Mapping of rabbit chromosome 1 markers generated from a microsatellite-enriched chromosome-specific library.

A genomic DNA library was produced from flow-sorted rabbit chromosome 1 and enriched for fragments containing CA-repeats. Clones containing CA-repeats were identified and primers for amplification of the microsatellite were developed after sequencing the clone. The degree of polymorphism was tested in rabbits from different breeds. This approach identified 12 microsatellite markers which could be used for studying linkage relationships in the progeny of an F(2)-intercross: (AX/JUxIIIVO/JU) F(2), and two backcrosses: (OS/JxX/J)X/J and (WH/JxX/J)X/J. Seven of these markers were mapped on chromosome 1.

The presence and influence of religion in American bioethics.

From the inception of the relatively short history of American bioethics in the mid-to-late 1960s, the place of religion in this field has been complex and controversial. It has also been a subject of more than casual interest and concern to bioethicists, and to an array of medical and non-medical groups in U.S. society for whom the activities and issues in which bioethics is engaged have ongoing import. The questions and the tensions linked to the status and influence of religion in the sphere of bioethics have ramifications that extend beyond bioethics and biomedicine into matters involving the relationship of religion to the institutional structure of American society--most particularly its political, legal foundations, and realm of public affairs--and to its cultural attributes and tradition. It is within this larger perspective that we will consider the association between American bioethics and religion. Our analysis includes two case studies: (1) how, in the early years of bioethics, a pioneering organization in the field dealt with the "redefinition of death" in its discussions and in a major medical journal publication; and (2) the way in which the most recently appointed federal bioethics commission, the National Bioethics Advisory Commission, involved religion in its work on cloning and stem cell research.

FVB/N: an inbred mouse strain preferable for transgenic analyses.

FVB/N mice offer a system suitable for most transgenic experiments and subsequent genetic analyses. The inbred FVB/N strain is characterized by vigorous reproductive performance and consistently large litters. Moreover, fertilized FVB/N eggs contain large and prominent pronuclei, which facilitate microinjection of DNA. The phenotype of large pronuclei in the zygote is a dominant trait associated with the FVB/N oocyte but not the FVB/N sperm. In experiments to generate transgenic mice, the same DNA constructs were injected into three different types of zygotes: FVB/N, C57BL/6J, and (C57BL/6J x SJL/J)F1. FVB/N zygotes survived well after injection, and transgenic animals were obtained with efficiencies similar to the F1 zygotes and much better than the C57BL/6J zygotes. Genetic markers of the FVB/N strain have been analyzed for 44 loci that cover 15 chromosomes and were compared with those of commonly used inbred strains. In addition to the albino FVB/N strain, pigmented congenic strains of FVB/N are being constructed. These features make the FVB/N strain advantageous to use for research with transgenic mice.

Genetics of a tissue esterase polymorphism (Est-6) in the rabbit (Oryctolagus cuniculus).

Genetic analysis of a polymorphic tissue esterase revealed a new locus (Est-6) with two alleles (Est-6a and Est-6b) on linkage group VI of the rabbit. Est-6 is closely linked to the Est-1,2,4 cluster. Esterase of Est-6 is found in many organs, particularly in liver and small intestine, but not in erythrocytes and serum. Est-6 esterase hydrolyzes alpha-naphthyl acetate and butyrate, naphthol AS-D acetate, indoxyl acetate, and butyrate as well as 5-bromoindoxyl acetate, N-acetyl-L-alanine-alpha-naphthyl ester but not 4-methylumbelliferyl acetate and fluorescein diacetate. The enzyme is inhibited by bis-p-nitrophenyl phosphate and eserine but not by p-chloromercuribenzoate. It was classified as a carboxylesterase (EC 3.1.1.1). Based on chromosomal localization, tissue distribution, substrate specificity, inhibitor sensitivity, and range of pI's, rabbit Est-6 is assumed to be homologous with mouse Es-7.

The osteopetrotic rabbit: general and skeletal features of a new outbred stock.

Stock of the lethal osteopetrotic mutation in the rabbit has been outbred to robust New Zealand White stock. Features of this new stock include skeletal sclerosis and failure of tooth eruption characteristic of osteopetrosis, together with hypocalcemia, hypophosphatemia, and osteoclasts of reduced numbers and abnormal cytology. No abnormalities of carbonic anhydrase II were detectable by electrophoresis. The commercial availability of mutants of this new stock makes them a potentially important source of information about both osteopetrosis and osteoclast biology.

Electron-microscopic analysis of nephroblastomas induced transplacentally in the IIIVO/J rabbit by a single dose of N-ethylnitrosourea.

Eight examples of nephroblastoma induced transplacentally in the partially inbred IIIVO/J strain of rabbit by a single intraperitoneal (60 mg/kg) dose of N-ethylnitrosourea (ENU) were studied by transmission electron-microscopy. At the light-microscopic level the ENU-induced tumors displayed the complex array of histotypic components described in a previous report (Hard GC, Fox RR: Histologic characterization of renal tumors (nephroblastomas) induced transplacentally in IIIVO/J and WH/J rabbits by N-ethylnitrosourea. Am J Pathol 1983, 113:8-18) namely, blastema, tubular profiles, "reniform" islands, glomeruloid bodies, squamoid foci, fascicles of mesenchymelike tumor cells, and an increasing fibrocollagenous stroma. Ultrastructurally, blastemalike cells were undifferentiated forms resembling metanephric blastema. Tubular profiles of varying development were typified by very prominent apical junctional complexes and a basal lamina, but no organized brush-border. "Reniform" islands were groups of more maturely formed tubules associated with the production of an interstitial matrix consisting almost solely of multilamellar basement membrane. Glomeruloid bodies were invaginations of small podocytelike cells, with a profusion of thin cytoplasmic processes resembling foot processes, and internal, homogeneous areas of basement membrane continuous with that surrounding the entire structure. As such, these structures were consistent with primitive attempts at glomerular differentiation but without vascular or mesangial elements. The squamoid foci were representative of true squamous differentiation in comprising cells filled with intermediate filaments and interconnected by lateral interdigitations and multiple, well-formed desmosomes. Spindle-shaped tumor cells disposed in fascicles, which could have been interpreted as bipotential differentiation into secondary mesenchyme at the histologic level, differed from the blastemal cell type only in shape. Ultrastructurally, the fascicles, in most cases, consisted of thin, basement-membrane-invested ribbons of attenuated cells enclosing small lumens sealed by intercellular junctions, suggestive of abortive tubule formation. Tumor cells disposed in fascicles therefore conformed to the same epithelial lineage as the other neoplastic components of these tumors. In contrast, cellular elements of the fibrocollagenous stroma--namely, fibroblasts, vascular endothelial cells, and scattered mononuclear inflammatory cells--could be clearly discriminated from the various tumor cell components on morphologic grounds, constituting a host reaction.(ABSTRACT TRUNCATED AT 400 WORDS)

A successful technique for the preservation of rabbit embryos.

A technique for successfully freezing, thawing and transferring rabbit embryos has been developed. Morula stage embryos were collected from super-ovulated female rabbits by flushing both oviducts and uterine horns with a tissue culture medium. Well developed, viable embryos were then transferred to freezing vials and a cryoprotectant, dimethyl sulfoxide (DMSO) was added in several steps to bring its final concentration to 1.6 molar. To freeze the embryos the temperature was lowered slowly (either 0.5 degrees C/min or 1.0 degrees C/min) to -80 degrees C at which point the vials were transferred directly to liquid nitrogen (-196 degrees C). Thawing was done at 8 degrees C/min. After thawing, phosphate buffered saline was added in a stepwise manner to dilute the DMSO. The thawed embryos were then cultured at 37 degrees C. Transfer of the embryos was accomplished by laparotomizing a pseudopregnant doe and introducing the embryos into the fimbriated ends of the oviducts. The 101 positively transferred embryos resulted in 45 implantations and 34 live born young.

Atropine esterase status of laboratory mice.

Fifty eight inbred strains of laboratory mice and two species of wild mice [Mus castaneus (inbred) and Peromyscus maniculatus bairdii (closed colony)] were tested for the presence of atropine esterase by the microhematocrit tube test of Tucker and Beattie. The results show clearly that laboratory mice do not have significant levels of atropine esterase as determined either by the microhematocrit tube test or by the agar plate test, and therefore that under these conditions atropine esterase is not a polymorphic variant as has been demonstrated clearly in the rabbit.

Genetic heterogeneity of rabbit alpha-1-antitrypsin.

Sixteen inbred or partially inbred strains of rabbits were investigated for electrophoretic and quantitative variations of alpha-1-antitrypsin (A-1-AT). We found interindividual differences in the electrophoretic A-1-AT patterns as well as quantitative differences in the concentrations of A-1-AT and the serum trypsin-inhibiting activity. Three electrophoretic phenotypes were distinguished: M, P and MP. M was characterized by a predominant anodal A-1-AT band, and P had a major cathodal component. The MP pattern can be explained by the occurrence of the M and P components in the same serum due to heterozygosity. The P pattern was associated with an A-1-AT concentration of approximately 56% of that in sera with the M phenotype. The levels of A-1-AT in sera with the MP phenotype were intermediate between those in M and P types. In addition to the type-specific quantitative variation, we found a quantitative sexual dimorphism of a moderate degree: Female rabbits had A-1-AT concentrations 16% less than males.

Spontaneous degenerative spinal disease in the laboratory rabbit.

The spines of 35 rabbits (32 New Zealand white and 3 AC/J), ranging in age from 3 months to 8 1/2 years, were investigated systematically for spontaneous degenerative changes. Three types of lesion were observed. (1) The nucleus pulposus underwent chondroid metaplasia throughout the length of the vertebral column by the age of 2 years. (2) Hydroxyapatite deposition was found in the nucleus pulposus in 12 of 20 animals examined roentgenographically. The lesion occurred principally in the distal thoracic segments and was first observed in 3-month-old rabbits. (3) Spondylosis occurred in each of four macerated spines from animals greater than 24 months old. Portions of the spine spared by disc calcification were affected.

Histologic characterization of renal tumors (nephroblastomas) induced transplacentally in IIIVO/J and WH/J rabbits by N-ethylnitrosourea.

The histologic features of 63 renal tumors induced in 39 rabbits of two partially inbred strains, IIIVO/J and WH/J, by transplacental exposure to N-ethylnitrosourea (ENU) were analyzed. All tumors in the series conformed to nephroblastoma, permitting the establishment of histologic standards for this neoplasm in the rabbit as well as observations on tumor progression. Essentially, nephroblastoma proved to be predominantly an epithelial tumor identifying with metanephrogenic blastema, which was presumed to be the tissue of origin during fetal development. The outstanding features comprised clusters or sheets of undifferentiated blastemalike tissue and differentiation along the epithelial pathway into tubular profiles and structures suggestive of primitive, nonvascularized glomeruli. The latter were frequently of a complex nature, with a papillary configuration. On the other hand, no definitive evidence of bipotential differentiation into malignant secondary mesenchyme was found, there being no recognizable areas of fibrosarcomatous elements or specialized connective tissue such as smooth or striated muscle, adipose tissue, cartilage, or osteoid. Mesenchymelike fascicular disposition of neoplastic cells between blastemal clusters was an acquired feature seen in advanced tumors but not in small early lesions. By light microscopy alone it was not possible to determine whether this represented a conformational change of tumor cells or true bipotential differentiation into neoplastic secondary mesenchyme. However, the reticulin pattern was not characteristic of sarcoma. A conspicuous feature accompanying the growth and development of tumors was the magnitude of host fibrous reaction discernible only as a simple ramifying stroma in the earliest lesions but attaining impressive proportions both within and around the tumor with advancing age. Increasing collagen formation appeared to be associated with ischemic necrosis of tumor tissue. Other features of advanced tumors were the presence of discrete foci of differentiated tubular structures suggestive of mature medullary elements and small islands of squamoid differentiation. Metastases occurred only in rabbits of strain IIIVO/J, which had been subjected to a single dose of the carcinogen, representing an incidence in this subgroup of 25%. Nephroblastomas resulting from transplacental induction in IIIVO/J rabbits, particularly by single, high doses of ENU, appear to provide a suitable model for the predominant histologic form of the Wilms' tumor complex in man.

Genetics of two tissue esterase polymorphisms (Est-4 and Est-5) in the rabbit.

Two polymorphic esterase systems were found after electrophoresis of rabbit tissue homogenates. Each of these systems is controlled by an autosomal locus with two alleles. Est-4 determines the absence (Est-4a) or presence (Est-4b) of two bands of esterase activity with intermediate anodcal mobility and broad substrate specificity. This polymorphism was found to be present in liver, small intestine, and spleen but not in kidney, heart, and testis. Est-5 is coding for cathodally migrating esterases which differ in mobility (Est-5a and Est-5b). This polymorphism was found only in kidney and testis homogenates. Est-5 esterases are more active against alpha-naphthyl acetate than against beta-naphthyl acetate and have no activity against alpha-naphthyl butyrate. Linkage analysis indicated that Est-4 is localized on rabbit LG VI as part of a cluster of esterase loci, whereas Est-5 segregates independently, Rabbits from two inbred and nine partly inbred strains were tested for these polymorphisms.

Rabbit linkage group VIII: the alleles of the prt gene.

The prt gene which is linked to the rabbit immunoglobulin kappa-light chain gene, ab, has two phenotypes, PRT+ and PRT-. These phenotypes can be distinguished only when serum proteins from different rabbits are separated by polyacrylamide gel electrophoresis. The serum protein profiles for PRT+ rabbits show a band that is located on the anodal side of transferrin. This band is missing in the serum profiles of PRT- rabbits. However, the PRT protein is present in these rabbits. An antiserum which reacts with PRT from PRT+ rabbits detects two electrophoretic variants of PRT which are located in areas of the polyacrylamide gel obscured by other serum proteins. These results and other suggest that the prt gene has three alleles, the prta allele encoding the protein found in PRT+ rabbits and the prtb and prtc alleles encoding the two electrophoretic variants found in PRT- rabbits.

Separation of beta-D-galactosidases in rabbit tissues: genetics of neutral beta-D-galactosidase.

Three different types of beta-D-galactosidase (EC 3.2.1.23) could be distinguished in rabbit tissues using electrophoretic procedures. (1) Acid beta-D-galactosidase with a low mobility and maximal activity at pH 3-5 was found in the particulate fraction of various tissue homogenates. This enzyme hydrolyzed 4-methylumbelliferyl-D-galactoside, but no activity against other glycoside substrates could be demonstrated. The enzyme was inhibited by galactono-(1 leads to 4)-lactone. (2) Lactose-hydrolyzing beta-D-galactosidase with an intermediate mobility was found only in juvenile small intestine. Most of the activity was found in the particulate fraction of the cell. The enzyme hydrolyzed several other synthetic glycoside substrates besides lactose. It was most active at pH 5-6 and strongly inhibited by glucono-(1 leads to 5)-lactone but not much affected by galactono-(1 leads to 4)-lactone. (3) Neutral beta-D-galactosidase with a fast mobility and maximal activity at pH 6-8 was found in the soluble fraction of homogenates from liver, kidney, and small intestine. This enzyme also showed a broad substrate specificity; it possessed activity against aryl-beta-D-glucoside, -fucoside, and -galactoside substrates but not against lactose. The enzyme was strongly inhibited by glucono-(1 leads to 5)-lactone and (less) by galactone-(1 leads to 4)-lactone. Neutral beta-D-galactosidase and neutral beta-D-glucosidase (EC 3.2.1.21) are probably identical enzymes in the rabbit. Individual variation, in both electrophoretic mobility and activity, was found for neutral beta-D-galactosidase. Genetic analysis of the electrophoretic variants revealed that two alleles at an autosomal locus are responsible for this variation.

Evidence for an hereditary defect in taurine transport in the ciliary epithelium of an inbred strain of rabbits.

The concentration of taurine in the aqueous humour and serum of 21 rabbits with hereditary buphthalmia (Bu rabbits-genotype bu/bu) was compared with the aqueous and serum taurine levels of eight strain-related normal rabbits (JAX) and nine non-strain-related normal rabbits (MCV). There was a significant difference in the mean aqueous taurine concentration in each of the three groups. The Bu rabbits had only 29% of the MCV rabbits' level while the JAX rabbits were intermediate with 56% of the MCV level. It is suggested that some of the JAX rabbits may be heterozygous and the Bu rabbits homozygous for a semi-dominant allele of a gene that is less efficient in taurine transport in the ciliary epithelium than the normal allele represented by the MCV animals.

Narrow axis: an inherited anomaly of the second cervical vertebra in the rabbit.

Narrow axis, an inherited anomaly resulting in a marked narrowing of the second cervical vertebra, has been observed in strain X/J rabbits. This condition is first recognizable on X rays at 32-33 days gestation. For size comparisons 21 measurements of the first five cervical vertebrae were taken on the skeletons of each of 14 strain X/J animals (7 normal and 7 with narrow axis) and 14 IIIC/J animals for control at two months of age and 27 strain X/J (11 normal and 16 narrow axis) and 14 strain IIIC/J at seven months of age. The primary effect appeared to be a premature fusion of the centrum with its neural arches. Expression is variable. The effect on the posterior articulation of the atlas appeared to be secondary and adaptive. The other cervical vertebrae and the foramen magnum were relatively unaffected. In the 20-year period encompassed in this report, X rays of 3244 rabbits were used for genetic analysis. Inheritance appears to be due to a single autosomal recessive gene with incomplete penetrance. The condition is neither sex-linked nor sex-limited. We propose the symbol nx for the gene responsible for narrow axis in the rabbit.

Identification of genetically homozygous rapid and slow acetylators of drugs and environmental carcinogens among established inbred rabbit strains.

Liver and gut mucosa N-acetyltransferase (NAT) cytosol (105,000 x g) was prepared from selected lines of New Zealand White rapid and slow acetylator rabbits bred and housed at the University of Michigan, and from inbred and partially inbred rabbits obtained from The Jackson Laboratory. Liver NAT activity was determined with p-aminobenzoic acid, p-aminosalicyclic acid, procainamide, sulfamethazine, isoniazid and 2-aminofluorene as substrates. Gut mucosal NAT activity was determined with 2-aminofluorene. A gene dose-response relationship was observed for both liver NAT and gut mucosa NAT with all substrates tested. Highest levels were always observed in homozygous rapid acetylator inbred strains (B/J, III/J, IIIC/J, III/DwJ, IIIEP/J and IIIVO/J), lower levels in obligate heterozygous rapid acetylator rabbits and lowest levels in homozygous slow acetylator inbred (ACEP/J, III/cdJ, IIIVO/ahJ, and IIIVO/vptJ) and outbred rabbits. The differences in magnitude of liver NAT activity level between acetylator genotypes was dependent on the substrate employed, progressively increasing in the following order: p-aminobenzoic acid, p-aminosalicyclic acid, procainamide, sulfamethazine, isoniazid, 2-aminofluorene. The determination of kinetic constants for liver p-aminosalicyclic acid NAT activity indicated a 2-fold difference in apparent Vmax between rapid acetylator genotypes and a 30-fold difference between rapid and slow acetylator phenotypes. In addition, the apparent Km for p-aminosalicyclic acid was significantly lower in the slow acetylators than in the rapid acetylators.