We studied temporal organization of both the photoreceptor (rod-like opsin, alpha subunit of the G protein transducin or alpha-TD) and melatonin generating (AANAT) proteins in the same pineal of a tropical surface dwelling free-living carp Catla catla, and analyzed possible correlation between them as well as with natural photo-thermal variables in an annual cycle. The pineal from individual fish was collected at four different time points (06.00 h, 12.00 h, 18.00 h, and 24.00 h) in a 24.00 h cycle and the same was repeated in four distinct seasons in an annual reproductive cycle to study each protein following Western blot and densitometric analyses of respective immunoblots. The rod-like opsin was represented by four distinct bands, a closely spaced doublet of 39 kDa and bands of 78 and 115 kDa. Two separate bands, one at 43 kDa and another at 65 kDa, were detected for alpha-TD, and a single band at 23 kDa for AANAT. Both the pineal photoreceptor proteins exhibited an identical pattern of diurnal variations with a peak at midday (12.00 h) and fall at midnight (24.00 h), while maximum band intensity of AANAT was noted in midnight (24.00 h) and minimum at midday (12.00 h) depicting a significant negative correlation (p<0.001) between them. Likewise, in an annual cycle, a significant (p<0.01) negative correlation was found between the expression of each pineal photoreceptor protein (being highest during the spawning phase) and AANAT (maximum during the post-spawning phase). Seasonal fluctuations of both the photoperiod and water temperature exhibited a significant (p<0.01) positive correlation with the expression of pineal photoreceptor proteins and a significant (p<0.05) negative correlation with the pineal AANAT. Collectively, the present phenological study is the first report on temporal organization of pineal photoreceptor proteins and their correlation with the melatonin rhythm-generating enzyme AANAT as well as environmental photo-thermal cues depicting their integrative role in the synthesis of proteins in the pineal in any fish. Nonetheless, importance of further experimental studies on carp is emphasized for a conclusive evidence of functional relationship between the studied variables in the pineal and the components of environment in which the fish live in.
