Limits...
Plasticity of Scarpa's Ganglion Neurons as a Possible Basis for Functional Restoration within Vestibular Endorgans.

Travo C, Gaboyard-Niay S, Chabbert C - Front Neurol (2012)

Bottom Line: When co-cultured with vestibular epithelia, primary vestibular neurons were able to establish de novo contacts with hair cells.Under the present paradigm, these contacts displayed morphological features of immature synaptic contacts.Preliminary observations using co-cultures of adult rodents suggest that this reparative capacity remained in older mice although to a lesser extent.

View Article: PubMed Central - PubMed

Affiliation: INSERM U1051, Institute for Neurosciences Montpellier, France.

ABSTRACT
In a previous study, we observed spontaneous restoration of vestibular function in young adult rodents following excitotoxic injury of the neuronal connections within vestibular endorgans. The functional restoration was supported by a repair of synaptic contacts between hair cells and primary vestibular neurons. This process was observed in 2/3 of the animals studied and occurred within 5 days following the synaptic damage. To assess whether repair capacity is a fundamental trait of vestibular endorgans and to decipher the cellular mechanisms supporting such a repair process, we studied the neuronal regeneration and synaptogenesis in co-cultures of vestibular epithelia and Scarpa's ganglion from young and adult rodents. We demonstrate that, under specific culture conditions, primary vestibular neurons from young mice or rats exhibit robust ability to regenerate nervous processes. When co-cultured with vestibular epithelia, primary vestibular neurons were able to establish de novo contacts with hair cells. Under the present paradigm, these contacts displayed morphological features of immature synaptic contacts. Preliminary observations using co-cultures of adult rodents suggest that this reparative capacity remained in older mice although to a lesser extent. Identifying the basic mechanisms underlying the repair process may provide a basis for novel therapeutic strategies to restore mature and functional vestibular synaptic contacts following damage or loss.

No MeSH data available.


Related in: MedlinePlus

Neurite outgrowth in 6–14 DIV co-cultures from P3–8 rodents. In co-cultures of vestibular ganglion, crista (Cr), and utricle (U) from P5 mouse (A) or rat (B) pups, neurons extend processes toward the sensory epithelia. Neurofilaments (green) were stained with anti-N52 antibodies, hair cells (red) with anti-calretinin serum, and nuclei (blue) with ToPro3. The importance of extrinsic factors was highlighted in these culture conditions (C,D). BDNF added to the culture medium [P5 mice; (C)] enhances neurite outgrowth, but its presence in the culture medium disrupts with the natural directional attraction of neurites by the sensory epithelium. Neurites spread all over the culture plate (arrows) without preferential orientation as observed in (A) or (B). Staining of Schwann cells [P3 rats; (D), red, arrow heads] demonstrates their contribution to the process of neurite outgrowth toward sensory epithelia. Scale bars 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3368229&req=5

Figure 1: Neurite outgrowth in 6–14 DIV co-cultures from P3–8 rodents. In co-cultures of vestibular ganglion, crista (Cr), and utricle (U) from P5 mouse (A) or rat (B) pups, neurons extend processes toward the sensory epithelia. Neurofilaments (green) were stained with anti-N52 antibodies, hair cells (red) with anti-calretinin serum, and nuclei (blue) with ToPro3. The importance of extrinsic factors was highlighted in these culture conditions (C,D). BDNF added to the culture medium [P5 mice; (C)] enhances neurite outgrowth, but its presence in the culture medium disrupts with the natural directional attraction of neurites by the sensory epithelium. Neurites spread all over the culture plate (arrows) without preferential orientation as observed in (A) or (B). Staining of Schwann cells [P3 rats; (D), red, arrow heads] demonstrates their contribution to the process of neurite outgrowth toward sensory epithelia. Scale bars 100 μm.

Mentions: Once removed from the inner ear of neonate mice or rats, isolated vestibular endorgans were cultured as 3D organotypic explants as previously reported (Gaboyard et al., 2005). Under these conditions hair cells rapidly recovered from the isolation procedure while an endolymphatic compartment containing enriched K+ liquid regenerated (Bartolami et al., 2011). In the present study, vestibular (Scarpa’s) ganglia were also dissected and co-cultured in the vicinity (within a distance of 0.5–1 mm) of the sensory epithelia (utricles and cristae ampullaris). During the first 3 days in vitro (DIV), both the proximal (not shown) and distal (Figure 2A) nerve processes of primary vestibular neurons degenerated after being severed from their targets or cell bodies respectively. Subsequently, de novo neurite outgrowth was observed. Bundles of fibers arising from Scarpa’s ganglion neurons with preferential orientation toward the sensory epithelia were clearly observable in all co-cultures after 6 DIV (mice n = 12; rats n = 10; Figures 1A,B). As BDNF was previously demonstrated to play a critical role during developmental innervation of sensory epithelia by vestibular primary neurons (Ernfors et al., 1995), we used the co-culture model from young rodents to examine the effect of BDNF addition on de novo neurite outgrowth. When BDNF (10 ng/mL; mice n = 4; rats n = 4) was added to the culture medium, the orientation of de novo neurite outgrowth was significantly altered (Figure 1C), forming a radiating pattern from Scarpa’s ganglia, and only occasionally still innervating the sensory epithelia. This observation confirms the role of BDNF in the guidance of neuritis during the innervation of sensory epithelia. It agrees with studies that show controlled release of BDNF by the targeted cells or their immediate environment is required for innervation of sensory epithelia by primary vestibular neurons (Brugeaud et al., 2007). Schwann cells have also been shown to play an important role in the guidance of neuronal processes toward their targets during peripheral nerve regeneration (Zhang and Yannas, 2005). Using immunostaining of the S100β protein, specifically expressed in vestibular glial cells (Bartolami et al., 2003), we confirmed that Schwann cells were part of the re-innervation process in the co-culture preparation. Indeed, the neuritic outgrowth of nerve fibers was never observed without accompanying Schwann cells (rats, n = 4; Figure 1D).


Plasticity of Scarpa's Ganglion Neurons as a Possible Basis for Functional Restoration within Vestibular Endorgans.

Travo C, Gaboyard-Niay S, Chabbert C - Front Neurol (2012)

Neurite outgrowth in 6–14 DIV co-cultures from P3–8 rodents. In co-cultures of vestibular ganglion, crista (Cr), and utricle (U) from P5 mouse (A) or rat (B) pups, neurons extend processes toward the sensory epithelia. Neurofilaments (green) were stained with anti-N52 antibodies, hair cells (red) with anti-calretinin serum, and nuclei (blue) with ToPro3. The importance of extrinsic factors was highlighted in these culture conditions (C,D). BDNF added to the culture medium [P5 mice; (C)] enhances neurite outgrowth, but its presence in the culture medium disrupts with the natural directional attraction of neurites by the sensory epithelium. Neurites spread all over the culture plate (arrows) without preferential orientation as observed in (A) or (B). Staining of Schwann cells [P3 rats; (D), red, arrow heads] demonstrates their contribution to the process of neurite outgrowth toward sensory epithelia. Scale bars 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3368229&req=5

Figure 1: Neurite outgrowth in 6–14 DIV co-cultures from P3–8 rodents. In co-cultures of vestibular ganglion, crista (Cr), and utricle (U) from P5 mouse (A) or rat (B) pups, neurons extend processes toward the sensory epithelia. Neurofilaments (green) were stained with anti-N52 antibodies, hair cells (red) with anti-calretinin serum, and nuclei (blue) with ToPro3. The importance of extrinsic factors was highlighted in these culture conditions (C,D). BDNF added to the culture medium [P5 mice; (C)] enhances neurite outgrowth, but its presence in the culture medium disrupts with the natural directional attraction of neurites by the sensory epithelium. Neurites spread all over the culture plate (arrows) without preferential orientation as observed in (A) or (B). Staining of Schwann cells [P3 rats; (D), red, arrow heads] demonstrates their contribution to the process of neurite outgrowth toward sensory epithelia. Scale bars 100 μm.
Mentions: Once removed from the inner ear of neonate mice or rats, isolated vestibular endorgans were cultured as 3D organotypic explants as previously reported (Gaboyard et al., 2005). Under these conditions hair cells rapidly recovered from the isolation procedure while an endolymphatic compartment containing enriched K+ liquid regenerated (Bartolami et al., 2011). In the present study, vestibular (Scarpa’s) ganglia were also dissected and co-cultured in the vicinity (within a distance of 0.5–1 mm) of the sensory epithelia (utricles and cristae ampullaris). During the first 3 days in vitro (DIV), both the proximal (not shown) and distal (Figure 2A) nerve processes of primary vestibular neurons degenerated after being severed from their targets or cell bodies respectively. Subsequently, de novo neurite outgrowth was observed. Bundles of fibers arising from Scarpa’s ganglion neurons with preferential orientation toward the sensory epithelia were clearly observable in all co-cultures after 6 DIV (mice n = 12; rats n = 10; Figures 1A,B). As BDNF was previously demonstrated to play a critical role during developmental innervation of sensory epithelia by vestibular primary neurons (Ernfors et al., 1995), we used the co-culture model from young rodents to examine the effect of BDNF addition on de novo neurite outgrowth. When BDNF (10 ng/mL; mice n = 4; rats n = 4) was added to the culture medium, the orientation of de novo neurite outgrowth was significantly altered (Figure 1C), forming a radiating pattern from Scarpa’s ganglia, and only occasionally still innervating the sensory epithelia. This observation confirms the role of BDNF in the guidance of neuritis during the innervation of sensory epithelia. It agrees with studies that show controlled release of BDNF by the targeted cells or their immediate environment is required for innervation of sensory epithelia by primary vestibular neurons (Brugeaud et al., 2007). Schwann cells have also been shown to play an important role in the guidance of neuronal processes toward their targets during peripheral nerve regeneration (Zhang and Yannas, 2005). Using immunostaining of the S100β protein, specifically expressed in vestibular glial cells (Bartolami et al., 2003), we confirmed that Schwann cells were part of the re-innervation process in the co-culture preparation. Indeed, the neuritic outgrowth of nerve fibers was never observed without accompanying Schwann cells (rats, n = 4; Figure 1D).

Bottom Line: When co-cultured with vestibular epithelia, primary vestibular neurons were able to establish de novo contacts with hair cells.Under the present paradigm, these contacts displayed morphological features of immature synaptic contacts.Preliminary observations using co-cultures of adult rodents suggest that this reparative capacity remained in older mice although to a lesser extent.

View Article: PubMed Central - PubMed

Affiliation: INSERM U1051, Institute for Neurosciences Montpellier, France.

ABSTRACT
In a previous study, we observed spontaneous restoration of vestibular function in young adult rodents following excitotoxic injury of the neuronal connections within vestibular endorgans. The functional restoration was supported by a repair of synaptic contacts between hair cells and primary vestibular neurons. This process was observed in 2/3 of the animals studied and occurred within 5 days following the synaptic damage. To assess whether repair capacity is a fundamental trait of vestibular endorgans and to decipher the cellular mechanisms supporting such a repair process, we studied the neuronal regeneration and synaptogenesis in co-cultures of vestibular epithelia and Scarpa's ganglion from young and adult rodents. We demonstrate that, under specific culture conditions, primary vestibular neurons from young mice or rats exhibit robust ability to regenerate nervous processes. When co-cultured with vestibular epithelia, primary vestibular neurons were able to establish de novo contacts with hair cells. Under the present paradigm, these contacts displayed morphological features of immature synaptic contacts. Preliminary observations using co-cultures of adult rodents suggest that this reparative capacity remained in older mice although to a lesser extent. Identifying the basic mechanisms underlying the repair process may provide a basis for novel therapeutic strategies to restore mature and functional vestibular synaptic contacts following damage or loss.

No MeSH data available.


Related in: MedlinePlus