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Characterization of the axon initial segment (AIS) of motor neurons and identification of a para-AIS and a juxtapara-AIS, organized by protein 4.1B.

Duflocq A, Chareyre F, Giovannini M, Couraud F, Davenne M - BMC Biol. (2011)

Bottom Line: We also identified in all α motor neurons a hemi-node-type organization, with a contactin-associated protein (Caspr)+ paranode-type, as well as a Caspr2+ and Kv1+ juxtaparanode-type compartment, referred to as a para-AIS and a juxtapara (JXP)-AIS, adjacent to the AIS, where the myelin sheath begins.We found that Kv1 channels appear in the AIS, para-AIS and JXP-AIS concomitantly with myelination and are progressively excluded from the para-AIS.Protein 4.1B plays a key role in ensuring the proper molecular compartmentalization of this hemi-node-type region.

View Article: PubMed Central - HTML - PubMed

Affiliation: INSERM UMRS 952, 9 Quai St Bernard, F-75005, Paris, France.

ABSTRACT

Background: The axon initial segment (AIS) plays a crucial role: it is the site where neurons initiate their electrical outputs. Its composition in terms of voltage-gated sodium (Nav) and voltage-gated potassium (Kv) channels, as well as its length and localization determine the neuron's spiking properties. Some neurons are able to modulate their AIS length or distance from the soma in order to adapt their excitability properties to their activity level. It is therefore crucial to characterize all these parameters and determine where the myelin sheath begins in order to assess a neuron's excitability properties and ability to display such plasticity mechanisms. If the myelin sheath starts immediately after the AIS, another question then arises as to how would the axon be organized at its first myelin attachment site; since AISs are different from nodes of Ranvier, would this particular axonal region resemble a hemi-node of Ranvier?

Results: We have characterized the AIS of mouse somatic motor neurons. In addition to constant determinants of excitability properties, we found heterogeneities, in terms of AIS localization and Nav composition. We also identified in all α motor neurons a hemi-node-type organization, with a contactin-associated protein (Caspr)+ paranode-type, as well as a Caspr2+ and Kv1+ juxtaparanode-type compartment, referred to as a para-AIS and a juxtapara (JXP)-AIS, adjacent to the AIS, where the myelin sheath begins. We found that Kv1 channels appear in the AIS, para-AIS and JXP-AIS concomitantly with myelination and are progressively excluded from the para-AIS. Their expression in the AIS and JXP-AIS is independent from transient axonal glycoprotein-1 (TAG-1)/Caspr2, in contrast to juxtaparanodes, and independent from PSD-93. Data from mice lacking the cytoskeletal linker protein 4.1B show that this protein is necessary to form the Caspr+ para-AIS barrier, ensuring the compartmentalization of Kv1 channels and the segregation of the AIS, para-AIS and JXP-AIS.

Conclusions: α Motor neurons have heterogeneous AISs, which underlie different spiking properties. However, they all have a para-AIS and a JXP-AIS contiguous to their AIS, where the myelin sheath begins, which might limit some AIS plasticity. Protein 4.1B plays a key role in ensuring the proper molecular compartmentalization of this hemi-node-type region.

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The transient axonal glycoprotein-1 (TAG-1)/contactin-associated protein-like 2 (Caspr2) complex is not required for voltage-gated potassium (Kv)1 channels expression at the axon initial segment (AIS) and juxtapara (JXP)-AIS. Triple immunostaining of ankyrin G (AnkG) (A, D), Kv1.1 (B, E) and Caspr2 (C, F) in motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown), in wild-type (WT) (A-C) and TAG-1-/- (D-F) mice. Inset in (E): Mean Kv1.1 immunofluorescence intensity at the AIS and JXP-AIS in WT and KO mice. Brackets indicate Kv1.1+ and Caspr2+ domains in the AIS and JXP-AIS. Scale bar = 5 μm.
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Figure 8: The transient axonal glycoprotein-1 (TAG-1)/contactin-associated protein-like 2 (Caspr2) complex is not required for voltage-gated potassium (Kv)1 channels expression at the axon initial segment (AIS) and juxtapara (JXP)-AIS. Triple immunostaining of ankyrin G (AnkG) (A, D), Kv1.1 (B, E) and Caspr2 (C, F) in motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown), in wild-type (WT) (A-C) and TAG-1-/- (D-F) mice. Inset in (E): Mean Kv1.1 immunofluorescence intensity at the AIS and JXP-AIS in WT and KO mice. Brackets indicate Kv1.1+ and Caspr2+ domains in the AIS and JXP-AIS. Scale bar = 5 μm.

Mentions: We first tested whether the TAG-1/Caspr2 complex also controls the clustering of Kv1 channels at the MN JXP-AIS, similarly to JXP-nodes. Caspr2 is indeed a good candidate since it was distributed in the MN JXP-AIS, as well as in the MN AIS, exactly like Kv1 channels: in adult MNs its distribution was restricted to the distal AIS, overlapping exactly with Kv1.1 distribution, and was found at the JXP-AIS with a higher level of expression, similarly to Kv1.1 (Figure 8B, C). In addition, Caspr2 was also expressed at the MN JXP-AIS and AIS at the very early stage of Kv1 channels expression in these compartments (data not shown). We thus used TAG-1-/- mice [38], in which Caspr2 expression was abolished at the MN JXP-AIS, as in JXP-nodes. Interestingly, Caspr2 expression was also missing at the MN AIS (Figure 8F). Surprisingly, in TAG-1-/- mice we found a normal distribution of Kv1.1 in 100% of MN JXP-AISs analyzed, similar to wild-type (WT) littermate controls (Figure 8B, E): Kv1.1 at the MN JXP-AIS displayed no statistically significant difference of immunofluorescence intensity in TAG-1-/- compared to WT mice (Figure 8E; from respectively 32 and 11 MN JXP-AISs analyzed from 3 TAG-1-/- and 3 WT mice). This result contrasts with the dramatic decrease in Kv1 channels expression found at JXP-nodes in sciatic nerves [20,21], which include both motor and sensory axons. In order to test whether this discrepancy could be due to motor axons being less vulnerable to the lack of TAG-1 and Caspr2 than sensory axons, we analyzed more specifically teased fibers from spinal cord ventral roots contributing to the sciatic nerve, which contain JXP-nodes exclusively from MNs. In TAG-1-/- mice, Kv1.1 expression was dramatically decreased in 88.1% of these MNs' JXP-nodes, as compared to WT mice (respectively 67 and 74 MN JXP-nodes analyzed from 3 TAG-1-/- and 3 WT mice; see Additional file 2, Figure S2), like at sciatic nerve JXP-nodes [20,21]. Spinal somatic MNs thus have a JXP-AIS that differs from their peripheral JXP-nodes and from the majority of JXP-nodes studied so far, in that it seems to be able to cluster Kv1 channels by a TAG-1/Caspr2-independent mechanism.


Characterization of the axon initial segment (AIS) of motor neurons and identification of a para-AIS and a juxtapara-AIS, organized by protein 4.1B.

Duflocq A, Chareyre F, Giovannini M, Couraud F, Davenne M - BMC Biol. (2011)

The transient axonal glycoprotein-1 (TAG-1)/contactin-associated protein-like 2 (Caspr2) complex is not required for voltage-gated potassium (Kv)1 channels expression at the axon initial segment (AIS) and juxtapara (JXP)-AIS. Triple immunostaining of ankyrin G (AnkG) (A, D), Kv1.1 (B, E) and Caspr2 (C, F) in motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown), in wild-type (WT) (A-C) and TAG-1-/- (D-F) mice. Inset in (E): Mean Kv1.1 immunofluorescence intensity at the AIS and JXP-AIS in WT and KO mice. Brackets indicate Kv1.1+ and Caspr2+ domains in the AIS and JXP-AIS. Scale bar = 5 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 8: The transient axonal glycoprotein-1 (TAG-1)/contactin-associated protein-like 2 (Caspr2) complex is not required for voltage-gated potassium (Kv)1 channels expression at the axon initial segment (AIS) and juxtapara (JXP)-AIS. Triple immunostaining of ankyrin G (AnkG) (A, D), Kv1.1 (B, E) and Caspr2 (C, F) in motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown), in wild-type (WT) (A-C) and TAG-1-/- (D-F) mice. Inset in (E): Mean Kv1.1 immunofluorescence intensity at the AIS and JXP-AIS in WT and KO mice. Brackets indicate Kv1.1+ and Caspr2+ domains in the AIS and JXP-AIS. Scale bar = 5 μm.
Mentions: We first tested whether the TAG-1/Caspr2 complex also controls the clustering of Kv1 channels at the MN JXP-AIS, similarly to JXP-nodes. Caspr2 is indeed a good candidate since it was distributed in the MN JXP-AIS, as well as in the MN AIS, exactly like Kv1 channels: in adult MNs its distribution was restricted to the distal AIS, overlapping exactly with Kv1.1 distribution, and was found at the JXP-AIS with a higher level of expression, similarly to Kv1.1 (Figure 8B, C). In addition, Caspr2 was also expressed at the MN JXP-AIS and AIS at the very early stage of Kv1 channels expression in these compartments (data not shown). We thus used TAG-1-/- mice [38], in which Caspr2 expression was abolished at the MN JXP-AIS, as in JXP-nodes. Interestingly, Caspr2 expression was also missing at the MN AIS (Figure 8F). Surprisingly, in TAG-1-/- mice we found a normal distribution of Kv1.1 in 100% of MN JXP-AISs analyzed, similar to wild-type (WT) littermate controls (Figure 8B, E): Kv1.1 at the MN JXP-AIS displayed no statistically significant difference of immunofluorescence intensity in TAG-1-/- compared to WT mice (Figure 8E; from respectively 32 and 11 MN JXP-AISs analyzed from 3 TAG-1-/- and 3 WT mice). This result contrasts with the dramatic decrease in Kv1 channels expression found at JXP-nodes in sciatic nerves [20,21], which include both motor and sensory axons. In order to test whether this discrepancy could be due to motor axons being less vulnerable to the lack of TAG-1 and Caspr2 than sensory axons, we analyzed more specifically teased fibers from spinal cord ventral roots contributing to the sciatic nerve, which contain JXP-nodes exclusively from MNs. In TAG-1-/- mice, Kv1.1 expression was dramatically decreased in 88.1% of these MNs' JXP-nodes, as compared to WT mice (respectively 67 and 74 MN JXP-nodes analyzed from 3 TAG-1-/- and 3 WT mice; see Additional file 2, Figure S2), like at sciatic nerve JXP-nodes [20,21]. Spinal somatic MNs thus have a JXP-AIS that differs from their peripheral JXP-nodes and from the majority of JXP-nodes studied so far, in that it seems to be able to cluster Kv1 channels by a TAG-1/Caspr2-independent mechanism.

Bottom Line: We also identified in all α motor neurons a hemi-node-type organization, with a contactin-associated protein (Caspr)+ paranode-type, as well as a Caspr2+ and Kv1+ juxtaparanode-type compartment, referred to as a para-AIS and a juxtapara (JXP)-AIS, adjacent to the AIS, where the myelin sheath begins.We found that Kv1 channels appear in the AIS, para-AIS and JXP-AIS concomitantly with myelination and are progressively excluded from the para-AIS.Protein 4.1B plays a key role in ensuring the proper molecular compartmentalization of this hemi-node-type region.

View Article: PubMed Central - HTML - PubMed

Affiliation: INSERM UMRS 952, 9 Quai St Bernard, F-75005, Paris, France.

ABSTRACT

Background: The axon initial segment (AIS) plays a crucial role: it is the site where neurons initiate their electrical outputs. Its composition in terms of voltage-gated sodium (Nav) and voltage-gated potassium (Kv) channels, as well as its length and localization determine the neuron's spiking properties. Some neurons are able to modulate their AIS length or distance from the soma in order to adapt their excitability properties to their activity level. It is therefore crucial to characterize all these parameters and determine where the myelin sheath begins in order to assess a neuron's excitability properties and ability to display such plasticity mechanisms. If the myelin sheath starts immediately after the AIS, another question then arises as to how would the axon be organized at its first myelin attachment site; since AISs are different from nodes of Ranvier, would this particular axonal region resemble a hemi-node of Ranvier?

Results: We have characterized the AIS of mouse somatic motor neurons. In addition to constant determinants of excitability properties, we found heterogeneities, in terms of AIS localization and Nav composition. We also identified in all α motor neurons a hemi-node-type organization, with a contactin-associated protein (Caspr)+ paranode-type, as well as a Caspr2+ and Kv1+ juxtaparanode-type compartment, referred to as a para-AIS and a juxtapara (JXP)-AIS, adjacent to the AIS, where the myelin sheath begins. We found that Kv1 channels appear in the AIS, para-AIS and JXP-AIS concomitantly with myelination and are progressively excluded from the para-AIS. Their expression in the AIS and JXP-AIS is independent from transient axonal glycoprotein-1 (TAG-1)/Caspr2, in contrast to juxtaparanodes, and independent from PSD-93. Data from mice lacking the cytoskeletal linker protein 4.1B show that this protein is necessary to form the Caspr+ para-AIS barrier, ensuring the compartmentalization of Kv1 channels and the segregation of the AIS, para-AIS and JXP-AIS.

Conclusions: α Motor neurons have heterogeneous AISs, which underlie different spiking properties. However, they all have a para-AIS and a JXP-AIS contiguous to their AIS, where the myelin sheath begins, which might limit some AIS plasticity. Protein 4.1B plays a key role in ensuring the proper molecular compartmentalization of this hemi-node-type region.

Show MeSH
Related in: MedlinePlus