Free nucleotides in pupal brains were examined in a bivoltine race (Daizo) of the silkworm, Bombyx mori, using high pressure liquid Chromatography (HPLC). Concentrations were nearly the same level as observed in rat brains. Major differences in monophosphates, diphosphates and triphosphates of purine and pyrimidine nucleotides, and an unidentified nucleotide derivative were observed between silkworms which were exposed to regimens of continuous light and warm conditions (27 °C) and those exposed to continuous darkness and cold conditions (15 °C) throughout maternal embryonic development. In particular, concentrations of ADP, ATP, GDP, GTP, IDP, ITP, UDP and UTP were more abundant in the pupal brains of insects exposed to continuous light and warm conditions. When brain-subesophageal ganglion complexes from non-diapause egg producers were cultured with mixtures of nucleotides, the resulting culture medium was highly effective in eliciting diapause in non-diapause egg producers. These results suggest that brain nucleotides have a role in diapause induction.