The present study reports significant rescue of SGNs in the deaf rat cochlea following chronic exogenous delivery of BDNF compared with AP-treated or untreated DC cochleae. The most extensive rescue of SGNs was evident in the basal region of the cochlea, although significant cell rescue did occur throughout all turns of the BDNF treated cochleae. This trophic advantage also resulted in a significant increase in SGN soma area compared with AP-treated or untreated DC cochleae. This is the first time, to our knowledge, that in vivo SGN rescue has been demonstrated in the rat - all previous studies of chronic exogenous neurotrophin delivery in the cochlea have been conducted using the guinea pig.
We also examined the longitudinal response of SGNs to gentamicin/furosemide-induced deafness in the rat. The profile of the SGN loss was similar to that seen in studies of other species, however the rate of SGN degeneration appeared to be slower in the rat than that of other rodents such as guinea pigs in which ~50% SGN loss is evident four weeks following deafening via co-administration of an aminoglycoside and loop diuretic (14
Exogenous application of BDNF in the rat cochlea rescues SGNs
BDNF-treated cochleae exhibited total loss of hair cells in all turns, consistent with untreated deafened cochleae. However, unlike DC or deafened AP-treated cochleae, the BDNF-treated cohort showed few signs of SGN degeneration when compared with NH control cochleae. Indeed, there was no statistically significant difference in SGN density between BDNF-treated and NH controls throughout most of the cochleae (LBT, UBT and LMT). Moreover, BDNF treatment produced a significant trophic effect on SGNs in all cochlear turns compared to untreated DC and AP-treated cochleae.
A significant reduction in SGN density, evident in the UMT of the BDNF cohort compared with NH controls, presumably reflects a reduction in the concentration of BDNF towards the apex of the cochlea, as the delivery cannula was located in the LBT. However, we cannot rule out the possibility that in the mature cochlea, basal SGNs are more sensitive to BDNF rescue compared with neurons located in more apical regions of the cochlea.
BDNF-treated cochleae also exhibited SGNs with large soma areas. In fact, in the basal region of the cochlea their somata were significantly larger than those in NH control cochleae. More apicalward the soma area of BDNF treated cochleae were similar to normal controls. While this observation must be interpreted cautiously due to the smaller number of control (n=4) versus BDNF treated cochleae (n=6), such an increase in SGN soma area associated with exogenous BDNF treatment is consistent with similar results using guinea pigs (21
). Although the mechanisms underlying this increase in soma area remain unclear, studies of striatal projection neurons have also demonstrated that exogenous BDNF results in an increase in soma area when associated with neural rescue (22
The present finding, that BDNF exerts a trophic effect on rat SGNs, demonstrates that this effect is not restricted to the guinea pig. This gives us confidence that the trophic effects of BDNF may be evident across a number of mammalian species, and that the use of this neurotrophin may have clinical applications in the future.
Safety and Clinical Implications
Since a broad objective of work in our laboratory is to develop therapies that may benefit cochlear implant patients, it is important to consider the possible clinical applications of neurotrophin therapy. The availability and functionality of SGNs are important factors that govern the benefits that patients can derive from cochlear implants. The number and integrity of these cells are considered to be of importance for the successful restoration of hearing for speech recognition (23
). Therefore, any agent that can protect or regenerate SGNs would be of great clinical benefit.
Neurotrophins have been shown, by this study and others, to act as survival factors for mature SGNs in animal models of SNHL (12
). Other research has demonstrated that neurotrophins and electrical stimulation act in synergy to prevent SGN degeneration (25
). Taken together, this work suggests that neurotrophins may have clinical application when combined with cochlear implants. However, further research is required before neurotrophins can be used clinically. First, a number of in vitro
studies have demonstrated synergistic effects on both SGN rescue and neuritogenesis when several growth factors are combined in treatment (16
). In contrast, there is no evidence of a synergistic effect in the few in vivo
studies that have combined growth factors (12
). This discrepancy may be related to age and/or species effects, however, it is important that the full potential of combining growth factors is explored prior to any clinical application of neurotrophins for rescue of mature SGNs.
Second, the long-term safety and effectiveness of neurotrophins must be addressed. It has recently been shown that removal of neurotrophin treatment leads to an accelerated decline in SGN survival (14
). This suggests that neurotrophins may need to be delivered long-term in order to be effective in the cochlea. The long-term delivery of neurotrophins into the cochlea is a major safety issue, particularly when exogenous neurotrophins are delivered at concentrations significantly above endogenous levels, as would be expected with the use of pumps. Under some circumstances neurotrophins are known to be neurotoxic as a result, for example, of free-radical induced necrosis (28
). The strategy for application of neurotrophins in the inner ear is therefore a major issue that needs to be considered. The standard delivery technique in experimental studies is to cannulate the scala tympani and infuse via an osmotic pump (29
). While osmotic pumps are effective in these studies, their finite lifespan, the potential for introducing infection into the inner ear, and the delivery of neurotrophins at concentrations well above physiological levels, make them unsuitable for long-term clinical use. A number of alternative delivery methods, including the use of viral vectors (25
) and cell-based therapies (30
), are currently being investigated. These techniques have the potential to provide long-term delivery of neurotrophins in a clinical setting.
Finally, because the scala tympani is connected to the cerebrospinal fluid via the cochlear aqueduct, future studies will also need to investigate the long-term safety implications of exogenous neurotrophin delivery in both the cochlea and the central nervous system.
We have shown that BDNF induces significant rescue of SGNs in the deafened rat cochlea. This finding demonstrates that the neurotrophic advantage that has previously been described in the guinea pig is also evident in the rat, and provides an important step in the development of neurotrophin therapy as a clinical intervention for cochlear implant patients.