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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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Identification of a juxtapara (JXP)-axon initial segment (AIS). Triple immunostaining of ankyrin G (AnkG) (A, E), contactin-associated protein (Caspr) (B, F) and voltage-gated potassium (Kv)1 channels (C, G) (Caspr and Kv1 are merged in (D, H)) along the axon of motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown). The Caspr+ segment is immediately followed by a Kv1+ segment: Kv1.1+ (D) and Kv1.2+ (H). Triple immunostaining of AnkG (I), Kvβ2 (J) and Kv1.1 (K) (Kvβ2 and Kv1.1 are merged in (L) along the axon of MNs. Brackets indicate the Kv1+ or Kvβ2+ AIS and JXP-AIS and the Caspr+ para-AIS. Scale bar = 5 μm.
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Figure 6: Identification of a juxtapara (JXP)-axon initial segment (AIS). Triple immunostaining of ankyrin G (AnkG) (A, E), contactin-associated protein (Caspr) (B, F) and voltage-gated potassium (Kv)1 channels (C, G) (Caspr and Kv1 are merged in (D, H)) along the axon of motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown). The Caspr+ segment is immediately followed by a Kv1+ segment: Kv1.1+ (D) and Kv1.2+ (H). Triple immunostaining of AnkG (I), Kvβ2 (J) and Kv1.1 (K) (Kvβ2 and Kv1.1 are merged in (L) along the axon of MNs. Brackets indicate the Kv1+ or Kvβ2+ AIS and JXP-AIS and the Caspr+ para-AIS. Scale bar = 5 μm.

Mentions: We then analyzed whether the AIS and para-AIS were further flanked by a JXP-node-like compartment, which would further support the presence of a hemi-node-type organization at the distal tip of the AIS. We analyzed the expression of JXP-node markers, namely Kv1.1 and Kv1.2. In 100% of somatic α MNs (n = 68) we found Kv1.1 and Kv1.2 expressed in a compartment contiguous to the Caspr+ para-AIS, on the opposite side of the distal AnkG+ AIS expressing Kv1.1 and Kv1.2 (Figure 6A-H). This result suggests the presence of a JXP-node-like compartment, which we here refer to as a 'juxtapara-AIS' ('JXP-AIS'). Kvβ2, which we had found to be expressed in the distal AIS of MNs, was also expressed in their JXP-AIS (Figure 6I-L). As in the AIS, Kvβ2 was found perfectly colocalized with Kv1.1 (Figure 6J-L) and Kv1.2 (data not shown), suggesting that heteromultimeric channels formed by the association of Kv1.1, Kv1.2 and Kvβ2 are present in the JXP-AIS. Of note, their expression levels in the JXP-AIS, in particular in its proximal domain adjacent to the para-AIS, always appeared higher than in the AIS (Figure 6C, G, K). In addition, the axon displays a significant widening in the JXP-AIS as compared to the AIS, which contributes to the higher expression level of Kv1.1, Kv1.2 and Kvβ2 observed in the JXP-AIS.


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)

Identification of a juxtapara (JXP)-axon initial segment (AIS). Triple immunostaining of ankyrin G (AnkG) (A, E), contactin-associated protein (Caspr) (B, F) and voltage-gated potassium (Kv)1 channels (C, G) (Caspr and Kv1 are merged in (D, H)) along the axon of motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown). The Caspr+ segment is immediately followed by a Kv1+ segment: Kv1.1+ (D) and Kv1.2+ (H). Triple immunostaining of AnkG (I), Kvβ2 (J) and Kv1.1 (K) (Kvβ2 and Kv1.1 are merged in (L) along the axon of MNs. Brackets indicate the Kv1+ or Kvβ2+ AIS and JXP-AIS and the Caspr+ para-AIS. Scale bar = 5 μm.
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Figure 6: Identification of a juxtapara (JXP)-axon initial segment (AIS). Triple immunostaining of ankyrin G (AnkG) (A, E), contactin-associated protein (Caspr) (B, F) and voltage-gated potassium (Kv)1 channels (C, G) (Caspr and Kv1 are merged in (D, H)) along the axon of motor neurons (MNs) labeled with the anti-Peripherin antibody (data not shown). The Caspr+ segment is immediately followed by a Kv1+ segment: Kv1.1+ (D) and Kv1.2+ (H). Triple immunostaining of AnkG (I), Kvβ2 (J) and Kv1.1 (K) (Kvβ2 and Kv1.1 are merged in (L) along the axon of MNs. Brackets indicate the Kv1+ or Kvβ2+ AIS and JXP-AIS and the Caspr+ para-AIS. Scale bar = 5 μm.
Mentions: We then analyzed whether the AIS and para-AIS were further flanked by a JXP-node-like compartment, which would further support the presence of a hemi-node-type organization at the distal tip of the AIS. We analyzed the expression of JXP-node markers, namely Kv1.1 and Kv1.2. In 100% of somatic α MNs (n = 68) we found Kv1.1 and Kv1.2 expressed in a compartment contiguous to the Caspr+ para-AIS, on the opposite side of the distal AnkG+ AIS expressing Kv1.1 and Kv1.2 (Figure 6A-H). This result suggests the presence of a JXP-node-like compartment, which we here refer to as a 'juxtapara-AIS' ('JXP-AIS'). Kvβ2, which we had found to be expressed in the distal AIS of MNs, was also expressed in their JXP-AIS (Figure 6I-L). As in the AIS, Kvβ2 was found perfectly colocalized with Kv1.1 (Figure 6J-L) and Kv1.2 (data not shown), suggesting that heteromultimeric channels formed by the association of Kv1.1, Kv1.2 and Kvβ2 are present in the JXP-AIS. Of note, their expression levels in the JXP-AIS, in particular in its proximal domain adjacent to the para-AIS, always appeared higher than in the AIS (Figure 6C, G, K). In addition, the axon displays a significant widening in the JXP-AIS as compared to the AIS, which contributes to the higher expression level of Kv1.1, Kv1.2 and Kvβ2 observed in the JXP-AIS.

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