Marion Bankstahl

Background: In rodent models of intracranial tumor development, evaluating the actual burden experienced by animals beyond procedural severity is essential for ethical and legal compliance. This study examined whether voluntary wheel running (VWR) could serve as a sensitive indicator of post-surgical burden following subcutaneous transmitter implantation, tumor cell injection, and tumor resection. It also assessed whether VWR supports the detection of humane endpoints. VWR outcomes were compared with body weight, clinical scores, heart rate, and activity levels recorded via telemetry. Methods: Fourteen male BDIX rats were housed individually in cages equipped with a running wheel. Under general anesthesia, telemetric devices to monitor heart rate and activity were subcutaneously implanted. After recovery, glioblastoma BT4Ca cells were stereotaxically injected into the right frontal cortex. Eight days later, the resulting tumors were microsurgically resected. Body weight, VWR, heart rate, and general activity were continuously monitored until the animals reached humane endpoint criteria, indicated by sudden weight loss and clinical deterioration. Results: On average, body weight and VWR declined significantly after all surgical procedures, with tumor resection causing the most pronounced effect. As animals approached the endpoint, a marked drop in these parameters was observed, along with an increased clinical score (p < 0.05). Activity measures supported these findings, though less consistently than weight and VWR. Conclusions: Monitoring body weight and VWR enables an effective assessment of the actual postoperative burden experienced by rats undergoing surgeries of different procedural complexity. Moreover, VWR is a valuable supplementary tool for identifying humane endpoints alongside body weight and clinical scoring.

Chicken embryos are able to process and receive noxious stimuli from embryonic day (ED) 13. In the context of animal welfare, analgesic refinement is highly recommended for painful procedures from this time onwards. In this first study, the pharmacokinetic and pharmacodynamic profiles of the opioid analgesics fentanyl (0.2 mg/kg) and buprenorphine (0.5 mg/kg) were studied in ED17 chicken embryos. Analgesic plasma concentrations were determined at different time points after administration via LC-MS/MS. In a validated model of mechanical nociception in chicken embryos, analgesic efficacy and potential side effects were determined based on mean arterial blood pressure, heart rate, beak, and body movements. Maximum mean plasma levels of 37.55 ng/ml (fentanyl) and 144.17 ng/ml (buprenorphine) were reached at 15 min with a plasma elimination half-life of around 75 or 60 min, respectively. No side effects on the cardiovascular system and no alterations in body movements, reflecting possible sedative effects, were detected. No significant reduction in mean arterial pressure after noxious stimulation of the base of the beak following opioid analgesic treatment was observed. However, medium to large effect sizes may suggest potential clinical importance. A significant reduction in wide beak opening, a nociceptive behavioral parameter, was observed 15 min after administration of fentanyl. In conclusion, both opioids may have potential as safe analgesic options for the use during potentially painful procedures in chicken embryos, although further research is needed to fully demonstrate their clinical efficacy.

Meloxicam is commonly used for analgesia, but no preparation approved specifically for use in mice. Here, we determined plasma concentrations, safety, and impact on home-cage behaviors for subcutaneous injection (5 mg/kg) and oral self-intake (sweetened drinking water, 20 mg/kg/24h, 5 days) for C57BL/6J mice (n = 21/sex). After injection, plasma concentrations measured by LC–MS/MS were higher in females (2h) and remained within an estimated therapeutic range (390–911 ng/mL) for up to 6h. T _1/2 was 2.32 h. Despite acceptance, plasma levels fluctuated strongly during oral self-intake. Side effects comprised reduced grip strength and increased vocalization (Irwin test), increased clinical score values (females, oral treatment) with two individuals reaching humane endpoint, increase in body temperature, body weight drop, decreased wheel-running activity, increased burrowing latency (males), and prolonged grooming activity (females). Grimace scale, nesting activity and plasma corticosterone remained unaffected. Red blood profile parameters and ALT were decreased and Ca ²⁺ and Cl ^- concentrations elevated. Histopathological analysis revealed inflammatory cells and hyperplasia in stomach and jejunum. In view of the unfavorable pharmacokinetics, meloxicam treatment via the drinking water is unsuitable in mice. The limited tolerability make prolonged treatment with the doses studied appear inappropriate.

As basis for evidence-based analgesia refinement, species-specific pharmacokinetic and tolerability profiles of carprofen were determined in rats for least aversive administration routes and prolonged treatment. Further, potential influence on behavioral pain indicators was evaluated. LC-MS/MS determined plasma concentrations in Sprague-Dawley rats (n = 21/sex) after subcutaneous (s.c.) injection (5 mg/kg) and during a 5-day treatment via the drinking water (d.w., 10 mg/kg/24 h). Irwin test parameters, clinical scoring, body weight, body temperature, fluid and food intake, grimace scale, burrowing, nesting, hematology, and histopathology were investigated. Plasma concentrations early after injection were higher in females, reached a maximum (C_max) of 39.16 ± 7.38 µg/ml at 3 h after injection and remained above an estimated in-vitro-derived therapeutic threshold (24.3 µg/ml) for at least 6 h with a T_1/2 of 7.06 h. Carprofen-medicated d.w. was readily consumed, with constant target dose intake over the 5-day treatment period reaching a C_max of 38.68 ± 8.67 µg/ml at 24 h. Tolerability and behavioral parameters revealed only minor changes, such as transient sedation (s.c.) and decreased body temperature (females). Gastrointestinal adverse effects were not detected. Carprofen’s pharmacokinetic profile allows for a practicable s.c. injection interval. Acceptance and tolerability during prolonged oral treatment with the assessed dose of 10 mg/kg/24 h makes its non-invasive administration promising for analgesia refinement in rats.

Unmet needs in the treatment of chronic otitis media and Eustachian tube dysfunction (ETD) triggered the development of stents for the Eustachian tube (ET). In this study, for the first time, stents were placed in an artificially blocked ET to evaluate stent function. Eight adult female sheep were injected with stabilized hyaluronic acid (HA) on both sides to induce ETD. Subsequently, a tapered nitinol ET stent was inserted on one side, and animals were examined bilaterally by endoscopy, tympanometry, cone beam computed tomography, and final histology. Seven of the stents were placed in the desired cartilaginous portion of the ET. At the end of the study, one stented side appeared slightly open; all other ET orifices were closed. Tympanometry revealed re-ventilation of the middle ear in four out of seven correctly stented animals within 3 to 6 weeks after stent insertion. The major amount of HA was found at the pharyngeal orifice of the ET anterior to the stent. Thus, the stent position did not completely align with the HA position. While a functional analysis will require refinement of the experimental setup, this study provides first promising results for stent insertion in a sheep model of ETD.

Introduction: Surgical interventions in mice require appropriate pain relief to ensure animal welfare and to avoid influence of pain on research findings. Carprofen is a non-steroidal anti-inflammatory drug commonly used as an analgesic for interventions inducing mild to moderate pain in laboratory rodents. Despite its frequent use, species-specific data on pharmacokinetics (PK), side effects, and potential impact on behavioral pain indicators are limited.Methods: We determined PK and tolerability profiles of carprofen in healthy male and female C57BL/6J mice (n = 42), administered at highest recommended doses via single subcutaneous (s.c.) injection (20 mg/kg) and oral self-administration (25 mg/kg/24 h) per drinking water (d.w.) for 5 days. Plasma concentrations were measured at various time points after the start of the treatment (n = 6 per time point), and side effects were evaluated using a modified Irwin test battery, hematology, and histopathology. Additionally, potential interference with cage-side behaviors commonly used for pain assessment, such as the mouse grimace scale, wheel running, burrowing, nesting, and grooming activity, was investigated.Results: Maximum plasma concentrations of 133.4 ± 11.3 μg/ml were reached 1 h after single s.c. injection with an elimination half-life of 8.52 h. Intake from d.w. resulted in a steady state within 24 h after the start of the treatment with plasma levels of around 60 μg/ml over 5 days in both sexes. The medicated water was well-accepted, and increased d.w. intake was observed in the first 24 h after exposure (p < 0.0001). The Irwin test revealed only minor influence on tested behavior and physiological functions. However, during treatment via d.w., an increase in body temperature (p < 0.0001) was observed, as well as a reduction in voluntary wheel running activity by 49–70% in male mice. Moreover, grooming behavior was slightly affected. Hematology and histopathology were without pathological findings that could be attributed to carprofen treatment. High-dose carprofen can be considered safe and of favorable PK for both administration routes assessed in healthy C57BL/6J mice of both sexes. Further efficacy evaluation of carprofen as monoanalgesic or component of multimodal post-surgical regimens is clearly encouraged; however, the impact on behavioral markers used for pain assessment should be considered in this context.

The Eustachian tube (ET) is a bottleneck when it comes to middle ear (ME) health. If its function is impaired, this can lead to serious consequences for the patient, such as hearing problems or deafness. Therefore, this study investigated a tapered nitinol stent (3–5 mm × 14 mm) for the human ET as a potential new permanent treatment for chronic Eustachian tube dysfunction (ETD) and thus ME ventilation disorders. The self-expanding stent was inserted unilaterally into the ET of 24 sheep with observation periods of 3, 6, and 12 months. Local tissue effects and the safety of the stent insertion were analyzed based on regular endoscopic checks, weekly tympanometry measurements, final imaging, and histological examinations. The animals showed no stent-related health restrictions. However, the individual anatomy and stenting procedure had an influence on the results. The tissue reaction in the endoscopic examinations was mild even though no concomitant antibiotics were administered. After all three monitoring periods, stented ETs had a significantly larger ET lumen than the non-stented contralateral ETs. However, tissue growth was detected in the stent. Overall, the first long-term study on an ET stent showed that the tapered ET stent could be a promising treatment option for ETD.

Surgical interventions in mice are prerequisite in various research fields and require appropriate pain relief, not only to ensure animal welfare but also to avoid influence of pain on research findings. Carprofen is a non-steroidal anti-inflammatory drug that is commonly used as an analgesic for interventions inducing mild to moderate pain in animals. Despite its frequent use also in laboratory rodents, data on pharmacokinetics and side effects, and on its potential impact on behavioral pain indicators are rare. This study aimed to determine pharmacokinetic and tolerability profiles of high dose carprofen in male and female C57Bl/6J mice, administered via single subcutaneous injection (s.c.) and oral self-administration per drinking water (d.w.). Plasma concentrations of carprofen were measured at various time points, and side effects were evaluated using a modified Irwin test protocol, hematology and histopathology. Additionally, potential effects on behavioral pain indicators commonly used to assess post-surgical pain, such as the mouse grimace scale, wheel running activity, burrowing, nesting and grooming behavior were investigated. Quantification of carprofen in plasma revealed maximum plasma concentrations of 133.4 ± 11.3 µg/ml after 1 hour and an elimination half-life of 8.52 hour after single s.c. injection of 20 mg/kg carprofen. Oral self-administration of carprofen (25 mg/kg/24 h) resulted in a steady-state < 24 hours over 5 days after treatment start with plasma levels of around 60 µg/ml. The carprofen-medicated water was highly accepted, and increased d.w. intake was observed in the first 24 hours after exposure for both sexes (p < 0.0001). Irwin test detected only minor side effects, and hematology and histopathology where without pathological findings that could be attributed to carprofen treatment. Except for a decrease of 49-70 % in wheel running activity in male mice, behavioral pain indicators were only very mildly affected. This study determined carprofen plasma levels in mice lying well above an estimated therapeutic concentration for both routes of administration. Carprofen was well tolerated at recommended high doses and may provide sufficient analgesia for minor interventions as well as be applied as a tolerable component in multimodal analgesic regimens.

Introduction: Sheep are frequently used in translational surgical orthopedic studies. Naturally, a good pain management is mandatory for animal welfare, although it is also important with regard to data quality. However, methods for adequate severity assessment, especially considering pain, are rather rare regarding large animal models. Therefore, in the present study, accompanying a surgical pilot study, telemetry and the Sheep Grimace Scale (SGS) were used in addition to clinical scoring for severity assessment after surgical interventions in sheep. Methods: Telemetric devices were implanted in a first surgery subcutaneously into four German black-headed mutton ewes (4–5 years, 77–115 kg). After 3–4 weeks of recovery, sheep underwent tendon ablation of the left M. infraspinatus. Clinical scoring and video recordings for SGS analysis were performed after both surgeries, and the heart rate (HR) and general activity were monitored by telemetry. Results: Immediately after surgery, clinical score and HR were slightly increased, and activity was decreased in individual sheep after both surgeries. The SGS mildly elevated directly after transmitter implantation but increased to higher levels after tendon ablation immediately after surgery and on the following day. Conclusion: In summary, SGS- and telemetry-derived data were suitable to detect postoperative pain in sheep with the potential to improve individual pain recognition and postoperative management, which consequently contributes to refinement.