting a Kd of approximately 32 nM with a Bmax of 550 fmol/mg protein in both animals. These sites are highly select as demonstrated by the inability of other types of signaling molecules to displace [3H]anandamide. Furthermore, this binding site is coupled to nitric oxide release. A deduced amino acid sequence (153 residues) analysis from a 480 pb amplified RT-PCR fragment cDNA exhibits a 49.3% and 47.2% sequence identity with human and rat cannabinoid receptors (CB1R), respectively. Thus, the leech cannabinoid receptor may be a G-protein coupled receptor with seven transmembrane domains as in CB1R. Moreover, this sequence exhibits highly conserved regions, particularly in the putative transmembrane domains 1 and 2. The presence of a cannabinoid receptor in these organisms indicates that this signaling system has been conserved during evolution.
Stefano
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GB
Neuroscience Research Institute, State University of New York at Old Westbury, 11568, USA. stefanog@surg.som.sunysb.edu
Salzet
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B
Salzet
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M
D000595:Amino Acid Sequence; D000818:Animals; D001095:Arachidonic Acids / Q000378:metabolism; D001483:Base Sequence; D002186:Cannabinoids / Q000378:metabolism; D002490:Central Nervous System / Q000378:metabolism*; D063388:Endocannabinoids; D006801:Humans; D007865:Leeches / Q000378:metabolism*; D008969:Molecular Sequence Data; D009569:Nitric Oxide / Q000378:metabolism*; D053284:Polyunsaturated Alkamides; D051381:Rats; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug / Q000235:genetics / Q000378:metabolism*
D001095:Arachidonic Acids; D002186:Cannabinoids; D063388:Endocannabinoids; D053284:Polyunsaturated Alkamides; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug; D009569:Nitric Oxide; C078814:anandamide