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Dorsoventral patterning of the Xenopus eye involves differential temporal changes in the response of optic stalk and retinal progenitors to Hh signalling.

Wang X, Lupo G, He R, Barsacchi G, Harris WA, Liu Y - Neural Dev (2015)

Bottom Line: In loss-of-function assays, inhibition of Hh signalling starting from neurula stages caused expansion of the dorsal retina at the expense of the ventral retina and the optic stalk, while the effects of Hh inhibition during optic vesicle stages were limited to the reduction of optic stalk size.Our results suggest the existence of two competence windows during which the Hh pathway differentially controls patterning of the eye region.We speculate that this temporal regulation is important to coordinate dorsoventral patterning with morphogenesis and differentiation processes during eye development.

View Article: PubMed Central - PubMed

Affiliation: The State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing, 100101, China. 13522764597@163.com.

ABSTRACT

Background: Hedgehog (Hh) signals are instrumental to the dorsoventral patterning of the vertebrate eye, promoting optic stalk and ventral retinal fates and repressing dorsal retinal identity. There has been limited analysis, however, of the critical window during which Hh molecules control eye polarity and of the temporal changes in the responsiveness of eye cells to these signals.

Results: In this study, we used pharmacological and molecular tools to perform stage-specific manipulations of Hh signalling in the developing Xenopus eye. In gain-of-function experiments, most of the eye was sensitive to ventralization when the Hh pathway was activated starting from gastrula/neurula stages. During optic vesicle stages, the dorsal eye became resistant to Hh-dependent ventralization, but this pathway could partially upregulate optic stalk markers within the retina. In loss-of-function assays, inhibition of Hh signalling starting from neurula stages caused expansion of the dorsal retina at the expense of the ventral retina and the optic stalk, while the effects of Hh inhibition during optic vesicle stages were limited to the reduction of optic stalk size.

Conclusions: Our results suggest the existence of two competence windows during which the Hh pathway differentially controls patterning of the eye region. In the first window, between the neural plate and the optic vesicle stages, Hh signalling exerts a global influence on eye dorsoventral polarity, contributing to the specification of optic stalk, ventral retina and dorsal retinal domains. In the second window, between optic vesicle and optic cup stages, this pathway plays a more limited role in the maintenance of the optic stalk domain. We speculate that this temporal regulation is important to coordinate dorsoventral patterning with morphogenesis and differentiation processes during eye development.

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CPM treatments cause stage dependent effects on eye DV polarity. (A) Lateral views of heads of st. 33 embryos treated with ethanol (mock) or 100 μM CPM from the indicated stages and hybridized with probes for Pax2, Vax1b, Vax2 or Tbx3. Compared to controls, CPM-treated embryos show stage-dependent changes in gene expression domains along the anteroposterior (AP; Pax2, Vax1b) or the DV (Vax2, Tbx3) axes of the eye. Scale bar, 200 μm. (B) Quantification of the mean AP (Pax2, Vax1b) or DV (Vax2, Tbx3) width/height of gene expression domains, normalized to total width/height of the eye, in the eyes of embryos treated as in (A). The number of eyes analysed for each probe and treatment condition is indicated within the corresponding histogram bar. Error bars show standard deviations. *P < 0.05; ns, non-significant (P ≥ 0.05) according to two-tailed Student’s t-test. (C) Histological sections of eyes of st. 33 embryos treated with ethanol or CPM from st. 13 and hybridized with the indicated probes. CPM-treated eyes show a reduction of Pax2, Vax1b and Vax2 expression domains along the eye proximodistal axis. Yellow brackets highlight the proximodistal extent of the whole ventral eye region. Scale bar, 100 μm.
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Fig6: CPM treatments cause stage dependent effects on eye DV polarity. (A) Lateral views of heads of st. 33 embryos treated with ethanol (mock) or 100 μM CPM from the indicated stages and hybridized with probes for Pax2, Vax1b, Vax2 or Tbx3. Compared to controls, CPM-treated embryos show stage-dependent changes in gene expression domains along the anteroposterior (AP; Pax2, Vax1b) or the DV (Vax2, Tbx3) axes of the eye. Scale bar, 200 μm. (B) Quantification of the mean AP (Pax2, Vax1b) or DV (Vax2, Tbx3) width/height of gene expression domains, normalized to total width/height of the eye, in the eyes of embryos treated as in (A). The number of eyes analysed for each probe and treatment condition is indicated within the corresponding histogram bar. Error bars show standard deviations. *P < 0.05; ns, non-significant (P ≥ 0.05) according to two-tailed Student’s t-test. (C) Histological sections of eyes of st. 33 embryos treated with ethanol or CPM from st. 13 and hybridized with the indicated probes. CPM-treated eyes show a reduction of Pax2, Vax1b and Vax2 expression domains along the eye proximodistal axis. Yellow brackets highlight the proximodistal extent of the whole ventral eye region. Scale bar, 100 μm.

Mentions: When embryos were treated with CPM starting from late gastrula/early neurula stages (st. 12.5 to 13 Figure 6A,B,C), changes to the overall DV organization of the eye were detectable. Vax2 expression domain in the VR was reduced along the eye DV axis, while Tbx3 domain in the DR was expanded ventrally (Figure 6A,B). In addition, the ventrally located OS, marked by Pax2 and Vax1b expression, was shortened (Figure 6A,B). Furthermore, transverse sections of embryos following whole-mount in situ hybridization revealed a proximo-distal reduction of Pax2, Vax1b and Vax2 expression domains in the ventral eye region of CPM-treated embryos (Figure 6C). CPM delivery from late neurula/early optic vesicle stages (st. 19 to 20) did not affect the expression domains of Vax2 and Tbx3 in the VR and the DR, respectively (Figure 6A,B). Notably, OS size was still reduced by these later treatments (Figure 6A,B). Finally, no changes on eye DV patterning were detected when CPM was applied starting from mid optic vesicle stages (stage 26, Figure 6A,B). These results are consistent with those from overexpression assays and, together, they support a model where Hh signalling controls the specification of both the VR and OS during neurula stages and supports maintenance of OS size during optic vesicle stages.Figure 6


Dorsoventral patterning of the Xenopus eye involves differential temporal changes in the response of optic stalk and retinal progenitors to Hh signalling.

Wang X, Lupo G, He R, Barsacchi G, Harris WA, Liu Y - Neural Dev (2015)

CPM treatments cause stage dependent effects on eye DV polarity. (A) Lateral views of heads of st. 33 embryos treated with ethanol (mock) or 100 μM CPM from the indicated stages and hybridized with probes for Pax2, Vax1b, Vax2 or Tbx3. Compared to controls, CPM-treated embryos show stage-dependent changes in gene expression domains along the anteroposterior (AP; Pax2, Vax1b) or the DV (Vax2, Tbx3) axes of the eye. Scale bar, 200 μm. (B) Quantification of the mean AP (Pax2, Vax1b) or DV (Vax2, Tbx3) width/height of gene expression domains, normalized to total width/height of the eye, in the eyes of embryos treated as in (A). The number of eyes analysed for each probe and treatment condition is indicated within the corresponding histogram bar. Error bars show standard deviations. *P < 0.05; ns, non-significant (P ≥ 0.05) according to two-tailed Student’s t-test. (C) Histological sections of eyes of st. 33 embryos treated with ethanol or CPM from st. 13 and hybridized with the indicated probes. CPM-treated eyes show a reduction of Pax2, Vax1b and Vax2 expression domains along the eye proximodistal axis. Yellow brackets highlight the proximodistal extent of the whole ventral eye region. Scale bar, 100 μm.
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Fig6: CPM treatments cause stage dependent effects on eye DV polarity. (A) Lateral views of heads of st. 33 embryos treated with ethanol (mock) or 100 μM CPM from the indicated stages and hybridized with probes for Pax2, Vax1b, Vax2 or Tbx3. Compared to controls, CPM-treated embryos show stage-dependent changes in gene expression domains along the anteroposterior (AP; Pax2, Vax1b) or the DV (Vax2, Tbx3) axes of the eye. Scale bar, 200 μm. (B) Quantification of the mean AP (Pax2, Vax1b) or DV (Vax2, Tbx3) width/height of gene expression domains, normalized to total width/height of the eye, in the eyes of embryos treated as in (A). The number of eyes analysed for each probe and treatment condition is indicated within the corresponding histogram bar. Error bars show standard deviations. *P < 0.05; ns, non-significant (P ≥ 0.05) according to two-tailed Student’s t-test. (C) Histological sections of eyes of st. 33 embryos treated with ethanol or CPM from st. 13 and hybridized with the indicated probes. CPM-treated eyes show a reduction of Pax2, Vax1b and Vax2 expression domains along the eye proximodistal axis. Yellow brackets highlight the proximodistal extent of the whole ventral eye region. Scale bar, 100 μm.
Mentions: When embryos were treated with CPM starting from late gastrula/early neurula stages (st. 12.5 to 13 Figure 6A,B,C), changes to the overall DV organization of the eye were detectable. Vax2 expression domain in the VR was reduced along the eye DV axis, while Tbx3 domain in the DR was expanded ventrally (Figure 6A,B). In addition, the ventrally located OS, marked by Pax2 and Vax1b expression, was shortened (Figure 6A,B). Furthermore, transverse sections of embryos following whole-mount in situ hybridization revealed a proximo-distal reduction of Pax2, Vax1b and Vax2 expression domains in the ventral eye region of CPM-treated embryos (Figure 6C). CPM delivery from late neurula/early optic vesicle stages (st. 19 to 20) did not affect the expression domains of Vax2 and Tbx3 in the VR and the DR, respectively (Figure 6A,B). Notably, OS size was still reduced by these later treatments (Figure 6A,B). Finally, no changes on eye DV patterning were detected when CPM was applied starting from mid optic vesicle stages (stage 26, Figure 6A,B). These results are consistent with those from overexpression assays and, together, they support a model where Hh signalling controls the specification of both the VR and OS during neurula stages and supports maintenance of OS size during optic vesicle stages.Figure 6

Bottom Line: In loss-of-function assays, inhibition of Hh signalling starting from neurula stages caused expansion of the dorsal retina at the expense of the ventral retina and the optic stalk, while the effects of Hh inhibition during optic vesicle stages were limited to the reduction of optic stalk size.Our results suggest the existence of two competence windows during which the Hh pathway differentially controls patterning of the eye region.We speculate that this temporal regulation is important to coordinate dorsoventral patterning with morphogenesis and differentiation processes during eye development.

View Article: PubMed Central - PubMed

Affiliation: The State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing, 100101, China. 13522764597@163.com.

ABSTRACT

Background: Hedgehog (Hh) signals are instrumental to the dorsoventral patterning of the vertebrate eye, promoting optic stalk and ventral retinal fates and repressing dorsal retinal identity. There has been limited analysis, however, of the critical window during which Hh molecules control eye polarity and of the temporal changes in the responsiveness of eye cells to these signals.

Results: In this study, we used pharmacological and molecular tools to perform stage-specific manipulations of Hh signalling in the developing Xenopus eye. In gain-of-function experiments, most of the eye was sensitive to ventralization when the Hh pathway was activated starting from gastrula/neurula stages. During optic vesicle stages, the dorsal eye became resistant to Hh-dependent ventralization, but this pathway could partially upregulate optic stalk markers within the retina. In loss-of-function assays, inhibition of Hh signalling starting from neurula stages caused expansion of the dorsal retina at the expense of the ventral retina and the optic stalk, while the effects of Hh inhibition during optic vesicle stages were limited to the reduction of optic stalk size.

Conclusions: Our results suggest the existence of two competence windows during which the Hh pathway differentially controls patterning of the eye region. In the first window, between the neural plate and the optic vesicle stages, Hh signalling exerts a global influence on eye dorsoventral polarity, contributing to the specification of optic stalk, ventral retina and dorsal retinal domains. In the second window, between optic vesicle and optic cup stages, this pathway plays a more limited role in the maintenance of the optic stalk domain. We speculate that this temporal regulation is important to coordinate dorsoventral patterning with morphogenesis and differentiation processes during eye development.

Show MeSH
Related in: MedlinePlus