Behavior and neurobehavioral teratology using the ferret

A behavioral profile of the ferret is presented for those who would like to use this animal in behavioral teratology and toxicology, or other disciplines involving behavior. We have reviewed neurobehavioral teratology of lisencephalic ferrets and neuropsychology of ferrets sustaining frontal lesions, as well as most of the studies of "normal" ferret behavior that have appeared in the research literature. Emphasis is placed on discussion of the tests used and how ferrets behaved on them. The behaviors discussed include spatial (maze) learning, delayed response, visual discrimination learning, discrimination learning sets, schedule maintained behavior, shock avoidance learning and spontaneously occurring behaviors, such as ambulation in open field, spontaneous alternation and species specific behaviors. Although the use of the ferret in behavioral experiments is not yet extensive and large gaps exist in our knowledge about the basic functional capacities of this animal, the ferret is unquestionably well suited for behavioral studies.     

Oseltamivir Population Pharmacokinetics in the Ferret: Model Application for Pharmacokinetic/Pharmacodynamic Study Design

The ferret is a suitable small animal model for preclinical evaluation of efficacy of antiviral drugs against various influenza strains, including highly pathogenic H5N1 viruses. Rigorous pharmacokinetics/pharmacodynamics (PK/PD) assessment of ferret data has not been conducted, perhaps due to insufficient information on oseltamivir PK. Here, based on PK data from several studies on both uninfected and influenza-infected groups (i.e., with influenza A viruses of H5N1 and H3N2 subtypes and an influenza B virus) and several types of anesthesia we developed a population PK model for the active compound oseltamivir carboxylate (OC) in the ferret. The ferret OC population PK model incorporated delayed first-order input, two-compartment distribution, and first-order elimination to successfully describe OC PK. Influenza infection did not affect model parameters, but anesthesia did. The conclusion that OC PK was not influenced by influenza infection must be viewed with caution because the influenza infections in the studies included here resulted in mild clinical symptoms in terms of temperature, body weight, and activity scores. Monte Carlo simulations were used to determine that administration of a 5.08 mg/kg dose of oseltamivir phosphate to ferret every 12 h for 5 days results in the same median OC area under the plasma concentration-time curve 0–12 h (i.e., 3220 mg h/mL) as that observed in humans during steady state at the approved dose of 75 mg twice daily for 5 days. Modeling indicated that PK variability for OC in the ferret model is high, and can be affected by anesthesia. Therefore, for proper interpretation of PK/PD data, sparse PK sampling to allow the OC PK determination in individual animals is important. Another consideration in appropriate design of PK/PD studies is achieving an influenza infection with pronounced clinical symptoms and efficient virus replication, which will allow adequate evaluation of drug effects.     

Responses to innocuous,but not noxious,somatosensory stimulation by neurons in the ferret superior colliculus

The intermediate and deep layers of the superior colliculus (SC) are known for their role in initiating orienting behaviors. To direct these orienting functions, the SC of some animals (e.g., primates, carnivores) is dominated by inputs from the distance senses (vision, audition). In contrast, the rodent SC relies more heavily on non-visual inputs, such as touch and nociception, possibly as an adaptive response to the proximity of dangers encountered during their somatosensory-dominant search behaviors. The ferret (a carnivore) seems to employ strategies of both groups: above ground they use visual/auditory cues, but during subterranean hunting ferrets must rely on non-visual signals to direct orienting. Therefore, the present experiments sought to determine whether the sensory inputs to the ferret SC reveal adaptations common to functioning in both environments. The results showed that the ferret SC is dominated (63%; 181/286) by visual/auditory inputs (like the cat), rather than by somatosensory inputs (as found in rodents). Furthermore, tactile responses were driven primarily from hair-receptors (like cats), not from the vibrissae (as in rodents). Additionally, while a majority of collicular neurons in rodents respond to brief noxious stimulation, no such neurons were encountered in the ferret SC. A small proportion (4%; 13/286) of the ferret SC neurons were responsive to long-duration (> 5s) noxious stimulation, but further tests could not establish these responses as nociceptive. Collectively, these data indicate that the ferret SC is best adapted for the animal's visual/acoustically guided activities and most closely resembles the SC of its phylogenetic relative, the cat.     

The ferret: an animal model to study influenza virus

There has been much critical influenza research conducted in a little-known laboratory animal--the ferret. The authors review some of these findings, discuss the reasons the ferret often becomes a model for influenza infection, and compare the ferret with other animal models.     

Responses to innocuous, but not noxious, somatosensory stimulation by neurons in the ferret superior colliculus

The intermediate and deep layers of the superior colliculus (SC) are known for their role in initiating orienting behaviors. To direct these orienting functions, the SC of some animals (e.g., primates, carnivores) is dominated by inputs from the distance senses (vision, audition). In contrast, the rodent SC relies more heavily on non-visual inputs, such as touch and nociception, possibly as an adaptive response to the proximity of dangers encountered during their somatosensory-dominant search behaviors. The ferret (a carnivore) seems to employ strategies of both groups: above ground they use visual/auditory cues, but during subterranean hunting ferrets must rely on non-visual signals to direct orienting. Therefore, the present experiments sought to determine whether the sensory inputs to the ferret SC reveal adaptations common to functioning in both environments. The results showed that the ferret SC is dominated (63%; 181/286) by visual/auditory inputs (like the cat), rather than by somatosensory inputs (as found in rodents). Furthermore, tactile responses were driven primarily from hair-receptors (like cats), not from the vibrissae (as in rodents). Additionally, while a majority of collicular neurons in rodents respond to brief noxious stimulation, no such neurons were encountered in the ferret SC. A small proportion (4%; 13/286) of the ferret SC neurons were responsive to long-duration (>5 s) noxious stimulation, but further tests could not establish these responses as nociceptive. Collectively, these data indicate that the ferret SC is best adapted for the animal's visuallacoustically guided activities and most closely resembles the SC of its phylogenetic relative, the cat.     

Anatomic and physiological characteristics of the ferret lateral rectus muscle and abducens nucleus.

The ferret has become a popular model for physiological and neurodevelopmental research in the visual system. We believed it important, therefore, to study extraocular whole muscle as well as single motor unit physiology in the ferret. Using extracellular stimulation, 62 individual motor units in the ferret abducens nucleus were evaluated for their contractile characteristics. Of these motor units, 56 innervated the lateral rectus (LR) muscle alone, while 6 were split between the LR and retractor bulbi (RB) muscle slips. In addition to individual motor units, the whole LR muscle was evaluated for twitch, tetanic peak force, and fatigue. The abducens nucleus motor units showed a twitch contraction time of 15.4 ms, a mean twitch tension of 30.2 mg, and an average fusion frequency of 154 Hz. Single-unit fatigue index averaged 0.634. Whole muscle twitch contraction time was 16.7 ms with a mean twitch tension of 3.32 g. The average fatigue index of whole muscle was 0.408. The abducens nucleus was examined with horseradish peroxidase conjugated with the subunit B of cholera toxin histochemistry and found to contain an average of 183 motoneurons. Samples of LR were found to contain an average of 4,687 fibers, indicating an LR innervation ratio of 25.6:1. Compared with cat and squirrel monkeys, the ferret LR motor units contract more slowly yet more powerfully. The functional visual requirements of the ferret may explain these fundamental differences.     

Progress toward generating a ferret model of cystic fibrosis by somatic cell nuclear transfer

Mammalian cloning by nuclear transfer from somatic cells has created new opportunities to generate animal models of genetic diseases in species other than mice. Although genetic mouse models play a critical role in basic and applied research for numerous diseases, often mouse models do not adequately reproduce the human disease phenotype. Cystic fibrosis (CF) is one such disease. Targeted ablation of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in mice does not adequately replicate spontaneous bacterial infections observed in the human CF lung. Hence, several laboratories are pursuing alternative animal models of CF in larger species such as the pig, sheep, rabbits, and ferrets. Our laboratory has focused on developing the ferret as a CF animal model. Over the past few years, we have investigated several experimental parameters required for gene targeting and nuclear transfer (NT) cloning in the ferret using somatic cells. In this review, we will discuss our progress and the hurdles to NT cloning and gene-targeting that accompany efforts to generate animal models of genetic diseases in species such as the ferret.     

Compendium of recent literature on the ferret

A survey of research publications since 1977 indicated that the ferret is a rather popular research animal. Using the BIOSIS and MEDLINE data bases, 569 citations of research involving the ferret were identified. Over 27% of these citations involved the use of ferrets in the field of physiology, and an additional 24% of the citations were in virology and immunology. The areas of pharmacology, toxicology and teratology accounted for 10.4, 8.4 and 4.0% of the citations, respectively.     

Minimal Hodgkin–Huxley type models for different classes of cortical and thalamic neurons

We review here the development of Hodgkin–Huxley (HH) type models of cerebral cortex and thalamic neurons for network simulations. The intrinsic electrophysiological properties of cortical neurons were analyzed from several preparations, and we selected the four most prominent electrophysiological classes of neurons. These four classes are “fast spiking” “regular spiking” “intrinsically bursting” and “low-threshold spike” cells. For each class, we fit “minimal” HH type models to experimental data. The models contain the minimal set of voltage-dependent currents to account for the data. To obtain models as generic as possible, we used data from different preparations in vivo and in vitro, such as rat somatosensory cortex and thalamus, guinea-pig visual and frontal cortex, ferret visual cortex, cat visual cortex and cat association cortex. For two cell classes, we used automatic fitting procedures applied to several cells, which revealed substantial cell-to-cell variability within each class. The selection of such cellular models constitutes a necessary step towards building network simulations of the thalamocortical system with realistic cellular dynamical properties.     

A comparison of experimental visceral leishmaniasis in the opossum, armadillo and ferret

Visceral leishmaniasis is a severe, chronic protozoal disease of humans and animals. Although chemotherapeutic agents are available for the treatment of this disease, problems such as drug toxicity, drug ineffectiveness and drug resistance of the parasite are responsible for treatment failures. To determine whether a drug is a potential antileishmanial agent, screening tests are performed using in vitro and in vivo models. Subsequently, a study using an appropriate animal model is performed to clearly determine the efficacy of a drug against Leishmania. Due to current public concerns regarding the use of companion animals in addition to the high costs of obtaining and maintaining these animals for research use, conventional animal models used in these chemotherapy studies, notably the dog and monkey, are becoming less acceptable. Therefore, new, less expensive and more accessible animal models are needed for the study of antileishmanial compounds. In this study, the armadillo, ferret and opossum were evaluated as possible new animal models for visceral leishmaniasis. The marked body weight loss, hepatomegaly, splenomegaly, large amastigote densities and the microscopic lesions observed in the infected opossums indicated that the opossum was more susceptible to visceral leishmaniasis than the armadillo or ferret.     

Camouflage,communication and thermoregulation: lessons from colour changing organisms

Organisms capable of rapid physiological colour change have become model taxa in the study of camouflage because they are able to respond dynamically to the changes in their visual environment. Here, we briefly review the ways in which studies of colour changing organisms have contributed to our understanding of camouflage and highlight some unique opportunities they present. First, from a proximate perspective, comparison of visual cues triggering camouflage responses and the visual perception mechanisms involved can provide insight into general visual processing rules. Second, colour changing animals can potentially tailor their camouflage response not only to different backgrounds but also to multiple predators with different visual capabilities. We present new data showing that such facultative crypsis may be widespread in at least one group, the dwarf chameleons. From an ultimate perspective, we argue that colour changing organisms are ideally suited to experimental and comparative studies of evolutionary interactions between the three primary functions of animal colour patterns: camouflage; communication; and thermoregulation.     

Macroscopic and roentgenographic anatomy of the skull of the ferret (Mustela putorius furo)

Normal macroscopic and roentgenographic features of the skull of the ferret (Mustela putorius furo) were examined and described. Data were based on a sample of 100 (50 male and 50 female) adult ferrets of known body weight and age. The skull was described macroscopically according to six standard views, i.e. dorsal, lateral, ventral, caudal, cranial and midsagittal. The mandible was described separately. The roentgenographic characteristics of the ferret skull were demonstrated only in lateral and dorsoventral projections. Furthermore, the skull length and width as well as the minimum frontal width were measured, and skull indices were derived from relevant measurements. Sexual dimorphism was examined both morphologically and craniometrically. Besides the common features of a carnivore skull, the ferret skull is relatively elongated and flat with a short facial region. The skulls of adult male ferrets are about 17% longer and 22% wider than those of the females. Significant sexual dimorphism also exists regarding certain skull indices. The general features and some dimensional parameters of the adult ferret skull support the contention that the ferret would be an interesting and workable alternative animal model in craniofacial research.     

Crossmodal Integration Improves Sensory Detection Thresholds in the Ferret

During the last two decades ferrets (Mustela putorius) have been established as a highly efficient animal model in different fields in neuroscience. Here we asked whether ferrets integrate sensory information according to the same principles established for other species. Since only few methods and protocols are available for behaving ferrets we developed a head-free, body-restrained approach allowing a standardized stimulation position and the utilization of the ferret’s natural response behavior. We established a behavioral paradigm to test audiovisual integration in the ferret. Animals had to detect a brief auditory and/or visual stimulus presented either left or right from their midline. We first determined detection thresholds for auditory amplitude and visual contrast. In a second step, we combined both modalities and compared psychometric fits and the reaction times between all conditions. We employed Maximum Likelihood Estimation (MLE) to model bimodal psychometric curves and to investigate whether ferrets integrate modalities in an optimal manner. Furthermore, to test for a redundant signal effect we pooled the reaction times of all animals to calculate a race model. We observed that bimodal detection thresholds were reduced and reaction times were faster in the bimodal compared to unimodal conditions. The race model and MLE modeling showed that ferrets integrate modalities in a statistically optimal fashion. Taken together, the data indicate that principles of multisensory integration previously demonstrated in other species also apply to crossmodal processing in the ferret.     

Factors affecting the electrofusion of mouse and ferret oocytes with ferret somatic cells

The domestic ferret, Mustela putorius furos, holds great promise as a genetic model for human lung disease, provided that key technologies for somatic cell nuclear transfer (SCNT) are developed. In this report, we extend our understanding of SCNT in this species by defining conditions for efficient cell fusion by electrical pulse. Two experimental systems were employed in this study. First, in vivo-matured mouse oocytes and ferret somatic cells were used to establish general parameters for fusion. One fibroblast, or cumulus cell, was agglutinated to nucleate, zona pellucida-free, mouse oocytes, and subjected to an electrical pulse. Similar electrical pulse conditions were also tested with 1 or 2 somatic cells inserted into the perivitelline space (PVS) of intact mouse oocytes. The fusion rate for a single fibroblast with a zona-free oocyte was 80.2%, significantly higher (P < 0.05) than that observed for 1, or 2, fibroblasts placed in the PVS (52.0% and 63.8%, respectively). The fusion rate (44.1%) following insertion of two cumulus cells was significantly higher (P < 0.05) than that following insertion of one cumulus cell (25.1%). Second, in vitro-matured ferret oocytes were enucleated, and one to three fibroblasts or cumulus cells were inserted into the PVS. Zona pellucida-free ferret oocytes were fragile and excluded from the study. The fusion rates with two or three fibroblasts were 71.4% and 76.8%, respectively; significantly higher (P < 0.05) than that for one fibroblast (48.6%). This cell number-dependent difference in fusion efficiency was also observed with cumulus cells. Fusion-derived (ferret-ferret) NT embryos cleaved, formed blastocysts in vitro, and underwent early-stage fetal development following embryo transfer. The rate of development was cell type-independent, in contrast to the cell type-dependent differences observed in fusion efficiency. In conclusion, fibroblasts fused more efficiently than cumulus cells and the efficiency of single cell fusions was improved when two or more cells were inserted into the PVS. These studies define conditions for efficient cell fusion with ferret oocytes and should facilitate SCNT and the development of genetically defined animal models in this species.     

Integrated data acquisition system for medical device testing and physiology research in compliance with good laboratory practices

In seeking approval from the US Food and Drug Administration (FDA) for clinical trial evaluation of an experimental medical device, a sponsor is required to submit experimental findings and support documentation to demonstrate device safety and efficacy that are in compliance with Good Laboratory Practices (GLP). The objective of this project was to develop an integrated data acquisition (DAQ) system and documentation strategy for monitoring and recording physiological data when testing medical devices in accordance with GLP guidelines mandated by the FDA. Data aquisition systems were developed as stand-alone instrumentation racks containing transducer amplifiers and signal processors, analog-to-digital converters for data storage, visual display and graphical user-interfaces, power conditioners, and test measurement devices. Engineering standard operating procedures (SOP) were developed to provide a written step-by-step process for calibrating, validating, and certifying each individual instrumentation unit and the integrated DAQ system. Engineering staff received GLP and SOP training and then completed the calibration, validation, and certification process for the individual instrumentation components and integrated DAQ system. Eight integrated DAQ systems have been successfully developed that were inspected by regulatory affairs consultants and determined to meet GLP guidelines. Two of these DAQ systems were used to support 40 of the pre-clinical animal studies evaluating the AbiCor artificial heart (ABIOMED, Danvers, MA). Based in part on these pre-clinical animal data, the AbioCor clinical trials began in July 2001. The process of developing integrated DAQ systems, SOP, and the validation and certification methods used to ensure GLP compliance are presented in this article.     

Reinforcement Learning on Slow Features of High-Dimensional Input Streams

Humans and animals are able to learn complex behaviors based on a massive stream of sensory information from different modalities. Early animal studies have identified learning mechanisms that are based on reward and punishment such that animals tend to avoid actions that lead to punishment whereas rewarded actions are reinforced. However, most algorithms for reward-based learning are only applicable if the dimensionality of the state-space is sufficiently small or its structure is sufficiently simple. Therefore, the question arises how the problem of learning on high-dimensional data is solved in the brain. In this article, we propose a biologically plausible generic two-stage learning system that can directly be applied to raw high-dimensional input streams. The system is composed of a hierarchical slow feature analysis (SFA) network for preprocessing and a simple neural network on top that is trained based on rewards. We demonstrate by computer simulations that this generic architecture is able to learn quite demanding reinforcement learning tasks on high-dimensional visual input streams in a time that is comparable to the time needed when an explicit highly informative low-dimensional state-space representation is given instead of the high-dimensional visual input. The learning speed of the proposed architecture in a task similar to the Morris water maze task is comparable to that found in experimental studies with rats. This study thus supports the hypothesis that slowness learning is one important unsupervised learning principle utilized in the brain to form efficient state representations for behavioral learning.     

Technical Restrictions of Computer-Manipulated Visual Stimuli and Display Units for Studying Animal Behaviour

Computer-manipulated visual stimuli are a well-established tool to experimentally study animal behaviour. They provide the opportunity to manipulate single or combined visual stimuli selectively, while other potentially confounding variables remain constant. A wide array of different presentation methods of artificial stimuli has been used in recent research. Furthermore, a wide range of basic hardware and software has been used to conduct experiments. The outcomes of experimental trials using computer-manipulated visual stimuli differed among studies. Failing or contradictory results were mostly discussed in a behavioural and ecological context. However, the results sometimes may be basically flawed due to methodological traps in the experimental design. Based on characteristics and restrictions of technical standards, we discuss which kinds of computer stimuli and visual display units are available today and their suitability for experimental trials when studying animal behaviour. A computer-manipulated stimulus displayed by a certain visual display unit may be accurate to investigate behaviour in a specific species, if various preconditions are met. However, simply due to technical characteristics of the visual display unit, the set-up may be unsuitable for other test species. Thus, future research should critically apply the included technique, with respect to both the intended kind of stimulus and the species under investigation. If these preconditions are met, computer-manipulated stimuli provide a high degree of standardization and the potential to display visual signals without losing a crucial amount of information from the native data.     

Predator stress induces behavioral inhibition and amygdala somatostatin receptor 2 gene expression

Psychological stressors precipitate and maintain stress-induced psychopathology, and it is likely that altered amygdala function underlies some of the deleterious effects of psychological stress. To understand the mechanisms underlying the linkage between the response to psychological stressors and maladaptive or psychopathological responses, we have focused on amygdala responsivity in animal models employing species-specific psychological stressors. In the present study, we characterized the effects of a 15-min exposure to a natural predator, the ferret, on rat behavior and the expression of the somatostatin family of genes in the amygdala. We examined the somatostatin family of genes because substantial evidence shows that central somatostatin systems are altered in various neuropsychiatric illnesses. We report that rats respond to acute ferret exposure with a significant increase in fearful and anxious behaviors that is accompanied by robust amygdala activation and an increase in somatostatin receptor 2 (sst2) messenger RNA expression within the amygdala and anterior cingulate cortex. These studies are the first to show stress-induced changes in amygdala sst2 expression and may represent one mechanism by which psychological stress is linked to adaptive and maladaptive behavioral responses.     

Tradeoff Between Travel Speed and Olfactory Food Detection in Ring‐Tailed Coatis (Nasua nasua)

The distance at which an animal can detect food has important ramifications for foraging behavior. Although some studies have investigated the factors which affect visual food detection, very little is known about what influences olfactory food detection abilities in wild animals. The food discovery behavior of ring‐tailed coatis (Nasua nasua) was studied using experimental fruit plots. Coatis primarily used olfaction to detect these new food sources, and visual food discovery appeared plausible in only five of 148 trials. Coatis detected the fruit from longer distances when traveling compared with when foraging for invertebrates in the leaf litter. Travel speed had a negative effect on discovery distance. Coatis traveling slowly detected the fruit plots from further away, which demonstrated a tradeoff between speed and food detection. If this tradeoff is biologically important, slower groups should have visited more fruit trees per day, so data taken during full‐day coati group follows were analyzed to determine whether this pattern occurred. Slower moving groups visited more fruit trees than faster groups once confounding factors such as daily travel distance, group identity, group spread, and year were controlled for. These results are consistent with the hypothesis that coatis exhibit a speed‐accuracy tradeoff for olfactory food detection. This tradeoff appears to be an important factor influencing the movement ecology of animal groups.