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Shox2 is required for the proper development of the facial motor nucleus and the establishment of the facial nerves.

Rosin JM, Kurrasch DM, Cobb J - BMC Neurosci (2015)

Bottom Line: Using a Nestin-Cre driver, we show that elimination of Shox2 throughout the brain results in elevated cell death in the facial motor nucleus at embryonic day 12.5 (E12.5) and E14.5, which correlates with impaired axonal projection properties of vMNs.We also observed changes in the spatial expression of the vMN cell fate factors Isl1 and Phox2b, and concomitant defects in Shh and Ptch1 expression in Shox2 mutants.Furthermore, we demonstrate that elimination of Shox2 results in the loss of dorsomedial and ventromedial subnuclei by postnatal day 0 (P0), which may explain the changes in physical activity and impaired feeding/nursing behavior in Shox2 mutants.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Calgary, 2500 University Drive N.W., BI286D, Calgary, AB, T2N 1N4, Canada. jmrosin2013@gmail.com.

ABSTRACT

Background: Axons from the visceral motor neurons (vMNs) project from nuclei in the hindbrain to innervate autonomic ganglia and branchial arch-derived muscles. Although much is known about the events that govern specification of somatic motor neurons, the genetic pathways responsible for the development of vMNs are less well characterized. We know that vMNs, like all motor neurons, depend on sonic hedgehog signaling for their generation. Similarly, the paired-like homeobox 2b (Phox2b) gene, which is expressed in both proliferating progenitors and post-mitotic motor neurons, is essential for the development of vMNs. Given that our previous study identified a novel role for the short stature homeobox 2 (Shox2) gene in the hindbrain, and since SHOX2 has been shown to regulate transcription of islet 1 (Isl1), an important regulator of vMN development, we sought to determine whether Shox2 is required for the proper development of the facial motor nucleus.

Results: Using a Nestin-Cre driver, we show that elimination of Shox2 throughout the brain results in elevated cell death in the facial motor nucleus at embryonic day 12.5 (E12.5) and E14.5, which correlates with impaired axonal projection properties of vMNs. We also observed changes in the spatial expression of the vMN cell fate factors Isl1 and Phox2b, and concomitant defects in Shh and Ptch1 expression in Shox2 mutants. Furthermore, we demonstrate that elimination of Shox2 results in the loss of dorsomedial and ventromedial subnuclei by postnatal day 0 (P0), which may explain the changes in physical activity and impaired feeding/nursing behavior in Shox2 mutants.

Conclusions: Combined, our data show that Shox2 is required for development of the facial motor nucleus and its associated facial (VII) nerves, and serves as a new molecular tool to probe the genetic programs of this understudied hindbrain region.

No MeSH data available.


Related in: MedlinePlus

Loss of Shox2 function in the facial motor nucleus interferes with the proper development of the medial subnuclei. a Diagram of a coronal section through the P0 brain highlights the region represented in panels b–v, particularly the 7 neuronal clusters that form the facial motor nucleus (nVII, dashed-circle) (image adapted from [23]). b X-gal stained coronal sections through P0 brains shows a loss of medial neuronal clusters in the facial motor nucleus of Nestin-Cre; Shox2lacZ/flox (B″) brains as compared to controls (B′). c–v ISH on P0 control (c, e, g, i, k, m, o, q, s, u) and Nestin-Cre; Shox2flox/− (d, f, h, j, l, n, p, r, t, v) coronal sections shows loss of Shox2 expression (compare c to d, dashed-circle), and decreases in Isl1 (compare e to f, dashed-circle), Phox2b (compare g to h, dashed-circle), Cntn2 (compare i to j, dashed-circle), Tubb3 (compare k to l, dashed-circle), Periph (compare m to n, dashed-circle), Slc18a3 (compare o to p, dashed-circle), Slit2 (compare q to r, dashed-circle), Shh (compare s to t, dashed-circle) and Ptch1 (compare u to v, dashed-circle) expression in the facial motor nucleus. DA dorsal accessory, DM dorsomedial, VM ventromedial, DI dorsal intermediate, VI ventral intermediate, DL dorsolateral, Lat lateral. Scale bar 250 μm.
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Fig7: Loss of Shox2 function in the facial motor nucleus interferes with the proper development of the medial subnuclei. a Diagram of a coronal section through the P0 brain highlights the region represented in panels b–v, particularly the 7 neuronal clusters that form the facial motor nucleus (nVII, dashed-circle) (image adapted from [23]). b X-gal stained coronal sections through P0 brains shows a loss of medial neuronal clusters in the facial motor nucleus of Nestin-Cre; Shox2lacZ/flox (B″) brains as compared to controls (B′). c–v ISH on P0 control (c, e, g, i, k, m, o, q, s, u) and Nestin-Cre; Shox2flox/− (d, f, h, j, l, n, p, r, t, v) coronal sections shows loss of Shox2 expression (compare c to d, dashed-circle), and decreases in Isl1 (compare e to f, dashed-circle), Phox2b (compare g to h, dashed-circle), Cntn2 (compare i to j, dashed-circle), Tubb3 (compare k to l, dashed-circle), Periph (compare m to n, dashed-circle), Slc18a3 (compare o to p, dashed-circle), Slit2 (compare q to r, dashed-circle), Shh (compare s to t, dashed-circle) and Ptch1 (compare u to v, dashed-circle) expression in the facial motor nucleus. DA dorsal accessory, DM dorsomedial, VM ventromedial, DI dorsal intermediate, VI ventral intermediate, DL dorsolateral, Lat lateral. Scale bar 250 μm.

Mentions: Although the loss of Shox2 in the brain resulted in drastic reductions in the size of the facial motor nucleus and the expression of the above-mentioned genes, the spared vMNs remaining in the facial motor nucleus of Shox2 mutants appeared to be mostly localized to the dorsal regions of the nucleus (Figure 6B″, f, n). This suggested that there could be a region-specific loss of distinct subnuclei in the facial motor nucleus in Shox2-mutant animals. Therefore we examined the expression patterns of these same genes in coronal sections of the facial motor nucleus, as this would allow us to more clearly observe the six subnuclei that comprise the main facial motor nucleus; specifically the lateral, dorsolateral, dorsal intermediate, ventral intermediate, dorsomedial and ventromedial nuclei [23, 24]. LacZ staining of P0 control and Nestin-Cre; Shox2lacZ/flox coronal sections of the facial motor nucleus demonstrated that elimination of Shox2 resulted in the loss of medially-localized subnuclei, specifically the dorsomedial and ventromedial nuclei, in addition to severe disruptions to the development of the intermediate nuclei, especially the dorsal intermediate (Figure 7b). Similar to what we observed at P0 in sagittal sections of the facial motor nucleus (Figure 6), the expression of Isl1, Phox2b, Cntn2, Tubb3, Periph, Slc18a3, Slit2, Shh and Ptch1 were restricted to the lateral or intermediate regions in coronal sections of the facial motor nucleus, or completely lost in Nestin-Cre; Shox2flox/− mutant animals as compared to controls (Figure 7e–v, dashed-circle). While the regional changes in expression were variable between the genes examined, in general the dorsomedial, ventromedial and dorsal intermediate subnuclei of the facial motor nucleus appeared to be most strongly affected in Shox2-mutant animals (Figure 7), which may reflect a regional specific requirement for Shox2 in the facial motor nucleus or may indicate that there are populations of vMNs within the facial motor nucleus that do not express Shox2 during development. Together, the results suggest that Shox2 is required for the development of specific subcompartments within the broader facial motor nucleus.Figure 7


Shox2 is required for the proper development of the facial motor nucleus and the establishment of the facial nerves.

Rosin JM, Kurrasch DM, Cobb J - BMC Neurosci (2015)

Loss of Shox2 function in the facial motor nucleus interferes with the proper development of the medial subnuclei. a Diagram of a coronal section through the P0 brain highlights the region represented in panels b–v, particularly the 7 neuronal clusters that form the facial motor nucleus (nVII, dashed-circle) (image adapted from [23]). b X-gal stained coronal sections through P0 brains shows a loss of medial neuronal clusters in the facial motor nucleus of Nestin-Cre; Shox2lacZ/flox (B″) brains as compared to controls (B′). c–v ISH on P0 control (c, e, g, i, k, m, o, q, s, u) and Nestin-Cre; Shox2flox/− (d, f, h, j, l, n, p, r, t, v) coronal sections shows loss of Shox2 expression (compare c to d, dashed-circle), and decreases in Isl1 (compare e to f, dashed-circle), Phox2b (compare g to h, dashed-circle), Cntn2 (compare i to j, dashed-circle), Tubb3 (compare k to l, dashed-circle), Periph (compare m to n, dashed-circle), Slc18a3 (compare o to p, dashed-circle), Slit2 (compare q to r, dashed-circle), Shh (compare s to t, dashed-circle) and Ptch1 (compare u to v, dashed-circle) expression in the facial motor nucleus. DA dorsal accessory, DM dorsomedial, VM ventromedial, DI dorsal intermediate, VI ventral intermediate, DL dorsolateral, Lat lateral. Scale bar 250 μm.
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Fig7: Loss of Shox2 function in the facial motor nucleus interferes with the proper development of the medial subnuclei. a Diagram of a coronal section through the P0 brain highlights the region represented in panels b–v, particularly the 7 neuronal clusters that form the facial motor nucleus (nVII, dashed-circle) (image adapted from [23]). b X-gal stained coronal sections through P0 brains shows a loss of medial neuronal clusters in the facial motor nucleus of Nestin-Cre; Shox2lacZ/flox (B″) brains as compared to controls (B′). c–v ISH on P0 control (c, e, g, i, k, m, o, q, s, u) and Nestin-Cre; Shox2flox/− (d, f, h, j, l, n, p, r, t, v) coronal sections shows loss of Shox2 expression (compare c to d, dashed-circle), and decreases in Isl1 (compare e to f, dashed-circle), Phox2b (compare g to h, dashed-circle), Cntn2 (compare i to j, dashed-circle), Tubb3 (compare k to l, dashed-circle), Periph (compare m to n, dashed-circle), Slc18a3 (compare o to p, dashed-circle), Slit2 (compare q to r, dashed-circle), Shh (compare s to t, dashed-circle) and Ptch1 (compare u to v, dashed-circle) expression in the facial motor nucleus. DA dorsal accessory, DM dorsomedial, VM ventromedial, DI dorsal intermediate, VI ventral intermediate, DL dorsolateral, Lat lateral. Scale bar 250 μm.
Mentions: Although the loss of Shox2 in the brain resulted in drastic reductions in the size of the facial motor nucleus and the expression of the above-mentioned genes, the spared vMNs remaining in the facial motor nucleus of Shox2 mutants appeared to be mostly localized to the dorsal regions of the nucleus (Figure 6B″, f, n). This suggested that there could be a region-specific loss of distinct subnuclei in the facial motor nucleus in Shox2-mutant animals. Therefore we examined the expression patterns of these same genes in coronal sections of the facial motor nucleus, as this would allow us to more clearly observe the six subnuclei that comprise the main facial motor nucleus; specifically the lateral, dorsolateral, dorsal intermediate, ventral intermediate, dorsomedial and ventromedial nuclei [23, 24]. LacZ staining of P0 control and Nestin-Cre; Shox2lacZ/flox coronal sections of the facial motor nucleus demonstrated that elimination of Shox2 resulted in the loss of medially-localized subnuclei, specifically the dorsomedial and ventromedial nuclei, in addition to severe disruptions to the development of the intermediate nuclei, especially the dorsal intermediate (Figure 7b). Similar to what we observed at P0 in sagittal sections of the facial motor nucleus (Figure 6), the expression of Isl1, Phox2b, Cntn2, Tubb3, Periph, Slc18a3, Slit2, Shh and Ptch1 were restricted to the lateral or intermediate regions in coronal sections of the facial motor nucleus, or completely lost in Nestin-Cre; Shox2flox/− mutant animals as compared to controls (Figure 7e–v, dashed-circle). While the regional changes in expression were variable between the genes examined, in general the dorsomedial, ventromedial and dorsal intermediate subnuclei of the facial motor nucleus appeared to be most strongly affected in Shox2-mutant animals (Figure 7), which may reflect a regional specific requirement for Shox2 in the facial motor nucleus or may indicate that there are populations of vMNs within the facial motor nucleus that do not express Shox2 during development. Together, the results suggest that Shox2 is required for the development of specific subcompartments within the broader facial motor nucleus.Figure 7

Bottom Line: Using a Nestin-Cre driver, we show that elimination of Shox2 throughout the brain results in elevated cell death in the facial motor nucleus at embryonic day 12.5 (E12.5) and E14.5, which correlates with impaired axonal projection properties of vMNs.We also observed changes in the spatial expression of the vMN cell fate factors Isl1 and Phox2b, and concomitant defects in Shh and Ptch1 expression in Shox2 mutants.Furthermore, we demonstrate that elimination of Shox2 results in the loss of dorsomedial and ventromedial subnuclei by postnatal day 0 (P0), which may explain the changes in physical activity and impaired feeding/nursing behavior in Shox2 mutants.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Calgary, 2500 University Drive N.W., BI286D, Calgary, AB, T2N 1N4, Canada. jmrosin2013@gmail.com.

ABSTRACT

Background: Axons from the visceral motor neurons (vMNs) project from nuclei in the hindbrain to innervate autonomic ganglia and branchial arch-derived muscles. Although much is known about the events that govern specification of somatic motor neurons, the genetic pathways responsible for the development of vMNs are less well characterized. We know that vMNs, like all motor neurons, depend on sonic hedgehog signaling for their generation. Similarly, the paired-like homeobox 2b (Phox2b) gene, which is expressed in both proliferating progenitors and post-mitotic motor neurons, is essential for the development of vMNs. Given that our previous study identified a novel role for the short stature homeobox 2 (Shox2) gene in the hindbrain, and since SHOX2 has been shown to regulate transcription of islet 1 (Isl1), an important regulator of vMN development, we sought to determine whether Shox2 is required for the proper development of the facial motor nucleus.

Results: Using a Nestin-Cre driver, we show that elimination of Shox2 throughout the brain results in elevated cell death in the facial motor nucleus at embryonic day 12.5 (E12.5) and E14.5, which correlates with impaired axonal projection properties of vMNs. We also observed changes in the spatial expression of the vMN cell fate factors Isl1 and Phox2b, and concomitant defects in Shh and Ptch1 expression in Shox2 mutants. Furthermore, we demonstrate that elimination of Shox2 results in the loss of dorsomedial and ventromedial subnuclei by postnatal day 0 (P0), which may explain the changes in physical activity and impaired feeding/nursing behavior in Shox2 mutants.

Conclusions: Combined, our data show that Shox2 is required for development of the facial motor nucleus and its associated facial (VII) nerves, and serves as a new molecular tool to probe the genetic programs of this understudied hindbrain region.

No MeSH data available.


Related in: MedlinePlus