Vol. 285, Issue 3, 1239-1245, June 1998

GR196429: A Nonindolic Agonist at High-Affinity Melatonin¹ Receptors

Isabel J. M. Beresford, Christopher Browning, Sarah J. Starkey, Jason Brown, Steven M. Foord, Josephine Coughlan, Peter C. North, Margarita L. Dubocovich² and Russell M. Hagan

Glaxo Wellcome Research and Development, Ltd., Medicines Research Centre, Stevenage, Hertfordshire, United Kingdom

N-[2-[2,3,7,8-tetrahydro-1H-furo(2,3-g)indol-1-yl]ethyl]acetamide (GR196429) is a novel, nonindolic melatonin receptor agonist. GR196429 had high affinity for human mt₁ (pK_i 9.9) and MT₂ (pK_i 9.8) receptors expressed in Chinese hamster ovary cells and for 2-[¹²⁵I]-iodomelatonin binding sites in human cerebellum, guinea pig superior colliculus and hypothalamus and chicken retina and tectum (pK_i 8.8-9.5). GR196429 was inactive at a wide range of other hormone and neurotransmitter receptors. In Chinese hamster ovary cells expressing human mt₁ or MT₂ receptors, both melatonin and GR196429 dose-dependently inhibited forskolin-stimulated cAMP accumulation. In rabbit isolated retina, GR196429 inhibited calcium-dependent [³H]-dopamine release with potency (IC₅₀ 30 pM) and maximum effect (76 ± 5% at 1 nM) similar to those of melatonin. The response was antagonized by the melatonin receptor antagonist luzindole (1 µM). In slices of rat brain suprachiasmatic nucleus, perfusion (1 h) with GR196429 at zeitgeber time 10 phase advanced the circadian peak in neuronal activity measured on the following day, with a maximum phase advance of 2.7 ± 0.3 h at 10 pM and an EC₅₀ of 0.6 pM, results that indicated a melatonin-like action on the phase of the circadian clock. CNS penetration and duration of receptor occupancy was determined in an ex vivo radioligand binding assay. In membranes of guinea pig superior colliculus prepared 30 min after administration of GR196429 (s.c.), 2-[¹²⁵I]-iodomelatonin binding was inhibited with an ED₅₀ of 0.04 mg/kg. After a dose of 1 mg/kg, binding was significantly inhibited for at least 3 h. Thus GR196429 is a potent and selective agonist at high-affinity melatonin receptors, which modulates circadian rhythms in an in vitro model of the circadian clock and which readily penetrates the CNS.