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Abundant expression of guidance and synaptogenic molecules in the injured spinal cord.

Jacobi A, Schmalz A, Bareyre FM - PLoS ONE (2014)

Bottom Line: To assign the expression of these molecules to distinct populations of interneurons we labeled short and long propriospinal neurons by retrograde tracing and glycinergic neurons using a transgenically expressed fluorescent protein.Overall the expression pattern of guidance and synaptogenic molecules in the cervical spinal cord appeared to be stable over time and was not substantially altered following a midthoracic spinal cord injury.Taken together, our study indicates that many of the guidance and synaptogenic cues that regulate neuronal circuit formation in development are also present in the adult CNS and therefore likely contribute to the remodelling of axonal connections in the injured spinal cord.

View Article: PubMed Central - PubMed

Affiliation: Institute of Clinical Neuroimmunology, Ludwig-Maximilians University Munich, Munich, Germany.

ABSTRACT

Background: Spinal interneurons have emerged as crucial targets of supraspinal input during post-injury axonal remodelling. For example, lesioned corticospinal projections use propriospinal neurons as relay stations to form intraspinal detour circuits that circumvent the lesion site and contribute to functional recovery. While a number of the molecules that determine the formation of neuronal circuits in the developing nervous system have been identified, it is much less understood which of these cues are also expressed in the injured spinal cord and can thus guide growing collaterals and initiate synaptogenesis during circuit remodelling.

Methodology/principal findings: To address this question we characterized the expression profile of a number of guidance and synaptogenic molecules in the cervical spinal cord of healthy and spinal cord-injured mice by in situ hybridization. To assign the expression of these molecules to distinct populations of interneurons we labeled short and long propriospinal neurons by retrograde tracing and glycinergic neurons using a transgenically expressed fluorescent protein. Interestingly, we found that most of the molecules studied including members of slit-, semaphorin-, synCAM-, neuroligin- and ephrin- families as well as their receptors are also present in the adult CNS. While many of these molecules were abundantly expressed in all interneurons examined, some molecules including slits, semaphorin 7a, synCAM4 and neuroligin 1 showed preferential expression in propriospinal interneurons. Overall the expression pattern of guidance and synaptogenic molecules in the cervical spinal cord appeared to be stable over time and was not substantially altered following a midthoracic spinal cord injury.

Conclusions: Taken together, our study indicates that many of the guidance and synaptogenic cues that regulate neuronal circuit formation in development are also present in the adult CNS and therefore likely contribute to the remodelling of axonal connections in the injured spinal cord.

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In Situ hybridization profile of EphB2 and EphrinB1 in cervical interneurons.In situ hybridization of EphB2 (A), EphrinB1 (B) mRNA in the cortex. No signal is detected with the sense probes for EphB2 (B) and EphrinB1 (E). Dotted lines in A and B represent layer V of the cortex. (C, F) Confocal images of double-labeled CST projection neurons (C; retrogradely-labeled CST projection neurons: green; EphB2: red and F; retrogradely-labeled CST projection neurons: green; EphrinB1: red). Scale bars in A, B, D, E: 100 µm; Scale bars C,F: 50 µm (25 µm in insets).
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pone-0088449-g009: In Situ hybridization profile of EphB2 and EphrinB1 in cervical interneurons.In situ hybridization of EphB2 (A), EphrinB1 (B) mRNA in the cortex. No signal is detected with the sense probes for EphB2 (B) and EphrinB1 (E). Dotted lines in A and B represent layer V of the cortex. (C, F) Confocal images of double-labeled CST projection neurons (C; retrogradely-labeled CST projection neurons: green; EphB2: red and F; retrogradely-labeled CST projection neurons: green; EphrinB1: red). Scale bars in A, B, D, E: 100 µm; Scale bars C,F: 50 µm (25 µm in insets).

Mentions: Ephrins and their receptors (Eph) are pleiotropic molecules involved in cell migration, axon guidance [32] and synapse formation [33] during nervous system development. Interestingly, eph-ephrin interactions can mediate both repulsive and attractive forces between cells [34]. EphB-ephrinB interaction has been shown to be important for proper ipsilateral targeting of CST and retinal axons [35]–[37]. In situ hybridization revealed that both ephB2 and ephrinB1mRNA are expressed throughout all laminae of the gray matter of the cervical spinal cord (Fig. 8A,B,G,H) with EphrinB1 having a dimmer expression in laminae I-IV (Table 2). Morphology analysis after counterstaining with NeuN suggests that both molecules are primarily expressed by neurons including by interneurons and at least in the case of EphB2 also motoneurons (Fig. 8C,I). Further characterization indeed showed that both EphB2 and Ephrin B1 are expressed in different interneuronal population located in the cervical spinal cord including LPSN, SPSN and glycinergic interneurons (Fig. 8D-F, J-L). While EphB2 mRNA is expressed by a similar proportion of these interneurons, Ephrin B1 was particularly prominently expressed in short propriospinal neurons in the healthy cervical spinal cord. Mostly, these expression patterns in the cervical spinal cord were not affected by the presence of a thoracic spinal cord injury (Fig. 8 M,N). The only exception is that the percentage of glycinergic interneurons expressing EphB2 was increased in the cervical spinal cord at 3 weeks post injury and decreased at 12 weeks after injury. Further expression analysis of cortical sections showed that both, Ephrin B1 and EphB2, were also expressed in layer II-III and layer V of the cortex (Fig. 9 A,B,D,E and Table 3). Retrograde labelling of layer V CST projection neurons revealed that 84, 6±3,5% of CST projection neurons express EphrinB1 and 74,1±2,2% EphB2 (Fig. 9 C,F).


Abundant expression of guidance and synaptogenic molecules in the injured spinal cord.

Jacobi A, Schmalz A, Bareyre FM - PLoS ONE (2014)

In Situ hybridization profile of EphB2 and EphrinB1 in cervical interneurons.In situ hybridization of EphB2 (A), EphrinB1 (B) mRNA in the cortex. No signal is detected with the sense probes for EphB2 (B) and EphrinB1 (E). Dotted lines in A and B represent layer V of the cortex. (C, F) Confocal images of double-labeled CST projection neurons (C; retrogradely-labeled CST projection neurons: green; EphB2: red and F; retrogradely-labeled CST projection neurons: green; EphrinB1: red). Scale bars in A, B, D, E: 100 µm; Scale bars C,F: 50 µm (25 µm in insets).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3921160&req=5

pone-0088449-g009: In Situ hybridization profile of EphB2 and EphrinB1 in cervical interneurons.In situ hybridization of EphB2 (A), EphrinB1 (B) mRNA in the cortex. No signal is detected with the sense probes for EphB2 (B) and EphrinB1 (E). Dotted lines in A and B represent layer V of the cortex. (C, F) Confocal images of double-labeled CST projection neurons (C; retrogradely-labeled CST projection neurons: green; EphB2: red and F; retrogradely-labeled CST projection neurons: green; EphrinB1: red). Scale bars in A, B, D, E: 100 µm; Scale bars C,F: 50 µm (25 µm in insets).
Mentions: Ephrins and their receptors (Eph) are pleiotropic molecules involved in cell migration, axon guidance [32] and synapse formation [33] during nervous system development. Interestingly, eph-ephrin interactions can mediate both repulsive and attractive forces between cells [34]. EphB-ephrinB interaction has been shown to be important for proper ipsilateral targeting of CST and retinal axons [35]–[37]. In situ hybridization revealed that both ephB2 and ephrinB1mRNA are expressed throughout all laminae of the gray matter of the cervical spinal cord (Fig. 8A,B,G,H) with EphrinB1 having a dimmer expression in laminae I-IV (Table 2). Morphology analysis after counterstaining with NeuN suggests that both molecules are primarily expressed by neurons including by interneurons and at least in the case of EphB2 also motoneurons (Fig. 8C,I). Further characterization indeed showed that both EphB2 and Ephrin B1 are expressed in different interneuronal population located in the cervical spinal cord including LPSN, SPSN and glycinergic interneurons (Fig. 8D-F, J-L). While EphB2 mRNA is expressed by a similar proportion of these interneurons, Ephrin B1 was particularly prominently expressed in short propriospinal neurons in the healthy cervical spinal cord. Mostly, these expression patterns in the cervical spinal cord were not affected by the presence of a thoracic spinal cord injury (Fig. 8 M,N). The only exception is that the percentage of glycinergic interneurons expressing EphB2 was increased in the cervical spinal cord at 3 weeks post injury and decreased at 12 weeks after injury. Further expression analysis of cortical sections showed that both, Ephrin B1 and EphB2, were also expressed in layer II-III and layer V of the cortex (Fig. 9 A,B,D,E and Table 3). Retrograde labelling of layer V CST projection neurons revealed that 84, 6±3,5% of CST projection neurons express EphrinB1 and 74,1±2,2% EphB2 (Fig. 9 C,F).

Bottom Line: To assign the expression of these molecules to distinct populations of interneurons we labeled short and long propriospinal neurons by retrograde tracing and glycinergic neurons using a transgenically expressed fluorescent protein.Overall the expression pattern of guidance and synaptogenic molecules in the cervical spinal cord appeared to be stable over time and was not substantially altered following a midthoracic spinal cord injury.Taken together, our study indicates that many of the guidance and synaptogenic cues that regulate neuronal circuit formation in development are also present in the adult CNS and therefore likely contribute to the remodelling of axonal connections in the injured spinal cord.

View Article: PubMed Central - PubMed

Affiliation: Institute of Clinical Neuroimmunology, Ludwig-Maximilians University Munich, Munich, Germany.

ABSTRACT

Background: Spinal interneurons have emerged as crucial targets of supraspinal input during post-injury axonal remodelling. For example, lesioned corticospinal projections use propriospinal neurons as relay stations to form intraspinal detour circuits that circumvent the lesion site and contribute to functional recovery. While a number of the molecules that determine the formation of neuronal circuits in the developing nervous system have been identified, it is much less understood which of these cues are also expressed in the injured spinal cord and can thus guide growing collaterals and initiate synaptogenesis during circuit remodelling.

Methodology/principal findings: To address this question we characterized the expression profile of a number of guidance and synaptogenic molecules in the cervical spinal cord of healthy and spinal cord-injured mice by in situ hybridization. To assign the expression of these molecules to distinct populations of interneurons we labeled short and long propriospinal neurons by retrograde tracing and glycinergic neurons using a transgenically expressed fluorescent protein. Interestingly, we found that most of the molecules studied including members of slit-, semaphorin-, synCAM-, neuroligin- and ephrin- families as well as their receptors are also present in the adult CNS. While many of these molecules were abundantly expressed in all interneurons examined, some molecules including slits, semaphorin 7a, synCAM4 and neuroligin 1 showed preferential expression in propriospinal interneurons. Overall the expression pattern of guidance and synaptogenic molecules in the cervical spinal cord appeared to be stable over time and was not substantially altered following a midthoracic spinal cord injury.

Conclusions: Taken together, our study indicates that many of the guidance and synaptogenic cues that regulate neuronal circuit formation in development are also present in the adult CNS and therefore likely contribute to the remodelling of axonal connections in the injured spinal cord.

Show MeSH
Related in: MedlinePlus