Cannabinomimetic behavioral effects of and adenylate cyclase inhibition by two new endogenous anandamides

We have previously shown that the endogenous putative cannabinoid ligand arachidonylethanolamide (anandamide, 20:4, n - 6) induces in vivo and in vitro effects typical of a cannabinoid agonist. We now report that two other endogenous anandamides, docosatetraenylethanolamide (anandamide, 22:4, n - 6) and homo-γ-linolenylethanolamide (anandamide, 20:3, n - 6), have similar activities. The new anandamides bind to SV40-transformed African green monkey kidney cells transfected with the rat brain cannabinoid receptor cDNA and display K_I values of 253.4 ± 41.1 and 244.8 ± 38.7, respectively. The value found for arachidonylethanolamide was 155.1 ± 13.8 nM. In addition, the new anandamides inhibit prostaglandin E₁-stimulated adenylate cyclase activity in Chinese hamster ovary-K₁ cells transfected with the cannabinoid receptor, as well as in N₁₈TG₂ mouse neuroblastoma cells that express the cannabinoid receptor naturally. The IC₅₀ values for the inhibition of adenylate cyclase in transfected Chinese hamster ovary-K₁ cells were 116.8 ± 8.7 and 109.3 ± 8.6 nM for docosatetraenylethanolamide and homo-γ-linolenylethanolamide, respectively. These values were similar to that obtained with arachidonylethanolamide (100.5 ± 7.7 nM), but were significantly higher than the IC₅₀ value observed with the plant cannabinoid Δ⁹-tetrahydrocannabinol (9.2 ± 8.6 nM). The inhibitory effects of the anandamides on adenylate cyclase activity were blocked by pertussis toxin, indicating the involvement of pertussis toxin-sensitive GTP-binding protein(s). In a tetrad of behavioral assays for cannabinoid-like effects, the two new anandamides exerted similar behavioral effects to those observed with Δ⁹-tetrahydrocannabinol and arachidonylethanolamide: inhibition of motor activity in an open field, hypothermia, catalepsy on a ring, and analgesia on a hot plate.