Overexpression of Bcl‐2 or Bcl‐xL prevents spiral ganglion neuron death and inhibits neurite growth

Abstract

Spiral ganglion neurons (SGNs) provide afferent innervation to the cochlea and rely on contact with hair cells (HCs) for their survival. Following deafferentation due to hair cell loss, SGNs gradually die. In a rat culture model, we explored the ability of prosurvival members of the Bcl-2 family of proteins to support the survival and neurite outgrowth of SGNs. We found that overexpression of either Bcl-2 or Bcl-xL significantly increases SGN survival in the absence of neurotrophic factors, establishing that the Bcl-2 pathway is sufficient for SGN cell survival and that SGN deprived of trophic support die by an apoptotic mechanism. However, in contrast to observations in central neurons and PC12 cells where Bcl-2 appears to promote neurite growth, both Bcl-2 and Bcl-xL overexpression dramatically inhibit neurite outgrowth in SGNs. This inhibition of neurite growth by Bcl-2 occurs in nearly all SGNs even in the presence of multiple neurotrophic factors implying that Bcl-2 directly inhibits neurite growth rather than simply rescuing a subpopulation of neurons incapable of extending neurites without additional stimuli. Thus, although overexpression of prosurvival members of the Bcl-2 family prevents SGN loss following trophic factor deprivation, the inhibition of neurite growth by these molecules may limit their efficacy for support of auditory nerve maintenance or regeneration following hair cell loss.

A Critical Time Window for the Survival of Neural Progenitor Cells by HDAC Inhibitors in the Hippocampus

“…3B and 3C; VPA 6 h, 146.9 ± 6.7%, VPA 24 h, 146.9 ± 1.6%; SBt 6 h, 193.4.8 ± 13.6%, SBt 12 h, 167.3 ± 5.5%, n = 3 per group). Although Bcl-2 may play a major role in promoting the survival of immature neurons, Bcl-xL, another anti-apoptotic Bcl-2 family member, may also be involved (Dietz et al, 2007;Hansen et al, 2007;Le et al, 2008). The presented results show that the expression of Bcl-xL increased by VPA and SBt (Figs.…”

Membrane depolarization inhibits spiral ganglion neurite growth via activation of multiple types of voltage sensitive calcium channels and calpain

“…Six to eight hours after plating (to allow time for attachment) the cultures were transfected with green fluorescent protein (GFP) -tagged autocamtide-2-related inhibitory peptide (GFP-AIP) or GFP-tagged control (scrambled) peptide (GFP-CON) (Bok et al, 2007) expression plasmids using calcium-phosphate precipitation as previously described , Hansen et al, 2003. Typically, this resulted in the transfection of 10-15% of the SGNs yielding approximately 130 transfected SGNs per well (Hansen et al, 2007). Twelve hours after transfection the medium was removed and replaced with NT-3-containing culture medium (depolarizing or control nondepolarizing) for an additional 48 hr.…”

“…If the longest neurite/branch of the chosen cell extended beyond the image border, additional images were acquired to include and measure the entire length of the process. Measurement of neurites in transfected SGNs was as above except that only GFP-expressing SGNs were scored as previously described (Hansen et al, 2007).…”

“…This completely fills the somata and neurites allowing clear visualization of SGN morphology in live or fixed cells (Fig. 3) (Hansen et al, 2003,Hansen et al, 2007. To do so, we infected spiral ganglion cultures with a lentiviral (feline immunodeficiency virus, FIV) vector expressing GFP (FIV-GFP).…”

Latanoprost Promotes Neurite Outgrowth in Differentiated RGC-5 Cells via the PI3K-Akt-mTOR Signaling Pathway

“…These results indicated that latanoprost promoted cell viability via the FP receptor. To investigate the neuroregenerative effect of latanoprost, the mean total neurite length (an average of all neurite outgrowth of neurons within each well) of differentiated RGC-5 cells was quantified after the application of 0.1 lM latanoprost for 24 h. Neurite Outgrowth HCS Reagent Hit Kit was widely used to assay the neurite outgrowth of variety neural cells (Hansen et al 2007;Radio et al 2008). By using the kit, cells were immunostained using the Hoechst dye and primary antibody with corresponding DyLight 488-conjugated secondary antibody.…”