FUNCTION OF LEPTIN AS A CARBOHYRDATE CATABOLIC STRESS HORMONE IN FISH: INSIGHTS FROM TRANSCRIPTOMIC AND PHYSIOLOGICAL STUDIES IN THE TILAPIA

Leptin is a cytokine that in mammals is thought to work primarily as an adipostat, whereby the hormone circulates in proportion to fat deposition and inhibits appetite and stimulates lipolysis and fatty acid oxidation to prevent excessive lipid accumulation.  Its function on energy homeostasis in fish is poorly understood despite leptin's well-conserved anorexigenic actions. We show that hepatic expression of leptin increases in response to seawater challenge and other catabolic stressors (fasting) and induces hyperglycemia and glycogenolysis in tilapia, Oreochromis mossambicus. A transcriptomic analysis of the tilapia pituitary rostral pars distalis (RPD, > 95% prolactin cells) was performed in order to identify novel cellular actions of leptin, and to characterize leptin effects on central genes involved in glucose metabolic pathways. Advanced clustering of the RNAseq data revealed that leptin stimulates the expression of the glycolytic enzyme, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), in a covariable manner to hypoxic stress response gene networks. Orthogonal tests show that recombinant tilapia leptin A (LepA), the dominant leptin paralog in fishes, increases the mRNA level of GAPDH and phosphofructokinase (PFK), the rate-limiting enzyme of glycolysis, after 6 h incubation of RPD. Likewise, leptin stimulates total glycolytic activity (lactate secretion) and PFK activity within 6 h. Glycolytic activity correlated significantly to both PFK mRNA levels and enzymatic activity. The potential signaling mechanisms for leptin action were also assessed. LepA stimulates STAT3 and ERK1 phosphorylation (activation) in the RPD. The stimulatory effect of leptin on glycolysis and PFK activity was suppressed by a STAT3 (Stattic) but not an ERK (PD98059) blocker, indicating the hormone stimulates glycolysis through a STAT3 mediated increases in glycolytic enzyme gene expression. Leptin stimulation of ERK signaling is most likely linked to leptin action in stimulating acute PRL release. To assess whether LepA might broadly regulate glycolysis, we tested its effects in another cell type, namely hepatocytes, where LepA is predominantly produced and may exert autocrine/paracrine effects. In hepatocyte incubations, LepA stimulates total glycolytic activity and PFK mRNA levels, but had little effect on GAPDH gene expression. Thus, leptin stimulation of GAPDH gene expression may be unique to the pituitary, however leptin stimulation of glycolysis as a whole, and PFK gene expression specifically, appear well conserved in multiple tissue types. These results identify a novel action of leptin as a direct stimulator of glycolysis through a STAT3-mediated mechanism in tilapia. Collectively, the results suggest that leptin enhances carbohydrate expenditure in tilapia by stimulating glycogenolysis and glycolysis.  This finding may represent an important function for leptin to enhance energy expenditure as a metabolic coping mechanism during the vertebrate adaptive stress response.  Understanding leptin's function in maintaining energy homeostasis will be important to enhancing growth and production of this, and other important cultivars.