**1. Introduction**

Rabbits are commonly used as animal models in biomedical research, e.g., orthopedic studies concerning bone healing, cartilage repair and infection studies [1–3]. In these experimental studies, the animals often undergo invasive surgeries which require systemic analgesia for several days. Looking into the literature, the administration of systemic analgesia in invasive experimental studies involving rabbits increased significantly from 16% (1995–997) to 50% (2005–2007) [4,5]. Additionally, despite improvement in the last 20 years it is reported that pain managemen<sup>t</sup> is still inadequate for pet rabbits [6]. Nevertheless, optimization of postoperative analgesia protocols is still highly needed as studies investigating effective analgesia for surgical interventions have been rarely reported for this species [7]. Among opioids, the most commonly used drug was buprenorphine as the first-line therapy over this period [4].

Buprenorphine, a partial μ-receptor agonist opioid, is commonly used for perioperative pain control in laboratory animal species [4]. Despite the long duration of action, multiple injections are required daily to guarantee a continuous analgesic coverage [8]. As rabbits are highly susceptible to stress and anxiety [9,10], repeated handling and medical interventions should be kept to a minimum to improve animal welfare. As an alternative, titrating analgesic drugs to effect would carry the risk of letting pain go undertreated, as pain recognition in a prey species like the rabbit represents a true challenge [10,11], even though the rabbit grimace scale is a promising tool to assess postoperative pain [12].

An opportunity to ensure a noninvasive and "stress free" analgesia is through the use of transdermal patches which provide a continuous release of medications into the bloodstream over a considerable period. Fentanyl, a potent m-agonist synthetic opioid characterized by a short half time, would not be adequate for postoperative analgesia in rabbits as an injectable formulation, but becomes interesting as a transdermal patch. Fentanyl patches were developed by the Alza Corporation and came on the market in 1991. These patches provide a sustained and constant drug release into the systemic circulation for at least 72 h, and are commonly prescribed in human medicine to treat chronic cancer pain [13]. In 1995, the first animal study investigating these opioid releasing patches in dogs was published [14]. Later, their off-label use in the veterinary medicine, especially in the experimental fields for postoperative pain therapy, became popular due to their properties as a noninvasive long-term delivery form. In a study from Foley et al., the use of fentanyl patches placed on the neck of rabbits was investigated for the first time and considered to be efficacious [15]. In other species such as dogs, cats, sheep, horses and minipigs, the application of the analgesic patches has further been investigated [14,16–23]. It was shown that exogenous and endogenous factors such as the amount of subcutaneous fat, skin integrity, structure and arrangemen<sup>t</sup> of hair follicles, possible dermal depots, body core temperature, skin thickness, first-pass cutaneous biotransformation as well as the environmental temperature caused strong intra- and inter-individual differences in the uptake of fentanyl into the bloodstream. Therefore, the prediction of fentanyl plasma concentrations is difficult [5,8,14,18,24]. Based on this described variability of drug absorption depending on the application locations, it was hypothesized that there is a variation in the uptake of fentanyl into the blood when different application locations are used [24]. The goal of this study was to investigate different locations for fentanyl patch application (neck, inner and outer surfaces of the ear) in rabbits. We assumed that the application on the inner or outer surface of the ear would provide a more reliable absorption of fentanyl with less variability due to a more homogenous anatomy and easier site preparation.

### **2. Materials and Methods**
