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Effects of intermittent 60-Hz high voltage electric fields on metabolism, activity, and temperature in mice.

Transient effects of 100-kV/m extremely low frequency electric fields were studied in the white footed deermouse, Peromyscus leucopus. Gross motor activity, carbon dioxide production, oxygen consumption, and core body temperature were monitored before, during, and after intermittent field exposures (four hour-long exposures, at one-hour intervals). Thirty-four mice were exposed in cages with plastic floors floating above ground potential, and 21 mice were exposed in cages with grounded metal floor plates. The first field exposure produced an immediate, transient increase of activity and gas measures during the inactive phase of the circadian cycle. All measures returned to baseline levels before the second exposure and were not significantly changed throughout the remainder of the exposures. The rapid habituation of field-induced arousal suggests that significant metabolic changes will not be measured in experiments in which the interval between exposure and measurement is greater than two hours.

Animals↗

Relationship between field strength and arousal response in mice exposed to 60-Hz electric fields.

White-footed mice, Peromyscus leucopus, were exposed to 60-Hz electric fields to study the relationship between field strength and three measures of the transient arousal response previously reported to occur with exposures at 100 kV/m. Five groups of 12 mice each were given a series of four 1-h exposures, separated by an hour, with each group exposed at one of the following field strengths: 75, 50, 35, 25, and 10 kV/m; 8 additional mice were sham-exposed with no voltage applied to the field generator. All mice were experimentally naive before the start of the experiment, and all exposures occurred during the inactive (lights-on) phase of the circadian cycle. The first exposure produced immediate increases in arousal measures, but subsequent exposures had no significant effect on any measure. These arousal responses were defined by significant increases of gross motor activity, carbon dioxide production, and oxygen consumption, and were frequently recorded with field strengths of 50 kV/m or higher. Significant arousal responses rarely occurred with exposures at lower field strengths. Responses of mice exposed at 75 and 50 kV/m were similar to previously described transient arousal responses in mice exposed to 100-kV/m electric fields. Less than half of the mice in each of the field strength groups below 50 kV/m showed arousal responses based on Z (standard) scores, but the arousals of the mice that did respond were similar to those of mice exposed at higher field strengths. Polynomial regression was used to calculate the field strength producing the greatest increases for each of the arousal measures. The results show that the amplitude of the transient arousal response is related to the strength of the electric field, but different measures of arousal may have different relationships to field strength.

Animals↗

Reproductive development in male deer mice exposed to aggressive behavior.

Male deer mice (Peromyscus maniculatus bairdii) were reared in a long photoperiod and housed individually from 3 weeks of age until they were killed 2, 4, or 6 weeks later. Males that were exposed to aggressive females for 2 min, three times per week, were of normal body weight and healthy, but had smaller reproductive organs than did males exposed to a control treatment. Reproductive development in male deer mice is responsive to at least two classes of social stimuli: pheromones and aggressive behavior.

Aggression↗

Genetic evidence for a maternal effect locus controlling genomic imprinting and growth.

Crosses between two species of deer mouse (Peromyscus) yield dramatic parent-of-origin effects. Female P. maniculatus (BW) crossed with male P. polionotus (PO) produce animals smaller than either parent. PO females crossed with BW males yield lethal overgrowth that has been associated with loss-of-imprinting (LOI). Previously, we mapped two loci influencing fetal growth. These two loci, however, do not account for the LOI, nor for the dysmorphic phenotypes. Here we report that maternal genetic background strongly influences the LOI. Analyses of crosses wherein maternal genetic background is varied suggest that this effect is likely due to the action of a small number of loci. We have termed these putative loci Meil. Estimation of Meil loci number was confounded by skewed allelic ratios in the intercross line employed. We show that the Meil loci are not identical to any of the DNA methyltransferases shown to be involved in regulation of genomic imprinting.

Animals↗

Hepatotoxicity due to allyl alcohol in deermice depends on alcohol dehydrogenase.

The role of alcohol dehydrogenase in the hepatic necrosis due to allyl alcohol was studied in two strains of the deermouse, Peromyscus maniculatus. Mice of the alcohol dehydrogenase-negative (AdhN) strain which lack alcohol dehydrogenase activity were resistant to allyl alcohol toxicity. In contrast, dose-dependent necrosis of periportal regions of the liver and increases in plasma levels of lactate dehydrogenase, sorbitol dehydrogenase and SGOT were observed in plasma from alcohol dehydrogenase-positive deermice (AdhF) 24 hr following administration of allyl alcohol (21 to 84 mg per kg). Half-maximal damage to periportal areas was observed with about 52 mg per kg allyl alcohol. Thus, these data demonstrate that metabolism of allyl alcohol to acrolein by alcohol dehydrogenase is obligatory for the hepatotoxicity of allyl alcohol.

1-Propanol↗

Influence of photoperiod, ambient temperature and melatonin on testosterone synthesis and release during reproductive maturation in male deer mice.

Four experiments were designed to investigate the influence of photoperiod and other environmental factors on androgen production and reproductive maturation in deer mice. Male prairie deer mice (Peromyscus maniculatus), born in a light/dark cycle of 6L:18D, either remained in this short photoperiod or were switched to a long day regimen of 16L:8D at weaning. In a cross-sectional experiment, the deer mice were killed between 3 and 8 weeks of age for measurement of serum testosterone concentration and reproductive organ weights. In a second experiment, blood was collected from each mouse at weekly intervals between 3 and 9 weeks of age. This repeated measures design was used to reduce the high variability in testosterone values observed in the first experiment. Reproductive organs were weighed at the termination of the experiment. Testosterone concentrations and reproductive organ weights were greater in males reared in the long photoperiod in both experiments. In a third experiment, the animals were housed under five different conditions to test the influence of high ambient temperature and melatonin as well as photoperiod. At 7 weeks of age, they received an injection of hCG or saline. More testosterone was released in deer mice reared in 16L:8D and 27 C than in those reared in short days (6L:18D) or those reared in high ambient temperature (35 C) or those treated with exogenous melatonin. One week later, animals were sacrificed. The single hCG treatment caused significant reversal of the suppression of accessory sex organ weights following melatonin, short days or 35 C temperature. In a fourth experiment, the additive influence of melatonin and 35 C temperature was tested. Animals treated with 35 C or both melatonin and 35 C had lower serum testosterone at 7 weeks of age, released less testosterone after hCG, and had smaller organ weights with or without hCG than long day controls. The influence of melatonin treatment and 35 C temperature appears to be additive for testicular weight and testosterone release after hCG. Thus, the attenuation of reproductive development that accompanied short days, melatonin treatment and high ambient temperature occurred via diminished testosterone secretion, which can be overcome at least in part by gonadotropin treatment.

Animals↗

On the activity and fidelity of chromatin-associated hepatic DNA polymerase-beta in aging murine species of different life spans.

Activity and accuracy of chromatin-directed DNA replication have been compared in young and aged Mus musculus and Peromyscus leucopus, two murine species with contrasting maximum lifespans. Chromatin isolated from livers of mature adults of both species copied efficiently exogenous DNA templates using predominantly DNA polymerase-beta. The DNA synthetic activity of liver chromatin remained constant in both species throughout their lifetimes. The fidelity of chromatin-directed poly [d(A-T)] synthesis was similar for the comparatively short-lived M. musculus and the relatively long-lived P. leucopus and remained unaltered in old animals. The fidelity of poly [d(A-T)] copying catalyzed by DNA polymerase-beta-dissociated from liver chromatin was comparable to that of the chromatin-directed synthesis. The dissociation enzymes did not exhibit diminished fidelity of poly [d(A-T)] synthesis with age. In all ages of both species examined, the murine liver DNA polymerase-beta, both chromatin-associated and solubilized, exhibited high error frequencies; approximately one dGMP was incorporated for every 500-1,000 complementary nucleotides polymerized. The relationship of these results to the accuracy of DNA replication and repair as a determinant of aging is considered.

Aging↗

In vitro fertilization of two species of deer mouse eggs by homologous or heterologous sperm and penetration of laboratory mouse eggs by deer mouse sperm.

Newly ovulated eggs from immature deer mice (Peromyscus maniculatus and P. polionotus) and mature laboratory mice (Mus musculus) treated with PMSG and HCG were inseminated in vitro with spermatozoa recovered from the cauda epididymidis of mature males. The time required for capacitation of deer mouse sperm in culture was estimated to be about two to five hours based on the dispersal of sperm agglutination and increase of sperm motility. The rate of sperm penetration through the zona pellucida of deer mouse eggs by homologous or heterologous sperm was relatively high (72-91%) but that of laboratory mouse eggs by deer mouse sperm was low (20-21%). After penetration through the zona pellucida, a high proportion of deer mouse eggs (79-93%) were fertilized by homologous or heterologous deer mouse sperm but no laboratory mouse eggs were fertilized by sperm of two species of deer mice. The zona pellucida was dissolved in a higher proportion of laboratory mouse eggs cultured with P. maniculatus (45%) than with P. polionotus sperm (3.4%), but this did not happen by incubation of deer mouse eggs with homologous or heterologous sperm. It seems that there is little difference in sperm penetration and fertilization between these two closely related species of deer mice but the reactions between the mouse eggs and deer mouse sperm are quite different.

Animals↗

A study of hetero-specific sperm-egg interactions in the rat, mouse, and deer mouse using in vitro fertilization and sperm injection.

Hetero-specific fertilization of zone-free eggs is used in these experiments as a tool to analyze the barriers to hybridization and to gain insight into the mechanisms of normal fertilization. When the zonae of rat eggs, which are a barrier to hetero-specific fertilization, are removed with pronase, the eggs can be fertilized by mouse sperm and the zygotes start to develop normally. A rat egg fertilized with mouse sperm completes meiosis and forms both male and female pronuclei. Chromosomes from both parents are found on he spindle at the metaphase stage of the first cleavage division. Under present culture conditions, embryos develop only to the two-cell stage, but this initial development of the hybrid is apparently normal. The question of whether sperm and egg membrane fusion is requisite for normal development is addressed by injecting sperm directly into the cytoplasm of unfertilized eggs. The injection of mouse sperm into rat eggs frequently leads to activation and formation of male and female pronuclei. The first cleavage division is indistinguishable from that following hetero-specific fertilization. Capacitated and uncapacitated sperm react alike when injected into eggs. Egg activation, however, is necessary for male pronucleus formation. Sperm from the deer mouse Peromyscus maniculatus bairdii, which are incapable of fertilizing even zonea-free eggs, respond like mouse sperm when injected into rat eggs. These data indicate that sperm interactions with the egg cytoplasm are less species-specific than interactions at the egg surface. Furthermore, the normal surface interactions of sperm and eggs are not essential for the start of development.

Animals↗

Assessment of mercury concentrations in small mammals collected near Las Vegas, Nevada, USA.

Mercury concentrations in liver and hair tissue were determined for five species of small mammals captured near Las Vegas, Nevada, USA. These data were then used to evaluate the suitability of using hair as a noninvasive technique for determining body burdens of mercury. A total of 104 small mammals were captured and analyzed; four main species were examined and included: Dipodomys merriami, Chaetodipus penicillatus, Peromyscus eremicus, and Neotoma lepida. Mean mercury concentrations were highest in N. lepida, followed by D. merriami, C. penicillatus, and P. eremicus respectively. Positive associations were found between hair and liver taken from D. merriami (r = 0.647) and C. penicillatus (r = 0.533) indicating that hair may be a suitable indicator of body burdens in these two species.

Animals↗

Sequence and expression analysis of deer mouse interferon-gamma, interleukin-10, tumor necrosis factor, and lymphotoxin-alpha.

Deer mice (Peromyscus maniculatus) are the principal host species of Sin Nombre (SN) virus, the primary etiologic agent of hantavirus cardiopulmonary syndrome in North America. The disease is a cytokine-mediated immunopathology characterized by pulmonary mononuclear infiltrates without discernible viral pathology. Infected deer mice remain life-long carriers and virus is found in many organs, including the lungs, but without pathology. It is unclear how deer mice respond to SN virus because no tools exist to examine the immune response in infected animals. As an initial step in examining host responses to SN virus, we have cloned partial cDNAs of deer mouse interferon-gamma (IFN-gamma), interleukin-10 (IL-10), tumor necrosis factor (TNF) and lymphotoxin-alpha (LTalpha). IL-10, TNF and LTalpha sequences are highly conserved compared to orthologs of other mammalian species, while IFN-gamma is substantially less conserved. Phylogenetic analyses indicate that the amino acid sequences of IFN-gamma and TNF may be useful in resolving relationships at the subfamily level within the rodent family Muridae. While all four sets of analyses were able to reconstruct clade Rodentia, they were not able to resolve the relationships among the mammalian orders represented in this study. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis of concanavalin A-stimulated splenocytes determined that maximal IFN-gamma and TNF expression occurred rapidly while IL-10 and LTalpha expression was maximal at 24 h.

Amino Acid Sequence↗

Isolation and initial characterization of a newfound hantavirus from California.

A fatal case of hantavirus pulmonary syndrome (HPS) in northern California prompted our attempt to isolate viruses from local rodents. From tissues of two deer mice, Peromyscus maniculatus, two hantaviruses (Convict Creek virus 107 and 74, CC107 and CC74) were established in cell culture. Viral antigens, proteins, and RNAs of the first and archetypical isolate (CC107) were examined, and portions of the medium (M) and small (S) genome segments of both isolates were sequenced. Antigenically, CC107 virus and the second isolate, CC74 virus, were more closely related to Puumala virus than Hantaan (HTN) virus, though distinct from both. Northern blots of viral RNAs showed the large and M segments of CC107 to be the same size as those of HTN virus, whereas the S segment was larger. Protein gels did not reveal CC107 to have a substantially larger nucleocapsid protein than HTN virus. Partial nucleotide sequence comparisons of CC107 and CC74 viruses revealed their M segments to be highly similar to one another, while their S segments differed by more than 10%. Nucleotide and deduced amino acid sequence comparisons showed the California isolates to be closely related to the newfound hantaviruses first detected in the Four Corners area and since incriminated in HPS through wide areas of the United States.

Amino Acid Sequence↗

Complete nucleotide sequences of the M and S segments of two hantavirus isolates from California: evidence for reassortment in nature among viruses related to hantavirus pulmonary syndrome.

We report the complete nucleotide sequence of the M and the S genome segments and a portion of the L segments of two hantavirus isolates from Peromyscus maniculatus trapped in eastern California. The isolates, Convict Creek 107 and 74 (CC107 and CC74) are genetically similar to viruses known to cause hantavirus pulmonary syndrome in New Mexico. CC107 and CC74 each have an M segment consisting of 3696 nucleotides with a coding potential of 1140 amino acids in the virus complementary-sense RNA (cRNA). The S segments of CC107 and CC74 are 2083 and 2047 nucleotides long, respectively, and each has an ORF in the cRNA capable of encoding a protein of 428 amino acids. Unusually long 3' noncoding regions of 757 and 721 nucleotides follow the S segment ORF of CC107 and CC74, respectively, and include numerous imperfect repetitive sequences. Whereas the M and S segments of any given hantavirus typically appear to diverge at comparable rates from homologous genes of any other hantavirus, CC107 and CC74 have M segments that differ by only 1% from one another but S segments that differ by 13%. After trivial explanations are rendered improbable, i.e., by consideration of the genetics of closely and distantly related hantaviruses, the most likely explanation for our data is that hantavirus genome segment reassortment occurred within rodent populations in California.

Animals↗

Coexistence of several novel hantaviruses in rodents indigenous to North America.

Three genetically distinct members of the Hantavirus genus have been detected in Nevada rodents by RT-PCR and nucleotide sequence analysis. These include Sin Nombre (SN), El Moro Canyon (ELMC), and Prospect Hill (PH)-like viruses which are primarily associated with Peromyscus maniculatus (deer mouse), Reithrodontomys megalotis (western harvest mouse), and Microtus spp. (voles), respectively. Although this region of the United States is ecologically diverse, rodents infected with different hantaviruses appear to coexist in several different geographical and ecological zones. In two widely separated states, Nevada and North Dakota, PH-like viruses are present in three different species of vole. In addition, ELMC-like virus has been detected in both R. megalotis and M. montanus (mountain vole). SN virus is a cause of hantavirus pulmonary syndrome throughout much of the United States. SN virus RNA is found in 12.5% of P. maniculatus in Nevada and eastern California. Two lineages of SN virus coexist in this region and differ from SN viruses originally found in infected rodents in New Mexico, Arizona, and Colorado. These data show the complexity of hantavirus maintenance in rodents. Distinct hantaviruses or virus lineages can coexist either in different or the same rodent species and in either different or the same geographic or ecological zones.

Animals↗

Genetic reassortment among viruses causing hantavirus pulmonary syndrome.

In order to determine the frequency and characteristics of reassortment among viruses causing hantavirus pulmonary syndrome (HPS), mixed infections were initiated in tissue culture by using two closely related strains of Sin Nombre virus, CC107 (from eastern California) and NMR11 (from New Mexico), which share the same species of rodent host in nature, the deer mouse (Peromyscus maniculatus). Potential reassortant virus plaques were screened by multiplex RT-PCR, using primers specific for individual genome segments of each strain. Reassortant viruses involving the M and S segments and, to a lesser extent, the L segment were detected in 8.5% of 294 progeny plaques tested. In addition, approximately 30% of the progeny virus plaques appeared to contain S or M segments originating from both parental virus strains, i.e., they were diploid. Most of these diploid virus genotypes were not stable, becoming either reassortant or parental virus strains upon plaque-to-plaque virus passage. In contrast to the results above, only one virus reassortant and four diploids were observed among 163 progeny virus plaques from mixed infections between Sin Nombre virus NMR11 and the genetically more distant Black Creek Canal virus, an HPS-causing virus from Florida, which has the cotton rat (Sigmodon hispidus) as its natural host.

Animals↗

Field and laboratory studies on the timing of oviposition and hatching of the western black-legged tick, Ixodes pacificus (Acari: Ixodidae).

The timing of oviposition and hatching of Ixodes pacificus was investigated in the field and at constant temperatures in the laboratory. Replete females held at temperatures between 9 and 29 degrees C began depositing eggs a mean of 9-70 days after drop off. Egg masses held between 12 and 25 degrees C commenced hatching 25-178 days after the onset of oviposition. Eggs held at 9 or 29 degrees C did not hatch. The lower temperature thresholds for development (LTD) for oviposition and hatching were 6.5 and 9 degrees C, respectively. The number of degree days required for oviposition and hatching was 173 and 588, respectively. Replete females placed in the field on 2 December through to 8 March deposited eggs from 2 February through to 24 April; the eggs commenced hatching between 2 July and 21 August. Unfed larvae from two of 20 egg masses survived through the winter and fed readily when exposed to deer mice (Peromyscus maniculatus) on 22 April. Replete larvae were returned to the field and moulted between 9 and 21 August. Larvae exposed to deer mice in August, 4 weeks after hatching, also fed readily. Although further studies are needed to clarify the timing of nymphal development, the present study suggests that I. pacificus requires more than 1 year to complete its life cycle.

Animals↗

Differential reproductive response to short photoperiod in deer mice: role of melatonin.

Inhibitory photoperiod differentially effects reproduction in deer mice (Peromyscus maniculatus nebrascensis). Pituitary-testicular function is arrested in about one-third of short-day exposed males (reproductively responsive mice), while an equal number remain fertile (reproductively nonresponsive mice). Both phenotypes are found in natural populations and their disparate reproductive responses have a genetic basis. To assess whether this difference is attributable to a prepineal/pineal or post-pineal mechanism, we compared spermatogenic responses of known and unknown phenotype to exogenous melatonin. Melatonin significantly reduced mean sperm number in long-day housed mice of unknown phenotype. But, individual responses ranged from azoospermia to normal spermatogenesis, and this range was not significantly different from that previously recorded for short-day exposed mice. Reproductively nonresponsive males were unaffected by melatonin administration when housed under long or short daylength. In contrast, melatonin significantly suppressed sperm production in reproductively responsive males housed under long photoperiod, but had no additional suppressive effect in short-day housed mice with regressed testes. These data demonstrate that melatonin is only effective in eliciting testicular regression in reproductively responsive males. Taken together, these results suggest that differential testicular response to photoperiod are caused by a post-pineal mechanism.

Animals↗

Bioavailability of metals and arsenic to small mammals at a mining waste-contaminated wetland.

In support of a baseline ecological risk assessment evaluating the impacts of mining wastes at the Milltown Reservoir Sediments Superfund site in Montana, a food chain transfer analysis was performed for resident small mammals. Deer mice (Peromyscus maniculatus) and meadow voles (Microtus pennsylvanicus) were trapped from a 200 A portion of a mixed upland and palustrine wetland, and concentrations of As, Cd, Cu, Pb, and Zn in carcass, liver, kidney, and testes were quantified. Concurrent to small mammal trapping, samples of grasses, forbs, and soils were collected and analyzed for metal and As residues. Using a linear multimedia food-chain model, assuming a forage base of wetland vegetation from the site, ingestion of local surface water, and incidental ingestion of soils with vegetation, body burdens of the metals and As in the herbivores were estimated. As a means of estimating potential element bioavailability at the site, the modeled body burdens were compared with measured element concentrations in herbivore tissues. The analysis indicates that the bioavailabilities of As, Cd, Cu, Pb, and Zn were equal to or less than 0.2% for internal organs and 0.1% for carcasses, on a microgram/g tissue wet weight basis. Available site data on soil pH, cation exchange capacity, and extractable elements fraction supported the limited release of soil elements. These results suggest that the bioavailable fraction of mining waste metals in riparian wetland soils may be quite small, and, for the Milltown Reservoir site, lower than originally anticipated.

Animals↗