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Evidence for a role of srGAP3 in the positioning of commissural axons within the ventrolateral funiculus of the mouse spinal cord.

Bacon C, Endris V, Andermatt I, Niederkofler V, Waltereit R, Bartsch D, Stoeckli ET, Rappold G - PLoS ONE (2011)

Bottom Line: Interestingly we observed a significant thickening of the ventral funiculus and a thinning of the lateral funiculus in the srGAP3 KO spinal cord, which has also recently been reported in the Robo2 KO.However, axons in the enlarged ventral funiculus of the srGAP3 KO are Robo1 positive but do not express Robo2, indicating that the thickening of the ventral funiculus in the srGAP3 KO is not a Robo2 mediated effect.We suggest a role for srGAP3 in the lateral positioning of post crossing axons within the ventrolateral funiculus.

View Article: PubMed Central - PubMed

Affiliation: Department of Human Molecular Genetics, University of Heidelberg, Heidelberg, Germany. Claire.bacon@med.uni-heidelberg.de

ABSTRACT
Slit-Robo signaling guides commissural axons away from the floor-plate of the spinal cord and into the longitudinal axis after crossing the midline. In this study we have evaluated the role of the Slit-Robo GTPase activating protein 3 (srGAP3) in commissural axon guidance using a knockout (KO) mouse model. Co-immunoprecipitation experiments confirmed that srGAP3 interacts with the Slit receptors Robo1 and Robo2 and immunohistochemistry studies showed that srGAP3 co-localises with Robo1 in the ventral and lateral funiculus and with Robo2 in the lateral funiculus. Stalling axons have been reported in the floor-plate of Slit and Robo mutant spinal cords but our axon tracing experiments revealed no dorsal commissural axon stalling in the floor plate of the srGAP3 KO mouse. Interestingly we observed a significant thickening of the ventral funiculus and a thinning of the lateral funiculus in the srGAP3 KO spinal cord, which has also recently been reported in the Robo2 KO. However, axons in the enlarged ventral funiculus of the srGAP3 KO are Robo1 positive but do not express Robo2, indicating that the thickening of the ventral funiculus in the srGAP3 KO is not a Robo2 mediated effect. We suggest a role for srGAP3 in the lateral positioning of post crossing axons within the ventrolateral funiculus.

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Commissural axons do not stall in srGAP3 KO spinal cords.WT and srGAP3 KO spinal cords were prepared in an open book configuration and dorsolateral commissural axons were traced by the injection of DiI into the dorsal spinal cord. A: Injections sites were classified as being either normal, containing >50% of axons stalling in the floor-plate, at the contralateral exit site, or both. No significant differences in the % occurrence of each phenotype were observed between WT and srGAP3 KO cord sections, using the Independent Samples T-test. (P value comparing the % of normal injection sites in WT and srGAP3 KO cords  = 0.12. P value comparing the % of injection sites with floor plate stalling in WT and srGAP3 KO cords  = 0.08). B: The number of embryos and the number of injection sites analysed for each genotype is provided in the table. C: Representative images of open book preparations injected with DiI showing that axons cross the floor plate normally and project longitudinally after crossing the contralateral exit site (marked by the white line).
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pone-0019887-g003: Commissural axons do not stall in srGAP3 KO spinal cords.WT and srGAP3 KO spinal cords were prepared in an open book configuration and dorsolateral commissural axons were traced by the injection of DiI into the dorsal spinal cord. A: Injections sites were classified as being either normal, containing >50% of axons stalling in the floor-plate, at the contralateral exit site, or both. No significant differences in the % occurrence of each phenotype were observed between WT and srGAP3 KO cord sections, using the Independent Samples T-test. (P value comparing the % of normal injection sites in WT and srGAP3 KO cords  = 0.12. P value comparing the % of injection sites with floor plate stalling in WT and srGAP3 KO cords  = 0.08). B: The number of embryos and the number of injection sites analysed for each genotype is provided in the table. C: Representative images of open book preparations injected with DiI showing that axons cross the floor plate normally and project longitudinally after crossing the contralateral exit site (marked by the white line).

Mentions: Commissural axons stall during crossing in Robo1 KO, Robo1/Robo2 double KO and Slit triple KO spinal cords [2], [4]. We wanted to determine whether commissural axons also stall in the spinal cord of srGAP3 KO mice. To do this we injected DiI into the area of the cell bodies of commissural neurons in the dorsal spinal cord of srGAP3 KO and WT spinal cords. We did not observe axons stalling within the floor plate of KO spinal cords (7% of injection sites, n = 97) significantly more often than in WT spinal cords (0% of injection sites, n = 51, P = 0.077) (Figure 3A). Additionally we found no significant difference in the number of axons stalling at the floor plate exit site in srGAP3 KO (21%) compared to WT (19%) spinal cords (Figure 3A). Post-crossing trajectories of these subpopulations of commissural neurons did not differ between WT and KO (Figure 3C).


Evidence for a role of srGAP3 in the positioning of commissural axons within the ventrolateral funiculus of the mouse spinal cord.

Bacon C, Endris V, Andermatt I, Niederkofler V, Waltereit R, Bartsch D, Stoeckli ET, Rappold G - PLoS ONE (2011)

Commissural axons do not stall in srGAP3 KO spinal cords.WT and srGAP3 KO spinal cords were prepared in an open book configuration and dorsolateral commissural axons were traced by the injection of DiI into the dorsal spinal cord. A: Injections sites were classified as being either normal, containing >50% of axons stalling in the floor-plate, at the contralateral exit site, or both. No significant differences in the % occurrence of each phenotype were observed between WT and srGAP3 KO cord sections, using the Independent Samples T-test. (P value comparing the % of normal injection sites in WT and srGAP3 KO cords  = 0.12. P value comparing the % of injection sites with floor plate stalling in WT and srGAP3 KO cords  = 0.08). B: The number of embryos and the number of injection sites analysed for each genotype is provided in the table. C: Representative images of open book preparations injected with DiI showing that axons cross the floor plate normally and project longitudinally after crossing the contralateral exit site (marked by the white line).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3104994&req=5

pone-0019887-g003: Commissural axons do not stall in srGAP3 KO spinal cords.WT and srGAP3 KO spinal cords were prepared in an open book configuration and dorsolateral commissural axons were traced by the injection of DiI into the dorsal spinal cord. A: Injections sites were classified as being either normal, containing >50% of axons stalling in the floor-plate, at the contralateral exit site, or both. No significant differences in the % occurrence of each phenotype were observed between WT and srGAP3 KO cord sections, using the Independent Samples T-test. (P value comparing the % of normal injection sites in WT and srGAP3 KO cords  = 0.12. P value comparing the % of injection sites with floor plate stalling in WT and srGAP3 KO cords  = 0.08). B: The number of embryos and the number of injection sites analysed for each genotype is provided in the table. C: Representative images of open book preparations injected with DiI showing that axons cross the floor plate normally and project longitudinally after crossing the contralateral exit site (marked by the white line).
Mentions: Commissural axons stall during crossing in Robo1 KO, Robo1/Robo2 double KO and Slit triple KO spinal cords [2], [4]. We wanted to determine whether commissural axons also stall in the spinal cord of srGAP3 KO mice. To do this we injected DiI into the area of the cell bodies of commissural neurons in the dorsal spinal cord of srGAP3 KO and WT spinal cords. We did not observe axons stalling within the floor plate of KO spinal cords (7% of injection sites, n = 97) significantly more often than in WT spinal cords (0% of injection sites, n = 51, P = 0.077) (Figure 3A). Additionally we found no significant difference in the number of axons stalling at the floor plate exit site in srGAP3 KO (21%) compared to WT (19%) spinal cords (Figure 3A). Post-crossing trajectories of these subpopulations of commissural neurons did not differ between WT and KO (Figure 3C).

Bottom Line: Interestingly we observed a significant thickening of the ventral funiculus and a thinning of the lateral funiculus in the srGAP3 KO spinal cord, which has also recently been reported in the Robo2 KO.However, axons in the enlarged ventral funiculus of the srGAP3 KO are Robo1 positive but do not express Robo2, indicating that the thickening of the ventral funiculus in the srGAP3 KO is not a Robo2 mediated effect.We suggest a role for srGAP3 in the lateral positioning of post crossing axons within the ventrolateral funiculus.

View Article: PubMed Central - PubMed

Affiliation: Department of Human Molecular Genetics, University of Heidelberg, Heidelberg, Germany. Claire.bacon@med.uni-heidelberg.de

ABSTRACT
Slit-Robo signaling guides commissural axons away from the floor-plate of the spinal cord and into the longitudinal axis after crossing the midline. In this study we have evaluated the role of the Slit-Robo GTPase activating protein 3 (srGAP3) in commissural axon guidance using a knockout (KO) mouse model. Co-immunoprecipitation experiments confirmed that srGAP3 interacts with the Slit receptors Robo1 and Robo2 and immunohistochemistry studies showed that srGAP3 co-localises with Robo1 in the ventral and lateral funiculus and with Robo2 in the lateral funiculus. Stalling axons have been reported in the floor-plate of Slit and Robo mutant spinal cords but our axon tracing experiments revealed no dorsal commissural axon stalling in the floor plate of the srGAP3 KO mouse. Interestingly we observed a significant thickening of the ventral funiculus and a thinning of the lateral funiculus in the srGAP3 KO spinal cord, which has also recently been reported in the Robo2 KO. However, axons in the enlarged ventral funiculus of the srGAP3 KO are Robo1 positive but do not express Robo2, indicating that the thickening of the ventral funiculus in the srGAP3 KO is not a Robo2 mediated effect. We suggest a role for srGAP3 in the lateral positioning of post crossing axons within the ventrolateral funiculus.

Show MeSH
Related in: MedlinePlus