Quantitative trait analysis of the development of pulmonary tolerance to inhaled zinc oxide in mice

Background

Individuals may develop tolerance to the induction of adverse pulmonary effects following repeated exposures to inhaled toxicants. Previously, we demonstrated that genetic background plays an important role in the development of pulmonary tolerance to inhaled zinc oxide (ZnO) in inbred mouse strains, as assessed by polymorphonuclear leukocytes (PMNs), macrophages, and total protein in bronchoalveolar lavage (BAL) phenotypes. The BALB/cByJ (CBy) and DBA/2J (D2) strains were identified as tolerant and non-tolerant, respectively. The present study was designed to identify candidate genes that control the development of pulmonary tolerance to inhaled ZnO.

Methods

Genome-wide linkage analyses were performed on a CByD2F2 mouse cohort phenotyped for BAL protein, PMNs, and macrophages following 5 consecutive days of exposure to 1.0 mg/m³ inhaled ZnO for 3 hours/day. A haplotype analysis was carried out to determine the contribution of each quantitative trait locus (QTL) and QTL combination to the overall BAL protein phenotype. Candidate genes were identified within each QTL interval using the positional candidate gene approach.

Results

A significant quantitative trait locus (QTL) on chromosome 1, as well as suggestive QTLs on chromosomes 4 and 5, for the BAL protein phenotype, was established. Suggestive QTLs for the BAL PMN and macrophage phenotypes were also identified on chromosomes 1 and 5, respectively. Analysis of specific haplotypes supports the combined effect of three QTLs in the overall protein phenotype. Toll-like receptor 5 (Tlr5) was identified as an interesting candidate gene within the significant QTL for BAL protein on chromosome 1. Wild-derived Tlr5-mutant MOLF/Ei mice were tolerant to BAL protein following repeated ZnO exposure.

Conclusion

Genetic background is an important influence in the acquisition of pulmonary tolerance to BAL protein, PMNs, and macrophages following ZnO exposure. Promising candidate genes exist within the identified QTL intervals that would be good targets for additional studies, including Tlr5. The implications of tolerance to health risks in humans are numerous, and this study furthers the understanding of gene-environment interactions that are likely to be important factors from person-to-person in regulating the development of pulmonary tolerance to inhaled toxicants.     

Genetic determinants of resistance to ectromelia (mousepox) virus-induced mortality.

Resistance to ectromelia (mousepox) virus-induced mortality was examined in crosses between susceptible DBA/2J, A/J, and BALB/cByJ mice and resistant C57BL/6J and AKR/J mice. Depending on the cross, resistance to mousepox virus was shown to be determined by one or more independently assorting autosomal loci with dominant alleles for resistance in AKR/J and C57BL/6J mice and recessive alleles in A/J, BALB/cByJ, and DBA/2J mice. A sexual dimorphism in resistance to disease was also observed.     

Hepatic triglyceride contents are genetically determined in mice: results of a strain survey

To assess whether genetic factor(s) determine liver triglyceride (TG) levels, a 10-mouse strain survey of liver TG contents was performed. Hepatic TG contents were highest in BALB/cByJ, medium in C57BL/6J, and lowest in SWR/J in both genders. Ninety and seventy-six percent of variance in hepatic TG in males and females, respectively, was due to strain (genetic) effects. To understand the physiological/biochemical basis for differences in hepatic TG among the three strains, studies were performed in males of the BALB/cByJ, C57BL/6J, and SWR/J strains. In vivo hepatic fatty acid (FA) synthesis rates and hepatic TG secretion rates ranked BALB/cByJ approximately C57BL/6J > SWR/J. Hepatic 1-(14)C-labeled palmitate oxidation rates and plasma beta-hydroxybutyrate concentrations ranked in reverse order: SWR/J > BALB/cByJ approximately C57BL/6J. After 14 h of fasting, plasma-free FA and hepatic TG contents rose most in BALB/cByJ and least in SWR/J. beta-Hydroxybutyrate concentrations rose least in BALB/cByJ and most in SWR/J. Adaptation to fasting was most effective in SWR/J and least in BALB/cByJ, perhaps because BALB/cByJ are known to be deficient in SCAD, a short-chain FA oxidizing enzyme. To assess the role of insulin action, glucose tolerance test (GTT) was performed. GTT-glucose levels ranked C57BL/6J > BALB/cByJ approximately SWR/J. Thus strain-dependent (genetic) factors play a major role in setting hepatic TG levels in mice. Processes such as FA production and hepatic export in VLDL on the one hand and FA oxidation on the other, explain some of the strain-related differences in hepatic TG contents. Additional factor(s) in the development of fatty liver in BALB/cByJ remain to be demonstrated.     

Kinetics of ectromelia virus (mousepox) transmission and clinical response in C57BL/6j, BALB/cByj and AKR/J inbred mice

Research was undertaken to answer basic questions on susceptibility, clinical response and transmission of ectromelia virus in selected strains of inbred mice. C57BL/6J and AKR/J were found to be markedly more resistant to a virulent strain of ectromelia virus (isolated during the 1979-80 outbreak at the National Institutes of Health) than C57LJ, BALB/cByJ, DBA/2J, A.By/SNJ and C3H/HeJ when infected by footpad inoculation. In C57BL/6J and AKR/J the LD50 was about 7 logs higher than the ID50. With one exception, C57LJ, the LD50 and ID50 titers in the other strains were about equal. In C57LJ the LD50 titer was intermediate. Following intragastric inoculation, virus was isolated from feces of C57BL/6J mice for as long as 46 days and up to 29 days from BALB/cByJ mice. Transmission to cage mates from intragastrically infected C57BL/6J and BALB/cByJ occurred up to 36 and 30 days respectively after infection. Virus was isolated from the spleen in 2 of 5 BALB/cByJ mice and 1 of 7 C57BL/6J mice tested 95 days after gastric inoculation. Following footpad inoculation, BALB/cByJ mice consistently transmitted virus to cage mates before death at 10-12 days. C57BL/6J mice transmitted between days 8 and 17, but not beyond. Virus was maintained in C57BL/6J mice by exposure to infected cage mates for seven passages, which was the most attempted. Clinical signs in infected C57BL/6J mice were usually subtle or inapparent.     

Spink2 Modulates Apoptotic Susceptibility and Is a Candidate Gene in the Rgcs1 QTL That Affects Retinal Ganglion Cell Death after Optic Nerve Damage

The Rgcs1 quantitative trait locus, on mouse chromosome 5, influences susceptibility of retinal ganglion cells to acute damage of the optic nerve. Normally resistant mice (DBA/2J) congenic for the susceptible allele from BALB/cByJ mice exhibit susceptibility to ganglion cells, not only in acute optic nerve crush, but also to chronic inherited glaucoma that is characteristic of the DBA/2J strain as they age. SNP mapping of this QTL has narrowed the region of interest to 1 Mb. In this region, a single gene (Spink2) is the most likely candidate for this effect. Spink2 is expressed in retinal ganglion cells and is increased after optic nerve damage. This gene is also polymorphic between resistant and susceptible strains, containing a single conserved amino acid change (threonine to serine) and a 220 bp deletion in intron 1 that may quantitatively alter endogenous expression levels between strains. Overexpression of the different variants of Spink2 in D407 tissue culture cells also increases their susceptibility to the apoptosis-inducing agent staurosporine in a manner consistent with the differential susceptibility between the DBA/2J and BALB/cByJ strains.     

Experimental allergic orchitis in mice

Inbred strains of mice were studied for their susceptibility to the induction of experimental allergic orchitis after sensitization with mouse testicular homogenate in complete Freund's adjuvant accompanied by injections of extract from Bordetella pertussis. Susceptibility to autoimmune orchitis was found to be linked to the major histocompatibility complex in BALB/c and C57BL/10 mice and mapped to genes encoded within the H-2D ^dregion. In five of six groups of bidirectional (susceptible × resistant) F₁ hybrids, H-2D ^d-linked susceptibility was inherited as a dominant autosomal trait. However, in (BALB/cByJ × DBA/2J)F₁ and (DBA/2J × BALB/cByJ)F₁ hybrids, dominant autosomal resistance to the induction of autoimmune orchitis was observed. Backcross analysis between the resistant F₁ hybrid and the susceptible BALB/cByJ parent suggests that a single independently segregating DBA/2J locus is capable of negating H-2D ^d-linked susceptibility, and controls resistance to the induction of autoimmune orchitis.Abbreviations used in this paper BP extract Bordetella pertussis extract - CFA complete Freund's adjuvant - EAO experimental allergic orchitis - Ir immune response - MHC major histocompatibility complex - MLH mouse liver homogenate - MTH mouse testis homogenate - PI pathology index     

Visual detection, pattern discrimination and visual acuity in 14 strains of mice

Based on the procedure of Prusky et al. (2000, Vision Research, 40, 2201-2209), we used a computer-based, two-alternative swim task to evaluate visual detection, pattern discrimination and visual acuity in 14 strains of mice from priority groups A and B of the JAX phenome project (129S1/SvImJ, A/J, AKR/J, BALB/cByJ, BALB/cJ, C3H/HeJ, C57BL/6J, CAST/Ei, DBA/2J, FVB/NJ, MOLF/Ei, SJL/J, SM/J and SPRET/Ei). Each mouse was tested for eight trials/day for 8 days on each of the three tests. There was a significant strain difference in visual ability in all three tests. Mice with reported normal vision (129S1/SvImJ, C57BL/6J and DBA/2J) and one albino strain (AKR/J) performed very well in these tasks. The other albino strains (A/J, BALB/cByJ and BALB/cJ) took longer to learn the tasks than mice with normal vision and did not reach the criterion of 70% correct. Mice with retinal degeneration (C3H/HeJ, FVB/NJ, MOLF/Ei and SJL/J) performed only at chance levels as did the three strains with unknown visual abilities (CAST/Ei, SM/J and SPRET/Ei). Because many behavioral tasks for rodents rely on visual cues, we suggest that the visual abilities of mice should be evaluated before they are tested in commonly used visuo-spatial learning and memory tasks.     

Acute locomotor responses to cocaine in adolescents vs. adults from four divergent inbred mouse strains

Growing evidence suggests that adolescent mice display differential sensitivity to the acute locomotor activating effects of cocaine as compared to adults, but the direction of the difference varies across studies and the reasons are not clear. Few studies have directly examined genetic contributions to age differences in locomotor stimulation from cocaine. The goal of this study was to determine the extent to which reduced stimulation in C57BL/6J adolescents as compared to adults generalizes to other strains. Therefore, we examined male and female mice from four genetically divergent inbred stains (BALB/cByJ, C57BL/6J, DBA/2J and FVB/NJ) at two ages, postnatal day 30 and postnatal day 65. Mice received either saline or cocaine (15 or 30 mg/kg), and then immediately were placed back into their home cages. Locomotor activity was recorded continuously in the home cage by video tracking. Adolescents displayed reduced stimulation as compared to adults for C57BL/6J, BALB/cByJ and female FVB/NJ mice. No age differences were observed for DBA/2J or male FVB/NJ. No main effects of sex were observed. Strain differences in pharmacokinetics, neural development or physiology could contribute to the observed differences between ages across strains. Future comparative studies could discover biological differences between strains that explain age differences in cocaine sensitivity.     

The Genetic Basis of Preference for Sweet Substances Among Inbred Strains of Mice: Preference Ratio Phenotypes and the Alleles of the Sac and dpa Loci

Five inbred strains (129/J, BALB/cByJ, C3HeB/FeJ, C57BI/6J andDBA/2J) were examined with two-bottle (48 h) preference ratiotesting across concentrations of sodium saccharin (3 x 10^–4M, 10^–3 M, 3 x 10^–3 M and 10^–2 M), d-phenylalanine(10^–3 M, 10^–2 M and 10^–1 M), and l-glutamine(10^–2 M, 3 x 10^–2 M, 10^–1 M and 3 x 10^–1M). Three consistent groupings of strains were observed acrosssubstances and concentrations:

1. C57BI/6J (preference at low andhigh concentrations);
2. BALB/cByJ and C3HeB/FeJ (preferenceat high concentrations);
3. 129/J and DBA/2J (preference at highconcentration for sodiumsaccharin and indifference to d-phenylalanineand l-glutamine).

If a single locus (presumably dpa or Sac) determines these phenotypes,there are likely to be three alleles. If two independent loci(presumably dpa and Sac) determine these phenotypes, an allelicassignment of Sac^b/dpa^+s for the C57BI/6J strain, Sac^b/dpa^–sfor the BALB/cByJ and C3HeB/FeJ strains, and either Sac^d/dpa^+sor Sac^d/dpa^–s for the 129/J and DBA/2J strains is suggested.Chem. Senses 20: 291–298, 1995.     

Immunologic tolerance to heterologous immunoglobulin: its relation to in vitro filtration by macrophages.

The cellular and molecular basis for the difference in ability of BCG to induce tolerance in BALB/c and DBA/2 mice has been examined by in vitro biofiltration. It was found that incubation with the adherent cells from BALB/c but not DBA/2 spleens could remove the material from BGG which inhibited tolerance induction in BALB/c mice. This material was shown to represent only a trace component in BGG, was present in only certain commercial batches of BGG, and was apparently unrelated to the presence of aggregates or endotoxin.     

Inhibition of nitric oxide synthesis increases mortality in Sindbis virus encephalitis.

Sindbis virus (SV) is an alphavirus that causes acute encephalomyelitis in mice. The outcome is determined by the strain of virus and by the age and genetic background of the host. The mortality rates after infection with NSV, a neurovirulent strain of SV, were as follows v: 81% (17 of 21) in BALB/cJ mice; 20% (4 of 20) in BALB/cByJ mice (P < 0.001); 100% in A/J, C57BL/6J, SJL, and DBA mice; and 79% (11 of 14) in immunodeficient scid/CB17 mice. Treatment with Nomega-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthetase (NOS) inhibitor, increased mortality to 100% (P < 0.05) in NSV-infected BALB/cJ mice, to 95% (P < 0.001) in BALB/cByJ mice, and to 100% in scid/CB17 mice. BALB/cJ and BALB/cByJ mice had similar levels of inducible NOS mRNA in their brains, which were not affected by L-NAME or NSV infection. Brain NOS activity was similar in BALB/cJ and BALB/cByJ mice before and after infection and was markedly inhibited by L-NAME. NSV replication in the brains of BALB/cJ mice, BALB/cByJ mice, and mice treated with L-NAME was similar. Treatment of N18 neuroblastoma cells with NO donors S-nitroso-N-acetylpenicillamine or sodium nitroprusside in vitro before infection increased cell viability at 42 to 48 h compared with untreated NSV-infected N18 cells with little effect on virus replication. These data suggest that NO protects mice from fatal encephalitis by a mechanism that does not directly involve the immune response or inhibition of virus growth but rather may enhance survival of the infected neuron until the immune response can control virus replication.     

Ambient ozone primes pulmonary innate immunity in mice

Exposure to ozone in air pollution in urban environments is associated with increases in pulmonary-related hospitalizations and mortality. Because ozone also alters clearance of pulmonary bacterial pathogens, we hypothesized that inhalation of ozone modifies innate immunity in the lung. To address our hypothesis, we exposed C57BL/6J mice to either free air or ozone, and then subsequently challenged with an aerosol of Escherichia coli LPS. Pre-exposure to ozone resulted in enhanced airway hyperreactivity, higher concentrations of both total protein and proinflammatory cytokines in lung lavage fluid, enhanced LPS-mediated signaling in lung tissue, and higher concentrations of serum IL-6 following inhalation of LPS. However, pre-exposure to ozone dramatically reduced inflammatory cell accumulation to the lower airways in response to inhaled LPS. The reduced concentration of cells in the lower airways was associated with enhanced apoptosis of both lung macrophages and systemic circulating monocytes. Moreover, both flow cytometry and confocal microscopy indicate that inhaled ozone causes altered distribution of TLR4 on alveolar macrophages and enhanced functional response to endotoxin by macrophages. These observations indicate that ozone exposure increases both the pulmonary and the systemic biologic response to inhaled LPS by priming the innate immune system.     

Postnatal episodic ozone results in persistent attenuation of pulmonary and peripheral blood responses to LPS challenge

Early life is a dynamic period of growth for the lung and immune system. We hypothesized that ambient ozone exposure during postnatal development can affect the innate immune response to other environmental challenges in a persistent fashion. To test this hypothesis, we exposed infant rhesus macaque monkeys to a regimen of 11 ozone cycles between 30 days and 6 mo of age; each cycle consisted of ozone for 5 days (0.5 parts per million at 8 h/day) followed by 9 days of filtered air. Animals were subsequently housed in filtered air conditions and challenged with a single dose of inhaled LPS at 1 yr of age. After completion of the ozone exposure regimen at 6 mo of age, total peripheral blood leukocyte and polymorphonuclear leukocyte (PMN) numbers were reduced, whereas eosinophil counts increased. In lavage, total cell numbers at 6 mo were not affected by ozone, however, there was a significant reduction in lymphocytes and increased eosinophils. Following an additional 6 mo of filtered air housing, only monocytes were increased in blood and lavage in previously exposed animals. In response to LPS challenge, animals with a prior history of ozone showed an attenuated peripheral blood and lavage PMN response compared with controls. In vitro stimulation of peripheral blood mononuclear cells with LPS resulted in reduced secretion of IL-6 and IL-8 protein in association with prior ozone exposure. Collectively, our findings suggest that ozone exposure during infancy can result in a persistent effect on both pulmonary and systemic innate immune responses later in life.     

Macronutrient diet selection in thirteen mouse strains

The strain distribution for macronutrient diet selection was described in 13 mouse strains (AKR/J, NZB/B1NJ, C57BL/6J, C57BL/6ByJ, DBA/2J, SPRET/Ei, CD-1, SJL/J, SWR/J, 129/J, BALB/cByJ, CAST/Ei, and A/J) with the use of a self-selection protocol in which separate carbohydrate, fat, and protein diets were simultaneously available for 26-30 days. Relative to carbohydrate, nine strains consumed significantly more calories from the fat diet; two strains consumed more calories from carbohydrate than from fat (BALB/cByJ, CAST/Ei). Diet selection by SWR/J mice was variable over time, resulting in a lack of preference. One strain (A/J) failed to adapt to the diet paradigm due to inadequate protein intake. Comparisons of proportional fat intake across strains revealed that fat selection/consumption ranged from 26 to 83% of total energy. AKR/J, NZB/B1NJ, and C67BL/6J mice self-selected the highest proportion of dietary fat, whereas the CAST/Ei and BALB/cByJ strains chose the lowest. Finally, epididymal fat depot weight was correlated with fat consumption. There were significant positive correlations in AKR/J and C57BL/6J mice, which are highly sensitive to dietary obesity. However, absolute fat intake was inversely correlated with epididymal fat in two of the lean strains: SWR/J and CAST/Ei. We hypothesize that the SWR/J and CAST/Ei strains are highly sensitive to a negative feedback signal generated by increasing body fat, but the AKR/J and C67BL/6J mice are not. The variation in dietary fat selection across inbred strains provides a tool for dissecting the complex genetics of this trait.     

Pulmonary biocompatibility assessment of inhaled single-wall and multiwall carbon nanotubes in BALB/c mice

With the widespread application of carbon nanotubes (CNTs) in diverse commercial processes, scientists are now concerned about the potential health risk of occupational exposures. In this study, CNT-induced pulmonary toxicity was investigated by exposing BALB/c mice to aerosolized single-wall (SW) CNT and multiwall (MW) CNT (5 μg/g of mice) for 7 consecutive days in a nose-only exposure system. Microscopic studies showed that inhaled CNTs were homogeneously distributed in the mouse lung. The total number of bronchoalveolar lavage polymorphonuclear leukocytes recovered from the mice exposed to SWCNT and MWCNT (1.2 × 10(6) ± 0.52 and 9.87 × 10(5) ± 1.45; respectively) was significantly greater than control mice (5.46 × 10(5) ± 0.78). Rapid development of pulmonary fibrosis in mice that inhaled CNT was also confirmed by significant increases in the collagen level. The lactate dehydrogenase levels were increased nearly 2- and 2.4-fold in mice that inhaled SWCNT and MWCNT, respectively, as compared with control mice. In addition, exposure of CNTs to mice showed a significant (p < 0.05) reduction of antioxidants (glutathione, superoxide dismutase, and catalase) and induction of oxidants (myloperoxidase, oxidative stress, and lipid peroxidation) compared with control. Apoptosis-related proteins such as caspase-3 and -8 activities were also significantly increased in mice that inhaled CNT than in control mice. Together, this study shows that inhaled CNTs induce inflammation, fibrosis, alteration of oxidant and antioxidant levels, and induction of apoptosis-related proteins in the lung tissues to trigger cell death.     

Control of fetal survival in CBA x DBA/2 mice by lymphokine therapy

In this study, we examined the effect of injecting various cytokines. We report here that tumour necrosis factor (TNF)alpha, gamma-interferon and interleukin 2 (IL-2) can, in some circumstances, increase fetal resorption rates in abortion-prone (CBA/J x DBA/2) and non-abortion prone (CBA/J x BALB/c,C3H x DBA/2) matings: 1000 units TNF enhanced resorptions from 43 to 79% in CBA x DBA/2, from 7 to 89% in CBA x BALB/c, from 5 to 47% in C3H x DBA/2. The effect was both gestational age- and dose-dependent. Gamma interferon and R-IL-2 enhanced resorptions from 38 to 68% and 76% respectively in the CBA/J x DBA/2 mating combination, whereas the rates in CBA/J x BALB/c matings were enhanced from 6 to 44% and 55%. Lipopolysaccharide (LPS), which is known to lead to the release of TNF-alpha, had a similar effect, leading to gestational age- and dose-dependent enhancement of resorptions up to 100%. However, cytokines of the CSF family, including IL-3 and GM-CSF, increased the chances of fetal survival when injected into abortion-prone mice, e.g. reducing resorption rates in the abortion-prone CBA/J x DBA/2 mating combination from 55 to 22% (IL-3), and 47 to 8% (GM-CSF). They also increased fetal and placental weight and, in particular, expanded the spongiotrophoblast zone in the placenta. The latter observations may be due to a direct trophic influence on placental cells, perhaps through a cytokine cascade, or an indirect effect due to inhibition of natural killer (NK)-like cells, or both. Whatever the mechanism, these results may find practical application in influencing reproductive outcome in women and other species.     

NaCl taste thresholds in 13 inbred mouse strains

Molecular mechanisms of salty taste in mammals are not completely understood. We use genetic approaches to study these mechanisms. Previously, we developed a high-throughput procedure to measure NaCl taste thresholds, which involves conditioning mice to avoid LiCl and then examining avoidance of NaCl solutions presented in 48-h 2-bottle preference tests. Using this procedure, we measured NaCl taste thresholds of mice from 13 genealogically divergent inbred stains: 129P3/J, A/J, BALB/cByJ, C3H/HeJ, C57BL/6ByJ, C57BL/6J, CBA/J, CE/J, DBA/2J, FVB/NJ, NZB/BlNJ, PWK/PhJ, and SJL/J. We found substantial strain variation in NaCl taste thresholds: mice from the A/J and 129P3/J strains had high thresholds (were less sensitive), whereas mice from the BALB/cByJ, C57BL/6J, C57BL/6ByJ, CE/J, DBA/2J, NZB/BINJ, and SJL/J had low thresholds (were more sensitive). NaCl taste thresholds measured in this study did not significantly correlate with NaCl preferences or amiloride sensitivity of chorda tympani nerve responses to NaCl determined in the same strains in other studies. To examine whether strain differences in NaCl taste thresholds could have been affected by variation in learning ability or sensitivity to toxic effects of LiCl, we used the same method to measure citric acid taste thresholds in 4 inbred strains with large differences in NaCl taste thresholds but similar acid sensitivity in preference tests (129P3/J, A/J, C57BL/6J, and DBA/2J). Citric acid taste thresholds were similar in these 4 strains. This suggests that our technique measures taste quality-specific thresholds that are likely to represent differences in peripheral taste responsiveness. The strain differences in NaCl taste sensitivity found in this study provide a basis for genetic analysis of this phenotype.     

Genetic Determination of the Developmental Program for Mouse Liver β-GALACTOSIDASE: Involvement of Sites Proximate to and Distant from the Structural Gene

The identification and mode of action of genetic loci that program gene expression during development are important for understanding differentiation in higher organisms. Previous work from this laboratory has identified two patterns for the postnatal development of liver beta-galactosidase among inbred mouse strains: type I, where activity levels remain constant after about 30 days of age, is found in strains DBA/2J, CBA/J, and BALB/cJ, among others; type II, where activity levels increase between 25 and 50 days of age to reach a new adult level, is found in strain C57BL/6J and related strains. It has been shown that the type I vs. type II developmental difference between strains C57BL/6J and DBA/2J is due to variation at a locus, Bgl-t, that maps with the beta-galactosidase complex, [Bgl], on chromosome 9. In the present study, we have confirmed the existence of Bgl-t as a temporal locus within [Bgl] by analysis of both a congenic strain carrying the beta-galactosidase complex of strain CBA/J in the C57BL/6J genetic background and a cross of strains CBA/J and C57BL/6J. The existence of additional temporal loci for beta-galactosidase that segregate independently of the structural gene and participate in determination of the type I vs. type II difference was revealed by analysis of: (1) a congenic strain containing the beta-galactosidase complex of strain BALB/cJ in the C57BL/10Sn background; (2) recombinant inbred lines derived from progenitor strains C57BL/6ByJ and BALB/cByJ; and (3) a genetic cross between strains C57BL/6ByJ and BALB/cByJ. Thus, for these pairs of strains, the type I vs. type II developmental difference is due to variation at a temporal locus (or loci) unlinked to the enzyme structural gene, and not at Bgl-t. These facts, together with information gathered from an examination of the distribution of beta-galactosidase phenotypes among over 100 inbred strains (Breen, Lusis and Paigen 1977), have led us to conclude that the postnatal developmental pattern for liver beta-galactosidase is determined by a set of interacting temporal genes. One of these, Bgl-t, is located within [Bgl], and one or more are separable from [Bgl] by recombination. A possible mode of interaction among the temporal and instructural loci is suggested.     

Taste discrimination between NaCl and KCl is disrupted by amiloride in inbred mice with amiloride-insensitive chorda tympani nerves

The amiloride-sensitive salt transduction pathway is thought to be critical for the discrimination between sodium and nonsodium salts in rodents. In rats, lingual application of amiloride appears to render NaCl qualitatively indistinguishable from KCl. In this study, we tested four strains of mice for salt discriminability. In one strain (C57BL/6J), chorda tympani nerve (CT) responses to NaCl are attenuated by amiloride, and in the other three strains (BALB/cByJ, 129P3/J, DBA/2J) they are not. Under water-restriction conditions, these mice (7 mice/strain) were trained in a gustometer to lick for water from one reinforcement spout in response to a five-lick presentation of NaCl and to lick from another in response to KCl [salt concentration was varied (0.1-1 M) to render intensity irrelevant]. Mice were then tested with the stimuli dissolved in amiloride hydrochloride, and the latter was used as the reinforcer as well. Each concentration of amiloride (0.1-100 microM) was used on 2 separate days with control sessions interposed. Mice from all four strains were able to discriminate NaCl from KCl reliably. Amiloride impaired this discrimination in a dose-dependent fashion. Moreover, performance on NaCl trials appeared to be more affected by amiloride than that on KCl trials in all four strains. Thus, in contrast to the predictions based on CT recordings, discrimination in all four strains appeared to depend on the amiloride-sensitive transduction pathway, which, in the case of BALB/cByJ, 129P3/J, and DBA/2J (and perhaps C57BL/6 as well), may exist in taste buds innervated by nerves other than the CT.     

Genetic variability in forced and voluntary endurance exercise performance in seven inbred mouse strains.

The goal of this study was to characterize the genetic contribution to both forced and voluntary exercise performance and to determine whether performance in these two paradigms is controlled by similar genetic influences. There were marked strain differences in treadmill exercise performance, with Swiss Webster (SW) and FVB/NJ mice showing elevated performance and C57BL/6J animals showing decreased performance compared with all other strains. There was no apparent relationship between treadmill performance and voluntary wheel performance, with the exception of SW mice, which demonstrated high performances on both the treadmill and the voluntary wheel. Numerous properties were measured to attempt to understand the basis for these differences in exercise performance. DBA/1J and SW mice exhibited significantly greater cardiac contractility than all other analyzed strains. Conversely, BALB/cByJ mice exhibited significantly reduced cardiac contractility compared with all other strains. Expression of molecular indicators of hypertrophy (atrial natriuretic factor and beta-myosin heavy chain) was significantly elevated in DBA/2J myocardium compared with all other analyzed strains.