Methylene blue inhibits function of the 5-HT transporter

Authors

  • Murat Oz,

    Corresponding author
    1. Functional Lipidomics Branch, Department of Pharmacology
    2. Integrative Neuroscience Section, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, U.S. Department of Health and Human Services, Baltimore MD, USA
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  • Dmytro Isaev,

    1. Functional Lipidomics Branch, Department of Pharmacology
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  • Dietrich E Lorke,

    1. Department of Anatomy, Faculty of Medicine and Health Sciences, UAE University, Al Ain, UAE
    2. Department of Cellular Biology & Pharmacology, College of Medicine, Florida International University, Miami, FL, USA
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  • Muhammed Hasan,

    1. Functional Lipidomics Branch, Department of Pharmacology
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  • Georg Petroianu,

    1. Department of Cellular Biology & Pharmacology, College of Medicine, Florida International University, Miami, FL, USA
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  • Toni S Shippenberg

    1. Integrative Neuroscience Section, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, U.S. Department of Health and Human Services, Baltimore MD, USA
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Dr Murat Oz, Department of Pharmacology, Functional Lipidomics Branch, Faculty of Medicine & Health Sciences, United Arab Emirates University, Al Ain, Abu Dhabi, UAE. E-mail: Murat_Oz@uaeu.ac.ae

Abstract

BACKGROUND AND PURPOSE Methylene blue (MB) is commonly employed as a treatment for methaemoglobinaemia, malaria and vasoplegic shock. An increasing number of studies indicate that MB can cause 5-HT toxicity when administered with a 5-HT reuptake inhibitor. MB is a potent inhibitor of monoamine oxidases, but other targets that may contribute to MB toxicity have not been identified. Given the role of the 5-HT transporter (SERT) in the regulation of extracellular 5-HT concentrations, the present study aimed to characterize the effect of MB on SERT.

EXPERIMENTAL APPROACH Live cell imaging, in conjunction with the fluorescent SERT substrate 4-(4-(dimethylamino)-styryl)-N-methylpyridinium (ASP+), [3H]5-HT uptake and whole-cell patch-clamp techniques were employed to examine the effects of MB on SERT function.

KEY RESULTS In EM4 cells expressing GFP-tagged human SERT (hSERT), MB concentration-dependently inhibited ASP+ accumulation (IC50: 1.4 ± 0.3 µM). A similar effect was observed in N2A cells. Uptake of [3H]5-HT was decreased by MB pretreatment. Furthermore, patch-clamp studies in hSERT expressing cells indicated that MB significantly inhibited 5-HT-evoked ion currents. Pretreatment with 8-Br-cGMP did not alter the inhibitory effect of MB on hSERT activity, and intracellular Ca2+ levels remained unchanged during MB application. Further experiments revealed that ASP+ binding to cell surface hSERT was reduced after MB treatment. In whole-cell radioligand experiments, exposure to MB (10 µM; 10 min) did not alter surface binding of the SERT ligand [125I]RTI-55.

CONCLUSIONS AND IMPLICATIONS MB modulated SERT function and suggested that SERT may be an additional target upon which MB acts to produce 5-HT toxicity.

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