Thyroid hormone receptor isoform content in cultured type 1 and type 2 astrocytes

Immunohistochemical studies previously reported from this laboratory showed that astrocytes in adult rat brain appear devoid of all thyroid hormone receptor (TR) isoforms. These findings, however, contrast with reports of measurable nuclear T-3 binding in astrocytes in cell culture. To address this discrepancy, TR protein and messenger RNA (mRNA) content of type 1 and type 2 astrocytes in culture were assayed. Type 1 cells represent astrocytes present in brain in vivo. Type 2 astrocytes differentiate in culture from bipotential progenitor 0-2A cells in the presence of serum. Under serum-free conditions, these progenitor cells differentiate into oligodendroglia. Total nuclear T, binding capacity in both type 1 and type 2 astrocytes was approximately 3000 sites/cell. Northern blots showed the presence of mRNA for TR beta 1, TR alpha 1, and TR alpha 2 in type 2 cells but failed to reveal the presence of these mRNAs in type 1 astrocytes. Moreover, Northern blots also failed to reveal TR beta 2 mRNA in both type 1 and type 2 astrocytes. These findings, therefore, raised a question as to which receptor isoform was responsible for the nuclear binding capacity observed in type 1 astrocytes. As anticipated, immunocytochemical analysis demonstrated prominent nuclear signals for TR beta 1, TR alpha 1, and TR alpha 2 mRNA in type 2 astrocytes but failed to demonstrate TR beta 1, TR alpha 1, or TR alpha 2 in type I astrocytes. Application of RT-PCR, however, revealed the presence of low levels of TR beta 2 mRNA in type 1 astrocytes. When stained with a specific anti-TR beta 2 antiserum, both type 1 and type 2 astrocytes showed a strong fluorescent signal concentrated in the nucleus. These data indicate that under the special conditions of cell culture, expression of the TR beta 2 isoform in type 1 accounts for the measured nuclear T-2 binding capacity.