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Localization of SSeCKS in unmyelinated primary sensory neurons.

Irmen CP, Siegel SM, Carr PA - J Brachial Plex Peripher Nerve Inj (2008)

Bottom Line: In capsaicin treated rats, SSeCKS immunoreactivity was essentially obliterated in the dorsal horn while in dorsal root ganglia quantitative analysis revealed a 43% reduction in the number of SSeCKS-labeled cells.This attenuation is concomitant with a decrease in fluoride-resistant acid phosphatase labeled fibers in the spinal cord dorsal horn and small neuronal somata in sensory ganglia.These results demonstrate that SSeCKS is primarily localized within a distinct subpopulation of small diameter, largely unmyelinated C-fiber primary sensory neurons putatively involved in nociception.

View Article: PubMed Central - HTML - PubMed

Affiliation: Dept. of Anatomy and Cell Biology, University of North Dakota, Grand Forks, ND 58202, USA. cirmen@medicine.nodak.edu

ABSTRACT

Background: SSeCKS (Src SupprEssed C Kinase Substrate) is a proposed protein kinase C substrate/A kinase anchoring protein (AKAP) that has recently been characterized in the rat peripheral nervous system. It has been shown that approximately 40% of small primary sensory neurons contain SSeCKS-immunoreactivity in a population largely separate from substance P (95.2%), calcitonin gene related peptide (95.3%), or fluoride resistant acid phosphatase (55.0%) labeled cells. In the spinal cord, it was found that SSeCKS-immunoreactive axon collaterals terminate in the dorsal third of lamina II outer in a region similar to that of unmyelinated C-, or small diameter myelinated Adelta-, fibers. However, the precise characterization of the anatomical profile of the primary sensory neurons containing SSeCKS remains to be determined. Here, immunohistochemical labeling at the light and ultrastructural level is used to clarify the myelination status of SSeCKS-containing sensory neuron axons and to further clarify the morphometric, and provide insight into the functional, classification of SSeCKS-IR sensory neurons.

Methods: Colocalization studies of SSeCKS with myelination markers, ultrastructural localization of SSeCKS labeling and ablation of largely unmyelinated sensory fibers by neonatal capsaicin administration were all used to establish whether SSeCKS containing sensory neurons represent a subpopulation of unmyelinated primary sensory C-fibers.

Results: Double labeling studies of SSeCKS with CNPase in the dorsal horn and Pzero in the periphery showed that SSeCKS immunoreactivity was observed predominantly in association with unmyelinated primary sensory fibers. At the ultrastructural level, SSeCKS immunoreactivity was most commonly associated with axonal membrane margins of unmyelinated fibers. In capsaicin treated rats, SSeCKS immunoreactivity was essentially obliterated in the dorsal horn while in dorsal root ganglia quantitative analysis revealed a 43% reduction in the number of SSeCKS-labeled cells. This attenuation is concomitant with a decrease in fluoride-resistant acid phosphatase labeled fibers in the spinal cord dorsal horn and small neuronal somata in sensory ganglia.

Conclusion: These results demonstrate that SSeCKS is primarily localized within a distinct subpopulation of small diameter, largely unmyelinated C-fiber primary sensory neurons putatively involved in nociception.

No MeSH data available.


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Electron micrographs of SSeCKS-IR in the lumbar spinal cord dorsal horn. (A) SSeCKS labeling in the dorsal horn of the spinal cord demonstrating the zone within lamina II outer in which the band of SSeCKS labeling (upper left of micrograph) is separated (dashed line) from the area of the dorsal horn lacking SSeCKS labeling (lower right of micrograph). (B) SSeCKS labeling of thinly myelinated fibers (arrows) located next to larger diameter, more heavily myelinated axons lacking SSeCKS labeling. (C) SSeCKS labeling of a bundle of unmyelinated fibers. (D) Enlargement of portion of (C) reveals labeling around the axonal plasma membrane (black arrow) of individual unmyelinated fibers, as well as labeling around vesicular structures (white arrow).
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Figure 2: Electron micrographs of SSeCKS-IR in the lumbar spinal cord dorsal horn. (A) SSeCKS labeling in the dorsal horn of the spinal cord demonstrating the zone within lamina II outer in which the band of SSeCKS labeling (upper left of micrograph) is separated (dashed line) from the area of the dorsal horn lacking SSeCKS labeling (lower right of micrograph). (B) SSeCKS labeling of thinly myelinated fibers (arrows) located next to larger diameter, more heavily myelinated axons lacking SSeCKS labeling. (C) SSeCKS labeling of a bundle of unmyelinated fibers. (D) Enlargement of portion of (C) reveals labeling around the axonal plasma membrane (black arrow) of individual unmyelinated fibers, as well as labeling around vesicular structures (white arrow).

Mentions: At the ultrastructural level, SSeCKS labeling was observed in the outer laminae of the lumbar dorsal horn (Fig. 2A). Specifically, SSeCKS-IR was associated with small diameter processes aggregated between larger diameter, myelinated axons. Occasionally, SSeCKS-IR could be seen within the cytoplasm of thinly myelinated axons (Fig. 2B). Comparison revealed these axons to be substantially smaller in diameter than those SSeCKS-negative axons with heavy myelination. Of the SSeCKS labeled cross-sectional profiles quantified, 15% appeared myelinated (1484 profiles counted). In those areas of the dorsal horn (ventral to lamina II) in which small diameter myelinated fibers were more abundant, myelinated SSeCKS labeled fibers were more commonly observed than non-myelinated SSeCKS labeled profiles. At the subcellular level, SSeCKS-IR was consistently localized to the plasma membrane and was frequently observed as a granular deposition throughout axonal cytoplasm (Fig. 2C, enlargement). Occasional labeling of membrane-associated vesicles was also detected. SSeCKS-IR was not found in association with any glial profiles but was found in endothelial cells as had been previously reported [12].


Localization of SSeCKS in unmyelinated primary sensory neurons.

Irmen CP, Siegel SM, Carr PA - J Brachial Plex Peripher Nerve Inj (2008)

Electron micrographs of SSeCKS-IR in the lumbar spinal cord dorsal horn. (A) SSeCKS labeling in the dorsal horn of the spinal cord demonstrating the zone within lamina II outer in which the band of SSeCKS labeling (upper left of micrograph) is separated (dashed line) from the area of the dorsal horn lacking SSeCKS labeling (lower right of micrograph). (B) SSeCKS labeling of thinly myelinated fibers (arrows) located next to larger diameter, more heavily myelinated axons lacking SSeCKS labeling. (C) SSeCKS labeling of a bundle of unmyelinated fibers. (D) Enlargement of portion of (C) reveals labeling around the axonal plasma membrane (black arrow) of individual unmyelinated fibers, as well as labeling around vesicular structures (white arrow).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2277419&req=5

Figure 2: Electron micrographs of SSeCKS-IR in the lumbar spinal cord dorsal horn. (A) SSeCKS labeling in the dorsal horn of the spinal cord demonstrating the zone within lamina II outer in which the band of SSeCKS labeling (upper left of micrograph) is separated (dashed line) from the area of the dorsal horn lacking SSeCKS labeling (lower right of micrograph). (B) SSeCKS labeling of thinly myelinated fibers (arrows) located next to larger diameter, more heavily myelinated axons lacking SSeCKS labeling. (C) SSeCKS labeling of a bundle of unmyelinated fibers. (D) Enlargement of portion of (C) reveals labeling around the axonal plasma membrane (black arrow) of individual unmyelinated fibers, as well as labeling around vesicular structures (white arrow).
Mentions: At the ultrastructural level, SSeCKS labeling was observed in the outer laminae of the lumbar dorsal horn (Fig. 2A). Specifically, SSeCKS-IR was associated with small diameter processes aggregated between larger diameter, myelinated axons. Occasionally, SSeCKS-IR could be seen within the cytoplasm of thinly myelinated axons (Fig. 2B). Comparison revealed these axons to be substantially smaller in diameter than those SSeCKS-negative axons with heavy myelination. Of the SSeCKS labeled cross-sectional profiles quantified, 15% appeared myelinated (1484 profiles counted). In those areas of the dorsal horn (ventral to lamina II) in which small diameter myelinated fibers were more abundant, myelinated SSeCKS labeled fibers were more commonly observed than non-myelinated SSeCKS labeled profiles. At the subcellular level, SSeCKS-IR was consistently localized to the plasma membrane and was frequently observed as a granular deposition throughout axonal cytoplasm (Fig. 2C, enlargement). Occasional labeling of membrane-associated vesicles was also detected. SSeCKS-IR was not found in association with any glial profiles but was found in endothelial cells as had been previously reported [12].

Bottom Line: In capsaicin treated rats, SSeCKS immunoreactivity was essentially obliterated in the dorsal horn while in dorsal root ganglia quantitative analysis revealed a 43% reduction in the number of SSeCKS-labeled cells.This attenuation is concomitant with a decrease in fluoride-resistant acid phosphatase labeled fibers in the spinal cord dorsal horn and small neuronal somata in sensory ganglia.These results demonstrate that SSeCKS is primarily localized within a distinct subpopulation of small diameter, largely unmyelinated C-fiber primary sensory neurons putatively involved in nociception.

View Article: PubMed Central - HTML - PubMed

Affiliation: Dept. of Anatomy and Cell Biology, University of North Dakota, Grand Forks, ND 58202, USA. cirmen@medicine.nodak.edu

ABSTRACT

Background: SSeCKS (Src SupprEssed C Kinase Substrate) is a proposed protein kinase C substrate/A kinase anchoring protein (AKAP) that has recently been characterized in the rat peripheral nervous system. It has been shown that approximately 40% of small primary sensory neurons contain SSeCKS-immunoreactivity in a population largely separate from substance P (95.2%), calcitonin gene related peptide (95.3%), or fluoride resistant acid phosphatase (55.0%) labeled cells. In the spinal cord, it was found that SSeCKS-immunoreactive axon collaterals terminate in the dorsal third of lamina II outer in a region similar to that of unmyelinated C-, or small diameter myelinated Adelta-, fibers. However, the precise characterization of the anatomical profile of the primary sensory neurons containing SSeCKS remains to be determined. Here, immunohistochemical labeling at the light and ultrastructural level is used to clarify the myelination status of SSeCKS-containing sensory neuron axons and to further clarify the morphometric, and provide insight into the functional, classification of SSeCKS-IR sensory neurons.

Methods: Colocalization studies of SSeCKS with myelination markers, ultrastructural localization of SSeCKS labeling and ablation of largely unmyelinated sensory fibers by neonatal capsaicin administration were all used to establish whether SSeCKS containing sensory neurons represent a subpopulation of unmyelinated primary sensory C-fibers.

Results: Double labeling studies of SSeCKS with CNPase in the dorsal horn and Pzero in the periphery showed that SSeCKS immunoreactivity was observed predominantly in association with unmyelinated primary sensory fibers. At the ultrastructural level, SSeCKS immunoreactivity was most commonly associated with axonal membrane margins of unmyelinated fibers. In capsaicin treated rats, SSeCKS immunoreactivity was essentially obliterated in the dorsal horn while in dorsal root ganglia quantitative analysis revealed a 43% reduction in the number of SSeCKS-labeled cells. This attenuation is concomitant with a decrease in fluoride-resistant acid phosphatase labeled fibers in the spinal cord dorsal horn and small neuronal somata in sensory ganglia.

Conclusion: These results demonstrate that SSeCKS is primarily localized within a distinct subpopulation of small diameter, largely unmyelinated C-fiber primary sensory neurons putatively involved in nociception.

No MeSH data available.


Related in: MedlinePlus