Variation in Thyroid Hormone Concentration in Brain Microdialysates from Freely-moving Adult Rats

Thyroid hormone has genomic and non-genomic actions in the developing and mature human. The focus of this research was to find if there was a measurable amount of thyroid hormone in the extracellular fluid of the adult rat brain and how it varied over time. To study the levels of thyroid hormone concentration in the adult brain, stereotaxic surgery was performed to implant a microdialysis cannula and electrodes for the study of electroencephalogram (EEG), electromyogram (EMG). The stylet blocking the microdialysis cannula was removed and the probe was inserted at the beginning of the study for sample collection. EEG and EMG waves were recorded concurrently. Artificial cerebrospinal fluid (aCSF) was run through the inlet tube of the probe and sample was obtained from the outlet using a refrigerated fraction collector. Collected samples were analyzed by high pressure liquid chromatography (HPLC), liquid chromatography mass spectrometry (LC/MS) and radioimmunoassay (RIA) for presence of the thyroid hormone. Samples run through the C18 column on the HPLC gave a peak for thyroid hormone at a retention time of 4.1minutes. Similar results were generated when the microdialysate samples were analyzed using LC/MS. The LC of the microdialysate gave the peak at the retention time of 4.1 minutes. This peak was further studied for confirmation by MS ionization. MS gave a fragmentation pattern which corresponded to the breakdown products of the thyroid hormone. The RIA kit with thyroid hormone iii standards prepared in aCSF was used for the detection of the thyroid hormone concentration. The level of thyroid hormone in samples varies over time. The variation in thyroid hormone level over time was compared to the sleep data obtained concurrently. The variation seen in the thyroid hormone concentration over time was detected by HPLC, LC/MS and RIA. This variation suggests some possibility of a mechanism of release of thyroid hormone in adult brain. This finding would be consistent with the potential signaling role of TH suggested by earlier experiments performed in our lab. iv Acknowledgement I owe my gratitude to Dr. J.V.Martin for guiding me from working in lab to writing this thesis. I am also thankful to Dr. P.J. McIlroy and Dr. W. M. Saidel for being my committee member and offering me their wisdom and expertise with utmost kindness and compassion. I would also like to thank James Boulden for his expert guidance with the software needed for my research. My appreciation is given …