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Staurosporine-induced collapse of cochlear hair bundles.

Goodyear RJ, Ratnayaka HS, Warchol ME, Richardson GP - J. Comp. Neurol. (2014)

Bottom Line: Staurosporine exposure results in the fusion of the hair bundle's stereocilia, a resorption of the parallel actin bundles of the stereocilia into the cytoplasm of the hair cell, a detachment of the apical, non-stereociliary membrane of the hair cell from the underlying cuticular plate, and a severing of the hair-bundle's rootlets from the actin cores of the stereocilia.It does not block membrane retrieval at the apical pole of the hair cells, nor does it elicit the externalization of phosphatidylserine.Staurosporine treatment causes a reduction in levels of the phosphorylated forms of ezrin, radixin, and moesin in cochlear cultures during the period of hair-bundle loss, indicating the integrity of the hair bundle may be actively maintained by the phosphorylation status of these proteins.

View Article: PubMed Central - PubMed

Affiliation: Sussex Neuroscience and School of Life Sciences, University of Sussex, Brighton, BN1 9QG, UK.

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Staurosporine dose–response. Confocal images from mid-apical (A,C,E,G) and basal (B,D,F,H) coil cochlear cultures grown for 16 hours in control medium (A,B) or in medium containing staurosporine at a concentration of 1 nM (C,D), 10 nM (E,F), or 100 nM (G,H). At concentrations of 1 and 10 nM staurosporine, hair bundles appear to partially collapse or splay (E,F), while at 100 nM staurosporine exposure results in severe disruption of the organ of Corti and fusion of stereocilia (G,H). Arrowheads and arrows point to disrupted IHCs and OHCs, respectively, in parts C–H. Scale bar = 20 μm in H (applies to all).
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fig02: Staurosporine dose–response. Confocal images from mid-apical (A,C,E,G) and basal (B,D,F,H) coil cochlear cultures grown for 16 hours in control medium (A,B) or in medium containing staurosporine at a concentration of 1 nM (C,D), 10 nM (E,F), or 100 nM (G,H). At concentrations of 1 and 10 nM staurosporine, hair bundles appear to partially collapse or splay (E,F), while at 100 nM staurosporine exposure results in severe disruption of the organ of Corti and fusion of stereocilia (G,H). Arrowheads and arrows point to disrupted IHCs and OHCs, respectively, in parts C–H. Scale bar = 20 μm in H (applies to all).

Mentions: Figure 1 illustrates the morphology of control cultures and the effects of kinase inhibitors that caused disruption of hair-bundle structure without a general toxic effect on the culture. The appearance of phalloidin-stained hair bundles in the middle of apical and basal-coil cochlear cultures following 24 hours in control medium is illustrated in Figure 1A,B. At a concentration of 10 μM, the protein kinase A (PKA) inhibitor H89 caused a considerable expansion of the apical surface of hair cells and a distortion of the typical V-shaped pattern of the outer hair cell (OHC) bundles (Fig. 1C). This effect, however, was restricted to cells in the apical-coil cultures and not observed in basal-coil hair cells (Fig. 1D). Two inhibitors of the lipid kinase phosphatidylinositol 3-kinase (PI-3 kinase), LY294002 and the fungal metabolite wortmannin, also preferentially affected the shape of apical-coil (Fig. 1E,G), as opposed to basal-coil (Fig. 1F,H) cochlear hair bundles. LY294002 resulted in changes to hair-bundle morphology at a concentration of 10 μM (Fig. 1E). At a concentration of 20 nM wortmannin also resulted in some disruption of hair-bundle structure; hair bundles of OHC became more compact in appearance, and those of the inner hair cells (IHC) became severely distorted (Fig. 1G). Only one of the inhibitors tested, staurosporine, caused the collapse of hair bundles throughout the entire length of the cochlea (Fig. 2). Its effects were therefore examined in more detail.


Staurosporine-induced collapse of cochlear hair bundles.

Goodyear RJ, Ratnayaka HS, Warchol ME, Richardson GP - J. Comp. Neurol. (2014)

Staurosporine dose–response. Confocal images from mid-apical (A,C,E,G) and basal (B,D,F,H) coil cochlear cultures grown for 16 hours in control medium (A,B) or in medium containing staurosporine at a concentration of 1 nM (C,D), 10 nM (E,F), or 100 nM (G,H). At concentrations of 1 and 10 nM staurosporine, hair bundles appear to partially collapse or splay (E,F), while at 100 nM staurosporine exposure results in severe disruption of the organ of Corti and fusion of stereocilia (G,H). Arrowheads and arrows point to disrupted IHCs and OHCs, respectively, in parts C–H. Scale bar = 20 μm in H (applies to all).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4321040&req=5

fig02: Staurosporine dose–response. Confocal images from mid-apical (A,C,E,G) and basal (B,D,F,H) coil cochlear cultures grown for 16 hours in control medium (A,B) or in medium containing staurosporine at a concentration of 1 nM (C,D), 10 nM (E,F), or 100 nM (G,H). At concentrations of 1 and 10 nM staurosporine, hair bundles appear to partially collapse or splay (E,F), while at 100 nM staurosporine exposure results in severe disruption of the organ of Corti and fusion of stereocilia (G,H). Arrowheads and arrows point to disrupted IHCs and OHCs, respectively, in parts C–H. Scale bar = 20 μm in H (applies to all).
Mentions: Figure 1 illustrates the morphology of control cultures and the effects of kinase inhibitors that caused disruption of hair-bundle structure without a general toxic effect on the culture. The appearance of phalloidin-stained hair bundles in the middle of apical and basal-coil cochlear cultures following 24 hours in control medium is illustrated in Figure 1A,B. At a concentration of 10 μM, the protein kinase A (PKA) inhibitor H89 caused a considerable expansion of the apical surface of hair cells and a distortion of the typical V-shaped pattern of the outer hair cell (OHC) bundles (Fig. 1C). This effect, however, was restricted to cells in the apical-coil cultures and not observed in basal-coil hair cells (Fig. 1D). Two inhibitors of the lipid kinase phosphatidylinositol 3-kinase (PI-3 kinase), LY294002 and the fungal metabolite wortmannin, also preferentially affected the shape of apical-coil (Fig. 1E,G), as opposed to basal-coil (Fig. 1F,H) cochlear hair bundles. LY294002 resulted in changes to hair-bundle morphology at a concentration of 10 μM (Fig. 1E). At a concentration of 20 nM wortmannin also resulted in some disruption of hair-bundle structure; hair bundles of OHC became more compact in appearance, and those of the inner hair cells (IHC) became severely distorted (Fig. 1G). Only one of the inhibitors tested, staurosporine, caused the collapse of hair bundles throughout the entire length of the cochlea (Fig. 2). Its effects were therefore examined in more detail.

Bottom Line: Staurosporine exposure results in the fusion of the hair bundle's stereocilia, a resorption of the parallel actin bundles of the stereocilia into the cytoplasm of the hair cell, a detachment of the apical, non-stereociliary membrane of the hair cell from the underlying cuticular plate, and a severing of the hair-bundle's rootlets from the actin cores of the stereocilia.It does not block membrane retrieval at the apical pole of the hair cells, nor does it elicit the externalization of phosphatidylserine.Staurosporine treatment causes a reduction in levels of the phosphorylated forms of ezrin, radixin, and moesin in cochlear cultures during the period of hair-bundle loss, indicating the integrity of the hair bundle may be actively maintained by the phosphorylation status of these proteins.

View Article: PubMed Central - PubMed

Affiliation: Sussex Neuroscience and School of Life Sciences, University of Sussex, Brighton, BN1 9QG, UK.

Show MeSH
Related in: MedlinePlus