Maintenance of pig brain function under extracorporeal pulsatile circulatory control (EPCC).

Selective vascular access to the brain is desirable in metabolic tracer, pharmacological and other studies aimed to characterize neural properties in isolation from somatic influences from chest, abdomen or limbs. However, current methods for artificial control of cerebral circulation can abolish pulsatility-dependent vascular signaling or neural network phenomena such as the electrocorticogram even while preserving individual neuronal activity. Thus, we set out to mechanically render cerebral hemodynamics fully regulable to replicate or modify native pig brain perfusion. To this end, blood flow to the head was surgically separated from the systemic circulation and full extracorporeal pulsatile circulatory control (EPCC) was delivered via a modified aorta or brachiocephalic artery. This control relied on a computerized algorithm that maintained, for several hours, blood pressure, flow and pulsatility at near-native values individually measured before EPCC. Continuous electrocorticography and brain depth electrode recordings were used to evaluate brain activity relative to the standard offered by awake human electrocorticography. Under EPCC, this activity remained unaltered or minimally perturbed compared to the native circulation state, as did cerebral oxygenation, pressure, temperature and microscopic structure. Thus, our approach enables the study of neural activity and its circulatory manipulation in independence of most of the rest of the organism.

Recording of pig neuronal activity in the comparative context of the awake human brain.

Gyriform mammals display neurophysiological and neural network activity that other species exhibit only in rudimentary or dissimilar form. However, neural recordings from large mammals such as the pig can be anatomically hindered and pharmacologically suppressed by anesthetics. This curtails comparative inferences. To mitigate these limitations, we set out to modify electrocorticography, intracerebral depth and intracortical recording methods to study the anesthetized pig. In the process, we found that common forms of infused anesthesia such as pentobarbital or midazolam can be neurophysiologic suppressants acting in dose-independent fashion relative to anesthetic dose or brain concentration. Further, we corroborated that standard laboratory conditions may impose electrical interference with specific neural signals. We thus aimed to safeguard neural network integrity and recording fidelity by developing surgical, anesthesia and noise reduction methods and by working inside a newly designed Faraday cage, and evaluated this from the point of view of neurophysiological power spectral density and coherence analyses. We also utilized novel silicon carbide electrodes to minimize mechanical disruption of single-neuron activity. These methods allowed for the preservation of native neurophysiological activity for several hours. Pig electrocorticography recordings were essentially indistinguishable from awake human recordings except for the small segment of electrical activity associated with vision in conscious persons. In addition, single-neuron and paired-pulse stimulation recordings were feasible simultaneously with electrocorticography and depth electrode recordings. The spontaneous and stimulus-elicited neuronal activities thus surveyed can be recorded with a degree of precision similar to that achievable in rodent or any other animal studies and prove as informative as unperturbed human electrocorticography.

Indicators of Postoperative Pain in Syrian Hamsters (Mesocricetus auratus).

Despite the use of Syrian hamsters (Mesocricetus auratus) in research, little is known about the evaluation of pain in this species. This study investigated whether the frequency of certain behaviors, a grimace scale, the treat-take-test proxy indicator, body weight, water consumption, and coat appearance could be monitored as signs of postoperative pain in hamsters in a research setting. Animals underwent no manipulation, anesthesia only or laparotomy under anesthesia. An ethogram was constructed and used to determine the frequencies of pain, active and passive behaviors by in-person and remote videorecording observation methods. The Syrian Hamster Grimace Scale (SHGS) was developed for evaluation of facial expressions before and after the surgery. The treat-take-test assessed whether surgery would affect the animals' motivation to take a high-value food item from a handler. The hypothesis was that behavior frequency, grimace scale, treat-take-test score, body weight, water consumption, and coat appearance would change from baseline in the surgery group but not in the no-intervention and anesthesia-only groups. At several time points, pain and passive behaviors were higher than during baseline in the surgery group but not the anesthesia-only and no-intervention groups. The SHGS score increased from baseline scores in 3 of the 9 animals studied after surgery. The frequency of pain behaviors and SHGS scores were highly specific but poorly sensitive tools to identify animals with pain. Behaviors in the pain category were exhibited by chiefly, but not solely, animals that underwent the laparotomy. Also, many animals that underwent laparotomy did not show behaviors in the pain category. Treat-take-test scores, body weight, water consumption, and coat appearance did not change from baseline in any of the 3 groups. Overall, the methods we tested for identifying Syrian hamsters experiencing postoperative pain were not effective. More research is needed regarding clinically relevant strategies to assess pain in Syrian hamsters.

Zwitterionic amino acid-based Poly(ester urea)s suppress adhesion formation in a rat intra-abdominal cecal abrasion model.

Hernia repair outcomes have improved with more robust material options for surgeons and optimized surgical techniques. However, ventral hernia repairs remain challenging with an inherent risk of post-surgical adhesions in the peritoneal space which can occur regardless of interventional material or its surgical placement. Herein, amino acid-based poly(ester urea)s (PEUs) with varied amount of an allyl ether side chains were modified post polymerization modification with the zwitterionic sulfnate group (3-((3-((3-mercaptopropanoyl)oxy)propyl) dimethylammonio)propane-1-sulfonate) to promote anti-adhesive properties. These alloc-PEUs were processed using roll-to-roll fabrication methods to afford films that were amenable to surface functionalization via a zwitterion-thiol. Functional group availability on the surface was confirmed via fluorescence microscopy, x-ray photoelectron spectroscopy (XPS), and quartz crystal microbalance (QCM) measurements. Zwitterionic treated PEUs exhibited reduced fibrinogen adsorption in vitro when compared to unfunctionalized control polymer. A rat intrabdominal cecal abrasion adhesion model was used to assess the extent and tenacity of adhesion formation in the presence of the PEUs. The 10% alloc-PEU zwitterion functionalized material was found to reduce the extent and tenacity of adhesions when compared to adhesion controls and the unfunctionalized PEU controls.

Preliminary Evaluation of Sustained-release Compared with Conventional Formulations of Meloxicam in Sheep (Ovis aries).

Sustained-release (SR) drugs refine current analgesic regimens by alleviating the need for multiple sessions of handling and restraint and by reducing the local tissue irritation that can occur due to repeated injections. Although a variety of SR drugs are already used in lab animal medicine, no studies exist that evaluate the suitability of an SR NSAID in sheep. This study used HPLC-MS to measure the plasma concentrations of 2 formulations of meloxicam-conventional and SRM- after subcutaneous administration in 6 adult ewes. Blood was collected at 0, 4, 12, 24, 36, 48, 60, 72, 84, 96, 120, 144, and 168 h after injection. In addition, physical exams, urinalysis, and biochemical analysis were performed at 0, 24, 48, and 120 h after dosage. Peak plasma concentrations were 1057 ± 433 ng/mL at 4 ± 0 h for conventional meloxicam and 3238 ± 1480 ng/mL at 6.7 ± 4.1 h for SR meloxicam (SRM). Elimination half-lives were 12.1 4.2 for CM and 15.2 ± 2.4 h for SRM. One sheep had an episode of acute renal azotemia starting 24 h after SRM administration; the episode resolved over time, and the definitive relationship to SRM administration was not determined. Plasma levels of SRM were higher than CM throughout the initial 24 h, remained variably elevated until 60 h after injection, but failed to sustain presumed therapeutic levels of 400 ng/mL for the full 72 h across all animals in this study. Further investigation is warranted to determine the safety and clinical efficacy of SRM in sheep. Currently, when SRM is used in sheep, we recommend the combination of a preemptive and multimodal analgesia regimen with clinical assessments throughout the postoperative period.

Doxycycline induces dysbiosis in female C57BL/6NCrl mice.

OBJECTIVE: This study aims to demonstrate the effect of oral doxycycline on fecal microbiota of mice. Doxycycline is a common effector for control of gene expression using the tet-inducible system in transgenic mice. The effect of oral doxycycline on murine gut microbiota has not been reported. We evaluated the effect of doxycycline treatment by sequencing the V4 hypervariable region of the 16S rRNA gene from fecal samples collected during a 4 week course of treatment at a dose of 2 mg/ml in the drinking water. RESULTS: The fecal microbiota of treated animals were distinct from control animals; the decreased richness and diversity were characterized primarily by Bacteroides sp. enrichment. These effects persisted when the treatment was temporarily discontinued for 1 week. These data suggest that doxycycline treatment can induce significant dysbiosis, and its effects should be considered when used in animal models that are or maybe sensitive to perturbation of the gut microbiota.

E-selectin inhibition with GMI-1271 decreases venous thrombosis without profoundly affecting tail vein bleeding in a mouse model.

Selectins, such as E-selectin (CD62E), function in venous thrombosis by binding and activating immune cells to initiate the coagulation cascade. GMI-1271 is a small molecule antagonist that inhibits E-selectin activity. Here we determine whether inhibition of E-selectin is sufficient to decrease acute venous thrombosis and associated inflammatory events in both prophylactic and treatment protocols without significantly affecting haemostasis. Male C57BL/6 mice underwent surgery for experimental thrombosis induction and were harvested at peak thrombus formation in our animal model, two days post induction. Groups included non-thrombosed true controls, shams, controls, and prophylactic or treatment groups of GMI-1271 (10 mg/kg intraperitoneal BID (twice a day) and low-molecular-weight heparin (LMWH, Lovenox 6 mg/kg subcutaneously (SC), once a day (SID). Compared with control animals, prophylaxis or treatment with LMWH and GMI-1271 in a dose-dependent manner significantly decreased thrombosis. GMI-1271 significantly lowered tail bleeding times when compared to LMWH. GMI-1271 and LMWH prophylactically administered significantly decreased vein wall neutrophil cell extravasation. However, all treatment and prophylactic therapies significantly decreased vein wall monocyte extravasation versus controls. GMI-1271 prophylactic therapy significantly decreased intra-thrombus cell counts versus control animals and other treatment groups. Immunohistochemistry confirmed that both treatments with GMI-1271 and LMWH significantly decreased activated leukocyte migration. GMI-1271 therapy significantly decreased thrombus weight and resulted in significantly lower bleeding times than LMWH. GMI-1271 treated mice showed decreased local and systemic inflammatory effects while modulating neutrophil activation, suggesting that GMI-1271 is a viable therapeutic candidate for venous thrombosis prophylaxis and treatment.

Validation of a Behavioral Ethogram for Assessing Postoperative Pain in Guinea Pigs (Cavia porcellus).

Although guinea pigs (Cavia porcellus) have been used in research for more than a century and remain the most prevalent USDA-covered species, little has been elucidated regarding the recognition of clinical pain or analgesic efficacy in this species. We sought to assess pain in guinea pigs by using newer, clinically relevant methods that have been validated in other rodent species: the behavioral ethogram and cageside proxy indicator. In this study, 10 male guinea pigs underwent electronic von Frey testing of nociception, remote videorecording of behavior, and cageside assessment by using time-to-consumption (TTC) of a preferred treat test. These assessments were performed across 2 conditions (anesthesia only and castration surgery under anesthesia) at 3 time points (2, 8, and 24 h after the event). The anesthesia only condition served to control for the nonpainful but potentially distressing components of the surgical experience. Compared with those after anesthesia only conditions, subtle body movements were increased and nociceptive thresholds were decreased at 2 and 8 h after surgery. At 24 h, neither subtle body movement behaviors nor nociceptive thresholds differed between the 2 conditions. In contrast, TTC scores did not differ between the anesthesia only and surgery conditions at any time point, underscoring the challenge of identifying pain in this species through cageside evaluation. By comparing ethogram scores with measures of nociception, we validated select behaviors as pain-specific. Therefore, our novel ethogram allowed us to assess postoperative pain and may further serve as a platform for future analgesia efficacy studies in guinea pigs.

Assessment of an Orofacial Operant Pain Assay as a Preclinical Tool for Evaluating Analgesic Efficacy in Rodents.

A model system capable of providing clinically relevant analgesic doses with minimal trauma has been elusive in laboratory animal medicine. Our laboratory has developed an orofacial operant pain system that effectively discriminates between non-noxious and noxious thermal stimuli in rats and mice. Male and female rats (Crl:SD) and mice (Crl:SKR-HR(hr)) were trained to perform a task (placing their face through an opening and having their cheeks stay in contact with thermodes) to receive a reward (a solution of sweetened condensed milk). Currently accepted doses of buprenorphine were tested by using a crossover design. Pain was induced in both species by sensitizing the depilated skin over both cheeks with capsaicin cream or by creating a surgical incision (rats only) and then allowing the animals to contact a temperature-regulated thermode while obtaining a reward. Optimal antinociceptive doses included 0.05 and 0.1 mg/kg in male mice but only 0.05 mg/kg in female mice. In rats, optimal antinociceptive doses included 0.03 and 0.05 mg/kg for male rats but only 0.03 mg/kg for female rats. The 2 pain-induction models in rats (capsaicin cream and surgical incision) did not differ. Our orofacial operant pain assay can determine clinically relevant analgesic doses for rodents in a preclinical assay. The automated, investigator-independent nature of the assay, in conjunction with its high sensitivity, makes this method an improvement over traditional noninvasive methods, providing better data for developing optimal analgesic recommendations for rats and mice.