Age-related changes in 2-[(125)I]-iodomelatonin binding sites in the brain of sea breams (Sparus aurata, L.)

The pineal organ of fish, through its 24h rhythmic release of melatonin, acts as a transducer of the photoperiod, influencing different physiological functions (e.g., reproduction, growth). The target sites for melatonin are poorly known in fish, especially marine species. A radioligand study was undertaken using the gilthead sea bream (Sparus aurata) maintained under natural temperature and photoperiod (at 28°N latitude). This species exhibits the property of changing sex during growth. Brains of one year-old males were collected at 16:00h and brains of three year-old females at 03:00, 10:00, 16:00 and 23:00h. Membrane homogenate receptor assays were run using 2-[(125)I]iodomelatonin as a ligand. Binding sites were detected in brains of young and old fish. In the younger, the exhibited a Bmax between 3.52 and 4.29 fmol mg protein(-1) and a KD between 358-380 pmol l(-1). In the older fish, the KD varied according to a daily pattern: values were three times higher at 03:00 and 10:00h (500-600 pmol l(-1)) than at 16:00 and 23:00h (150-300 pmol l(-1)). The number of sites also were higher at 03:00 and 10:00h (180-200 fmol mg protein(-1)) than at 16:00 and 23:00h (95-110 fmol mg protein(-1)). Melatonin and iodomelatonin displaced 2-[(125)I]iodomelatonin binding in a dose dependent manner, the second being more potent than the first. Binding was also inhibited by GTP. The results provide the first evidence for the presence of membrane melatonin binding sites in the brain of an exclusively marine fish. They suggest that their number and affinity varies during growth and throughout a light/dark cycle. Future experiments will aim to precise the anatomical location and role of these binding sites.