Inhibition of agouti-related peptide expression by glucose in a clonal hypothalamic neuronal cell line is mediated by glycolysis, not oxidative phosphorylation

The regulation of neuroendocrine electrical activity and gene expression by glucose is mediated through several distinct metabolic pathways. Many studies have implicated AMP and ATP as key metabolites mediating neuroendocrine responses to glucose, especially through their effects on AMP-activated protein kinase (AMPK), but other studies have suggested that glycolysis, and in particular the cytoplasmic conversion of nicotinamide adenine dinucleotide (NAD +) to reduced NAD (NADH), may play a more important role than oxidative phosphorylation for some effects of glucose. To address these molecular mechanisms further, we have examined the regulation of agouti-related peptide (AgRP) in a clonal hypothalamic cell line, N-38. AgRP expression was induced monotonically as glucose concentrations decreased from 10 to 0.5 mM glucose and with increasing concentrations of glycolytic inhibitors. However, neither pyruvate nor 3-beta-hydroxybutyrate mimicked the effect of glucose to reduce AgRP mRNA, but on the contrary, produced the opposite effect of glucose and actually increased AgRP mRNA. Nevertheless, 3 beta-hydroxybutyrate mimicked the effect of glucose to increase ATP and to decrease AMPK phosphorylation. Similarly, inhibition of AMPK by RNA interference increased, and activation of AMPK decreased, AgRP mRNA. Additional studies demonstrated that neither the hexosamine nor the pentose/ carbohydrate response element-binding protein pathways mediate the effects of glucose on AgRP expression. These studies do not support that either ATP or AMPK mediate effects of glucose on AgRP in this hypothalamic cell line but support a role for glycolysis and, in particular, NADH. These studies support that cytoplasmic or nuclear NADH, uniquely produced by glucose metabolism, mediates effects of glucose on AgRP expression.