The activity of low voltage-activated Ca 2+ (Ca V3) channels is tightly coupled to neurotransmitter and hormone secretion. Previous studies have shown that Ca V3 channels are regulated by glucocorticoids (GCs), though the mechanism underlying channel regulation remains unclear. Here, using the pituitary GH 3 cell line as a model, we investigated whether Ca V3 channel expression is under the control of GCs, and if their actions are mediated by transcriptional and/or post-transcriptional mechanisms. RT-PCR and western blot analyses showed that Ca V3.1 but not Ca V3.2 and Ca V3.3 channels is expressed in the GH 3 cells, and patch clamp recordings confirmed that Ca 2+ currents through low voltage-activated channels were decreased after chronic treatment with GCs. Consistent with this, total plasma membrane expression of Ca V3.1 protein as analyzed by cell-surface biotinylation assays and semi-quantitative western blotting was also down-regulated, while quantitative real-time RT-PCR analysis revealed a significant decrease of Ca V3.1 mRNA expression in the treated cells. In contrast, patch-clamp recordings on HEK-293 cells stably expressing recombinant Ca V3.1 channels showed that Ca 2+ currents were not affected by GC treatment. These results suggest that decreased transcription is a likely mechanism to explain the inhibitory actions of GCs on the functional expression of native Ca V3.1 channels.
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Acknowledgments This work was supported by funds from Cona-cyt to RF. We thank Drs. M. E. Mendoza (Cinvestav-IPN, Mexico) and J. C. Gomora (IFC-UNAM, Mexico) for the generous gift of the cell lines and Dr. M. Hernandez (Cinvestav-IPN, Mexico) and D. Mornet (INSERM ERI 25 Muscle et Pathologies, France) for the anti-actin and anti-β-dystroglycan antibodies, respectively. We are also indebted to J. Arikkath for critically reviewing the manuscript.
Copyright 2010 Elsevier B.V., All rights reserved.
All Science Journal Classification (ASJC) codes
- Cellular and Molecular Neuroscience
- Cell Biology