l3

Question 1

what can SEM show in Shh KO mouse?

Answer 1

-frontal SEM view shows absence of hypothalamus in Shh -/- mouse
- holoprosencephaly
- cyclopia-lateral mesenchyme fails to resolve bilaterally
(mutations in nodal/tgf - Shh signalling components)

Question 2

What happens when Shh is pharmacologically blocked in PM using an Shh antagonist?

Answer 2

• no Nkx2.1 (hypothalamic marker) induced in lateral neural plate
• no signs of hypthalamic develpment
• Shh is REQUIRED for hypothalamic development

(lateral neural plate doesnt usually form hypothalamus but does when Shh PM is planted underneath it)

Question 3

Why is there few possible in vivo GOF studies which can be done in mice?

Answer 3

-bead cannot be used as in head - don’t have anything (e.g. somites) to keep in place - will move around

(can examine Shh in vitro)

Question 4

how is Shh sufficiency in hypothalamic development measured?

Answer 4

• in vitro
• neural tissue exposed to Shh (Shh + culture)
• do not see hypothalamic markers (e.g. nkx2.1)

-SHH IS NOT SUFFICIENT TO INDUCE pre-hypothalamic cell

Question 5

Is Shh required and sufficient to induce hypothalamus?

Answer 5

• Shh is required

- Shh is not sufficient

Question 6

what are co-expressed in the PM and what does this do?

Answer 6

• Shh and nodal are coexpressed in PM
• form pre-hypthalamic cell (Shh+)
• Shh and nodal together induce Shh expression

Question 7

what do cells responding to Shh produce?

Answer 7

-Gli-1 and Ptc-1

Question 8

what happens in the PM after Shh+ prehypothalamic cells are produced?

Answer 8

PM upregulates BMP7

- for a brief period nodal and bmp7 are coexpressed in the PM

Question 9

why is it critical that Nodal and BMP7 are coexpressed briefly in PM?

Answer 9

• because this coexpression induces a Fgf+ hypothalamic progenitor cell

Question 10

other than Fgf10 what else does bmp7 expression induce?

Answer 10

psmad
tbx2
nkx2.1
shh

(+fgf - markers of hypothalamic progenitor cells)

Question 11

give some features/properties of hypothalamic progenitor cells

Answer 11

• proliferate
• stay alive
• migrate
• differentiate
• are multipotent

(similar properties to NCCs (BMP and Fgf also give NCCs these properties)

Question 12

what properties does Fgf10 expression induce and how is this shown?

Answer 12

• survival and proliferation

- shown by apoptotic cell cycle and proliferation markers

Question 13

what properties does tbx2 expression induce and how is this shown?

Answer 13

• ability to migrate and differentiate
• is tbx2 is ectopically expressed
• tbx2 is KD by siRNA

Question 14

what happens to anterior progenitors once they are induced?

Answer 14

• first hypothalamic progenitors grow and differentiate anteriorly - anterior hypthalamus
• then they differentiate and grow posteriorly - posterior/mamilary hypothalamus
• some fgf10+ progenitors are retained - tuberal hypothalamus

Question 15

what is eDu/BrdU?

Answer 15

• analogues of thymidine
• incorperated into newly synthesized DNA during DNA synthesis
• labels newly formed DNA/cells

Question 16

How does Shh drive progression from a hypothalamic progenitor to an anterior progenitor?

Answer 16

• it upregulates TF RX2

Question 17

what happens if Shh signalling is blocked as soon as fgf10+ progenitors form?
(cre-ERT/tamoxifen???)

Answer 17

• Fgf10+ progenitors accumulate
• prevent aterior progenitors forming
• prevent anterior neurons differentiating

Question 18

what are neurons that derive from anterior progenitors responsible for?

Answer 18

core homeostasis

Question 19

what happens in foetal alcohol syndrome?

Answer 19

Shh expression in PM/ealry hypothalamus is blocked

- disrupts hypothalamus formation?

Question 20

what happens if Shh is overexpessed too much?

Answer 20

could lead to cancer