First published online 6 August 2008
doi: 10.1242/dev.025163
Development 135, 3007-3011 (2008)
Published by The Company of Biologists 2008
Non-cell-autonomous effects of Ret deletion in early enteric neurogenesis
Silvia Bogni1,
Paul Trainor2,
Dipa Natarajan1,
Robb Krumlauf2 and
Vassilis Pachnis1,*
1 Division of Molecular Neurobiology, National Institute for Medical Research,
The Ridgeway, Mill Hill, London NW7 1AA, UK.
2 Stowers Institute for Medical Research, 1000 East 50th Street, Kansas City, MO
64110, USA.
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Fig. 1. Normal development of the peripheral nervous system in cultured
embryos. E8.5 mouse embryos (A) were cultured for 3 days and
processed for whole-mount in situ hybridisation with a Ret-specific
riboprobe (B). In situ hybridisation of a freshly dissected E10.5
embryo (C). In both cases, Ret expression is observed in the
VII, IX and X cranial sensory ganglia and in the stomach (st) and midgut (mg).
The asterisk in A marks the approximate site of cell grafting (see text, and
legend to Fig. 2). Arrows
indicate enteric neural crest cells (NCCs) within the gut of cultured (B) and
freshly dissected (C) embryos.
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Fig. 2. Colonisation of the gut of E8.5+3 embryos transplanted with
Ret+ ENS progenitors. (A) DiI labelling of NCCs at the
level of somites 2-4 results in efficient colonisation of the gut by
fluorescent cells. (B) E8.5+3 wild-type mouse embryos transplanted with
Ret+ cells isolated from the intestine of E11.5 Rosa26bgeo
embryos. At the end of the culture period, β-geo+ cells
(arrow) are restricted to the gastrointestinal tract. (C,D)
Examples of the two classes of embryos classified according to the extent of
gut colonisation by grafted cells. st, stomach; mg, midgut; cae, caecum.
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Fig. 3. Ret+ cells grafted to the vagal NCC pathway of E8.5 embryos
are capable of colonising the entire length of the gastrointestinal tract.
(A) Whole-mount X-Gal staining of gut from E8.5+3 mouse embryos
transplanted with Ret+ cells from the intestine of E11.5
Rosa26bgeo embryos. Note the efficient colonisation of the entire gut
by β-geo+ cells. (B) Higher magnification of the gut
shown in A. Arrow points to a cluster of β-geo+ cells.
(C) Cryosections of E8.5+3+7 gut show that the progeny of grafted cells
are localised within the myenteric plexus (mp, arrows). (D,E)
Similar cryosections from embryos transplanted with YFP-expressing
Ret+ cells were double immunostained for GFP and TuJ1 (D) or for
GFP and B-FABP (E). st, stomach; mg, midgut; cae, caecum.
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Fig. 4. Non-cell-autonomous effect of the Ret- mutation on
the colonisation of E8.5 by Ret+ ENS progenitors. Whole-mount
X-Gal staining of E8.5+3+7 guts from Ret+/- (A) and
Ret-/- (B) mouse embryos grafted with
Ret+ cells from the intestine of E11.5 Rosa26bgeo embryos.
Contrary to control gut (A), no β-gal+ cells were detected
beyond the anterior foregut of Ret-deficient gut (B). Representative images of
Ret+/- (C) and Ret-/-
(D) E8.5+3 embryos grafted at E8.5 with Ret+ cells isolated
from the intestine of E11.5 embryos. Grafted embryos (three homozygous mutant
and 12 heterozygous or wild-type) were cultured for 3 days and then processed
for β-gal histochemistry. Arrows in C and D point to
β-geo+ cells in the foregut (stomach and duodenum anlage). st,
stomach; mg, midgut.
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© The Company of Biologists Ltd 2008
