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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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Voltage-gated sodium (Nav) channel distribution in the axon initial segments (AISs) in motor neurons (MNs). Quadruple immunostaining of Peripherin (A, G), ankyrin G (AnkG) (B, H), Nav1.1 (C, I) and Nav1.6 (D, J) (Nav1.1 and Nav1.6 are merged in (E, K)) showing two populations of AISs: expressing Nav1.1 in a proximal compartment complementary to Nav1.6 expression (A-F) or expressing Nav1.6 alone throughout the AIS (G-L). (F, L) The mean immunofluorescence intensity profile (shown by the line) ± SEM from n = 6 AISs is shown for AnkG, Nav1.1 and Nav1.6. For each AIS and each antibody, immunofluorescence intensities were normalized relative both to its maximum intensity along the AIS and to the length of the AIS. The beginning and the end of the AnkG+ AIS in B and H are shown (also in F and L) by blue arrowheads. The bracket indicates the proximal AIS Nav1.1 expression domain. Scale bar = 5 μm.
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Figure 3: Voltage-gated sodium (Nav) channel distribution in the axon initial segments (AISs) in motor neurons (MNs). Quadruple immunostaining of Peripherin (A, G), ankyrin G (AnkG) (B, H), Nav1.1 (C, I) and Nav1.6 (D, J) (Nav1.1 and Nav1.6 are merged in (E, K)) showing two populations of AISs: expressing Nav1.1 in a proximal compartment complementary to Nav1.6 expression (A-F) or expressing Nav1.6 alone throughout the AIS (G-L). (F, L) The mean immunofluorescence intensity profile (shown by the line) ± SEM from n = 6 AISs is shown for AnkG, Nav1.1 and Nav1.6. For each AIS and each antibody, immunofluorescence intensities were normalized relative both to its maximum intensity along the AIS and to the length of the AIS. The beginning and the end of the AnkG+ AIS in B and H are shown (also in F and L) by blue arrowheads. The bracket indicates the proximal AIS Nav1.1 expression domain. Scale bar = 5 μm.

Mentions: In order to characterize the MN AIS excitability properties, we analyzed the AIS composition in terms of ion channels, and started with Nav channels. We investigated the expression of Nav1 channels in the AnkG+ AIS of Peripherin+ motor axons. We did not find any AIS expression of Nav1.2 in MNs (data not shown). We found that 79.8% of MNs (from n = 35) expressed both Nav1.1 and Nav1.6 in a rather complementary fashion, with Nav1.1 expressed in the proximal part of the AIS, close to the soma, and Nav1.6 found more strongly expressed towards the distal AIS (Figure 3A-F). Analysis of Nav1.1 and Nav1.6 immunofluorescence intensity profiles along the AIS, as compared to that of AnkG, confirmed these two complementary distributions: intensity of Nav1.1 decreased when that of Nav1.6 increased (Figure 3F). In the remaining 20.2% of MNs, Nav1.1 was not expressed at the AIS and Nav1.6 was expressed along the entire AIS (Figure 3G-L), with an immunofluorescence intensity profile displaying a slightly lower level in the proximal AIS (Figure 3L).


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)

Voltage-gated sodium (Nav) channel distribution in the axon initial segments (AISs) in motor neurons (MNs). Quadruple immunostaining of Peripherin (A, G), ankyrin G (AnkG) (B, H), Nav1.1 (C, I) and Nav1.6 (D, J) (Nav1.1 and Nav1.6 are merged in (E, K)) showing two populations of AISs: expressing Nav1.1 in a proximal compartment complementary to Nav1.6 expression (A-F) or expressing Nav1.6 alone throughout the AIS (G-L). (F, L) The mean immunofluorescence intensity profile (shown by the line) ± SEM from n = 6 AISs is shown for AnkG, Nav1.1 and Nav1.6. For each AIS and each antibody, immunofluorescence intensities were normalized relative both to its maximum intensity along the AIS and to the length of the AIS. The beginning and the end of the AnkG+ AIS in B and H are shown (also in F and L) by blue arrowheads. The bracket indicates the proximal AIS Nav1.1 expression domain. Scale bar = 5 μm.
© Copyright Policy - open-access
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Figure 3: Voltage-gated sodium (Nav) channel distribution in the axon initial segments (AISs) in motor neurons (MNs). Quadruple immunostaining of Peripherin (A, G), ankyrin G (AnkG) (B, H), Nav1.1 (C, I) and Nav1.6 (D, J) (Nav1.1 and Nav1.6 are merged in (E, K)) showing two populations of AISs: expressing Nav1.1 in a proximal compartment complementary to Nav1.6 expression (A-F) or expressing Nav1.6 alone throughout the AIS (G-L). (F, L) The mean immunofluorescence intensity profile (shown by the line) ± SEM from n = 6 AISs is shown for AnkG, Nav1.1 and Nav1.6. For each AIS and each antibody, immunofluorescence intensities were normalized relative both to its maximum intensity along the AIS and to the length of the AIS. The beginning and the end of the AnkG+ AIS in B and H are shown (also in F and L) by blue arrowheads. The bracket indicates the proximal AIS Nav1.1 expression domain. Scale bar = 5 μm.
Mentions: In order to characterize the MN AIS excitability properties, we analyzed the AIS composition in terms of ion channels, and started with Nav channels. We investigated the expression of Nav1 channels in the AnkG+ AIS of Peripherin+ motor axons. We did not find any AIS expression of Nav1.2 in MNs (data not shown). We found that 79.8% of MNs (from n = 35) expressed both Nav1.1 and Nav1.6 in a rather complementary fashion, with Nav1.1 expressed in the proximal part of the AIS, close to the soma, and Nav1.6 found more strongly expressed towards the distal AIS (Figure 3A-F). Analysis of Nav1.1 and Nav1.6 immunofluorescence intensity profiles along the AIS, as compared to that of AnkG, confirmed these two complementary distributions: intensity of Nav1.1 decreased when that of Nav1.6 increased (Figure 3F). In the remaining 20.2% of MNs, Nav1.1 was not expressed at the AIS and Nav1.6 was expressed along the entire AIS (Figure 3G-L), with an immunofluorescence intensity profile displaying a slightly lower level in the proximal AIS (Figure 3L).

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