Cochlear microdialysis for quantification of dexamethasone and fluorescein entry into scala tympani during round window administration

Abstract

Before new drugs for the treatment of inner ear disorders can be studied in controlled clinical trials, it is important that their pharmacokinetics be established in inner ear fluids. Microdialysis allows drug levels to be measured in perilymph without the volume disturbances and potential cerebrospinal fluid contamination associated with fluid sampling. The aims of this study were to show: (i) that despite low recovery rates from miniature dialysis probes, significant amounts of drug are removed from small fluid compartments, (ii) that dialysis sampling artifacts can be accounted for using computer simulations and (iii) that microdialysis allows quantification of the entry rates through the round window membrane (RWM) into scala tympani (ST). Initial experiments used microdialysis probes in small compartments in vitro containing sodium fluorescein. Stable concentrations were observed in large compartments (1000 μl) but significant concentration declines were observed in smaller compartments (100, 10 and 5.6 μl) comparable to the size of the inner ear. Computer simulations of these experiments closely approximated the experimental data. In in vivo experiments, sodium fluorescein 10 mg/ml and dexamethasone-dihydrogen-phosphate disodium salt 8 mg/ml were simultaneously applied to the RWM of guinea pigs. Perilymph concentration in the basal turn of ST was monitored using microdialysis. The fluorescein concentration reached after 200 min application (585 ± 527 μg/ml) was approximately twice that of dexamethasone phosphate (291 ± 369 μg/ml). Substantial variation in concentrations was found between animals by approximately a factor of 34 for fluorescein and at least 41 for dexamethasone phosphate. This is, to a large extent, thought to be the result of the RWM permeability varying in different animals. It was not caused by substance analysis variations, because two different analytic methods were used and the concentration ratio between the two substances remained nearly constant across the experiments and because differences were apparent for the repeated samples obtained in each animal. Interpretation of the results using computer simulations allowed RWM permeability to be quantified. It also demonstrated, however, that cochlear clearance values could not be reliably obtained with microdialysis because of the significant contribution of dialysis to clearance. The observed interanimal variation, e.g., in RWM permeability, is likely to be clinically relevant to the local application of drugs in patients.

Dexamethasone Concentration Gradients Along Scala Tympani After Application to the Round Window Membrane

“…The method of animal preparation and drug application is described in detail elsewhere (19). In brief, nine specific pathogen free guinea pigs (mean weight: 423g; 300-600g; Charles River, Kißlegg, Germany) were anesthetized by intraperitoneal injection of an initial dose of 8 mg/ kg xylazine (Bayer, Leverkusen, Germany) plus 140 mg/kg ketamine hydrochloride (Pharmacia & Upjohn, Erlangen, Germany).…”

“…Anesthesia was maintained by repeated intramuscular injections of 25-33% of the initial dose. Further preparation followed as described in Hahn et al (2006) and in Figure 1. The animal experiments were approved by the animal studies committee of the University of Tübingen.…”

“…For further HPLC-details see Hahn et al (19). The peaks for Dex-P and Dex appeared at 7.0 min and 26.1 min, respectively.…”

Perilymph Pharmacokinetics of Markers and Dexamethasone Applied and Sampled at the Lateral Semi-Circular Canal

“…There have been numerous pharmacokinetic studies of locally-applied corticosteroids in animals (Nomura 1984;Parnes et al 1999;Chandrasekhar et al 2000;Bachmann et al 2001;Hahn et al 2006Hahn et al , 2012Liu et al 2006;Plontke et al 2008b;Yang et al 2008;Wang et al 2009;Borden et al 2011;Salt et al 2011a) and humans (Bird et al 2007;Bird et al 2011), but only a subset of these studies has been analyzed quantitatively to derive an elimination rate. Plontke and analyzed prednisoline data of Bachmann et al (2001) from guinea pigs and found it to be consistent with an elimination half-time from perilymph of 130 min.…”

Marker retention in the cochlea following injections through the round window membrane

“…Although many substances can be applied to the round window membrane (RWM) and from there diffuse into perilymph, large or anionic molecules do not easily permeate the RWM (Goycoolea and Lundman, 1997). In addition, pharmacokinetic studies suggest that there is considerable variation in the permeability of the RWM in different animals, resulting in highly variable drug levels in perilymph (Chelikh et al, 2003,Hahn et al, 2006,Mynatt et al, 2006,Plontke et al, 2007. To overcome these problems, an alternative approach is to inject the drug or other substance directly into the inner ear fluids.…”