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Factors influencing the efficacy of round window dexamethasone protection of residual hearing post-cochlear implant surgery
Hearing Research 255 (2009) 67–72
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Research papers
Factors influencing the efficacy of round window dexamethasone protection of residual hearing post-cochlear implant surgery Andrew Chang a, Hayden Eastwood a, David Sly a, David James a, Rachael Richardson a,b, Stephen O’Leary a,b,* a b
University of Melbourne, Department of Otolaryngology, Royal Victorian Eye and Ear Hospital, 32 Gisborne St, East Melbourne, Victoria 3002, Australia Bionic Ear Institute, 384 Albert St., East Melbourne, Victoria 3002, Australia
a r t i c l e
i n f o
Article history: Received 12 December 2008 Received in revised form 1 May 2009 Accepted 18 May 2009 Available online 17 June 2009 Keywords: Cochlear implantation Round window membrane Dexamethasone Pharmacokinetic Residual hearing
a b s t r a c t Aim: To protect hearing in an experimental model of cochlear implantation by the application of dexamethasone to the round window prior to surgery. The present study examined the dosage and timing relationships required to optimise the hearing protection. Methods: Dexamethasone or saline (control) was absorbed into a pledget of the carboxymethylcellulose and hyaluronic acid and applied to the round window of the guinea pig prior to cochlear implantation. The treatment groups were 2% w/v dexamethasone for 30, 60 and 120 min; 20% dexamethasone applied for 30 min. Auditory sensitivity was determined pre-operatively, and at 1 week after surgery, with puretone auditory brainstem response audiometry (2–32 kHz). Cochlear implantation was performed via a cochleostomy drilled into the basal turn of the cochlea, into which a miniature cochlear implant dummy electrode was inserted using soft-surgery techniques. Results: ABR thresholds were elevated after cochlear implantation, maximally at 32 kHz and to a lesser extent at lower frequencies. Thresholds were less elevated after dexamethasone treatment, and the hearing protection improved when 2% dexamethasone was applied to the round window for longer periods of time prior to implantation. The time that dexamethasone need be applied to achieve hearing protection could be reduced by increasing the concentration of steroid, with a 20% application for 30 min achieving similar levels of protection to a 60 min application of 2% dexamethasone. Conclusions: Hearing protection is improved by increasing the time that dexamethasone is applied to the round window prior to cochlear implantation, and the waiting time can be reduced by increasing the steroid concentration. These results suggest that the diffusion dexamethasone through the cochlea is the prime determinant of the extent of hearing protection. Crown Copyright Ó 2009 Published by Elsevier B.V. All rights reserved.
1. Introduction Cochlear implant patients with residual (acoustic) hearing in the implanted ear can combine both electrical and acoustic stimulation (EAS) to improve speech perception, particularly in the presence of background noise (Gifford et al., 2007; James et al., 2006; Turner et al., 2008). Consequently, the preservation of hearing in the implanted ear has become a goal of cochlear implant surgery. But even with the introduction of minimally-traumatic ‘‘soft” surgical techniques (Lehnhardt, 1993) (Rogowski et al., 1995) and electrodes that have been modified to reduce intracochlear trauma during their insertion (Gantz and Turner, 2004; Lenarz et al., 2006), residual hearing is lost or incompletely preserved in a third of cases
* Corresponding author. Address: University of Melbourne, Department of Otolaryngology, Royal Victorian Eye and Ear Hospital, 32 Gisborne St, East Melbourne, Victoria 3002, Australia. E-mail address: [email protected] (S. O’Leary).
(Gstoettner et al., 2004, 2008). Some researchers have begun to explore the possibility that better hearing preservation may be achieved by the application of protective pharmacological agents to the inner ear at the time of surgery (Eshraghi et al., 2007; James et al., 2008; Ye et al., 2007), and here we examine the dose-time relations for hearing preservation when the glucocorticosteroid, dexamethasone is applied to the round window in an experimental model of cochlear implantation. Cochlear protection with steroids is presumably achieved primarily through suppression of the inflammatory response initiated by cochlear implant surgery (Cope and Bova, 2008), and possibly also through direct anti-apoptotic effects (Sasson and Amsterdam, 2003) on cochlear tissues. Clinically, residual hearing may be lost progressively over the first few days after cochlear implant surgery (Adunka et al., 2006). Over this period of time there will be progression of the inflammatory response initiated by the surgical trauma, and there is experimental evidence to suggest that the permanent hearing loss may be due in part to secondary oxidative
0378-5955/$ - see front matter Crown Copyright Ó 2009 Published by Elsevier B.V. All rights reserved. doi:10.1016/j.heares.2009.05.010
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A. Chang et al. / Hearing Research 255 (2009) 67–72
stress and apoptosis of hair cells (Eshraghi, 2006). In addition, cochlear inflammation has other effects known to be detrimental to hearing, such as disturbance of inner ear fluid homeostasis (Ichimiya et al., 2000; Maeda et al., 2005) and the promotion of fibrosis and/or osteoneogenesis (Li et al., 2007). That suppression of the cochlear inflammatory response, by the intra-operative application of steroids to the inner ear, reduces hearing loss has been demonstrated experimentally in models of cochlear implantation (Eshraghi et al., 2007; James et al., 2008; Ye et al., 2007). The authors have demonstrated that high frequency hearing can be protected in the guinea pig when a 2% solution of dexamethasone is applied to the round window on a polymeric sponge (SeprapackTM, Genzyme) 30 min prior to cochlear implantation. The round window was chosen as the route of delivery because it is accessible during surgery, and locally delivered steroids are known to achieve a higher intracochlear drug concentrations than with systemic administration. SeprapackTM (Genzyme, Cambridge, MA, USA), a carboxymethylcellulose hyaluronic acid polymer was chosen, as it allowed for an extended delivery of the drug. Dexamethasone delivered via a Seprapack sponge was as still detectable within the cochlea 24 h after the initial round window application(James et al., 2008) Local delivery also has the advantage of avoiding the potential risk of systemic complications associated with the parenteral administration of steroids, and there is a clinical precedent for its use in the treatment of sudden sensorineural hearing loss, intractable tinnitus and Meniere’s disease (Chandrasekhar, 2001; Dodson et al., 2004; Gouveris et al., 2005; Silverstein et al., 1996). In order to determine whether the delivery of steroids to the round window has the potential to be a useful strategy for hearing protection in the clinical setting, its duration of local delivery must be optimised. In our previous study with 2% dexamethasone applied for 30 min, protection was achieved in the lower basal turn of the cochlea, at the site of electrode insertion. However, the residual hearing to be protected in cochlear implant recipients is in the lower frequencies originating in the second and upper cochlear turns. Therefore, for this strategy to be clinically useful, round window delivery of steroids must be shown to protect lower frequency hearing. Better hearing protection might be achieved by either increasing the time that the steroid is applied to the round window prior to implantation, or by increasing the concentration of the drug applied to the round window. Here, we examine these dose-time relations, drawing upon theoretical models of cochlear pharmacokinetics to inform the experimental design (Salt, 2002). Such models are based upon drug diffusion, and predict the most efficient way of increasing the spatial extent over which the drug is distributed, is to increase the application time. Secondly, it is predicted that the duration required to achieve a therapeutic effect may be shortened (off-set) by increasing the concentration of the drug (Salt, 2002; Salt and Plontke, 2005). It is anticipated that the results from this study will assist in the translation of this approach to clinical application.
2. Methods
2.2. Materials Dexamethasone (Dexamethasone 21-Phosphate disodium salt, Sigma–Aldrich, USA) was made into solutions of 2%(w/v) and 20%(w/v) in sterile water and adsorbed into 2 mm discs of the carboxymethylcellulose hyaluronic acid polymer, Seprapack (Genzyme Biosurgery, Framingham), for 1 min prior to round window application. The non-stimulating electrodes used in these studies consisted of three platinum rings spaced 0.75 mm apart (centre-to-centre) on a Silastic carrier. The rings were used to help gauge the depth of insertion. The diameter at the tip of the electrode was 0.41 mm with a maximum shaft diameter of 0.43 mm. A 25 lm platinum wire was welded to the platinum rings to reinforce the electrode. The electrodes were ultrasonically cleaned and sterilised with ethylene oxide and sealed in a plastic envelope. 2.3. Experimental design Hearing guinea pigs were implanted with a dummy cochlear implant electrode, following application of the dexamethasone, or controls, to the round window. The main outcome measure was a frequency-specific change in auditory brainstem response (ABR) threshold 1 week after implantation. This timeline was chosen because hearing thresholds were stable after 1 week of observation in our previous study (James et al., 2008). A secondary outcome measure was the histological appearance of the cochlea at the end of the experiment. Animals were randomly allocated into saline control (n = 7) and treatment (dexamethasone) groups. Dexamethasone treated animals were further divided into groups receiving different durations of application of drug prior to implantation, [30 min (n = 4), 60 min (n = 6) and 120 min (n = 4) all with a dexamethasone concentration of 2%(w/v)] and animals receiving 20%(w/v) dexamethasone concentration for 30 min (n = 6). The prediction of intracochlear dexamethasone concentrations derived from Salt’s Cochlear Fluid Model is found in Table 1. Note that this model predicts a similar concentration of dexamethasone at the site of cochlear implantation (32 kHz region) when 2%(w/v) dexamethasone is applied for 60 min and when a 20%(w/v) solution is applied for 30 min. Therefore, similar levels of hearing protection may be predicted from both of these groups. 2.4. Auditory brainstem response recordings ABR recordings were taken from all guinea pigs to assess hearing across the experiment timeframe. Guinea pig ABR recordings were made immediately pre- and post-surgery and 7 days after surgery, immediately prior to perfusion of the animals and harvesting of cochleae for histology. Once anesthetised, the animal was placed inside an acoustically shielded room. To isolate the implanted ear acoustically, the contralateral ear was filled with Otoform-K2 (DLT, West Yorkshire), a vulcaning silicone. Computer-generated acoustic stimuli (5 ms tone pips with 1 ms rise/fall times) were delivered via a
2.1. Animals Thirty-four adult, normal hearing guinea pigs (Dunkin–Hartley strain), weighing between 750 and 900 g were used in this project. The project was approved by the Royal Victorian Eye and Ear Hospital Animal Ethics Committee (Ethics Approval #07/140A). Both intramuscular and inhalation anaesthesia were used during auditory brainstem response (ABR) recordings and during surgery. The intramuscular agents were 4 mg/kg ketamine and 60 mg/kg xylazine. The inhalation anaesthesia, ****isoflourane, was administered with oxygen at a concentration of 0.5–1% and at a rate of 500 ml/ min. Respiration rate was used to monitor the depth of anaesthesia.
Table 1 The predicted concentration of dexamethasone (mM) present in the perilymph at various frequencies along the cochlea at the time of surgery. The concentrations were calculated using the Washington University Cochlear fluid Simulator V1.6 with round window delivery and no middle ear clearance.
32 kHz 24 kHz 16 kHz 8 kHz 2 kHz
2% Dex 30 min
2% Dex 60 min
2% Dex 120 min
2% Dex 30 min
0.224 0.028 0.001 >.001 >.001
2.360 0.803 0.132 0.002 >.001
8.125 4.763 1.895 0.251 0.001
2.239 0.276 0.009 >.001 >.001
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loudspeaker (Richard Allen DT-20, UK) placed 10 cm from the implanted ear in line with the external auditory meatus. These acoustic stimuli were delivered at frequencies of 2, 8, 16, 24 and 32 kHz. Stimulus intensity was increased in 5 dB steps from subthreshold levels. The recordings were made from hypodermic needle electrodes inserted through the skin at the vertex and the nape, with a ground electrode inserted through the skin of the hindquarters in the midline. The recordings were amplified by a factor of 100,000 (DAM-5A, W-P Instruments Inc., New Haven, Conn, USA) and band-pass filtered (Krohn-Hite 3750, Avon, Mass., USA) between 150 Hz and 3 kHz (6db/Octave). The output signals were sent to a 16-bit analogue to digital converter (Tucker Davis Technologies, USA). These signals were sampled at 20 kHz for 10 ms following the stimulus onset and were averaged over 250 stimulus repetitions. The recorded waveforms were analysed using custom program (software program written by Dr. James Fallon, adapted by Prof. Stephen O’Leary) in Igor Pro 5.02 (Wavemetrics Inc.), with the researcher blind to the treatment group. Input–output curves were generated from data analysis and threshold was defined as the lowest intensity stimulus which evoked a response to wave III greater than 0.4 lV in amplitude. 2.5. Cochlear implantation A dorsolateral, posterior auricular approach was used to gain access to the bulla and the cochlea. The surgical field behind the pinna was shaved and the skin was washed with BetadineÒ (Mundipharma B.V.). All surgeries were performed on the left cochlea. A ‘‘soft” surgical technique similar to human implantation surgery was used to reduce surgical trauma (Rogowski et al., 1995). At the time points studied (30, 60 and 120 min) before cochleostomy a bead of SeprapakTM soaked in either Dexamethsone or saline, was place on the round window. This bead was left in situ after the operation and allowed to dissolve naturally. After the indicated time, a cochleostomy was made with a 0.7 mm diameter drill over the basal turn at a low speed to prevent excessive heat building from the drill tip. There was no aspiration of perilymph from the cochleostomy. The electrode was inserted with an atraumatic technique to the first point of resistance; this corresponded to the third platinum ring which gives a total insertion length of 1.75 mm. The cochleostomy was plugged with fascia to prevent perilymph leak. The extra-cochlear portion of the electrode was secured into muscle with sutures to prevent extrusion of the electrode. 2.6. Histology Intracardiac perfusion was performed with heparinized normal saline followed by 10% neutral buffered formalin. The cochleae were removed and decalcified in 4%(w/v) ethylenediaminetetraacetic acid (EDTA). Cochleae were subsequently trimmed, oriented in agar, and then embedded in paraffin. Cochleae were cut at 5 lm and mid-modiolar sections were mounted on slides and stained with haematoxylin and eosin. The final specimens were analysed by an experienced histologist in a blinded manner, for evidence of acute inflammatory responses. 2.7. Data analysis For statistical analysis the ABR thresholds were grouped into lower basal (24 and 32 kHz), upper basal (8 and 16 kz) and second turn (2 kHz) regions to increase the sensitivity of the analyses. The difference between pre and post-cochlear implant (at 1 week) ABR thresholds (threshold shift) were compared using One-Way Analysis of Variance with threshold-shift as the independent variable and either duration of pre-implant treatment, or drug concentration as
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the dependent variable. Pair wise multiple comparisons (Holm–Sidak method) were performed when appropriate. All calculations were performed using Sigmastat, version 3.2 (Systat Software Inc.) 3. Results ABR thresholds are reported 1 week after cochlear implantation, following a partial recovery of thresholds averaging 10 dB that occurred over the first 7 days following surgery. This time point was chosen because our previous studies have demonstrated that threshold shifts are stable after 1 week (James et al., 2008) with no further changes to threshold shifts up to 3 months post implantation (unpublished data). In control animals (‘‘Control” in Fig. 1), threshold shifts were greatest in the lower basal turn (24 and 32 kHz), near where the electrode is inserted, and at 16 kHz which corresponds with a cochlear region in the upper basal turn, just apical to the tip of the electrode. The threshold shift at 2 kHz (second turn) was <5 dB for all groups. These findings are consistent with observations made previously (James et al., 2008) with this experimental model of cochlear implantation. Increasing the time that dexamethasone was applied to the round window prior to implantation significantly reduced the threshold shifts in the lower basal turn (Fig. 1, 24 and 32 kHz) when compared to saline controls (DF = 3, SS = 1940.774, MS = 646.925, F = 6.884, p < 0.001). On subgroup analysis (with multiple pair wise comparisons) threshold shifts were found to be significantly smaller after 60 and 120 min treatment with 2% dexamethasone groups compared to controls (Diff of Means 13.274, t = 3.481, p < 0.05 and Diff of Means 16.607, t = 3.865, p < 0.05, respectively). In the upper basal region the reduction in threshold shift was modest after a 60 min application of 2% dexamethasone, but marked after 120 min (120 min vs. Control, t = 2.993, p = 0.032) when the protection was complete. These results are consistent with there being a ‘‘threshold” period of pre-treatment (of 120 min) required in order to protect hearing in the upper basal region. In the second turn there was no significant difference between the threshold shifts of dexamethasone-treated animals and controls (DF = 2, SS = 42.262, MS = 14.087, F = 0.244, p = 0.864). When a higher concentration of dexamethasone (20%) was applied to the round window for 30 min, threshold shifts in the lower basal turn were smaller than in controls (DF = 2, SS = 1315.821, MS = 657.910, F = 8.792, p < 0.001). On subgroup analysis (using pair wise comparisons), both 2% dexamethasone applied for 60 min and the 20% dexamethasone applied for 30 min led to a significant reduction in threshold shift when compared to controls (60 min 2%: Diff of Means 13.274, t = 3.901, p < 0.025 and 30 min 20%: Diff of Means 10.774, t = 3.166, p < 0.05). Drug concentration in the 32 kHz region of the cochlea was predicted by the model to be similar in each group (grey bars on Fig. 1) and the threshold shifts were similar. Twelve cochleae were selected at random for histological analysis; 8 dexamethasone-treated and 4 controls. The cochleae were examined for evidence of inflammation and fibrosis (James et al., 2008). Minimal fibrosis was present along the osseous spiral lamina, basilar membrane and upper outer wall of the scala tympani in some animals. The amount of tissue response was insufficient to quantify using our cochlear tissue response scale (James et al., 2008), and not extensive enough to distinguish between the treatment and control groups. 4. Discussion The application of dexamethasone to the round window prior to cochlear implant surgery can protect hearing across the frequency
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Fig. 1. The mean threshold shifts (black diamonds ± SEM) for all groups at the indicated frequencies 1 week post implantation. The grey bars indicate the predicted concentration of dexamethasone present in the perilymph at the time of implantation. All dexamethasone groups displayed a level of protection at 32 kHz, 1 week post implantation compared to control and had predicted concentrations dexamethasone greater than 0.2 mM at the time of implantation. At 16 and 24 kHz, all dexamethasone cohorts except the 2% 30 min dexamethasone still displayed a level of protection compared to the control and these three cohort still had predicted dexamethasone concentrations greater that 0.2 mM at 24 kHz. At frequencies below 16 kHz there was very little threshold shift occurring in the saline controls or dexamethasone cohorts. At 2 kHz the highest concentration of dexamethasone predicted was 0.001 mM in the 2% 120 min cohort.
range. Protection improves with longer application times, and the time required to achieve a therapeutic effect can be shortened by increasing the drug dose. These results demonstrate that round
window steroid delivery has the potential for protecting the residual hearing of cochlear implant patients. There was a good correspondence between the University of Washington’s Cochlear Fluid Simulator (Dr. Alec Salt) predictions of dexamethasone distribution within the cochlea and the extent of hearing protection. The simulation predicted higher drug levels and more extensive intracochlear drug distribution by increasing the time that dexamethasone was applied to the round window, and this corresponded experimentally with hearing protection over a more extended region of the cochlea. The simulation also predicted that the concentration of dexamethasone in the basal turn of the cochlea (at the site of implantation) would be equivalent for both a 60 min application of 2% dexamethasone and a 30 min application of 20% dexamethasone, and this corresponded experimentally with similar levels of hearing protection observed at 32KHz for both treatments. However, this interpretation may be confounded by the knowledge that the higher concentration of dexamethasone may change the round window permeability (Goycoolea, 2001; Hahn et al., 2006; Plontke et al., 2007), and this has not been controlled for in the present studies. These results demonstrate the utility of modelling in helping to design therapeutic strategies, but there are limitations to this approach. Foremost is the lack of a method for relating directly the simulated cochlear drug concentrations and the hearing protection. This would require knowledge of the perilymphatic steroid concentration required to protect the organ of Corti (and possibly the lateral cochlear structures) in vivo. The latter could potentially be subject to a cochleotopic gradient because the susceptibility of hair cells to injury is greater in the basal than the apical regions of the cochlea (Wu et al., 2002). Until these matters are better understood, simulation is likely to be to provide qualitative, rather than quantitative information to guide the development of therapeutic strategies. But these limitations aside, simulation provides the best method available to guide the clinical translation of the experimental results to a protective therapy for cochlear implantation, and to help account for inter-species differences such as the cochlear size and shape. Consistent with our previous observations (James et al., 2008), hearing loss was maximal at the site of cochleostomy and electrode array insertion, but also present at lower frequencies originating at cochlear sites apical to the site of surgery. The hearing loss at the site of surgery may relate to direct cochlear trauma, and it has been speculated that oxidative stress may contribute to the higher-frequency hearing loss. Dexamethasone protected against hearing loss across both frequency domains. Other studies have shown a similar trend in hearing protection after electrode insertion and withdrawal followed by continuous administration of dexamethasone for 1 month with a mini-osmotic pump (Eshraghi et al., 2007; Ye et al., 2004, 2007). The main therapeutic action of a steroid in the context of injury prevention is to suppress the post-operative an inflammatory insult (Cope and Bova, 2008). The present experiment supports this interpretation. There is histological evidence to suggest that a single application of dexamethasone to the round window may reduce the foreign body reaction to the cochlear implant (James et al., 2008). The absence of inflammatory reaction the present study is a contrary finding, and could potentially be due to operator-dependent factors, but equally it might be a consequence of the shorter period of observation in this study (1 week) compared with the previous investigation (1 month). The one-week period of study reported here is insufficient time for the development of a mature fibrous reaction to the implant. Therefore, a longer period of observation may be required to examine the fibrous tissue reaction. The duration of treatment required for optimal hearing protection could present practical challenges for the clinical translation of the experimental method presented here. The round window application times required to achieve protection (1–2 h) are very
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long compared with the duration of cochlear implant surgery, which is typically 90–120 min. It seems likely that surgeons would find the extended time of surgery required to achieve optimal hearing protection impractical. It must also be acknowledged that even longer waiting times might be required clinically than we required experimentally, because the human cochlea has a larger volume than that of the guinea pig. Given this possibility it is worthwhile reviewing the relative strengths of limitations of the present and alternative methods for cochlear drug delivery. The use of SeprapackTM, as a delivery vehicle applied to the round window had some advantages over intra-tympanic administration. The SeprapackTM can be positioned over the round window membrane; therefore, there is no loss of dexamethasone within the middle ear space, and minimal risk of absorption of steroid directly through the otic capsule into the cochlea. Whilst, the SeprapackTM is on the round window before the surgery, it retains the dexamethasone and permits a slow release after the surgery. While the release profile has not yet been fully characterized, previous work in our laboratories has shown dexamethasone and its breakdown products for a period of no more than 24 h as estimated by perilymph sampling in vivo (James et al., 2008). Other local delivery strategies are currently under investigation, but all have potential short-comings that await further study (Borenstein et al., 2007; Chen et al., 2005; Kopke et al., 2006). These methods include drug-eluting electrode array, use of microinjection and micro-osmotic pump (Borkholder, 2008). The drug-eluting electrode array may provide dexamethasone to the area of trauma but its efficacy may be limited because the dexamethasone is not delivered before surgery (Mathivanar et al., 1990; Mercanzini et al., 2007). The use of microinjection may itself result in trauma due to an increase in the hydraulic pressure from the injection, unless it is done in a controlled fashion and the latter is likely to be difficult to achieve reproducibly in surgery. Micro-fluidic systems have been developed which both deliver, and simultaneously remove quantities of fluid in order to reduce the hydraulic effects on the cochlea (Chen et al., 2005), and these systems can increase the concentration in the basal turn at a greater rate than diffusion alone. However, these may not be necessary for dexamethasone because it diffuses rapidly through the RW (noting that RW is quite a large surface area compared with a micro-fluidic channel). Micro-osmotic pumps can provide medium to long term infusion of dexamethasone, however, these may require additional surgical procedures and increase the risk of cochlear infection (Pettingill et al., 2008). The current hearing protection strategy is applicable clinically, but the parameters for clinical drug delivery need to account for the differences in cochlear size, and the practical constraints of surgical practice. To overcome the potentially long periods of application that may be required for dexamethasone to diffuse through the larger human cochlea, we speculate that a combined approach (systemic and local) may be required to ensure that sufficient drug concentration is achieved in the cochlea prior to surgery. Acknowledgements Project grants from the National Health and Medical Research Council Australia and Otolaryngology Scholarship from the Garnett Passe and Rodney Williams Memorial Foundation. Helen Feng for manufacturing electrode arrays. Maria Clark and Prudence Nielsen for preparing histological materials. Dr. Stuart Galloway and St. Vincent’s Hospital Anatomical Pathology Department for histological analysis. Dr. James Fallon for providing the ABR analysis program (NIDCD Contract HHS-N-263-2007-00053-C; PI: RK Shepherd). Salt A, Washington University Cochlear Fluids Simulator: Version 1.6 available from http://oto.wustl.edu/cochlea/model.htm. (This program was developed and is made available by Grant funding (DC 01368) from the National Institute on Deafness
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Salt, A.N., 2002. Simulation of methods for drug delivery to the cochlear fluids. Adv. Otorhinolaryngol. 59, 140–148. Salt, A.N., Plontke, S.K., 2005. Local inner-ear drug delivery and pharmacokinetics. Drug Discov. Today 10, 1299–1306. Sasson, R., Amsterdam, A., 2003. Pleiotropic anti-apoptotic activity of glucocorticoids in ovarian follicular cells. Biochem. Pharmacol. 66, 1393– 1401. Silverstein, H., Choo, D., Rosenberg, S.I., Kuhn, J., Seidman, M., Stein, I. 1996. Intratympanic steroid treatment of inner ear disease and tinnitus (preliminary report). Ear Nose Throat J. 75, 468–471, 474, 476 passim.
Turner, C.W., Reiss, L.A., Gantz, B.J., 2008. Combined acoustic and electric hearing: preserving residual acoustic hearing. Hear. Res. 242, 164–171. Wu, W.J., Sha, S.H., Schacht, J., 2002. Recent advances in understanding aminoglycoside ototoxicity and its prevention. Audiol. Neurootol. 7, 171–174. Ye, Q., Kiefer, J., Tillein, J., Klinke, R., Gstoettner, W., 2004. Intracochlear application of steroids: an experimental study in guinea pigs. Cochlear Implants Int. 5 (Suppl. 1), 17–18. Ye, Q., Tillein, J., Hartmann, R., Gstoettner, W., Kiefer, J., 2007. Application of a corticosteroid (Triamcinolon) protects inner ear function after surgical intervention. Ear Hear. 28, 361–369.
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Research papers
Factors influencing the efficacy of round window dexamethasone protection of residual hearing post-cochlear implant surgery Andrew Chang a, Hayden Eastwood a, David Sly a, David James a, Rachael Richardson a,b, Stephen O’Leary a,b,* a b
University of Melbourne, Department of Otolaryngology, Royal Victorian Eye and Ear Hospital, 32 Gisborne St, East Melbourne, Victoria 3002, Australia Bionic Ear Institute, 384 Albert St., East Melbourne, Victoria 3002, Australia
a r t i c l e
i n f o
Article history: Received 12 December 2008 Received in revised form 1 May 2009 Accepted 18 May 2009 Available online 17 June 2009 Keywords: Cochlear implantation Round window membrane Dexamethasone Pharmacokinetic Residual hearing
a b s t r a c t Aim: To protect hearing in an experimental model of cochlear implantation by the application of dexamethasone to the round window prior to surgery. The present study examined the dosage and timing relationships required to optimise the hearing protection. Methods: Dexamethasone or saline (control) was absorbed into a pledget of the carboxymethylcellulose and hyaluronic acid and applied to the round window of the guinea pig prior to cochlear implantation. The treatment groups were 2% w/v dexamethasone for 30, 60 and 120 min; 20% dexamethasone applied for 30 min. Auditory sensitivity was determined pre-operatively, and at 1 week after surgery, with puretone auditory brainstem response audiometry (2–32 kHz). Cochlear implantation was performed via a cochleostomy drilled into the basal turn of the cochlea, into which a miniature cochlear implant dummy electrode was inserted using soft-surgery techniques. Results: ABR thresholds were elevated after cochlear implantation, maximally at 32 kHz and to a lesser extent at lower frequencies. Thresholds were less elevated after dexamethasone treatment, and the hearing protection improved when 2% dexamethasone was applied to the round window for longer periods of time prior to implantation. The time that dexamethasone need be applied to achieve hearing protection could be reduced by increasing the concentration of steroid, with a 20% application for 30 min achieving similar levels of protection to a 60 min application of 2% dexamethasone. Conclusions: Hearing protection is improved by increasing the time that dexamethasone is applied to the round window prior to cochlear implantation, and the waiting time can be reduced by increasing the steroid concentration. These results suggest that the diffusion dexamethasone through the cochlea is the prime determinant of the extent of hearing protection. Crown Copyright Ó 2009 Published by Elsevier B.V. All rights reserved.
1. Introduction Cochlear implant patients with residual (acoustic) hearing in the implanted ear can combine both electrical and acoustic stimulation (EAS) to improve speech perception, particularly in the presence of background noise (Gifford et al., 2007; James et al., 2006; Turner et al., 2008). Consequently, the preservation of hearing in the implanted ear has become a goal of cochlear implant surgery. But even with the introduction of minimally-traumatic ‘‘soft” surgical techniques (Lehnhardt, 1993) (Rogowski et al., 1995) and electrodes that have been modified to reduce intracochlear trauma during their insertion (Gantz and Turner, 2004; Lenarz et al., 2006), residual hearing is lost or incompletely preserved in a third of cases
* Corresponding author. Address: University of Melbourne, Department of Otolaryngology, Royal Victorian Eye and Ear Hospital, 32 Gisborne St, East Melbourne, Victoria 3002, Australia. E-mail address: [email protected] (S. O’Leary).
(Gstoettner et al., 2004, 2008). Some researchers have begun to explore the possibility that better hearing preservation may be achieved by the application of protective pharmacological agents to the inner ear at the time of surgery (Eshraghi et al., 2007; James et al., 2008; Ye et al., 2007), and here we examine the dose-time relations for hearing preservation when the glucocorticosteroid, dexamethasone is applied to the round window in an experimental model of cochlear implantation. Cochlear protection with steroids is presumably achieved primarily through suppression of the inflammatory response initiated by cochlear implant surgery (Cope and Bova, 2008), and possibly also through direct anti-apoptotic effects (Sasson and Amsterdam, 2003) on cochlear tissues. Clinically, residual hearing may be lost progressively over the first few days after cochlear implant surgery (Adunka et al., 2006). Over this period of time there will be progression of the inflammatory response initiated by the surgical trauma, and there is experimental evidence to suggest that the permanent hearing loss may be due in part to secondary oxidative
0378-5955/$ - see front matter Crown Copyright Ó 2009 Published by Elsevier B.V. All rights reserved. doi:10.1016/j.heares.2009.05.010
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stress and apoptosis of hair cells (Eshraghi, 2006). In addition, cochlear inflammation has other effects known to be detrimental to hearing, such as disturbance of inner ear fluid homeostasis (Ichimiya et al., 2000; Maeda et al., 2005) and the promotion of fibrosis and/or osteoneogenesis (Li et al., 2007). That suppression of the cochlear inflammatory response, by the intra-operative application of steroids to the inner ear, reduces hearing loss has been demonstrated experimentally in models of cochlear implantation (Eshraghi et al., 2007; James et al., 2008; Ye et al., 2007). The authors have demonstrated that high frequency hearing can be protected in the guinea pig when a 2% solution of dexamethasone is applied to the round window on a polymeric sponge (SeprapackTM, Genzyme) 30 min prior to cochlear implantation. The round window was chosen as the route of delivery because it is accessible during surgery, and locally delivered steroids are known to achieve a higher intracochlear drug concentrations than with systemic administration. SeprapackTM (Genzyme, Cambridge, MA, USA), a carboxymethylcellulose hyaluronic acid polymer was chosen, as it allowed for an extended delivery of the drug. Dexamethasone delivered via a Seprapack sponge was as still detectable within the cochlea 24 h after the initial round window application(James et al., 2008) Local delivery also has the advantage of avoiding the potential risk of systemic complications associated with the parenteral administration of steroids, and there is a clinical precedent for its use in the treatment of sudden sensorineural hearing loss, intractable tinnitus and Meniere’s disease (Chandrasekhar, 2001; Dodson et al., 2004; Gouveris et al., 2005; Silverstein et al., 1996). In order to determine whether the delivery of steroids to the round window has the potential to be a useful strategy for hearing protection in the clinical setting, its duration of local delivery must be optimised. In our previous study with 2% dexamethasone applied for 30 min, protection was achieved in the lower basal turn of the cochlea, at the site of electrode insertion. However, the residual hearing to be protected in cochlear implant recipients is in the lower frequencies originating in the second and upper cochlear turns. Therefore, for this strategy to be clinically useful, round window delivery of steroids must be shown to protect lower frequency hearing. Better hearing protection might be achieved by either increasing the time that the steroid is applied to the round window prior to implantation, or by increasing the concentration of the drug applied to the round window. Here, we examine these dose-time relations, drawing upon theoretical models of cochlear pharmacokinetics to inform the experimental design (Salt, 2002). Such models are based upon drug diffusion, and predict the most efficient way of increasing the spatial extent over which the drug is distributed, is to increase the application time. Secondly, it is predicted that the duration required to achieve a therapeutic effect may be shortened (off-set) by increasing the concentration of the drug (Salt, 2002; Salt and Plontke, 2005). It is anticipated that the results from this study will assist in the translation of this approach to clinical application.
2. Methods
2.2. Materials Dexamethasone (Dexamethasone 21-Phosphate disodium salt, Sigma–Aldrich, USA) was made into solutions of 2%(w/v) and 20%(w/v) in sterile water and adsorbed into 2 mm discs of the carboxymethylcellulose hyaluronic acid polymer, Seprapack (Genzyme Biosurgery, Framingham), for 1 min prior to round window application. The non-stimulating electrodes used in these studies consisted of three platinum rings spaced 0.75 mm apart (centre-to-centre) on a Silastic carrier. The rings were used to help gauge the depth of insertion. The diameter at the tip of the electrode was 0.41 mm with a maximum shaft diameter of 0.43 mm. A 25 lm platinum wire was welded to the platinum rings to reinforce the electrode. The electrodes were ultrasonically cleaned and sterilised with ethylene oxide and sealed in a plastic envelope. 2.3. Experimental design Hearing guinea pigs were implanted with a dummy cochlear implant electrode, following application of the dexamethasone, or controls, to the round window. The main outcome measure was a frequency-specific change in auditory brainstem response (ABR) threshold 1 week after implantation. This timeline was chosen because hearing thresholds were stable after 1 week of observation in our previous study (James et al., 2008). A secondary outcome measure was the histological appearance of the cochlea at the end of the experiment. Animals were randomly allocated into saline control (n = 7) and treatment (dexamethasone) groups. Dexamethasone treated animals were further divided into groups receiving different durations of application of drug prior to implantation, [30 min (n = 4), 60 min (n = 6) and 120 min (n = 4) all with a dexamethasone concentration of 2%(w/v)] and animals receiving 20%(w/v) dexamethasone concentration for 30 min (n = 6). The prediction of intracochlear dexamethasone concentrations derived from Salt’s Cochlear Fluid Model is found in Table 1. Note that this model predicts a similar concentration of dexamethasone at the site of cochlear implantation (32 kHz region) when 2%(w/v) dexamethasone is applied for 60 min and when a 20%(w/v) solution is applied for 30 min. Therefore, similar levels of hearing protection may be predicted from both of these groups. 2.4. Auditory brainstem response recordings ABR recordings were taken from all guinea pigs to assess hearing across the experiment timeframe. Guinea pig ABR recordings were made immediately pre- and post-surgery and 7 days after surgery, immediately prior to perfusion of the animals and harvesting of cochleae for histology. Once anesthetised, the animal was placed inside an acoustically shielded room. To isolate the implanted ear acoustically, the contralateral ear was filled with Otoform-K2 (DLT, West Yorkshire), a vulcaning silicone. Computer-generated acoustic stimuli (5 ms tone pips with 1 ms rise/fall times) were delivered via a
2.1. Animals Thirty-four adult, normal hearing guinea pigs (Dunkin–Hartley strain), weighing between 750 and 900 g were used in this project. The project was approved by the Royal Victorian Eye and Ear Hospital Animal Ethics Committee (Ethics Approval #07/140A). Both intramuscular and inhalation anaesthesia were used during auditory brainstem response (ABR) recordings and during surgery. The intramuscular agents were 4 mg/kg ketamine and 60 mg/kg xylazine. The inhalation anaesthesia, ****isoflourane, was administered with oxygen at a concentration of 0.5–1% and at a rate of 500 ml/ min. Respiration rate was used to monitor the depth of anaesthesia.
Table 1 The predicted concentration of dexamethasone (mM) present in the perilymph at various frequencies along the cochlea at the time of surgery. The concentrations were calculated using the Washington University Cochlear fluid Simulator V1.6 with round window delivery and no middle ear clearance.
32 kHz 24 kHz 16 kHz 8 kHz 2 kHz
2% Dex 30 min
2% Dex 60 min
2% Dex 120 min
2% Dex 30 min
0.224 0.028 0.001 >.001 >.001
2.360 0.803 0.132 0.002 >.001
8.125 4.763 1.895 0.251 0.001
2.239 0.276 0.009 >.001 >.001
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loudspeaker (Richard Allen DT-20, UK) placed 10 cm from the implanted ear in line with the external auditory meatus. These acoustic stimuli were delivered at frequencies of 2, 8, 16, 24 and 32 kHz. Stimulus intensity was increased in 5 dB steps from subthreshold levels. The recordings were made from hypodermic needle electrodes inserted through the skin at the vertex and the nape, with a ground electrode inserted through the skin of the hindquarters in the midline. The recordings were amplified by a factor of 100,000 (DAM-5A, W-P Instruments Inc., New Haven, Conn, USA) and band-pass filtered (Krohn-Hite 3750, Avon, Mass., USA) between 150 Hz and 3 kHz (6db/Octave). The output signals were sent to a 16-bit analogue to digital converter (Tucker Davis Technologies, USA). These signals were sampled at 20 kHz for 10 ms following the stimulus onset and were averaged over 250 stimulus repetitions. The recorded waveforms were analysed using custom program (software program written by Dr. James Fallon, adapted by Prof. Stephen O’Leary) in Igor Pro 5.02 (Wavemetrics Inc.), with the researcher blind to the treatment group. Input–output curves were generated from data analysis and threshold was defined as the lowest intensity stimulus which evoked a response to wave III greater than 0.4 lV in amplitude. 2.5. Cochlear implantation A dorsolateral, posterior auricular approach was used to gain access to the bulla and the cochlea. The surgical field behind the pinna was shaved and the skin was washed with BetadineÒ (Mundipharma B.V.). All surgeries were performed on the left cochlea. A ‘‘soft” surgical technique similar to human implantation surgery was used to reduce surgical trauma (Rogowski et al., 1995). At the time points studied (30, 60 and 120 min) before cochleostomy a bead of SeprapakTM soaked in either Dexamethsone or saline, was place on the round window. This bead was left in situ after the operation and allowed to dissolve naturally. After the indicated time, a cochleostomy was made with a 0.7 mm diameter drill over the basal turn at a low speed to prevent excessive heat building from the drill tip. There was no aspiration of perilymph from the cochleostomy. The electrode was inserted with an atraumatic technique to the first point of resistance; this corresponded to the third platinum ring which gives a total insertion length of 1.75 mm. The cochleostomy was plugged with fascia to prevent perilymph leak. The extra-cochlear portion of the electrode was secured into muscle with sutures to prevent extrusion of the electrode. 2.6. Histology Intracardiac perfusion was performed with heparinized normal saline followed by 10% neutral buffered formalin. The cochleae were removed and decalcified in 4%(w/v) ethylenediaminetetraacetic acid (EDTA). Cochleae were subsequently trimmed, oriented in agar, and then embedded in paraffin. Cochleae were cut at 5 lm and mid-modiolar sections were mounted on slides and stained with haematoxylin and eosin. The final specimens were analysed by an experienced histologist in a blinded manner, for evidence of acute inflammatory responses. 2.7. Data analysis For statistical analysis the ABR thresholds were grouped into lower basal (24 and 32 kHz), upper basal (8 and 16 kz) and second turn (2 kHz) regions to increase the sensitivity of the analyses. The difference between pre and post-cochlear implant (at 1 week) ABR thresholds (threshold shift) were compared using One-Way Analysis of Variance with threshold-shift as the independent variable and either duration of pre-implant treatment, or drug concentration as
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the dependent variable. Pair wise multiple comparisons (Holm–Sidak method) were performed when appropriate. All calculations were performed using Sigmastat, version 3.2 (Systat Software Inc.) 3. Results ABR thresholds are reported 1 week after cochlear implantation, following a partial recovery of thresholds averaging 10 dB that occurred over the first 7 days following surgery. This time point was chosen because our previous studies have demonstrated that threshold shifts are stable after 1 week (James et al., 2008) with no further changes to threshold shifts up to 3 months post implantation (unpublished data). In control animals (‘‘Control” in Fig. 1), threshold shifts were greatest in the lower basal turn (24 and 32 kHz), near where the electrode is inserted, and at 16 kHz which corresponds with a cochlear region in the upper basal turn, just apical to the tip of the electrode. The threshold shift at 2 kHz (second turn) was <5 dB for all groups. These findings are consistent with observations made previously (James et al., 2008) with this experimental model of cochlear implantation. Increasing the time that dexamethasone was applied to the round window prior to implantation significantly reduced the threshold shifts in the lower basal turn (Fig. 1, 24 and 32 kHz) when compared to saline controls (DF = 3, SS = 1940.774, MS = 646.925, F = 6.884, p < 0.001). On subgroup analysis (with multiple pair wise comparisons) threshold shifts were found to be significantly smaller after 60 and 120 min treatment with 2% dexamethasone groups compared to controls (Diff of Means 13.274, t = 3.481, p < 0.05 and Diff of Means 16.607, t = 3.865, p < 0.05, respectively). In the upper basal region the reduction in threshold shift was modest after a 60 min application of 2% dexamethasone, but marked after 120 min (120 min vs. Control, t = 2.993, p = 0.032) when the protection was complete. These results are consistent with there being a ‘‘threshold” period of pre-treatment (of 120 min) required in order to protect hearing in the upper basal region. In the second turn there was no significant difference between the threshold shifts of dexamethasone-treated animals and controls (DF = 2, SS = 42.262, MS = 14.087, F = 0.244, p = 0.864). When a higher concentration of dexamethasone (20%) was applied to the round window for 30 min, threshold shifts in the lower basal turn were smaller than in controls (DF = 2, SS = 1315.821, MS = 657.910, F = 8.792, p < 0.001). On subgroup analysis (using pair wise comparisons), both 2% dexamethasone applied for 60 min and the 20% dexamethasone applied for 30 min led to a significant reduction in threshold shift when compared to controls (60 min 2%: Diff of Means 13.274, t = 3.901, p < 0.025 and 30 min 20%: Diff of Means 10.774, t = 3.166, p < 0.05). Drug concentration in the 32 kHz region of the cochlea was predicted by the model to be similar in each group (grey bars on Fig. 1) and the threshold shifts were similar. Twelve cochleae were selected at random for histological analysis; 8 dexamethasone-treated and 4 controls. The cochleae were examined for evidence of inflammation and fibrosis (James et al., 2008). Minimal fibrosis was present along the osseous spiral lamina, basilar membrane and upper outer wall of the scala tympani in some animals. The amount of tissue response was insufficient to quantify using our cochlear tissue response scale (James et al., 2008), and not extensive enough to distinguish between the treatment and control groups. 4. Discussion The application of dexamethasone to the round window prior to cochlear implant surgery can protect hearing across the frequency
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Fig. 1. The mean threshold shifts (black diamonds ± SEM) for all groups at the indicated frequencies 1 week post implantation. The grey bars indicate the predicted concentration of dexamethasone present in the perilymph at the time of implantation. All dexamethasone groups displayed a level of protection at 32 kHz, 1 week post implantation compared to control and had predicted concentrations dexamethasone greater than 0.2 mM at the time of implantation. At 16 and 24 kHz, all dexamethasone cohorts except the 2% 30 min dexamethasone still displayed a level of protection compared to the control and these three cohort still had predicted dexamethasone concentrations greater that 0.2 mM at 24 kHz. At frequencies below 16 kHz there was very little threshold shift occurring in the saline controls or dexamethasone cohorts. At 2 kHz the highest concentration of dexamethasone predicted was 0.001 mM in the 2% 120 min cohort.
range. Protection improves with longer application times, and the time required to achieve a therapeutic effect can be shortened by increasing the drug dose. These results demonstrate that round
window steroid delivery has the potential for protecting the residual hearing of cochlear implant patients. There was a good correspondence between the University of Washington’s Cochlear Fluid Simulator (Dr. Alec Salt) predictions of dexamethasone distribution within the cochlea and the extent of hearing protection. The simulation predicted higher drug levels and more extensive intracochlear drug distribution by increasing the time that dexamethasone was applied to the round window, and this corresponded experimentally with hearing protection over a more extended region of the cochlea. The simulation also predicted that the concentration of dexamethasone in the basal turn of the cochlea (at the site of implantation) would be equivalent for both a 60 min application of 2% dexamethasone and a 30 min application of 20% dexamethasone, and this corresponded experimentally with similar levels of hearing protection observed at 32KHz for both treatments. However, this interpretation may be confounded by the knowledge that the higher concentration of dexamethasone may change the round window permeability (Goycoolea, 2001; Hahn et al., 2006; Plontke et al., 2007), and this has not been controlled for in the present studies. These results demonstrate the utility of modelling in helping to design therapeutic strategies, but there are limitations to this approach. Foremost is the lack of a method for relating directly the simulated cochlear drug concentrations and the hearing protection. This would require knowledge of the perilymphatic steroid concentration required to protect the organ of Corti (and possibly the lateral cochlear structures) in vivo. The latter could potentially be subject to a cochleotopic gradient because the susceptibility of hair cells to injury is greater in the basal than the apical regions of the cochlea (Wu et al., 2002). Until these matters are better understood, simulation is likely to be to provide qualitative, rather than quantitative information to guide the development of therapeutic strategies. But these limitations aside, simulation provides the best method available to guide the clinical translation of the experimental results to a protective therapy for cochlear implantation, and to help account for inter-species differences such as the cochlear size and shape. Consistent with our previous observations (James et al., 2008), hearing loss was maximal at the site of cochleostomy and electrode array insertion, but also present at lower frequencies originating at cochlear sites apical to the site of surgery. The hearing loss at the site of surgery may relate to direct cochlear trauma, and it has been speculated that oxidative stress may contribute to the higher-frequency hearing loss. Dexamethasone protected against hearing loss across both frequency domains. Other studies have shown a similar trend in hearing protection after electrode insertion and withdrawal followed by continuous administration of dexamethasone for 1 month with a mini-osmotic pump (Eshraghi et al., 2007; Ye et al., 2004, 2007). The main therapeutic action of a steroid in the context of injury prevention is to suppress the post-operative an inflammatory insult (Cope and Bova, 2008). The present experiment supports this interpretation. There is histological evidence to suggest that a single application of dexamethasone to the round window may reduce the foreign body reaction to the cochlear implant (James et al., 2008). The absence of inflammatory reaction the present study is a contrary finding, and could potentially be due to operator-dependent factors, but equally it might be a consequence of the shorter period of observation in this study (1 week) compared with the previous investigation (1 month). The one-week period of study reported here is insufficient time for the development of a mature fibrous reaction to the implant. Therefore, a longer period of observation may be required to examine the fibrous tissue reaction. The duration of treatment required for optimal hearing protection could present practical challenges for the clinical translation of the experimental method presented here. The round window application times required to achieve protection (1–2 h) are very
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long compared with the duration of cochlear implant surgery, which is typically 90–120 min. It seems likely that surgeons would find the extended time of surgery required to achieve optimal hearing protection impractical. It must also be acknowledged that even longer waiting times might be required clinically than we required experimentally, because the human cochlea has a larger volume than that of the guinea pig. Given this possibility it is worthwhile reviewing the relative strengths of limitations of the present and alternative methods for cochlear drug delivery. The use of SeprapackTM, as a delivery vehicle applied to the round window had some advantages over intra-tympanic administration. The SeprapackTM can be positioned over the round window membrane; therefore, there is no loss of dexamethasone within the middle ear space, and minimal risk of absorption of steroid directly through the otic capsule into the cochlea. Whilst, the SeprapackTM is on the round window before the surgery, it retains the dexamethasone and permits a slow release after the surgery. While the release profile has not yet been fully characterized, previous work in our laboratories has shown dexamethasone and its breakdown products for a period of no more than 24 h as estimated by perilymph sampling in vivo (James et al., 2008). Other local delivery strategies are currently under investigation, but all have potential short-comings that await further study (Borenstein et al., 2007; Chen et al., 2005; Kopke et al., 2006). These methods include drug-eluting electrode array, use of microinjection and micro-osmotic pump (Borkholder, 2008). The drug-eluting electrode array may provide dexamethasone to the area of trauma but its efficacy may be limited because the dexamethasone is not delivered before surgery (Mathivanar et al., 1990; Mercanzini et al., 2007). The use of microinjection may itself result in trauma due to an increase in the hydraulic pressure from the injection, unless it is done in a controlled fashion and the latter is likely to be difficult to achieve reproducibly in surgery. Micro-fluidic systems have been developed which both deliver, and simultaneously remove quantities of fluid in order to reduce the hydraulic effects on the cochlea (Chen et al., 2005), and these systems can increase the concentration in the basal turn at a greater rate than diffusion alone. However, these may not be necessary for dexamethasone because it diffuses rapidly through the RW (noting that RW is quite a large surface area compared with a micro-fluidic channel). Micro-osmotic pumps can provide medium to long term infusion of dexamethasone, however, these may require additional surgical procedures and increase the risk of cochlear infection (Pettingill et al., 2008). The current hearing protection strategy is applicable clinically, but the parameters for clinical drug delivery need to account for the differences in cochlear size, and the practical constraints of surgical practice. To overcome the potentially long periods of application that may be required for dexamethasone to diffuse through the larger human cochlea, we speculate that a combined approach (systemic and local) may be required to ensure that sufficient drug concentration is achieved in the cochlea prior to surgery. Acknowledgements Project grants from the National Health and Medical Research Council Australia and Otolaryngology Scholarship from the Garnett Passe and Rodney Williams Memorial Foundation. Helen Feng for manufacturing electrode arrays. Maria Clark and Prudence Nielsen for preparing histological materials. Dr. Stuart Galloway and St. Vincent’s Hospital Anatomical Pathology Department for histological analysis. Dr. James Fallon for providing the ABR analysis program (NIDCD Contract HHS-N-263-2007-00053-C; PI: RK Shepherd). Salt A, Washington University Cochlear Fluids Simulator: Version 1.6 available from http://oto.wustl.edu/cochlea/model.htm. (This program was developed and is made available by Grant funding (DC 01368) from the National Institute on Deafness
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