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Regulation of Wnt signaling by nociceptive input in animal models.

Shi Y, Yuan S, Li B, Wang J, Carlton SM, Chung K, Chung JM, Tang SJ - Mol Pain (2012)

Bottom Line: In addition, Wnt3a, a prototypic Wnt ligand that activates the canonical pathway, is also enriched in the superficial layers.Furthermore, Wnt5a, a prototypic Wnt ligand for non-canonical pathways, and its receptor Ror2 are also up-regulated in the SCDH of these models.Our results suggest that Wnt signaling pathways are regulated by nociceptive input.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA.

ABSTRACT

Background: Central sensitization-associated synaptic plasticity in the spinal cord dorsal horn (SCDH) critically contributes to the development of chronic pain, but understanding of the underlying molecular pathways is still incomplete. Emerging evidence suggests that Wnt signaling plays a crucial role in regulation of synaptic plasticity. Little is known about the potential function of the Wnt signaling cascades in chronic pain development.

Results: Fluorescent immunostaining results indicate that β-catenin, an essential protein in the canonical Wnt signaling pathway, is expressed in the superficial layers of the mouse SCDH with enrichment at synapses in lamina II. In addition, Wnt3a, a prototypic Wnt ligand that activates the canonical pathway, is also enriched in the superficial layers. Immunoblotting analysis indicates that both Wnt3a a β-catenin are up-regulated in the SCDH of various mouse pain models created by hind-paw injection of capsaicin, intrathecal (i.t.) injection of HIV-gp120 protein or spinal nerve ligation (SNL). Furthermore, Wnt5a, a prototypic Wnt ligand for non-canonical pathways, and its receptor Ror2 are also up-regulated in the SCDH of these models.

Conclusion: Our results suggest that Wnt signaling pathways are regulated by nociceptive input. The activation of Wnt signaling may regulate the expression of spinal central sensitization during the development of acute and chronic pain.

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Cellular localization of β-catenin in the SCDH.A-C: Double-staining of β-catenin (A) and neuronal marker NeuN (B). Small clusters of β-catenin immunoreaction product (red) are in the cytoplasm around NeuN-labeled neuronal nuclei (arrows). Low levels of β-catenin staining are also observed around non-neuronal cell bodies (arrowheads) deep in the SCDH. Little staining is seen around NeuN in deep SCDH regions (asterisks). D-F: Double-staining of β-catenin (D) and synapsin I (E), a pre-synaptic marker. The β-catenin staining substantially overlaps with synapsin I detected in lamina II (F, arrows). G-I: Double-staining of β-catenin (G) and PSD95 (H). β-catenin staining overlapped with PSD95 staining (I, arrows). J-L: Double-staining of β-catenin (J) and CD11b (K). β-catenin was barely detectable in CD11b-postive microglial cells (L, arrows). M-O: Double-staining of β-catenin (M) and GFAP (N). β-catenin staining did not overlapped with GFAP staining (O, arrows). Insets are images at a higher magnification of the areas indicated by arrowheads, to show more clearly the spatial relation of β-catenin and various molecular markers. DAPI (blue) staining was performed to visualize all cells. Scale bar: 50 μm.
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Figure 2: Cellular localization of β-catenin in the SCDH.A-C: Double-staining of β-catenin (A) and neuronal marker NeuN (B). Small clusters of β-catenin immunoreaction product (red) are in the cytoplasm around NeuN-labeled neuronal nuclei (arrows). Low levels of β-catenin staining are also observed around non-neuronal cell bodies (arrowheads) deep in the SCDH. Little staining is seen around NeuN in deep SCDH regions (asterisks). D-F: Double-staining of β-catenin (D) and synapsin I (E), a pre-synaptic marker. The β-catenin staining substantially overlaps with synapsin I detected in lamina II (F, arrows). G-I: Double-staining of β-catenin (G) and PSD95 (H). β-catenin staining overlapped with PSD95 staining (I, arrows). J-L: Double-staining of β-catenin (J) and CD11b (K). β-catenin was barely detectable in CD11b-postive microglial cells (L, arrows). M-O: Double-staining of β-catenin (M) and GFAP (N). β-catenin staining did not overlapped with GFAP staining (O, arrows). Insets are images at a higher magnification of the areas indicated by arrowheads, to show more clearly the spatial relation of β-catenin and various molecular markers. DAPI (blue) staining was performed to visualize all cells. Scale bar: 50 μm.

Mentions: Because the Wnt/β-catenin pathway plays important roles in synaptic plasticity such as long-term potentiation (LTP) [22,33], we were interested in testing if this pathway is involved in the regulation of central sensitization. As an initial step toward this goal, we performed fluorescent immunostaining in naïve mice to determine the spinal distribution of β-catenin and Wnt3a, two signaling proteins in the canonical pathway. We observed that β-catenin immunostaining formed a predominant band in the dorsal horn, although a low level of signal was detected throughout the gray matter of the spinal cord (Figure 1). To define further the laminar distribution of the protein in the SCDH, we used molecular markers to label the specific layers of the dorsal horn. We found that β-catenin was enriched in lamina II, both the inner (IIi) and outer (IIo) segments (Figure 1), while its staining in lamina I was relatively low (Figure 1 A1-2). Staining for isolectin B4 (IB4) and PKCγ, considered as specific markers for the outer and inner segments of lamina II, respectively, confirmed the presence of β-catenin in both the lamina IIi and IIo (Figure 1 B1-2 and C1-2). These observations indicate that the β-catenin is enriched in lamina II of the SCDH. Previous studies revealed that β-catenin is expressed in hippocampal neurons [33]. We performed double-staining experiments, using NeuN to label neuronal cell bodies. As shown in Figure 2 A-C, label for β-catenin in lamina II was observed in regions surrounding neuronal nuclei labeled by NeuN, indicating that the majority of β-catenin was in neuronal cytoplasm. On the other hand, β-catenin staining in non-neuronal cell bodies (NeuN-negative; DAPI-labeled) was detectable but relatively low.


Regulation of Wnt signaling by nociceptive input in animal models.

Shi Y, Yuan S, Li B, Wang J, Carlton SM, Chung K, Chung JM, Tang SJ - Mol Pain (2012)

Cellular localization of β-catenin in the SCDH.A-C: Double-staining of β-catenin (A) and neuronal marker NeuN (B). Small clusters of β-catenin immunoreaction product (red) are in the cytoplasm around NeuN-labeled neuronal nuclei (arrows). Low levels of β-catenin staining are also observed around non-neuronal cell bodies (arrowheads) deep in the SCDH. Little staining is seen around NeuN in deep SCDH regions (asterisks). D-F: Double-staining of β-catenin (D) and synapsin I (E), a pre-synaptic marker. The β-catenin staining substantially overlaps with synapsin I detected in lamina II (F, arrows). G-I: Double-staining of β-catenin (G) and PSD95 (H). β-catenin staining overlapped with PSD95 staining (I, arrows). J-L: Double-staining of β-catenin (J) and CD11b (K). β-catenin was barely detectable in CD11b-postive microglial cells (L, arrows). M-O: Double-staining of β-catenin (M) and GFAP (N). β-catenin staining did not overlapped with GFAP staining (O, arrows). Insets are images at a higher magnification of the areas indicated by arrowheads, to show more clearly the spatial relation of β-catenin and various molecular markers. DAPI (blue) staining was performed to visualize all cells. Scale bar: 50 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 2: Cellular localization of β-catenin in the SCDH.A-C: Double-staining of β-catenin (A) and neuronal marker NeuN (B). Small clusters of β-catenin immunoreaction product (red) are in the cytoplasm around NeuN-labeled neuronal nuclei (arrows). Low levels of β-catenin staining are also observed around non-neuronal cell bodies (arrowheads) deep in the SCDH. Little staining is seen around NeuN in deep SCDH regions (asterisks). D-F: Double-staining of β-catenin (D) and synapsin I (E), a pre-synaptic marker. The β-catenin staining substantially overlaps with synapsin I detected in lamina II (F, arrows). G-I: Double-staining of β-catenin (G) and PSD95 (H). β-catenin staining overlapped with PSD95 staining (I, arrows). J-L: Double-staining of β-catenin (J) and CD11b (K). β-catenin was barely detectable in CD11b-postive microglial cells (L, arrows). M-O: Double-staining of β-catenin (M) and GFAP (N). β-catenin staining did not overlapped with GFAP staining (O, arrows). Insets are images at a higher magnification of the areas indicated by arrowheads, to show more clearly the spatial relation of β-catenin and various molecular markers. DAPI (blue) staining was performed to visualize all cells. Scale bar: 50 μm.
Mentions: Because the Wnt/β-catenin pathway plays important roles in synaptic plasticity such as long-term potentiation (LTP) [22,33], we were interested in testing if this pathway is involved in the regulation of central sensitization. As an initial step toward this goal, we performed fluorescent immunostaining in naïve mice to determine the spinal distribution of β-catenin and Wnt3a, two signaling proteins in the canonical pathway. We observed that β-catenin immunostaining formed a predominant band in the dorsal horn, although a low level of signal was detected throughout the gray matter of the spinal cord (Figure 1). To define further the laminar distribution of the protein in the SCDH, we used molecular markers to label the specific layers of the dorsal horn. We found that β-catenin was enriched in lamina II, both the inner (IIi) and outer (IIo) segments (Figure 1), while its staining in lamina I was relatively low (Figure 1 A1-2). Staining for isolectin B4 (IB4) and PKCγ, considered as specific markers for the outer and inner segments of lamina II, respectively, confirmed the presence of β-catenin in both the lamina IIi and IIo (Figure 1 B1-2 and C1-2). These observations indicate that the β-catenin is enriched in lamina II of the SCDH. Previous studies revealed that β-catenin is expressed in hippocampal neurons [33]. We performed double-staining experiments, using NeuN to label neuronal cell bodies. As shown in Figure 2 A-C, label for β-catenin in lamina II was observed in regions surrounding neuronal nuclei labeled by NeuN, indicating that the majority of β-catenin was in neuronal cytoplasm. On the other hand, β-catenin staining in non-neuronal cell bodies (NeuN-negative; DAPI-labeled) was detectable but relatively low.

Bottom Line: In addition, Wnt3a, a prototypic Wnt ligand that activates the canonical pathway, is also enriched in the superficial layers.Furthermore, Wnt5a, a prototypic Wnt ligand for non-canonical pathways, and its receptor Ror2 are also up-regulated in the SCDH of these models.Our results suggest that Wnt signaling pathways are regulated by nociceptive input.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX 77555, USA.

ABSTRACT

Background: Central sensitization-associated synaptic plasticity in the spinal cord dorsal horn (SCDH) critically contributes to the development of chronic pain, but understanding of the underlying molecular pathways is still incomplete. Emerging evidence suggests that Wnt signaling plays a crucial role in regulation of synaptic plasticity. Little is known about the potential function of the Wnt signaling cascades in chronic pain development.

Results: Fluorescent immunostaining results indicate that β-catenin, an essential protein in the canonical Wnt signaling pathway, is expressed in the superficial layers of the mouse SCDH with enrichment at synapses in lamina II. In addition, Wnt3a, a prototypic Wnt ligand that activates the canonical pathway, is also enriched in the superficial layers. Immunoblotting analysis indicates that both Wnt3a a β-catenin are up-regulated in the SCDH of various mouse pain models created by hind-paw injection of capsaicin, intrathecal (i.t.) injection of HIV-gp120 protein or spinal nerve ligation (SNL). Furthermore, Wnt5a, a prototypic Wnt ligand for non-canonical pathways, and its receptor Ror2 are also up-regulated in the SCDH of these models.

Conclusion: Our results suggest that Wnt signaling pathways are regulated by nociceptive input. The activation of Wnt signaling may regulate the expression of spinal central sensitization during the development of acute and chronic pain.

Show MeSH
Related in: MedlinePlus