Melatonin Receptor‐Mediated Stimulation of Phosphoinositide Breakdown in Chick Brain Slices

The direct effect of melatonin and related agonists on Li+‐amplified phosphoinositide breakdown was studied in chick brain slices prelabeled with myo‐[2‐3H]‐inositol. The melatonin receptor agonist 6‐chloromelatonin (10–100 µM) increased, in a concentration‐dependent manner, the accumulation of inositol phosphates (IP) in chick brain slices. This effect of 6‐chloromelatonin (10 µM) was rapid as transient increases in IP3/IP4 (maximal increase, 29% at 20 s) and IP2 levels (maximal increase, 36% at 1 min) were observed, followed by a slower but sustained increase in IP1 level (30% at 5 min), when the amount of IP3/IP4 and IP2 had already been decreased to the control level. The phosphoinositide response elicited by 6‐chloromelatonin (10 µM) was dependent on the presence of extracellular calcium. Direct stimulation of membrane phospholipase C by 6‐chloromelatonin (10 µM) in isolated myo‐[2‐3H]inositol‐prelabeled optic tectum membranes was dependent on the presence of guanosine‐5′‐O‐(3‐thio)triphosphate (1 µM), thus suggesting that G protein(s) link melatonin receptor activation to phospholipase C stimulation. The competitive melatonin receptor antagonist luzindole (10–100 µM) inhibited in a concentration‐dependent manner the IP1 accumulation stimulated by 6‐chloromelatonin (10–100 µM); however, it did not affect the accumulation stimulated by 5‐hydroxytryptamine (10 µM). By contrast, methysergide (10 µM) completely inhibited 5‐hydroxytryptamine (10 µM)‐, but not 6‐chloromelatonin (10 µM)‐, induced IP1 accumulation. Melatonin receptor agonists increased IP1 accumulation in a concentration‐dependent manner reaching different maximal responses. N‐Acetyl‐5‐hydroxytryptamine was more potent than melatonin in increasing IP1 accumulation, suggesting activation of a melatonin receptor site other than the ML‐1 melatonin receptor (i.e., N‐acetyl‐5‐hydroxytryptamine ≥ melatonin). In conclusion, these results demonstrate that activation of a melatonin receptor with pharmacological characteristics different from those of the ML‐1 subtype leads to activation of the phospholipase C‐mediated signal transduction pathway.