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Why is it that with no sonic hedgehog from the prechordal mesoderm leads to cyclopia

The eye fields fail to split


Knockout itself doesn’t provide direct evidence that you need sonic hedgehog to induce the hypothalamus. What experiments have been done that do prove this

By preventing sonic hedgehog coming from the prechordal mesoderm when the embryo is alive. This can be done by soaking a bead with a sonic hedgehog antagonist to see if you still get hypothalamic progenitors. Similarly experiments where you culture naïve neural plate tissue with the prechordal mesoderm to see if this tissue begins to express hypothalamic markers.


What is significant about sonic hedgehog action in the ventral forebrain in terms of its ability to induce its own expression

Sonic hedgehog from the prechordal mesoderm in the ventral forebrain requires the additional Nodal signal in order to induce sonic hedgehog positive pre-hypothalamic progenitors. This is in contrast to the spinal cord where sonic hedgehog from the notochord alone is able to induce sonic hedgehog expression by the floor plate cells


Sonic hedgehog from the prechordal mesoderm is necessary for hypothalamic development T or F



It isn’t possible to carry gain of function studies with sonic hedgehog soaked beads in the forebrain because these beads wont be held in place. How else can we examine the sufficiency of sonic hedgehog alone in inducing the definitive hypothalamic progenitors

In vitro studies exposing neural tissue to sonic hedgehog. This doesn’t results in the expression of hypothalamic marks as nodal is absolutely required as well


What is the effect of the action of nodal and sonic hedgehog signalling by the prechordal mesoderm

Induction of sonic hedgehog expression in pre-hypothalamic cells


Nodal and sonic hedgehog from the prechordal mesoderm induce hypothalamic marker expression in the cells above it T or F

F – they only act to induce sonic hedgehog expression itself not the markers of hypothalamic progenitors


What is the term given to the sonic hedgehog positive cells induced by sonic hedgehog and nodal from the prechordal mesoderm

Pre-hypothalamic cells


If a cell it responding to sonic hedgehog signalling what sort of gene products would you expect it to be expressing

Ptc-1 and Gli-1


The cells that are receiving the sonic hedgehog signal from the prechordal mesoderm are also then expressing it T or F



Following induction of the pre-hypothalamic cell what rapid change(s) in expression are seen in the prechordal mesoderm

Immediately after Shh is induced and pre-hypothalamic cells appear BMP7 is rapidly upregulated in the prechordal mesoderm. Hence for a brief period Nodal and BMP7 are co-expressed in the prechordal mesoderm. This is unlike the floor plate of the spinal cord


What is the combined effect of both Nodal and BMP7 expression by the prechordal mesoderm

These signals act together to induce the expression of FGF10 by the newly formed pre-hypothalamic cell


What are the downstream targets as a result of BMP7 signalling in the pre-hypothalamic cell

BMP7 signalling leads to the phosphorylation of Smads1/5/8. pSmads1/5/8 then upregulate Tbx2 and Nkx2.1 in the sonic hedgehog positive pre-hypothalamic cells


In what ways are the pre-hypothalamic cells alike the neural crest cells of the spinal cord

There are multipotent proliferative and migratory


How many different types of neurons are there in the hypothalamus



What is the role of FGF10 expression by the pre-hypothalamic cells

Supports survival and proliferation


What is the role of Tbx2 expression by the pre-hypothalamic cells

Supports the ability of these cells to migrate and differentiate


The tbx2 and FGF10 expression patterns in the pre-hypothalamic cells differs T or F

F – they are expressed in the same pattern


How can we prove that FGF is required to drive proliferation in these cells

Use an FGF inhibitor such as SU5042 to block FGF signalling. Then look at markers of cell cycle to see the effects on proliferation


What could you use to look at proliferation in cells

A marker of the cell cycle such as anti-pH3 which indicates M phase or EdU which can be used to indicate S phase


How can we assay the effects of tbx2 on differentiation

Knockdown its expression via siRNA


Once the prechordal mesoderm has established an early multipotent hypothalamic stem-like cell describe how this gives rise to the different regions of the early hypothalamus

Hypothalamic progenitors grow and differentiate first to anterior hypothalamic neurons then to posterior/mammillary hypothalamic neurons. Some FGF10 positive progenitors are always retained in the so-called tuberal hypothalamus. These cells eventually enter a very very slow cycle and are retained as stem-like cells


Which signalling factors act to specify the hypothalamic stem like cell seen in the tuberal hypothalamus

Sonic hedgehog and BMP


FGF10 positive cells in the hypothalamus will show strong staining indicative of proliferation T or F

T – this will be in the tuberal region


The cells in the anterior hypothalamus will strain positive for EdU T or F

F – this is a marker of proliferation whereas the anterior hypothalamic cells will have begun to differentiate


Sonic hedgehog drives the progression of FGF10 positive hypothalamic progenitors to an anterior neuronal fate T or F



What would happen if we experimentally prevent sonic hedgehog signalling as soon as the hypothalamic progenitors form at the same time as injecting a dye to fate map the cells

After leaving the embryo to develop we would observe an accumulation of FGF10 positive progenitors as well as a failure of anterior progenitor formation and their subsequent failure to differentiate


Sonic hedgehog is acting as a spatial morphogen in the early hypothalamus to direct the 3 cell fates depending on the concentration of the ligand the cells are exposed to T or F

F – sonic hedgehog is acting to select a differentiation pathway and drive the progression of FGF10 positive hypothalamic progenitor cells to anterior fates


What transcription factor is upregulated as a result of sonic hedgehog signalling in the anterior hypothalamic progenitors



What neurons do the anterior hypothalamic progenitors ultimately give rise to

Neurons that mediate core body homeostasis