Postnatal Hypothyroidism Permanently Disrupts Neural Stem Cell Fate in the Murine Subventricular Zone

Abstract

The subventricular zone (SVZ) of the adult mammalian brain harbors neural stem cells (NSCs) that generate neurons and oligodendrocytes throughout life. Single-cell RNA-Seq analysis on mouse SVZ-NSCs isolated at different developmental stages established they gradually acquire their adult neurogliogenic identity between postnatal day (P) 7 and 20. However, the factors governing this transition remain elusive. As a key factor driving transcriptional responses during brain development, as well as NSC lineage commitment in the adult SVZ, we hypothesized that thyroid hormone (TH) could fulfil this role. TH serum levels rise postnatally and peak around P15. Re-analysis of single-cell data from the P2 and P20 SVZ revealed a dynamically increased expression of the TH transporters Mct8 and Oatp1c1, as well as the TH-(in)activating deiodinases Dio2 and Dio3 in NSCs, signs that local TH action is promoted. Immunostainings showed a concomitant burst in SVZ-neurogenesis between P4 and P21. Then, to study what occurs if TH synthesis is blocked, we fed dams a 0.15% propylthiouracil-enriched diet from embryonic day 15 to P21. Postnatal hypothyroidism decreased PH3-positive mitotic cell numbers at P4 and P21, whereas increased Sox2 expression coincided with a larger proportion SOX2-positive SVZ-NSCs and progenitors. In the dorsal SVZ, the main site of neurogliogenesis, less neuroblasts were detected at P21, while numbers of OLIG2-positive oligodendroglia precursors did not change significantly. Next, we prepared in vitro neurospheres from dissected SVZs of control and PTU-treated mice, and allowed them differentiate with or without exogenous T3. The neuro/glia balance in neurosphere cultures prepared from P4 animals of either condition did not change when T3 was added, suggesting perinatal NSCs are irresponsive to TH. The balance did change in T3-treated neurospheres prepared from control P21 animals, however, not in those from P21 PTU-exposed mice, suggesting hypothyroid NSCs are irresponsive to T3. Lastly, we examined 3-month-old mice that regained a normal diet following developmental PTU exposure. Fewer oligodendroglia precursors in the SVZ resulted in a lasting altered neuro/glia output. A reduced ability to remember earlier-presented odors indicates impaired olfaction, a behavior strongly depending on SVZ-neurogenesis. Taken together the data indicate that developmental hypothyroidism affects postnatal SVZ organization and permanently alters NSC lineage commitment. Our study allows to determine relevant new read-outs to identify adverse outcome events on brain development and will permit comparison with events following exposure to endocrine disruptors.