Increased glycogen synthase kinase-3 activity in diabetes- and obesity-prone C57BL/6J mice.

Although the precise mechanisms contributing to insulin resistance and type 2 diabetes are unknown, it is believed that defects in downstream components of the insulin signaling pathway may be involved. In this work, we hypothesize that a serine/threonine kinase, glycogen synthase kinase-3 (GSK-3), may be pertinent in this regard. To test this hypothesis, we examined GSK-3 activity in two inbred mouse strains known to be susceptible (C57BL/6J) or resistant (A/J) to diet-induced obesity and diabetes. Examination of GSK-3 in fat, liver, and muscle tissues of C57BL/6J mice revealed that GSK-3 activity increased twofold in the epididymal fat tissue and remained unchanged in muscle and liver of mice fed a high-fat diet, compared with their low-fat diet-fed counterparts. In contrast, GSK-3 activity did not change in the epididymal fat tissue of A/J mice, regardless of the type of diet they were fed. In addition, both basal and diet-induced GSK-3 activity was higher (2.3- and 3.2-fold, respectively) in the adipose tissue of C57BL/6J mice compared with that in A/J mice. Taken together, our studies suggest an unsuspected link between increased GSK-3 activity and development of insulin resistance and type 2 diabetes in fat tissue of C57BL/6J mice, and implicate GSK-3 as a potential factor contributing to susceptibility of C57BL/6J mice to diet-induced diabetes.

Genetic analysis of behavioral, neuroendocrine, and biochemical parameters in inbred rodents: initial studies in Lewis and Fischer 344 rats and in A/J and C57BL/6J mice.

Previous work has identified inherent behavioral, neuroendocrine, and biochemical differences among inbred rodent strains that have been related to the animals' differential responsiveness to drugs of abuse or stress. In the present study, we sought to determine (1) whether there are genetic correlations among particular phenotypic traits that differ between a pair of inbred rat strains (Lewis and Fischer 344) or a pair of inbred mouse strains (A/J and C57BL/6J); (2) which of these traits might be amenable to quantitative trait locus analysis; and (3) whether additional behavioral or biochemical differences relevant to drug- or stress-responsiveness could be identified in these strains. Specifically, we measured several behavioral, neuroendocrine, and biochemical traits in parental Lewis and Fischer 344 rats and in 298 members of an F2 intercross population, as well as in parental A/J and C57BL/6J mice and in 11 of the AXB/BXA recombinant inbred mouse strains. Traits measured included exploratory locomotor activity in a novel environment; amphetamine-induced locomotor activity; several specific protein levels in striatal regions, including inhibitory G protein subunits, the dopamine transporter, the Fos family member transcription factor DeltaFosB, and the protein phosphatase inhibitor DARPP-32; and late-afternoon plasma corticosterone concentrations. Each of the traits measured in F2 rats or recombinant inbred mice appears to be influenced by multiple genes, as well as by environmental factors. There were statistically significant, albeit relatively weak, correlations among several traits in an F2 intercross population bred from Lewis and Fischer rats. Among the traits studied in Lewis and Fischer rats, one seemed most amenable to quantitative trait locus analysis: the level of the inhibitory G-protein subunit, Galphai, in the nucleus accumbens. We also found a robust genetic correlation between levels of DeltaFosB and levels of the dopamine transporter in striatal regions in AXB/BXA recombinant inbred mouse strains. While these studies demonstrate the likely complexity of the genetic factors that influence the numerous phenotypes associated with altered responsiveness to drugs of abuse and stress, they represent an initial and necessary step toward identifying specific genetic factors involved.

Genetic regulation of lipopolysaccharide-induced interleukin 1 production by murine peritoneal macrophages.

The production of IL 1 by LPS-stimulated peritoneal macrophages from inbred mouse strains was studied. Macrophages from A/J (A) mice were deficient in IL 1 production, when compared with high IL 1-producing strains, including C57BL/6J (B). The difference between A and B macrophages was maintained over a wide LPS concentration range and throughout a 72-hr incubation period. Because of these differences, it was possible to investigate the mechanisms regulating IL 1 production by applying techniques of genetic analysis by using recombinant inbred (RI) strains derived from the A and B progenitors. A strain distribution pattern (SDP) of IL 1 production (low/high response) was obtained with the use of 15 AXB/BXA RI strains. This suggested the presence of a major gene locus controlling the production of IL 1 in response to LPS stimulation, with allelic differences presumably resulting in deficient or efficient IL 1 production. In addition, there appeared to be one or more other loci involved in determining the magnitude of the IL 1 response to LPS in the responder mice. The IL 1 response did not appear to be linked to the major histocompatibility complex, since B10.A mice (which share the same H-2a haplotype as A/J) were efficient IL 1 producers. There did not appear to be any correlation between the degree of IL 1 production and the magnitude of the peritoneal macrophage inflammatory response, or between IL 1 production and LPS responsiveness (as determined by splenocyte proliferation). SDP analysis also indicated that the IL 1 response was not linked to macrophage tumoricidal activity. A comparison of the SDP for IL 1 production with a library of SDP for other known genetic waits suggested linkage with at least four loci on chromosome 1.

Kinetics of arylamine N-acetyltransferase in tissues from rapid and slow acetylator mice.

Kinetic parameters for arylamine N-acetyltransferase activity in liver, blood, and bladder from C57BL/6J and A/J mouse strains were determined using an improved assay system, and some deviations were found from previously reported results. In the present studies, blood N-acetyltransferase activity with p-aminobenzoic acid and 2-aminofluorene as substrates was 20- and 10-fold greater, respectively, in C57BL/6J than in A/J mice. Urinary bladder possessed N-acetyltransferase activity for both 2-aminofluorene and p-aminobenzoic acid which differed 2-fold, and reflected the liver and blood phenotype. An apparent Km difference for 2-aminofluorene was observed between C57BL/6J and A/J liver N-acetyltransferase. Contrary to earlier studies, the liver N-acetyltransferase activity differed 3-fold between the A/J and C57BL/6J mouse strains, with either p-aminobenzoic acid or 2-aminofluorene as substrates. Dimethylsulfoxide at concentrations used in the 2-aminofluorene acetylation assay in earlier studies, inhibited the A/J liver N-acetyltransferase to a greater extent than the C57BL/6J enzyme, which may have contributed to the larger difference in liver NAT activity with 2-aminofluorene reported previously.

Metabolism and DNA binding of 2,6-dinitrotoluene in Fischer-344 rats and A/J mice.

2,6-Dinitrotoluene (2,6-DNT) is a potent hepatocarcinogen in Fischer-344 rats, while its 2,4-isomer is believed to be noncarcinogenic. Neither 2,6-DNT nor 2,4-DNT is carcinogenic in the strain A mouse lung tumor bioassay. To explore the possible reasons for these differences in tumor responses, we have studied the in vitro metabolism and DNA binding of 2,6-DNT in cultured hepatocytes of the Fischer-344 rat and the A/J mouse, and have also investigated the in vivo DNA binding of 2,6-DNT and 2,4-DNT in these two species. In vitro metabolism of 2,6-DNT by rat and mouse hepatocytes was similar and resulted mainly in the formation of 2,6-dinitrobenzyl alcohol, either unconjugated or as a glucuronide (57.5 to 85.5% of the total per fraction), with smaller amounts of polar, acidic metabolites (8.4 to 38.7%) and minor amounts (1.2 to 5.3%) of 2-amino-6-nitrotoluene. Anaerobic metabolism of 2,6-DNT by an extract of rat or mouse cecal contents resulted mainly in the formation of 2-amino-6-nitrotoluene and 2-(N-acetylamino)-6-nitrotoluene, and minor amounts of 2,6-diaminotoluene. Ip administration of 2,6-DNT or 2,4-DNT (150 mg/kg each) to Fischer-344 rats resulted, after 24 hr, in covalent binding to DNA of the liver (131.1 to 259.9 pmol 2,6-DNT/mg DNA; 215.4 to 226.8 pmol 2,4-DNT/mg DNA), and lower binding to DNA of the lungs and the intestine (14.9 to 22.7 pmol 2,6-DNT/mg DNA; 45.0 to 75.0 pmol 2,4-DNT/mg DNA). Similar treatment of A/J mice resulted in lower binding in the liver (25.9 to 31.9 pmol 2,6-DNT/mg DNA; 42.6 to 58.9 pmol 2,4-DNT/mg DNA), no detectable binding of 2,6-DNT in extrahepatic tissues and low amounts of binding of 2,4-DNT to lung and intestinal DNA (9.7 to 39.0 pmol/mg DNA). In vitro binding of 2,6-DNT to DNA of cultured hepatocytes from both A/J mice and Fischer-344 rats required prior metabolism of 2,6-DNT by the respective extracts from cecal contents. DNA binding was non-detectable in hepatocytes incubated with 2,6-DNT only. It is concluded that binding of 2,6-DNT to liver DNA requires its prior reductive metabolism, probably by intestinal microorganisms, and that the higher binding of 2,6-DNT in the Fischer-344 rat than in the A/J mouse may, in part, be responsible for the high susceptibility of the Fischer-344 rat to 2,6-DNT carcinogenesis.(ABSTRACT TRUNCATED AT 400 WORDS)

Strain differences among inbred mice in protein kinase C activity.

A protein kinase was identified in mouse organs whose activity is strictly dependent on the presence of Ca++ and phosphatidylserine when assayed at pH 6, and thus has the characteristics of protein kinase C. The relative order of specific activities was brain greater than spleen greater than lung greater than heart, an order similar to that found previously for rat organs. Mice from seven strains had the same level of protein kinase C activity, but strain A/J had half as much activity in each organ as did the other strains. F1 hybrid mice resulting from a cross between A/J and BALB/cByJ mice had levels of activity intermediate to the parental strains, indicating additive inheritance of this genetic difference.

Effects of genetically different strains of mice on the microsomal activation of 2-aminofluorene.

Five strains of mice were examined for microsomal cytochrome P450 (Cyt P450) activity and ability of the microsomal preparation to activate 2-aminofluorene (2-AF) to its mutagenic form. Each strain was assayed under normal and induced conditions. Group A strains known to be easily induced to carcinogenesis by chemicals (C57BL/6J and BALB/cJ) showed higher levels of Cyt P450 and increased mutagenesis activity of the metabolized 2-AF than Group B strains which are known to be high spontaneous tumor producers (CE/J, A/HeJ, C3H/HeJ). Induction of the hepatic microsomes with Aroclor 1254 increased the difference between the two groups.

Rodent models of the human isoniazid-acetylator polymorphism.

Inbred strains and subpopulations of rats, laboratory mice, and deer mice were examined for individual variation in the ability to metabolize several arylamines (p-aminobenzoic acid, sulfamethazine, aniline, alpha-naphthylamine, and aminofluorene) by N-acetylation. Individual differences within species were found to be dependent upon the tissue source of N-acetyltransferase activity and the acetyl acceptor employed. Long-Evans rats possessed about 2-fold more p-aminobenzoic acid N-acetyltransferase activity in blood and liver than Sprague-Dawley rats; no strain differences could be found with sulfamethazine. Nine strains of laboratory mice (Mus musculus) were found to have considerable liver p-aminobenzoic acid N-acetyltransferase activity but only slight activity towards sulfamethazine. No strain differences were apparent in regard to liver N-acetyltransferase activity. Blood p-aminobenzoic acid N-acetyltransferase activity was distinctly polymorphic in laboratory mice; of the nine strains tested, only A/J mice did not have this activity. Partially inbred deer mice (Peromyscus maniculatus) showed a narrower phenotypic range than random-bred stock from which they were obtained, which suggests the existence of distinct subpopulations with respect to N-acetylation capacity. Presumptive evidence for multiple forms of N-acetyltransferase in liver and blood was obtained through a study of substrate specificity.