Introduction
The peripheral nerve is composed of myelinated and unmyelinated nerve fibers. Different fibers originate from different neurons like motor neurons in the ventral horn of the spinal cord, sensory neurons from dorsal root ganglia and autonomic neurons. Most forms of hereditary neuropathies (HN) affect the myelinated motor and/or sensory neurons. Autonomic dys- function is seen in some special subforms of hereditary neuropathies like hereditary sensory and autonomic neuropathy (HSAN or HAN). Many genes encoding proteins which are located in the myelinated nerve fiber were identified as disease causing genes when mutated (reviewed [40]).
The function of some of these proteins has been elucidated over the last few years but the function of many of these genes is not understood yet. In the following, the focus is laid on the proteins for which the biological function has been shown in appropriate experiments.
The function of the peripheral nervous system (PNS) is to connect the central nervous system with the surrounding environment of the organism.
For this purpose the normal function of the PNS is fundamentally depen- dent on the correct morphological and molecular organization of the pe- ripheral nerve fiber.
1.1 Cellular components of the PNS
The two major cellular components of peripheral nerves are (1) axons orig- inating either from motor neurons located within the brainstem motor nuclei, from the ventral horn of the spinal cord or from sensory dorsal root ganglia and (2) glial cells, in the PNS represented by Schwann cells. In the PNS, two different kinds of Schwann cells can be found, unmyelinating and myelinat- ing cells. Unmyelinating Schwann cells are responsible for the correct en- sheathing of multiple axons which are smaller than 1 lm in diameter while myelinating Schwann cells ensheath single axons with a diameter of more than 1 lm with myelin. Myelinating Schwann cells align to a discrete part
P. Young, M. Boentert
of an axon in a 1:1 relation [12, 27]. The process of myelination is character- ized by the formation of a defined number of wraps of compacted cell mem- brane of a single Schwann cell along the discrete segment of the axon [31].
The nucleus of the myelinating Schwann cell is located finally outside the myelin sheath and a small collar of cytoplasm persists at this outer side of the myelin compartment which is defined as the abaxonal compartment while the adaxonal compartment of the myelin sheath is defined as the small residual rim of cytoplasm of the Schwann cell at the innermost myelin wrap adjacent to the axon (Fig. 1.1). The abaxonal and the adaxonal compartment are linked via cytoplasmic channels called Schmidt-Lanterman incisures which enable traffic of substances between the inner and outer compartments of the Schwann cell. The abaxonal compartment is characterized by the pres- ence of extracellular matrix receptors (ECM) [22]. The adaxonal compart- ment is characterized by the presence of molecules which mediate cell adhe- sion like the myelin associated glycoprotein (MAG) [35].
Besides Schwann cells and axons, fibroblasts are found in the PNS. Some immune cells are also found in the normal healthy nerve. The impact of immune cells and fibroblasts on inflammation, trauma, hereditary periph- eral nerve diseases and axonal degeneration is not fully elucidated.
Fig. 1.1. Cross section of a single myelinated nerve fiber. A single Schwann cell is ensheathing a single axon in a 1:1 relation. The nucleus of the Schwann cell (SCN) is in close contact to the axon while the whole axon (A) is surrounded bycompacted myelin (M). Arrow heads indi- cate the adaxonal compartment of the Schwann cell cytoplasm while the abaxonal compart- ment is indicated bySCC
1.2 Architecture of the myelin compartment
Myelin is a highly specified material which is required for insulation of the axon against its surroundings and myelin enables saltatory nerve conduc- tion along the axon. Besides insulating stretches along the axon, saltatory nerve conduction depends on gaps within the compacted myelin in which ion exchange is possible to maintain the electrical conduction. These gaps are called nodes of Ranvier. The node of Ranvier is characterized by a complex architecture comprising several proteins. Disruption of the com- pacted myelin sheath is further regularly seen in regions called Schmidt- Lanterman incisures consisting of uncompacted myelin. The myelin seg- ments which extend between two nodes of Ranvier are called internodal re- gions of the peripheral myelinated nerve fiber (Fig. 1.2).
Fig. 1.2. Morphologyand molecular architecture of the nodal, paranodal and juxtaparanodal region in myelinated nerve fibers. Longitudinal section of a myelinated nerve fiber showing a node of Ranvier. A and B The nodal region (N) of the axon (A) is flanked bythe paranodal re- gion (P). The flanking region is defined as the juxtaparanodal region (J). B Caspr is localized in the paranodal region of the axon while the potassium channel Kv1.2 is located in the juxtaparanodal region of the axon. Caspr and Kv1.2 are both expressed in the outer mesaxon (indicated byarrows)
1.2.1 The internode
The length of an internode depends on the axon diameter and is about 100 times the axon diameter. Internodes of 1 mm length are found in large fi- bers [13]. The internodes, making up most of the length of the myelinated nerve fibers, contain mainly compacted myelin lamellae. The compacted myelin sheath is formed by fusion of adjacent Schwann cell membranes re- sulting in a highly specific pattern of dark and light lines. Each period is separated into the dark major dense line and a bright line which is sepa- rated by a dark line called the intermediate line. The width of each period is strictly determined and is between 12 nm (in fixed tissue) up to a maxi- mum of 19 nm (in unfixed tissue). The number of periods is strictly re- lated to the axon diameter. Little is known about the mechanisms which lead to the specific axon diameter-dependent thickness of the myelin sheath. Axonal expression of neuregulins and glial expression of ERB-B re- ceptor 2 were shown to have an important impact on the thickness of the myelin sheath [20]. The compacted myelin compartment consists mainly of cholesterol and sphingolipids. Further some specialized lipids like galacto- cerebrosides and sulfatides are found. Proteins are a small fraction of the compacted myelin. The myelin protein zero (P0, encoded by the MPZ gene), the peripheral myelin protein 22 (PMP22), and myelin basic protein (MBP) represent the major proteins found in the compacted myelin com- partment of the PNS.
P0 is the most abundant protein in the compacted myelin of the PNS.
Its main function is to mediate and enable myelin compaction. It has been shown that P0 consists of a single immunoglobulin-like motif in its extra- cellular domain and has a highly positively charged intracellular domain [17]. This combination is postulated to be a prerequisite for myelin com- paction. With the aid of the extracellular domains homophilically interact- ing tetramers can be formed within the same membrane (cis position) and with the apposing membrane (trans position) [18, 28]. Functional studies in P0 deficient cells and mice underlined the function of P0 in compaction [7, 10]. Thus, it is postulated that major dense line compaction is mediated by P0.
PMP22 is a small tetraspan intrinsic membrane protein which has a ma- jor impact on myelination and myelin maintenance as well as Schwann cell proliferation. PMP22 function is regulated by many factors. The function of PMP22 is highly dosage dependent but the basis of this dosage depen- dency is far from being fully understood (reviewed in [33]).
MBP is a minor protein component of the compacted myelin sheath [32]. Although it is found to be expressed within the major dense line its function is still unclear. In contrast to P0 and PMP22, MBP is also ex- pressed in the myelin compartment of the central nervous system (CNS) [36]. Interactions between MBP and PMP22 or P0, respectively, have not been shown so far. The loss of MBP immunoreactivity is a reliable marker for demyelination in the PNS as well as in the CNS.
The periaxonal space between the myelin sheath and the axon is sealed by the inner mesaxon. The inner mesaxon runs along the whole internode.
The outer sealing of the myelin sheath, called the outer mesaxon, is achieved by the adhesion of two membrane loops of the Schwann cell forming two lips filled with cytoplasm and sealed to each other by adhe- rens junctions. Within the outer mesaxon, E-cadherin and beta-catenin are specifically localized (Fig. 1.3). E-cadherin deficiency causes a widening of the outer mesaxon in mice but has no impact on the compacted myelin formation in these mice. At the axon in the regions apposing the inner mesaxon, which is the adaxonal membrane part of the ensheathing Schwann cell, contactin±associated protein 1 (caspr1), contactin and Kv1.1 and Kv1.2 are localized [1, 26] (Fig. 1.2).
The compacted myelin compartment of the internode is regularly inter- rupted by uncompacted myelin bridges called Schmidt-Lanterman inci-
Fig. 1.3. Immunohistochemical staining with an antibody against E-cadherin on a myelinated nerve fiber. E-cadher- in is expressed in the myelated nerve fiber in the uncom- pacted myelin compartment at the paranodal region of the node of Ranvier (arrow), the Schmidt-Lanterman inci- sures (asterisks) and the outer mesaxon (arrow heads)
sures. These incisures show an accumulation of potassium channels, E-cad- herin, beta-catenin, caspr and several tight junction markers. The function of the incisures is yet not fully understood but the expression of gap junc- tion forming gap junction protein beta 1 (GJB1) in these structures sug- gests that they have an impact on the diffusion of several molecules since it was shown that radial dye radial transfer is mediated via diffusion across incisures [2].
1.2.2 The node of Ranvier
The main proteins which are found accumulated axonally at the node of Ranvier are voltage-gated sodium channels belonging to a multigene fami- ly. However, the sodium channel Nav1.6 [6] is the main representative.
Further sodium channels at the node of Ranvier are Nav1.2, Nav1.8 and Nav1.9 [4, 11, 15]. Sodium channels are anchored by two different splice variants of ankyrinG [16]. Furthermore spectrin is accumulated at the node.
In contrast to the nodes of Ranvier found in the CNS, the nodal region in the PNS is covered by interdigitating microvilli originating from the lat- eral end of myelinating Schwann cells. The microvilli are in close contact with the axonal cytoskeleton [14]. The diameter of the axon itself is re- duced at the nodal region. Microvilli contain F-actin and proteins like ez- rin, radixin and moesin which all belong to the family of F-actin binding ERM proteins (ERM stands for ezrin, radixin and moesin). ERM proteins can bind to merlin, the gene product of the neurofibromatosis 2 gene.
1.2.3 The paranodal region
The paranodal region is formed by uncompacted myelin loops formed out of the lateral edge of the myelin sheath. In thin fibers, each loop reaches the axon and forms close contacts to the axon. In large fibers not all loops reach the axon. Contacts between the paranodal loops and the axon are formed by septate-like junctions. Septate-like junctions contain contactin while the apposing axonal segment contains contactin associated protein (caspr) [8, 19, 25]. NF155, an isoform of neurofascin, is also expressed in the paranodal loops. Contactin and caspr heterodimers colocalize with NF155 within the paranodal region [34]. The paranodal region is also char- acterized by an accumulation of molecules which are involved in mediating adherens structures between the paranodal loops. Contactin and caspr are essential for the normal and stable architecture of the paranodal region as it was shown in mice deficient for these proteins [3, 5]. In these animals the spacing between the loops and the axon is enlarged and microvillar processes invade the periaxonal space and disturb the formation of the paranodal loops. Furthermore, these animals show a disturbed accumula-
tion of potassium channels in the juxtaparanodal region which is the re- gion adjacent to the paranodal region, providing evidence that the correct formation of septate like junctions is the basis for correct potassium chan- nel assembly during myelination [3, 5].
A further group of molecules expressed in the paranodal region are members of the cadherin/catenin complex. They are expressed between paranodal loops of the same Schwann cell and are known to be involved in the establishment of adherens junctions [9, 29]. E-cadherin and beta-cate- nin are colocalized in the paranodal loops [9]. E-cadherin deficiency does not result in disturbance of the formation of paranodal loops [39]. Thus, the function of the cadherin/catenin complex is not understood so far.
Other proteins expressed in the paranodal region are claudin and PAR3 which are associated with the formation of tight junctions [21].
1.2.4 The juxtaparanodal region
The juxtaparanodal region of the myelinated nerve fiber is specified by the accumulation of delayed rectifying potassium channels as Kv1.1 and Kv1.2 [23, 24, 37, 38]. These channels form tetramers and are located in the axon.
The distribution of these channels is tightly dependent on the correct lo- calization of paranodally expressed caspr1. Caspr2, a homologue to caspr1, is localized in the juxtaparanodal compartment. Caspr2 and Kv1.1 and Kv1.2 may be linked to each other by a PDZ domain. Functionally potas- sium channels at the juxtaparanodal region are necessary for normal im- pulse generation in the axon since the deficiency for Kv1.1 in mice showed abnormal impulse generation near the neuromuscular junctions [30, 41].
1.3 Unmyelinated nerve fibers
Unmyelinated nerve fibers have a diameter between 0.2 and 3 lm. In con- trast to the myelinated fiber bundles unmyelinated fiber bundles are ac- companied by a single Schwann cell. Unmyelinated fibers lack the above described organization of proteins found in myelinated fibers. Unmyeli- nated fibers are packed into bundles by unmyelinating Schwann cells. Ac- cumulation of ion channels is not observed and proteins which take part in fiber bundling are not well described so far.
References
1. Arroyo EJ, Xu YT, Zhou L, Messing A, Peles E, Chiu SY, Scherer SS (1999) Myeli- nating Schwann cells determine the internodal localization of Kv1.1, Kv1.2, Kvbe- ta2, and Caspr. J Neurocytol 28:333±347
2. Balice-Gordon RJ, Bone LJ, Scherer SS (1998) Functional gap junctions in the schwann cell myelin sheath. J Cell Biol 142:1095±1104
3. Bhat MA, Rios JC, Lu Y, Garcia-Fresco GP, Ching W, St Martin M, Li J, Einheber S, Chesler M, Rosenbluth J, Salzer JL, Bellen HJ (2001) Axon-glia interactions and the domain organization of myelinated axons requires neurexin IV/Caspr/Parano- din. Neuron 30:369±383
4. Boiko T, Rasband MN, Levinson SR, Caldwell JH, Mandel G, Trimmer JS, Mat- thews G (2001) Compact myelin dictates the differential targeting of two sodium channel isoforms in the same axon. Neuron 30:91±104
5. Boyle ME, Berglund EO, Murai KK, Weber L, Peles E, Ranscht B (2001) Contactin orchestrates assembly of the septate-like junctions at the paranode in myelinated peripheral nerve. Neuron 30:385±397
6. Caldwell JH, Schaller KL, Lasher RS, Peles E, Levinson SR (2000) Sodium channel Na(v)1.6 is localized at nodes of ranvier, dendrites, and synapses. Proc Natl Acad Sci USA 97:5616±5620
7. D'Urso D, Brophy PJ, Staugaitis SM, Gillespie CS, Frey AB, Stempak JG, Colman DR (1990) Protein zero of peripheral nerve myelin: biosynthesis, membrane inser- tion, and evidence for homotypic interaction. Neuron 4:449±460
8. Einheber S, Zanazzi G, Ching W, Scherer S, Milner TA, Peles E, Salzer JL (1997) The axonal membrane protein Caspr, a homologue of neurexin IV, is a component of the septate-like paranodal junctions that assemble during myelination. J Cell Biol 139:1495±1506
9. Fannon AM, Sherman DL, Ilyina-Gragerova G, Brophy PJ, Friedrich VL, Jr, Col- man DR (1995) Novel E-cadherin-mediated adhesion in peripheral nerve:
Schwann cell architecture is stabilized by autotypic adherens junctions. J Cell Biol 129:189±202
10. Filbin MT, Walsh FS, Trapp BD, Pizzey JA, Tennekoon GI (1990) Role of myelin P0 protein as a homophilic adhesion molecule. Nature 344:871±872
11. Fjell J, Hjelmstrom P, Hormuzdiar W, Milenkovic M, Aglieco F, Tyrrell L, Dib-Hajj S, Waxman SG, Black JA (2000) Localization of the tetrodotoxin-resistant sodium channel NaN in nociceptors. Neuroreport 11:199±202
12. Friede RL, Samorajski T (1968) Myelin formation in the sciatic nerve of the rat. A quantitative electron microscopic, histochemical and radioautographic study. J Neuropathol Exp Neurol 27:546±570
13. Hess A, Young JZ (1952) The nodes of Ranvier. Proc R Soc Lond B Biol Sci 140:301±320
14. Ichimura T, Ellisman MH (1991) Three-dimensional fine structure of cytoskeletal- membrane interactions at nodes of Ranvier. J Neurocytol 20:667±681
15. Kaplan MR, Cho MH, Ullian EM, Isom LL, Levinson SR, Barres BA (2001) Differ- ential control of clustering of the sodium channels Na(v)1.2 and Na(v)1.6 at de- veloping CNS nodes of Ranvier. Neuron 30:105±119
16. Kordeli E, Lambert S, Bennett V (1995) AnkyrinG. A new ankyrin gene with neu- ral-specific isoforms localized at the axonal initial segment and node of Ranvier. J Biol Chem 270:2352±2359
17. Lemke G, Axel R (1985) Isolation and sequence of a cDNA encoding the major structural protein of peripheral myelin. Cell 40:501±508
18. Martini R, Mohajeri MH, Kasper S, Giese KP, Schachner M (1995) Mice doubly deficient in the genes for P0 and myelin basic protein show that both proteins contribute to the formation of the major dense line in peripheral nerve myelin. J Neurosci 15:4488±4495
19. Menegoz M, Gaspar P, Le Bert M, Galvez T, Burgaya F, Palfrey C, Ezan P, Arnos F, Girault JA (1997) Paranodin, a glycoprotein of neuronal paranodal membranes.
Neuron 19:319±331
20. Michailov GV, Sereda MW, Brinkmann BG, Fischer TM, Haug B, Birchmeier C, Role L, Lai C, Schwab MH, Nave KA (2004) Axonal neuregulin-1 regulates myelin sheath thickness. Science 304:700±703
21. Poliak S, Matlis S, Ullmer C, Scherer SS, Peles E (2002) Distinct claudins and as- sociated PDZ proteins form different autotypic tight junctions in myelinating Schwann cells. J Cell Biol 159:361±372
22. Previtali SC, Feltri ML, Archelos JJ, Quattrini A, Wrabetz L, Hartung H (2001) Role of integrins in the peripheral nervous system. Prog Neurobiol 64:35±49 23. Rasband MN, Trimmer JS, Peles E, Levinson SR, Shrager P (1999) K+ channel
distribution and clustering in developing and hypomyelinated axons of the optic nerve. J Neurocytol 28:319±331
24. Rasband MN, Trimmer JS, Schwarz TL, Levinson SR, Ellisman MH, Schachner M, Shrager P (1998) Potassium channel distribution, clustering, and function in re- myelinating rat axons. J Neurosci 18:36±47
25. Rios JC, Melendez-Vasquez CV, Einheber S, Lustig M, Grumet M, Hemperly J, Peles E, Salzer JL (2000) Contactin-associated protein (Caspr) and contactin form a complex that is targeted to the paranodal junctions during myelination. J Neu- rosci 20:8354±8364
26. Rios JC, Melendez-Vasquez CV, Einheber S, Lustig M, Grumet M, Hemperly J, Peles E, Salzer JL (2000) Contactin-associated protein (Caspr) and contactin form a complex that is targeted to the paranodal junctions during myelination. J Neu- rosci 20:8354±8364
27. Samorajski T, Friede RL (1968) A quantitative electron microscopic study of mye- lination in the pyramidal tract of rat. J Comp Neurol 134:323±338
28. Shapiro L, Doyle JP, Hensley P, Colman DR, Hendrickson WA (1996) Crystal structure of the extracellular domain from P0, the major structural protein of pe- ripheral nerve myelin. Neuron 17:435±449
29. Shapiro L, Fannon AM, Kwong PD, Thompson A, Lehmann MS, Grubel G, Le- grand JF, Als-Nielsen J, Colman DR, Hendrickson WA (1995) Structural basis of cell-cell adhesion by cadherins. Nature 374:327±337
30. Smart SL, Lopantsev V, Zhang CL, Robbins CA, Wang H, Chiu SY, Schwartzkroin PA, Messing A, Tempel BL (1998) Deletion of the K(V)1.1 potassium channel causes epilepsy in mice. Neuron 20:809±819
31. Smith KJ, Blakemore WF, Murray JA, Patterson RC (1982) Internodal myelin vol- ume and axon surface area. A relationship determining myelin thickness? J Neurol Sci 55:231±246
32. Streicher R, Stoffel W (1989) The organization of the human myelin basic protein gene. Comparison with the mouse gene. Biol Chem Hoppe Seyler 370:503±510 33. Suter U, Scherer SS (2003) Disease mechanisms in inherited neuropathies. Nat
Rev Neurosci 4:714±726
34. Tait S, Gunn-Moore F, Collinson JM, Huang J, Lubetzki C, Pedraza L, Sherman DL, Colman DR, Brophy PJ (2000) An oligodendrocyte cell adhesion molecule at the site of assembly of the paranodal axo-glial junction. J Cell Biol 150:657±666 35. Trapp BD (1990) Myelin-associated glycoprotein. Location and potential functions.
Ann NY Acad Sci 605:29±43
36. Trapp BD, Hauer P, Lemke G (1988) Axonal regulation of myelin protein mRNA levels in actively myelinating Schwann cells. J Neurosci 8:3515±3521
37. Vabnick I, Shrager P (1998) Ion channel redistribution and function during devel- opment of the myelinated axon. J Neurobiol 37:80±96
38. Wang H, Kunkel DD, Martin TM, Schwartzkroin PA, Tempel BL (1993) Heteromul- timeric K+ channels in terminal and juxtaparanodal regions of neurons. Nature 365:75±79
39. Young P, Boussadia O, Berger P, Leone DP, Charnay P, Kemler R, Suter U (2002) E-cadherin is required for the correct formation of autotypic adherens junctions of the outer mesaxon but not for the integrity of myelinated fibers of peripheral nerves. Mol Cell Neurosci 21:341±351
40. Young P, Suter U (2003) The causes of Charcot-Marie-Tooth disease. Cell Mol Life Sci 60:2547±2560
41. Zhou L, Zhang CL, Messing A, Chiu SY (1998) Temperature-sensitive neuromus- cular transmission in Kv1.1 null mice: role of potassium channels under the mye- lin sheath in young nerves. J Neurosci 18:7200±7215
