title: Glucose Alters Per2 Rhythmicity Independent of AMPK, Whereas AMPK Inhibitor Compound C Causes Profound Repression of Clock Genes and AgRP in mHypoE-37 Hypothalamic Neurons.
abstract: Specific neurons in the hypothalamus are regulated by peripheral hormones and nutrients to maintain proper metabolic control. It is unclear if nutrients can directly control clock gene expression. We have therefore utilized the immortalized, hypothalamic cell line mHypoE-37, which exhibits robust circadian rhythms of core clock genes. mHypoE-37 neurons were exposed to 0.5 or 5.5 mM glucose, comparable to physiological levels in the brain. Per2 and Bmal1 mRNAs were assessed every 3 hours over 36 hours. Incubation with 5.5 mM glucose significantly shortened the period and delayed the phase of Per2 mRNA levels, but had no effect on Bmal1. Glucose had no significant effect on phospho-GSK3β, whereas AMPK phosphorylation was altered. Thus, the AMPK inhibitor Compound C was utilized, and mRNA levels of Per2, Bmal1, Cryptochrome1 (Cry1), agouti-related peptide (AgRP), carnitine palmitoyltransferase 1C (Cpt1c), and O-linked N-acetylglucosamine transferase (Ogt) were measured. Remarkably, Compound C dramatically reduced transcript levels of Per2, Bmal1, Cry1, and AgRP, but not Cpt1c or Ogt. Because AMPK was not inhibited at the same time or concentrations as the clock genes, we suggest that the effect of Compound C on gene expression occurs through an AMPK-independent mechanism. The consequences of inhibition of the rhythmic expression of clock genes, and in turn downstream metabolic mediators, such as AgRP, could have detrimental effects on overall metabolic processes. Importantly, the effects of the most commonly used AMPK inhibitor Compound C should be interpreted with caution, considering its role in AMPK-independent repression of specific genes, and especially clock gene rhythm dysregulation.
authors.affiliation: Department of Physiology, University of Toronto, Toronto, ON, Canada.|Department of Endocrinology and Metabolism, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Johanneke E
authors.initials: JE
authors.lastname: Oosterman
authors.affiliation: Department of Physiology, University of Toronto, Toronto, ON, Canada.|Department of Obstetrics and Gynecology, University of Toronto, Toronto, ON, Canada.|Department of Medicine, University of Toronto, Toronto, ON, Canada.|Division of Cellular and Molecular Biology, Toronto General Hospital Research Institute, University Health Network, Toronto, ON, Canada.
authors.forename: Denise D
authors.initials: DD
authors.lastname: Belsham
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D056930:ARNTL Transcription Factors / Q000235:genetics; D054369:Agouti-Related Protein / Q000235:genetics; D000818:Animals; D056926:CLOCK Proteins / Q000235:genetics; D002460:Cell Line; D002940:Circadian Rhythm / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects*; D005947:Glucose / Q000494:pharmacology*; D007031:Hypothalamus / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D056950:Period Circadian Proteins / Q000235:genetics*; D010766:Phosphorylation / Q000187:drug effects; D011720:Pyrazoles / Q000494:pharmacology*; D011743:Pyrimidines / Q000494:pharmacology*
chemical_list: D056930:ARNTL Transcription Factors; D054369:Agouti-Related Protein; C515199:Agrp protein, mouse; C538961:Bmal1 protein, mouse; C513312:Per2 protein, mouse; D056950:Period Circadian Proteins; D011720:Pyrazoles; D011743:Pyrimidines; C516138:dorsomorphin; D056926:CLOCK Proteins; D055372:AMP-Activated Protein Kinases; D005947:Glucose