Melatonin sensitizes human myometrial cells to oxytocin in a protein kinase C alpha/extracellular-signal regulated kinase-dependent manner.
BACKGROUND
Studies have shown that labor occurs primarily in the night/morning hours. Recently, we identified the human myometrium as a target for melatonin (MEL), the neuroendocrine output signal coding for circadian night.


OBJECTIVE
The purpose of this study was to determine the signaling pathway underlying the effects of MEL on contractility and the contractile machinery in immortalized human myometrial cells.


METHODS
To ascertain the signaling pathway of MEL leading to its effects on myometrial contractility in vitro, we performed gel retraction assays with cells exposed to iodo-MEL (I-MEL) with or without oxytocin and the Rho kinase inhibitor Y27632. I-MEL effects on inositol trisphosphate (IP(3))/diacylglycerol (DAG)/protein kinase C (PKC) signaling were also investigated. Additionally, we assayed for caldesmon phosphorylation and ERK1/2 activation.


RESULTS
I-MEL was found to activate PKC alpha via the phospholipase C/IP(3)/DAG signaling pathway, which was confirmed by PKC enzyme assay. I-MEL did not affect myosin light chain phosphatase activity, and its effects on contractility were insensitive to Rho kinase inhibition. I-MEL did increase phosphorylation of ERK1/2 and caldesmon, which was inhibited by the MAPK kinase inhibitor PD98059 or the PKC inhibitor C1.


CONCLUSIONS
MEL sensitizes myometrial cells to subsequent procontractile signals in vitro through activation of the phospholipase C/IP(3)/DAG signaling pathway, resulting in specific activation of PKC alpha and ERK1/2, thereby phosphorylating caldesmon, which increases actin availability for myosin binding and cross-bridging. In vivo, this sensitization would provide a mechanism for the increased nocturnal uterine contractility and labor that has been observed in late-term human pregnancy.
Sharkey
James T
JT
Florida State University College of Medicine, Department of Biomedical Sciences, 1115 West Call Street, Tallahassee Florida 32306-4300, USA.
Cable
Casey
C
Olcese
James
J
D015153:Blotting, Western; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D005260:Female; D006801:Humans; D047468:Immunoprecipitation; D007293:Inosine Triphosphate / Q000378:metabolism; D008550:Melatonin / Q000494:pharmacology*; D009119:Muscle Contraction / Q000502:physiology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D009215:Myometrium / Q000166:cytology / Q000187:drug effects*; D018994:Myosin Light Chains / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D009218:Myosins / Q000378:metabolism; D010121:Oxytocin / Q000494:pharmacology*; D010766:Phosphorylation; D051571:Protein Kinase C-alpha / Q000037:antagonists & inhibitors / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
D002148:Calmodulin-Binding Proteins; D018994:Myosin Light Chains; D007293:Inosine Triphosphate; D010121:Oxytocin; D054460:rho-Associated Kinases; D051571:Protein Kinase C-alpha; D009219:Myosin-Light-Chain Kinase; D048049:Extracellular Signal-Regulated MAP Kinases; D009218:Myosins; D008550:Melatonin
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