| Summary: | Food deprivation activates forkhead box O (FOXO), a transcription factor downstream of insulin receptors. In the pond snail <i>Lymnaea stagnalis</i>, insulin signaling and food deprivation improve memory consolidation following conditioned taste aversion (CTA) learning. We investigated the subcellular localization of FOXO in <i>Lymnaea</i> and changes in its expression levels following food deprivation, CTA learning, and insulin administration. Immunohistochemistry revealed that <i>Lymnaea</i> FOXO (LymFOXO) was located in the central nervous system (CNS) neuronal cytoplasm in food-satiated snails but was mainly in neuronal nuclei in food-deprived snails. Following CTA acquisition, LymFOXO translocated to the nuclei in food-satiated snails and remained in the nuclei in food-deprived snails. Contrary to our expectations, insulin administered to the CNS did not induce LymFOXO translocation into the nuclei in food-satiated snails. Real-time PCR was used to quantify LymFOXO mRNA levels, its target genes, and insulin signaling pathway genes and revealed that LymFOXO mRNA was upregulated in food-deprived snails compared to food-satiated snails. Insulin applied to isolated CNSs from food-satiated snails increased LymFOXO compared to vehicle-treated samples. Food deprivation prepares FOXO to function in the nucleus and enhances CTA learning in snails. Insulin application did not directly affect LymFOXO protein localization. Thus, insulin administration may stimulate pathways other than the LymFOXO cascade.
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