Maintaining Balance: Examining Deiodination of Thyroid Hormones in the Developing Brain
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Thyroid hormones (TH) regulate development, growth, and metabolism. Deiodinases (DIOs) are metabolizing enzymes that activate (Dio2) and deactivate (Dio3) THs, exerting the precise local control over TH action that is critical for brain development. In vitro assays have identified chemicals with DIO activity, yet little is known of their potential effects in intact mammalian systems. An LCMS-based assay we previously developed to assess DIO in liver, was optimized to examine DIO activity in developing rat brain. Microsomes were prepared from cortex of rat pups on postnatal days (PN)2, 14, and 60. To examine the selective action of Dio2, Dio3 was blocked with desethylamiodarone. Similarly, morin hydrate isolated action of Dio3 by blocking Dio2. Separate aliquots of equal protein content were spiked with T3, T4, or rT3, incubated in phosphate buffer and TH analytes measured by LCMS. We then investigated if alterations in DIO activation would be measurable ex vivo in cortical tissue after in vivo exposure to iopanoic acid (IOP), a pan-inhibitor of DIOs. Our three main findings include: successful isolation of functional microsomes from rat brain, demonstration of age-dependent differences in DIO activity, and observation of altered DIO activity in tissue derived from IOP-exposed rats. Dio2 activity peaked on PN14, presumably enhancing TH action during a period of rapid brain development. Inversely Dio3 activity was highest on PN2, limiting TH action at this young age. This assay enables the assessment of xenobiotics on DIO activity and potential associated neurotoxic impacts on the developing brain. Does not reflect EPA policy.