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A critical role for pannexin-1 in activation of innate immune cells of the choroid plexus.

Maslieieva V, Thompson RJ - Channels (Austin) (2014)

Bottom Line: Here we have developed a novel technique for studying epiplexus cells in acutely isolated, live and intact choroid plexus.We show that epiplexus cells are potently activated by exogenous ATP, increasing their motility within the tissue.Furthermore, ATP acts at least in part through the P2X4 ionotropic purinergic receptor.

View Article: PubMed Central - PubMed

Affiliation: Hotchkiss Brain Institute; Department of Cell Biology and Anatomy; University of Calgary; Calgary, AB Canada.

ABSTRACT
Epiplexus cells are a population of innate immune cells in the choroid plexus of the brain ventricles. They are thought to contribute to the immune component of the blood-cerebrospinal-fluid-barrier (BCSFB). Here we have developed a novel technique for studying epiplexus cells in acutely isolated, live and intact choroid plexus. We show that epiplexus cells are potently activated by exogenous ATP, increasing their motility within the tissue. This ATP-induced chemokinesis required activation of pannexin-1 channels, which are expressed by the epithelial cells of the choroid plexus and not the epiplexus cells themselves. Furthermore, ATP acts at least in part through the P2X4 ionotropic purinergic receptor. Thus, the resident immune cells of the choroid plexus appear to be in communication with the epithelial cells through pannexin-1 channels.

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Figure 4. P2X7 and P2X4 receptors are both expressed on epiplexus cells, but only P2X4 is involved in chemokinesis. (A) Immunofluorescent staining for P2X4 receptors in epiplexus cells and CP epithelium. Note that both the epithelial and the epiplexus cells are positive for P2X4. (B) Immunofluorescent staining for P2X7 receptors is evident only in epiplexus cells. (C, D, and E) The P2X7 receptor blocker 1 µM BBG did not prevent activation of epiplexus cell by ATP, and the P2X7 receptor agonist BzATP did not trigger an increase in epiplexus cell activity. Thirty µM 5-BDBD, a P2X4 receptor antagonist, alone or in combination with probenecid significantly decreased ATP-triggered chemokinesis. (C) Normalized average distance and statistical analysis where each symbol represents an individual cell; (D) normalized summative distance; (E) slope of normalized summative distance where each symbol represents the average of all cells in a given CP. Scale bars are 50 μm.
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Figure 4: Figure 4. P2X7 and P2X4 receptors are both expressed on epiplexus cells, but only P2X4 is involved in chemokinesis. (A) Immunofluorescent staining for P2X4 receptors in epiplexus cells and CP epithelium. Note that both the epithelial and the epiplexus cells are positive for P2X4. (B) Immunofluorescent staining for P2X7 receptors is evident only in epiplexus cells. (C, D, and E) The P2X7 receptor blocker 1 µM BBG did not prevent activation of epiplexus cell by ATP, and the P2X7 receptor agonist BzATP did not trigger an increase in epiplexus cell activity. Thirty µM 5-BDBD, a P2X4 receptor antagonist, alone or in combination with probenecid significantly decreased ATP-triggered chemokinesis. (C) Normalized average distance and statistical analysis where each symbol represents an individual cell; (D) normalized summative distance; (E) slope of normalized summative distance where each symbol represents the average of all cells in a given CP. Scale bars are 50 μm.

Mentions: Block of chemokinesis by Panx1 antagonists suggests that purinergic receptors on the epithelial cells may be involved, because this is the identified site of Panx1 expression in the CP. Several ionotropic purinergic receptors have been reported to be linked to Panx1, including P2X7 and P2X4.29,30 Furthermore, both P2X7 and P2X4 are expressed in monocytic cells and can facilitate their activation.24,48 We first examined the expression of P2X7 and P2X4 in the CP by immunohistochemistry. P2X7 strongly co-labeled with IB4, but did not convincingly label the epithelial cells above background (Fig. 4B). In contrast, P2X4 was detected in both the epithelial cells and in IB4 positive epiplexus cells (Fig. 4A). Based on this pattern of labeling we predicted that P2X4, and not P2X7, is important for activation of epiplexus cells by ATP.


A critical role for pannexin-1 in activation of innate immune cells of the choroid plexus.

Maslieieva V, Thompson RJ - Channels (Austin) (2014)

Figure 4. P2X7 and P2X4 receptors are both expressed on epiplexus cells, but only P2X4 is involved in chemokinesis. (A) Immunofluorescent staining for P2X4 receptors in epiplexus cells and CP epithelium. Note that both the epithelial and the epiplexus cells are positive for P2X4. (B) Immunofluorescent staining for P2X7 receptors is evident only in epiplexus cells. (C, D, and E) The P2X7 receptor blocker 1 µM BBG did not prevent activation of epiplexus cell by ATP, and the P2X7 receptor agonist BzATP did not trigger an increase in epiplexus cell activity. Thirty µM 5-BDBD, a P2X4 receptor antagonist, alone or in combination with probenecid significantly decreased ATP-triggered chemokinesis. (C) Normalized average distance and statistical analysis where each symbol represents an individual cell; (D) normalized summative distance; (E) slope of normalized summative distance where each symbol represents the average of all cells in a given CP. Scale bars are 50 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4048302&req=5

Figure 4: Figure 4. P2X7 and P2X4 receptors are both expressed on epiplexus cells, but only P2X4 is involved in chemokinesis. (A) Immunofluorescent staining for P2X4 receptors in epiplexus cells and CP epithelium. Note that both the epithelial and the epiplexus cells are positive for P2X4. (B) Immunofluorescent staining for P2X7 receptors is evident only in epiplexus cells. (C, D, and E) The P2X7 receptor blocker 1 µM BBG did not prevent activation of epiplexus cell by ATP, and the P2X7 receptor agonist BzATP did not trigger an increase in epiplexus cell activity. Thirty µM 5-BDBD, a P2X4 receptor antagonist, alone or in combination with probenecid significantly decreased ATP-triggered chemokinesis. (C) Normalized average distance and statistical analysis where each symbol represents an individual cell; (D) normalized summative distance; (E) slope of normalized summative distance where each symbol represents the average of all cells in a given CP. Scale bars are 50 μm.
Mentions: Block of chemokinesis by Panx1 antagonists suggests that purinergic receptors on the epithelial cells may be involved, because this is the identified site of Panx1 expression in the CP. Several ionotropic purinergic receptors have been reported to be linked to Panx1, including P2X7 and P2X4.29,30 Furthermore, both P2X7 and P2X4 are expressed in monocytic cells and can facilitate their activation.24,48 We first examined the expression of P2X7 and P2X4 in the CP by immunohistochemistry. P2X7 strongly co-labeled with IB4, but did not convincingly label the epithelial cells above background (Fig. 4B). In contrast, P2X4 was detected in both the epithelial cells and in IB4 positive epiplexus cells (Fig. 4A). Based on this pattern of labeling we predicted that P2X4, and not P2X7, is important for activation of epiplexus cells by ATP.

Bottom Line: Here we have developed a novel technique for studying epiplexus cells in acutely isolated, live and intact choroid plexus.We show that epiplexus cells are potently activated by exogenous ATP, increasing their motility within the tissue.Furthermore, ATP acts at least in part through the P2X4 ionotropic purinergic receptor.

View Article: PubMed Central - PubMed

Affiliation: Hotchkiss Brain Institute; Department of Cell Biology and Anatomy; University of Calgary; Calgary, AB Canada.

ABSTRACT
Epiplexus cells are a population of innate immune cells in the choroid plexus of the brain ventricles. They are thought to contribute to the immune component of the blood-cerebrospinal-fluid-barrier (BCSFB). Here we have developed a novel technique for studying epiplexus cells in acutely isolated, live and intact choroid plexus. We show that epiplexus cells are potently activated by exogenous ATP, increasing their motility within the tissue. This ATP-induced chemokinesis required activation of pannexin-1 channels, which are expressed by the epithelial cells of the choroid plexus and not the epiplexus cells themselves. Furthermore, ATP acts at least in part through the P2X4 ionotropic purinergic receptor. Thus, the resident immune cells of the choroid plexus appear to be in communication with the epithelial cells through pannexin-1 channels.

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