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Dec. 5, 2008

The right mix for neuron culture

Eliminating signaling molecules from tissue culture results in the generation of hypothalamic neurons from embryonic stem cells

image of vasopressin-producing neurons Figure 1:Vasopressin-producing neurons (green) in the embryonic hypothalamus.

The hypothalamus is a part of the brain containing groups of neurons called nuclei, which regulate many body functions including blood pressure, stress, hunger, and sexual development and function. Understanding how the different populations of hypothalamic neurons develop from stem cells may enable scientists to cure diseases that are associated with hypothalamic dysfunction.

A team of researchers, led by Yoshiki Sasai at the RIKEN Center for Developmental Biology in Kobe, has now discovered that embryonic stem (ES) cells can mature into hypothalamic neurons only when as many exogenous signaling factors as possible are removed from the liquid medium in which the cells are grown1.

Mouse and human embryonic stem cells are often grown in liquid medium containing multiple growth factors such as those in ‘knockout serum replacement’ (KSR). ES cells grown in KSR often express markers characteristic of the telencephalon, which gives rise to brain structures such as the cerebrum and striatum. The KSR still contains factors, such as insulin, that may affect the maturation of the ES cells. So, when the researchers cultured the ES cells without KSR, they observed maturation of the cells into hypothalamic progenitor cells.

Insulin was the factor that inhibited hypothalamic maturation of the ES cells, since very few ES cells cultured in insulin expressed hypothalamic progenitor cell markers, such as the protein Rax. In addition, the researchers determined that ES cells were most sensitive to insulin during days 4 and 5 in culture.

Sonic hedgehog (Shh) is a signaling factor known to affect the maturation of progenitor cells throughout the central nervous system. Sasai and colleagues found that treatment of Rax-expressing hypothalamic progenitor cells with Shh induces development of neurons from areas of the hypothalamus that are involved in controlling appetite. On the other hand, Rax-expressing hypothalamic progenitor cells that are not treated with Shh go on to mature into hypothalamic neurons that express arginine-vasopressin, a hormone known to be involved in regulating water retention in the kidneys, and blood pressure (Fig. 1).

Because these findings were all derived from work on mouse ES cells in tissue culture, additional experiments are needed to determine whether the mechanisms for hypothalamic development identified in the work would also apply in the setting of normal brain development and in humans.

In future experiments, Sasai says, “we will continue to study the molecular and cellular mechanism for the generation of other parts of the brain.”


  • 1. Wataya, N., Ando, S., Muguruma, K., Ikeda, H., Watanabe, K., Eiraku, M., Kawada, M., Takahashi, J., Hashimoto, N. & Sasai, Y. Minimization of exogenous signals in ES cell culture induces rostral hypothalamic differentiation. Proceedings of the National Academy of Sciences USA105, 11796–11801 (2008). (Link)