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Functional gap junctions in the schwann cell myelin sheath.

Balice-Gordon RJ, Bone LJ, Scherer SS - J. Cell Biol. (1998)

Bottom Line: Gap junctions are localized to periodic interruptions in the compact myelin called Schmidt-Lanterman incisures and to paranodes; these regions contain at least one gap junction protein, connexin32 (Cx32).The radial diffusion of low molecular weight dyes across the myelin sheath was not interrupted in myelinating Schwann cells from cx32- mice, indicating that other connexins participate in forming gap junctions in these cells.Owing to the unique geometry of myelinating Schwann cells, a gap junction-mediated radial pathway may be essential for rapid diffusion between the adaxonal and perinuclear cytoplasm, since this radial pathway is approximately one million times faster than the circumferential pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, The University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6074, USA. rbaliceg@mail.med.upenn.edu

ABSTRACT
The Schwann cell myelin sheath is a multilamellar structure with distinct structural domains in which different proteins are localized. Intracellular dye injection and video microscopy were used to show that functional gap junctions are present within the myelin sheath that allow small molecules to diffuse between the adaxonal and perinuclear Schwann cell cytoplasm. Gap junctions are localized to periodic interruptions in the compact myelin called Schmidt-Lanterman incisures and to paranodes; these regions contain at least one gap junction protein, connexin32 (Cx32). The radial diffusion of low molecular weight dyes across the myelin sheath was not interrupted in myelinating Schwann cells from cx32- mice, indicating that other connexins participate in forming gap junctions in these cells. Owing to the unique geometry of myelinating Schwann cells, a gap junction-mediated radial pathway may be essential for rapid diffusion between the adaxonal and perinuclear cytoplasm, since this radial pathway is approximately one million times faster than the circumferential pathway.

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Rapid diffusion of 5,6-carboxyfluorescein across incisures to the inner/adaxonal collar of Schwann cell cytoplasm.  Right panels, from the same fiber at the onset (0 s), and 15, 30, 80,  and 147 s after the onset of iontophoretic injection. The locations  of an incisure are indicated with an arrowhead and were confirmed by viewing the fiber with polarized light. At the onset of  injection, dye immediately fills the outer collar of Schwann cell  cytoplasm. By 15 s, an incisure is labeled and a faint train track is  apparent on one side of the myelin sheath. The train track pattern and incisure are more apparent at 30 s. At longer times (80  and 147 s), more incisures become filled and the train track pattern is seen further away from the injection site, but in this case is  only clearly visualized on the left side of the fiber. Left panels,  quantitative analysis of pattern of dye distribution. The intensity  of pixels in a line perpendicular to the long axis of the fiber was  mapped at the position indicated by the black arrowheads in the  corresponding photomicrograph. The light microscopic images of  dye diffusion were obtained using manual gain settings of the SIT  camera so that changes in the line histogram intensity mapped  across the same region of the fiber over time could be compared  in terms of absolute intensity (right panels; 0–255 intensity levels).  A doublet is apparent in the left side of the histogram by 30 s after  the onset of injection (black arrow; corresponding location in the  30-s image is indicated by white arrow). Within the doublet, the  intensity of the first peak in the doublet representing the outside  collar of cytoplasm and the intensity of the second peak increased  over the first minutes of injection in parallel. Thus, the train track  pattern of labeling is consistent with the diffusion of 5,6-carboxyfluorescein from the outer/abaxonal to the inner/adaxonal cytoplasm across incisures. Bars, 5 μm.
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Figure 4: Rapid diffusion of 5,6-carboxyfluorescein across incisures to the inner/adaxonal collar of Schwann cell cytoplasm. Right panels, from the same fiber at the onset (0 s), and 15, 30, 80, and 147 s after the onset of iontophoretic injection. The locations of an incisure are indicated with an arrowhead and were confirmed by viewing the fiber with polarized light. At the onset of injection, dye immediately fills the outer collar of Schwann cell cytoplasm. By 15 s, an incisure is labeled and a faint train track is apparent on one side of the myelin sheath. The train track pattern and incisure are more apparent at 30 s. At longer times (80 and 147 s), more incisures become filled and the train track pattern is seen further away from the injection site, but in this case is only clearly visualized on the left side of the fiber. Left panels, quantitative analysis of pattern of dye distribution. The intensity of pixels in a line perpendicular to the long axis of the fiber was mapped at the position indicated by the black arrowheads in the corresponding photomicrograph. The light microscopic images of dye diffusion were obtained using manual gain settings of the SIT camera so that changes in the line histogram intensity mapped across the same region of the fiber over time could be compared in terms of absolute intensity (right panels; 0–255 intensity levels). A doublet is apparent in the left side of the histogram by 30 s after the onset of injection (black arrow; corresponding location in the 30-s image is indicated by white arrow). Within the doublet, the intensity of the first peak in the doublet representing the outside collar of cytoplasm and the intensity of the second peak increased over the first minutes of injection in parallel. Thus, the train track pattern of labeling is consistent with the diffusion of 5,6-carboxyfluorescein from the outer/abaxonal to the inner/adaxonal cytoplasm across incisures. Bars, 5 μm.

Mentions: To determine directly whether the double train track pattern of dye diffusion resulted from dye passing from the outer/abaxonal collar of Schwann cell cytoplasm to the inner/adaxonal collar of cytoplasm, the path of dye diffusion was monitored continuously during injection. Selected planes from one such sequence are shown in Fig. 4. From the onset of injection, dye spread rapidly along the myelin sheath, initially remaining within the outer/abaxonal collar near the nucleus (Fig. 4; 0 s). In most fibers, the dye appeared to collect at nearby incisures (Fig. 4; 15 s, arrowhead), partially filling the inner/adaxonal collar of cytoplasm immediately and more completely within a few seconds (Fig. 4; 30 s, arrow). The dye continued to fill these compartments, with each line becoming progressively brighter in intensity (Fig. 4; 80 and 147 s), and also spread longitudinally, usually reaching the nodes of Ranvier within a few minutes. Injections that were monitored with continuous acquisition onto videotape confirmed the above sequence of events, and, in addition, showed that the incisures themselves were labeled as the dye front moved down the myelin sheath. In some fibers, including the one illustrated in Fig. 4, light microscopic images of dye diffusion were obtained using manual gain settings of the SIT camera, allowing quantitative comparisons of changes in intensity mapped across the same region of the fiber over time (Fig. 4; right-hand panels). This analysis confirmed that the intensity of the outer and inner cytoplasmic collars increased over time, and that the inner cytoplasmic collar filled after the outer one. Thus, the temporal sequence of the train track pattern indicates that 5,6-carboxyfluorescein diffuses from the outer/abaxonal to the inner/ adaxonal cytoplasm across incisures.


Functional gap junctions in the schwann cell myelin sheath.

Balice-Gordon RJ, Bone LJ, Scherer SS - J. Cell Biol. (1998)

Rapid diffusion of 5,6-carboxyfluorescein across incisures to the inner/adaxonal collar of Schwann cell cytoplasm.  Right panels, from the same fiber at the onset (0 s), and 15, 30, 80,  and 147 s after the onset of iontophoretic injection. The locations  of an incisure are indicated with an arrowhead and were confirmed by viewing the fiber with polarized light. At the onset of  injection, dye immediately fills the outer collar of Schwann cell  cytoplasm. By 15 s, an incisure is labeled and a faint train track is  apparent on one side of the myelin sheath. The train track pattern and incisure are more apparent at 30 s. At longer times (80  and 147 s), more incisures become filled and the train track pattern is seen further away from the injection site, but in this case is  only clearly visualized on the left side of the fiber. Left panels,  quantitative analysis of pattern of dye distribution. The intensity  of pixels in a line perpendicular to the long axis of the fiber was  mapped at the position indicated by the black arrowheads in the  corresponding photomicrograph. The light microscopic images of  dye diffusion were obtained using manual gain settings of the SIT  camera so that changes in the line histogram intensity mapped  across the same region of the fiber over time could be compared  in terms of absolute intensity (right panels; 0–255 intensity levels).  A doublet is apparent in the left side of the histogram by 30 s after  the onset of injection (black arrow; corresponding location in the  30-s image is indicated by white arrow). Within the doublet, the  intensity of the first peak in the doublet representing the outside  collar of cytoplasm and the intensity of the second peak increased  over the first minutes of injection in parallel. Thus, the train track  pattern of labeling is consistent with the diffusion of 5,6-carboxyfluorescein from the outer/abaxonal to the inner/adaxonal cytoplasm across incisures. Bars, 5 μm.
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Figure 4: Rapid diffusion of 5,6-carboxyfluorescein across incisures to the inner/adaxonal collar of Schwann cell cytoplasm. Right panels, from the same fiber at the onset (0 s), and 15, 30, 80, and 147 s after the onset of iontophoretic injection. The locations of an incisure are indicated with an arrowhead and were confirmed by viewing the fiber with polarized light. At the onset of injection, dye immediately fills the outer collar of Schwann cell cytoplasm. By 15 s, an incisure is labeled and a faint train track is apparent on one side of the myelin sheath. The train track pattern and incisure are more apparent at 30 s. At longer times (80 and 147 s), more incisures become filled and the train track pattern is seen further away from the injection site, but in this case is only clearly visualized on the left side of the fiber. Left panels, quantitative analysis of pattern of dye distribution. The intensity of pixels in a line perpendicular to the long axis of the fiber was mapped at the position indicated by the black arrowheads in the corresponding photomicrograph. The light microscopic images of dye diffusion were obtained using manual gain settings of the SIT camera so that changes in the line histogram intensity mapped across the same region of the fiber over time could be compared in terms of absolute intensity (right panels; 0–255 intensity levels). A doublet is apparent in the left side of the histogram by 30 s after the onset of injection (black arrow; corresponding location in the 30-s image is indicated by white arrow). Within the doublet, the intensity of the first peak in the doublet representing the outside collar of cytoplasm and the intensity of the second peak increased over the first minutes of injection in parallel. Thus, the train track pattern of labeling is consistent with the diffusion of 5,6-carboxyfluorescein from the outer/abaxonal to the inner/adaxonal cytoplasm across incisures. Bars, 5 μm.
Mentions: To determine directly whether the double train track pattern of dye diffusion resulted from dye passing from the outer/abaxonal collar of Schwann cell cytoplasm to the inner/adaxonal collar of cytoplasm, the path of dye diffusion was monitored continuously during injection. Selected planes from one such sequence are shown in Fig. 4. From the onset of injection, dye spread rapidly along the myelin sheath, initially remaining within the outer/abaxonal collar near the nucleus (Fig. 4; 0 s). In most fibers, the dye appeared to collect at nearby incisures (Fig. 4; 15 s, arrowhead), partially filling the inner/adaxonal collar of cytoplasm immediately and more completely within a few seconds (Fig. 4; 30 s, arrow). The dye continued to fill these compartments, with each line becoming progressively brighter in intensity (Fig. 4; 80 and 147 s), and also spread longitudinally, usually reaching the nodes of Ranvier within a few minutes. Injections that were monitored with continuous acquisition onto videotape confirmed the above sequence of events, and, in addition, showed that the incisures themselves were labeled as the dye front moved down the myelin sheath. In some fibers, including the one illustrated in Fig. 4, light microscopic images of dye diffusion were obtained using manual gain settings of the SIT camera, allowing quantitative comparisons of changes in intensity mapped across the same region of the fiber over time (Fig. 4; right-hand panels). This analysis confirmed that the intensity of the outer and inner cytoplasmic collars increased over time, and that the inner cytoplasmic collar filled after the outer one. Thus, the temporal sequence of the train track pattern indicates that 5,6-carboxyfluorescein diffuses from the outer/abaxonal to the inner/ adaxonal cytoplasm across incisures.

Bottom Line: Gap junctions are localized to periodic interruptions in the compact myelin called Schmidt-Lanterman incisures and to paranodes; these regions contain at least one gap junction protein, connexin32 (Cx32).The radial diffusion of low molecular weight dyes across the myelin sheath was not interrupted in myelinating Schwann cells from cx32- mice, indicating that other connexins participate in forming gap junctions in these cells.Owing to the unique geometry of myelinating Schwann cells, a gap junction-mediated radial pathway may be essential for rapid diffusion between the adaxonal and perinuclear cytoplasm, since this radial pathway is approximately one million times faster than the circumferential pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, The University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6074, USA. rbaliceg@mail.med.upenn.edu

ABSTRACT
The Schwann cell myelin sheath is a multilamellar structure with distinct structural domains in which different proteins are localized. Intracellular dye injection and video microscopy were used to show that functional gap junctions are present within the myelin sheath that allow small molecules to diffuse between the adaxonal and perinuclear Schwann cell cytoplasm. Gap junctions are localized to periodic interruptions in the compact myelin called Schmidt-Lanterman incisures and to paranodes; these regions contain at least one gap junction protein, connexin32 (Cx32). The radial diffusion of low molecular weight dyes across the myelin sheath was not interrupted in myelinating Schwann cells from cx32- mice, indicating that other connexins participate in forming gap junctions in these cells. Owing to the unique geometry of myelinating Schwann cells, a gap junction-mediated radial pathway may be essential for rapid diffusion between the adaxonal and perinuclear cytoplasm, since this radial pathway is approximately one million times faster than the circumferential pathway.

Show MeSH
Related in: MedlinePlus