Reelin is essential for neuronal migration but not for radial glial elongation in neonatal ferret cortex

Numerous functions related to neuronal migration are linked to the glycoprotein reelin. Reelin also elongates radial glia, which are disrupted in mutant reeler mice. Our lab developed a model of cortical dysplasia in ferrets that shares features with the reeler mouse, including impaired migration of neurons into the cerebral cortex and disrupted radial glia. Explants of normal ferret cortex in coculture with dysplastic ferret cortex restore the deficits in this model. To determine if reelin is integral to the repair, we used explants of P0 mouse cortex either of the wild type (WT) or heterozygous (het) for the reelin gene, as well as P0 reeler cortex (not containing reelin), in coculture with organotypic cultures of dysplastic ferret cortex. This arrangement revealed that all types of mouse cortical explants (WT, het, reeler) elongated radial glia in ferret cortical dysplasia, indicating that reelin is not required for proper radial glial morphology. Migration of cells into ferret neocortex, however, did not improve with explants of reeler cortex, but was almost normal after pairing with WT or het explants. We also placed an exogenous source of reelin in ferret cultures at the pial surface to reveal that migrating cells move toward the reelin source in dysplastic cortex; radial glia in these cultures were also improved toward normal. Our results demonstrate that the normotopic position of reelin is important for proper neuronal positioning, and that reelin is capable of elongating radial glial cells but is not the only radialization factor.     

Functional characterization of ferret CCL20 and CCR6 and identification of chemotactic inhibitors

CCL20 is currently the only known chemokine ligand for the receptor CCR6, and is a mucosal chemokine involved in normal and pathological immune responses. Although nucleotide sequence data are available for ccl20 and ccr6 sequences from multiple species, the ferret ccl20 and ccr6 sequences have not been determined. To increase our understanding of immune function in ferret models of infection and vaccination, we have used RT-PCR to obtain the ferret ccl20 and ccr6 cDNA sequences and functionally characterize the encoded proteins. The open reading frames of both genes were highly conserved across species and mostly closely related to canine sequences. For functional analyses, single cell clones expressing ferret CCR6 were generated, a ferret CCL20/mouse IgG_2a fusion protein (fCCL20-mIgG_2a) was produced, and fCCL20 was chemically synthesized. Cell clones expressing ferret CCR6 responded chemotactically to fCCL20-mIgG2a fusion protein and synthetic ferret CCL20. Chemotaxis inhibition studies identified the polyphenol epigallocatechin-3-gallate and the murine γ-herpesvirus 68 M3 protein as inhibitors of fCCL20. Surface plasmon resonance studies revealed that EGCG bound directly to fCCL20. These results provide molecular characterization of previously unreported ferret immune gene sequences and for the first time identify a broad-spectrum small molecule inhibitor of CCL20 and reveal CCL20 as a target for the herpesviral M3 protein.     

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.     

Epidemic and maintenance of rabies in chinese ferret badgers (Melogale moschata) indicated by epidemiology and the molecular signatures of rabies viruses

An epidemic of Chinese ferret badger-associated human rabies was investigated in Wuyuan county, Jiangxi province and rabies viruses isolates from ferret badgers in different districts in Jiangxi and Zhejiang provinces were sequenced with their nucleotides and amino acids and aligned for epidemiological analysis. The results showed that the human rabies in Wuyuan are only associated with ferret badger bites; the rabies virus can be isolated in a high percentage of ferret badgers in the epidemic areas in Jiangxi and Zhejiang provinces; the isolates share the same molecular features in nucleotides and have characteristic amino acid signatures, i.e., 2 sites in the nucleoprotein and 3 sites in the glycoprotein, that are distinct from virus isolates from dogs in the same region. We conclude that rabies in Chinese ferret badgers has formed an independent transmission cycle and ferret badgers may serve as another important rabies reservoir independent of dog rabies in China.     

Genomic homology of the domestic ferret with cats and humans

We used chromosome paints from both the domestic cat and humans to directly establish chromosomal homology between the genome of these species and the domestic ferret. The chromosome painting data indicate that the ferret has a highly conserved karyotype closer to the ancestral carnivore karyotype than that of the cat. The cat chromosome paints revealed 22 homologous autosomal regions in the ferret genome: 16 ferret chromosomes were hybridized by a single cat paint, while 3 ferret chromosomes were hybridized by two cat paints. In situ hybridization combined with banding showed that ferret Chromosome (Chr) 1 = cat A2p/C2, Chr 2 = F2/C1q, and Chr 3 = A2q/D2. Five ferret chromosomes are homologous to single arms of cat chromosomes: ferret 4 = A1q, 5 = B1q, 6 = C1p, 10 = A1p, and 12 = B1p. The human chromosome paints revealed 32 + XY homologous regions in the ferret genome: 9 ferret chromosomes were each hybridized by a single human paint, 7 by two paints, 3 by three paints. The 10 ferret chromosomes hybridized by multiple human paints produced the following associations: ferret 1 = human 19/3/21, 2 = 8q/2q, 3 = 10/7, 5 = 8/4, 8 = 15/14, 9 = 10/12/22, 11 = 20/2, 12 = 8/4, 14 = 12/22/18, 18 = 19/16. We present an index of genomic diversity, Z, based on the relative number of conserved whole chromosome and chromosome segments as a preliminary statistic for rapid comparison between species. The index of diversity between human-ferret (Z = 0.812) is slightly less than human-cat (Z = 0.843). The homology data presented here allow us to transfer gene mapping data from both cats and humans to the ferret. Received: 21 December 1999 / Accepted: 30 May 2000     

A comparison of sperm morphology and silver nitrate staining characteristics in the domestic ferret and the black-footed ferret

Ejaculated sperm from the domestic ferret (Mustela putorius furo) and the black-footed ferret (Mustela nigripes) were compared for differences in morphological abnormalities and argentophilic protein distribution. Thawed domestic ferret sperm was also compared to fresh sperm to determine whether there were any effects on cell morphology due to cryopreservation. There were statistically significant differences between the two species of ferret in two of the categories scored. The domestic ferret had a higher frequency of cells that were bent in the midpiece and in the principal piece, and a higher frequency of headless and tailless cells when compared to the black-footed ferret. There were no statistically significant differences in cell morphology between the fresh and cryopreserved ejaculates of the domestic ferret employing a standard egg yolk cryoextender. Silver nitrate staining distribution was different between the two species in both the head and tail region.     

Quantitative light and scanning electron microscopy of ferret sperm.

Sperm were obtained via electroejaculation from Domestic ferret, (Mustela putorius furo), Siberian ferret (M. eversmanni), Black-footed ferret (M. nigripes), and a hybrid between Siberian and Domestic, called the Fitch ferret (M. sp.). Comparisons of sperm were made by four different microscopy techniques to determine whether differences exist among species. First, Nomarski differential interference microscopy could be used to distinguish domestic ferret sperm from the others on the basis of the structure of the posterior part of the acrosome. Second, both silver staining, which demonstrates argentophilic protein distribution, and scanning electron microscopy (SEM), revealed differences among the morphology of sperm for each species; variation in the unique appearance of the acrosome in ferret sperm was detected especially well by SEM. To quantify differences in morphology, five sperm head parameters were measured using image analysis; light microscopy produced significantly larger values than did SEM (all parameters and all species but Fitch), and there were significant differences owing to species for all parameters but one. Generally, our data demonstrate the value of complementary techniques to distinguish among sperm of closely related species and more specifically may help establish evolutionary relationships among the ferret species studied. In addition, they provide baseline data important for the captive breeding of the endangered Black-footed ferret.     

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.     

Comparison of black-footed ferret (Mustela nigripes) and domestic ferret (M. putorius furo) courtship activity

Because of the scarcity of the endangered black-footed ferret (Mustela nigripes) and the amount of knowledge necessary for their conservation, surrogate research can play an important role in recovery. In this paper, we investigate surrogate behavioral research potential by comparing courtship behavior of the black-footed ferret to the congeneric domestic ferret (M. putorius furo). Ten female domestic ferrets were bred to five male domestic ferrets and eight female black-footed ferrets were bred to five black-footed ferret males. Courtship activities were defined, analyzed, and quantitatively compared between both groups. Lag sequential analysis of was used to prepare the behavioral matrices, and matrix cells were compared between groups with an equality of proportions test. Courtship patterns did not differ significantly between the two closely related species, and the domestic ferret would probably be an adequate surrogate for reproductive behavior research on the black-footed ferret.     

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.     

Immunologic comparisons of Pneumocystis carinii strains obtained from rats, ferrets, and mice using convalescent sera from the same sources.

Pneumocystis carinii (Pc) infections were developed in animals immunosuppressed by dexamethasone treatment either from activation of latent infection (ferret) or trans-tracheal inoculation of Pc obtained from infected lungs of the homologous species (rat, mouse). Convalescent antisera were obtained by stopping dexamethasone treatment after 2-4 wk and allowing 5-8 wk for recovery. Parasites from infected lungs were purified by differential filtration, solubilized in loading buffer, subjected to sodium dodecyl sulfate- polyacrylamide gel electrophoresis, and blotted to polyvinylidene fluoride sheets for Western analysis. Antisera from each animal species were reacted on Western blots of antigens from rat, ferret, and mouse. Each combination of antigen and antibody from the same species of animal showed reaction with 5 or more bands of Pc antigen. Convalescent mouse antibody did not react with rat or ferret antigens. Convalescent rat antibody reacted with a mouse antigen at about 66 kDa but not with ferret antigen, and convalescent ferret antibody showed minimal, probably non-specific reactions with both rat and mouse antigens. Variations in reactions indicate antigenic differences in Pc strains infecting these animals.     

The isolated, buffer-perfused ferret heart: a new model for the study of cardiac physiology and metabolism.

The isolated, buffer-perfused ferret heart is a new model for the study of cardiac physiology and metabolism. Compared to the more commonly used isolated heart preparation, the rat heart, the ferret has a lower rate-pressure product due to lower heart rate, a remarkably low coronary flow and almost complete oxygen extraction. The ferret heart remains in stable haemodynamic and metabolic conditions for a longer period of time than the rat heart. ATP contents of the two species are similar, but creatine phosphate content is higher in the ferret while NAD content is much lower.     

Comparative processing and function of human and ferret cystic fibrosis transmembrane conductance regulator

The most common cystic fibrosis transmembrane conductance regulator (CFTR) gene mutation is ΔF508, and this causes cystic fibrosis (CF). New CF models in the pig and ferret have been generated that develop lung, pancreatic, liver, and intestinal pathologies that reflect disease in CF patients. Species-specific biology in the processing of CFTR has demonstrated that pig and mouse ΔF508-CFTR proteins are more effectively processed to the apical membrane of airway epithelia than human ΔF508-CFTR. The processing behavior of ferret WT- and ΔF508-CFTR proteins remains unknown, and such information is important to predicting the utility of a ΔF508-CFTR ferret. To this end, we sought to compare processing, membrane stability, and function of human and ferret WT- and ΔF508-CFTR proteins in a heterologous expression system using HT1080, HEK293T, BHK21, and Cos7 cells as well as human and ferret CF polarized airway epithelia. Analysis of the protein processing and stability by metabolic pulse-chase and surface On-Cell Western blots revealed that WT-fCFTR half-life and membrane stability were increased relative to WT-hCFTR. Furthermore, in BHK21, Cos7, and CuFi cells, human and ferret ΔF508-CFTR processing was negligible, whereas low levels of processing of ΔF508-fCFTR could be seen in HT1080 and HEK293T cells. Only the WT-fCFTR, but not ΔF508-fCFTR, produced functional cAMP-inducible chloride currents in both CF human and ferret airway epithelia. Further elucidation of the mechanism responsible for elevated fCFTR protein stability may lead to new therapeutic approaches to augment CFTR function. These findings also suggest that generation of a ferret CFTR(ΔF508/ΔF508) animal model may be useful.     

A comparison of the composition and apolipoprotein content of the lipoproteins isolated from human and ferret (Mustela putorius furo L.) serum.

1. The lipoproteins isolated at densities of less than 1.006, 1.006-1.063 and 1.063-1.21 g/ml from human and ferret (Mustela putorius furo L.) serum were compared. 2. Ferret very low density lipoprotein contained proportionately less triglyceride and more phospholipid than human. 3. Ferret low density lipoprotein contained proportionately more triglyceride and less cholesterol than human. 4. High density lipoprotein was the major lipoprotein in ferret serum. 5. The gel electrophoretic patterns of lipoprotein apoproteins and the pattern of apoprotein solubility in tetramethylurea were similar for human and ferret fractions. 6. The ferret may provide a convenient animal for the study of serum lipoprotein structure, function and metabolism.     

Population Indices Versus Correlated Density Estimates of Black-Footed Ferret Abundance

ABSTRACT Estimating abundance of carnivore populations is problematic because individuals typically are elusive, nocturnal, and dispersed across the landscape. Rare or endangered carnivore populations are even more difficult to estimate because of small sample sizes. Considering behavioral ecology of the target species can drastically improve survey efficiency and effectiveness. Previously, abundance of the black-footed ferret (Mustela nigripes) was monitored by spotlighting and generating indices of relative abundance because reintroduced populations were slow to establish. Indices, however, lack variance estimates and are costly to generate for the black-footed ferret. We therefore used spotlight surveys and live-trapping in conjunction with a robust mark-recapture estimator to improve abundance monitoring for the black-footed ferret, one of North America's most endangered carnivores. We estimated abundance of the black-footed ferret at Shirley Basin, Wyoming, USA, using correlated density estimates and Program MARK. We compared our results to 2 indices of relative abundance, minimum number alive and predicted number of ferrets from litter counts. The correlated density estimate for the black-footed ferret (N̂_R = 229; 95% CI = 161–298) was similar to minimum number alive (N̂_R = 192) and predicted number of ferrets from litter counts (N̂_R= 235). The efficiency and effectiveness of survey methods we used for the black-footed ferret were high by carnivoran standards. Our results suggest that the sampling approach we utilized can be implemented for a fraction of the cost and effort required to generate 2 indices of relative abundance for the black-footed ferret. Although we recommend managers implement a similar survey approach to monitor abundance of reintroduced populations of the black-footed ferret, analysis with sparse data sets will be problematic. Until the black-footed ferret becomes widespread and abundant at a reintroduction site, spotlighting will remain preferable as a means to generate indices of distribution and relative abundance for the black-footed ferret.     

Establishment and characterization of ferret cells in culture

Cells derived from the brain of a 6 wk-old ferret have been subcultured over 100 times and have undergone over 400 population doublings in vitro. These cells, referred to as Mpf cells, have an absolute efficiency of colony formation in excess of 45%, exhibit a mean population doubling time of 12.5 h, possess ferret-specific antigens, and have isozymes with electrophoretic properties that are the same as those of isozymes found in ferret liver. The cells exhibit a cytopathic effect and support the synthesis of progeny virus when they are infected with the viruses of lymphocytic choriomeningitis, Newcastle disease, pseudorabies, Sindbis, vaccinia, and vesicular stomatitis. The passage level of the Mpf cells, their elapsed number of population doublings, their possession of ferret-specific antigens, and the comigration of four isozymes obtained from these cells and ferret liver define the cells as an established line of ferret cells.     

The electrocardiogram of normal ferrets and ferrets with right ventricular hypertrophy

Sixty-eight electrocardiograms were recorded on ferrets (Mustela putorius furo). These represent 29 normal weanling males, 19 normal adult males and 20 adult males with right ventricular hypertrophy (RVH). Analyses of rate, rhythm, axis and total voltage were used to define the normal electrocardiogram (ECG) and to identify changes seen in RVH. The normal ferret has a heart rate of about 300 beats per minute and a mean electrical axis of + 86 degrees +/- 6.6 (SD). A 56% increase in right ventricular weight to body weight ratio was not associated with right axis deviation. The overall voltage produced on the ECG was increased in the group with RVH as compared to the normal group (p less than 0.030).     

The ferret as a model organism to study influenza A virus infection

Influenza is a human pathogen that continues to pose a public health threat. The use of small mammalian models has become indispensable for understanding the virulence of influenza viruses. Among numerous species used in the laboratory setting, only the ferret model is equally well suited for studying both the pathogenicity and transmissibility of human and avian influenza viruses. Here, we compare the advantages and limitations of the mouse, ferret and guinea pig models for research with influenza A viruses, emphasizing the multifarious uses of the ferret in the assessment of influenza viruses with pandemic potential. Research performed in the ferret model has provided information, support and guidance for the public health response to influenza viruses in humans. We highlight the recent and emerging uses of this species in influenza virus research that are advancing our understanding of virus-host interactions.     

Antibodies to the N-terminus of human Gastrin-34 react with material in rat and ferret brain

Antibodies specific for the N-terminus of human big gastrin, or NT G34, reveal in immunohistochemistry extensive systems of nerve cell bodies and fibres in the rat and ferret brain. However, when the same antibodies were used in radioimmunoassay of rat and ferret brain tissue extracts they failed to reveal immunoreactive material. At least one of the antibodies used for radioimmunoassay could be shown to react with NT G34 in rat pyloric antral extracts. Antibodies specific for other peptides derived from progastrin failed to reveal the systems demonstrated with the N-terminal G34 antibodies. It is concluded that expression of the gastrin gene is unlikely to account for the present observations. Instead we suggest that a novel peptide with low affinity for NT G34 specific antibodies is found in rat and ferret central neurones.     

Establishment and characterization of ferret cells in culture

Summary Cells derived from the brain of a 6 wk-old ferret have been subcultured over 100 times and have undergone over 400 population doublings in vitro. These cells, referred to as Mpf cells, have an absolute efficiency of colony formation in excess of 45%, exhibit a mean population doubling time of 12.5 h, possess ferret-specific antigens, and have isozymes with electrophoretic properties that are the same as those of isozymes found in ferret liver. The cells exhibit a cytopathic effect and support the synthesis of progeny virus when they are infected with the viruses of lymphocytic choriomeningitis, Newcastle disease, pseudorabies, Sindbis, vaccinia, and vesicular stomatitis. The passage level of the Mpf cells, their elapsed number of population doublings, their possession of ferret-specific antigens, and the comigration of four isozymes obtained from these cells and ferret liver define the cells as an established line of ferret cells.