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Failure of fluid absorption in the endolymphatic sac initiates cochlear enlargement that leads to deafness in mice lacking pendrin expression.

Kim HM, Wangemann P - PLoS ONE (2010)

Bottom Line: Cochlear lumen formation was found to begin at the base of the cochlea between embryonic day (E) 13.5 and 14.5.Ligation or resection performed later, at E17.5, did not alter the cochlea lumen.In conclusion, the data suggest that cochlear lumen formation is initiated by fluid secretion in the vestibular labyrinth and temporarily controlled by fluid absorption in the endolymphatic sac.

View Article: PubMed Central - PubMed

Affiliation: Anatomy and Physiology Department, Kansas State University, Manhattan, Kansas, United States of America.

ABSTRACT
Mutations of SLC26A4 are among the most prevalent causes of hereditary deafness. Deafness in the corresponding mouse model, Slc26a4(-/-), results from an abnormally enlarged cochlear lumen. The goal of this study was to determine whether the cochlear enlargement originates with defective cochlear fluid transport or with a malfunction of fluid transport in the connected compartments, which are the vestibular labyrinth and the endolymphatic sac. Embryonic inner ears from Slc26a4(+/-) and Slc26a4(-/-) mice were examined by confocal microscopy ex vivo or after 2 days of organ culture. Culture allowed observations of intact, ligated or partially resected inner ears. Cochlear lumen formation was found to begin at the base of the cochlea between embryonic day (E) 13.5 and 14.5. Enlargement was immediately evident in Slc26a4(-/-) compared to Slc26a4(+/-) mice. In Slc26a4(+/-) and Slc26a4(-/-) mice, separation of the cochlea from the vestibular labyrinth by ligation at E14.5 resulted in a reduced cochlear lumen. Resection of the endolymphatic sacs at E14.5 led to an enlarged cochlear lumen in Slc26a4(+/-) mice but caused no further enlargement of the already enlarged cochlear lumen in Slc26a4(-/-) mice. Ligation or resection performed later, at E17.5, did not alter the cochlea lumen. In conclusion, the data suggest that cochlear lumen formation is initiated by fluid secretion in the vestibular labyrinth and temporarily controlled by fluid absorption in the endolymphatic sac. Failure of fluid absorption in the endolymphatic sac due to lack of Slc26a4 expression appears to initiate cochlear enlargement in mice, and possibly humans, lacking functional Slc26a4 expression.

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Fluid absorption in the endolymphatic sac ‘drains’ the cochlea during lumen formation.A, D and G: Luminal area measurements (average of the two basal cross-sections) in cochleae with and without endolymphatic sac. Cochleae were harvested at E14.5 and E17.5 from Slc26a4+/− and Slc26a4−/− mice and maintained two days in organ culture. The number next to the bars represents the N number of animals. Significant differences are marked (*). B and C, cochlear cross-sections of E14.5 Slc26a4+/− mice with and without endolymphatic sac. E and F, cochlear cross-sections of E14.5 Slc26a4−/− mice with and without endolymphatic sac. H and I, cochlear cross-sections of E17.5 Slc26a4+/− mice with and without endolymphatic sac. Scala media was highlighted in pink. Images without the pink overlay are available in Figure S1 (Supporting information). F-actin (green) and nuclei (blue) were labeled. Cochlear cross-sections were imaged by laser-scanning microscopy. Arrows point to the two basal cross-sections, which were used for digital area measurements.
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pone-0014041-g009: Fluid absorption in the endolymphatic sac ‘drains’ the cochlea during lumen formation.A, D and G: Luminal area measurements (average of the two basal cross-sections) in cochleae with and without endolymphatic sac. Cochleae were harvested at E14.5 and E17.5 from Slc26a4+/− and Slc26a4−/− mice and maintained two days in organ culture. The number next to the bars represents the N number of animals. Significant differences are marked (*). B and C, cochlear cross-sections of E14.5 Slc26a4+/− mice with and without endolymphatic sac. E and F, cochlear cross-sections of E14.5 Slc26a4−/− mice with and without endolymphatic sac. H and I, cochlear cross-sections of E17.5 Slc26a4+/− mice with and without endolymphatic sac. Scala media was highlighted in pink. Images without the pink overlay are available in Figure S1 (Supporting information). F-actin (green) and nuclei (blue) were labeled. Cochlear cross-sections were imaged by laser-scanning microscopy. Arrows point to the two basal cross-sections, which were used for digital area measurements.

Mentions: In a fourth series of experiments, we determined whether the endolymphatic sac contributes to cochlear lumen formation. Embryonic inner ears were again harvested at E14.5 and E17.5. Great care was taken to not injure the endolymphatic sac during dissection. The endolymphatic sac was then removed by manual resection (Fig. 8). Resected and non-resected inner ears were maintained in organ culture for two day, then sectioned and examined by confocal microscopy. The cochlear lumen of inner ears harvested at E14.5 from Slc26a4+/− mice was found to be significantly larger in resected compared to non-resected inner ears (Fig. 9). In contrast, the cochlear lumen of inner ears harvested at E14.5 from Slc26a4−/− mice was enlarged and no difference was apparent between resected and non-resected inner ears. Further, no difference in the cochlear lumina were found between resected and non-resected inner ears harvested at E17.5. These observations suggest that the onset of cochlear lumen formation is controlled in Slc26a4+/− mice by fluid absorption in the endolymphatic sac. This control is transient and found to be lost at E17.5. The observed enlargement in Slc26a4−/− suggests that loss of Slc26a4 expression impairs fluid absorption in the endolymphatic sac.


Failure of fluid absorption in the endolymphatic sac initiates cochlear enlargement that leads to deafness in mice lacking pendrin expression.

Kim HM, Wangemann P - PLoS ONE (2010)

Fluid absorption in the endolymphatic sac ‘drains’ the cochlea during lumen formation.A, D and G: Luminal area measurements (average of the two basal cross-sections) in cochleae with and without endolymphatic sac. Cochleae were harvested at E14.5 and E17.5 from Slc26a4+/− and Slc26a4−/− mice and maintained two days in organ culture. The number next to the bars represents the N number of animals. Significant differences are marked (*). B and C, cochlear cross-sections of E14.5 Slc26a4+/− mice with and without endolymphatic sac. E and F, cochlear cross-sections of E14.5 Slc26a4−/− mice with and without endolymphatic sac. H and I, cochlear cross-sections of E17.5 Slc26a4+/− mice with and without endolymphatic sac. Scala media was highlighted in pink. Images without the pink overlay are available in Figure S1 (Supporting information). F-actin (green) and nuclei (blue) were labeled. Cochlear cross-sections were imaged by laser-scanning microscopy. Arrows point to the two basal cross-sections, which were used for digital area measurements.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2984494&req=5

pone-0014041-g009: Fluid absorption in the endolymphatic sac ‘drains’ the cochlea during lumen formation.A, D and G: Luminal area measurements (average of the two basal cross-sections) in cochleae with and without endolymphatic sac. Cochleae were harvested at E14.5 and E17.5 from Slc26a4+/− and Slc26a4−/− mice and maintained two days in organ culture. The number next to the bars represents the N number of animals. Significant differences are marked (*). B and C, cochlear cross-sections of E14.5 Slc26a4+/− mice with and without endolymphatic sac. E and F, cochlear cross-sections of E14.5 Slc26a4−/− mice with and without endolymphatic sac. H and I, cochlear cross-sections of E17.5 Slc26a4+/− mice with and without endolymphatic sac. Scala media was highlighted in pink. Images without the pink overlay are available in Figure S1 (Supporting information). F-actin (green) and nuclei (blue) were labeled. Cochlear cross-sections were imaged by laser-scanning microscopy. Arrows point to the two basal cross-sections, which were used for digital area measurements.
Mentions: In a fourth series of experiments, we determined whether the endolymphatic sac contributes to cochlear lumen formation. Embryonic inner ears were again harvested at E14.5 and E17.5. Great care was taken to not injure the endolymphatic sac during dissection. The endolymphatic sac was then removed by manual resection (Fig. 8). Resected and non-resected inner ears were maintained in organ culture for two day, then sectioned and examined by confocal microscopy. The cochlear lumen of inner ears harvested at E14.5 from Slc26a4+/− mice was found to be significantly larger in resected compared to non-resected inner ears (Fig. 9). In contrast, the cochlear lumen of inner ears harvested at E14.5 from Slc26a4−/− mice was enlarged and no difference was apparent between resected and non-resected inner ears. Further, no difference in the cochlear lumina were found between resected and non-resected inner ears harvested at E17.5. These observations suggest that the onset of cochlear lumen formation is controlled in Slc26a4+/− mice by fluid absorption in the endolymphatic sac. This control is transient and found to be lost at E17.5. The observed enlargement in Slc26a4−/− suggests that loss of Slc26a4 expression impairs fluid absorption in the endolymphatic sac.

Bottom Line: Cochlear lumen formation was found to begin at the base of the cochlea between embryonic day (E) 13.5 and 14.5.Ligation or resection performed later, at E17.5, did not alter the cochlea lumen.In conclusion, the data suggest that cochlear lumen formation is initiated by fluid secretion in the vestibular labyrinth and temporarily controlled by fluid absorption in the endolymphatic sac.

View Article: PubMed Central - PubMed

Affiliation: Anatomy and Physiology Department, Kansas State University, Manhattan, Kansas, United States of America.

ABSTRACT
Mutations of SLC26A4 are among the most prevalent causes of hereditary deafness. Deafness in the corresponding mouse model, Slc26a4(-/-), results from an abnormally enlarged cochlear lumen. The goal of this study was to determine whether the cochlear enlargement originates with defective cochlear fluid transport or with a malfunction of fluid transport in the connected compartments, which are the vestibular labyrinth and the endolymphatic sac. Embryonic inner ears from Slc26a4(+/-) and Slc26a4(-/-) mice were examined by confocal microscopy ex vivo or after 2 days of organ culture. Culture allowed observations of intact, ligated or partially resected inner ears. Cochlear lumen formation was found to begin at the base of the cochlea between embryonic day (E) 13.5 and 14.5. Enlargement was immediately evident in Slc26a4(-/-) compared to Slc26a4(+/-) mice. In Slc26a4(+/-) and Slc26a4(-/-) mice, separation of the cochlea from the vestibular labyrinth by ligation at E14.5 resulted in a reduced cochlear lumen. Resection of the endolymphatic sacs at E14.5 led to an enlarged cochlear lumen in Slc26a4(+/-) mice but caused no further enlargement of the already enlarged cochlear lumen in Slc26a4(-/-) mice. Ligation or resection performed later, at E17.5, did not alter the cochlea lumen. In conclusion, the data suggest that cochlear lumen formation is initiated by fluid secretion in the vestibular labyrinth and temporarily controlled by fluid absorption in the endolymphatic sac. Failure of fluid absorption in the endolymphatic sac due to lack of Slc26a4 expression appears to initiate cochlear enlargement in mice, and possibly humans, lacking functional Slc26a4 expression.

Show MeSH
Related in: MedlinePlus