Day/night fluctuations in melatonin content, arylalkylamine N-acetyltransferase activity and NAT mRNA expression in the CNS, peripheral tissues and hemolymph of the cockroach, Periplaneta americana

Melatonin content measured by a radioenzymatic assay in the brain of the American cockroach (Periplaneta americana) showed a day/night fluctuation with higher levels at night under LD 12:12. The activity of arylalkylamine N-acetyltransferase (NAT) in brain was also higher at night and this pattern continued in constant darkness. The results suggest that the rhythmicity in melatonin content can be caused by NAT. Melatonin content in hemolymph showed an even greater day/night difference, more than 12 times that in brain under LD 12:12. Melatonin levels in retina were also higher at night while NAT activity was not significantly higher at night than at daytime. Using a probe designed from NAT cloned from testes we performed Northern blot analysis of total RNA, which revealed that the level of NAT mRNA was higher in midgut, ovary and female accessory glands than in fat body and brain. The level of transcript in midgut was higher at night, but the levels in ovary and female accessory reproductive gland showed the opposite pattern. We also used the antibody to whole Drosophila melanogaster aaNAT1 protein, seeking a homologous antigen in the cephalic ganglia. NAT-like antigen was detected in several restricted populations of cells in the brain that were partially co-localized with PER-like antigen. The results suggest that NAT exists in multiple forms in various tissues of the cockroach and that its functions and regulations can vary among tissues. The results in the brain led to the conclusion that NAT could be a clock-controlled gene functioning as an output regulator of the circadian clock.