The effect of exercise training in a cold environment on thermoregulation in adult and aged C57BL/6J mice

We studied the effect of exercise training in cold environment (six weeks of daily, one-hour runs on a treadmill at ambient temperature of 6 ± I°C at 60±65% of VO_2max) on cold-induced metabolic heat production, heat loss, and cold tolerance in adult and aged C57BL/6J male mice. In adult mice, exercise training in cold environment resulted in greater cold-induced heat production and cold tolerance without changes in heat loss, similar to the effects of daily cold exposure without exercise. In aged mice, daily cold exposures did not affect cold tolerance and cold-induced heat production, but exercise training in the cold resulted in greater cold-induced heat production and cold tolerance. Heat loss in aged mice increased similarly after both repeated cold exposures and exercise training in the cold. Therefore, mechanisms of effect of exercise training on cold tolerance are different in adult and aged animals. Exercise training in cold environment does not affect cold-induced heat production and cold tolerance in adult mice, but improves them in aged animals.     

The effect of exercise training in cold on shivering and nonshivering thermogenesis in adult and aged C57BL/6J mice

To understand the mechanisms of improvement of cold-induced heat production in aged mice following exercise training, the relative contributions of shivering and nonshivering thermogenesis to cold-induced metabolic responses were assessed in adult and aged C57BL/6J male mice, which inhabited sedentarily at room temperature, or were subjected either to a regimen of moderate intensity exercise training at 6°C, or to sedentary repeated exposures to the same temperature. The main findings were that (1) aged mice had greater cold-induced nonshivering thermogenesis, but lower shivering than adult mice; (2) exercise training in a cold environment enhanced cold-induced nonshivering thermogenesis in adult mice, but suppressed it in aged animals; (3) exercise training in a cold environment increased shivering thermogenesis in both age groups, but this increase was much greater in aged mice; (4) the increase of cold-induced shivering thermogenesis was mainly responsible for increased cold tolerance in aged mice after exercise training in a cold environment.     

Nonshivering thermogenesis during acute cold exposure in adult and aged C57BL/6J mice

In C57BL/6J adult and aged mice, housed at room temperature (22.5 ± 1°C), we measured O₂ consumption and CO₂ production and calculated metabolic heat production under conditions of anesthesia and myorelaxation during acute cold stimulation when body temperature was lowered 7.5°C below control level. An independent group of mice was subjected to a three hour partial physical restraint at 6°C and concentration of uncoupling protein (thermogenin) was measured in interscapular brown adipose tissue mitochondria at different times after cold exposure. Heat production under anesthesia and myorelaxation was about 57–66% lower than in nonanesthetized conditions, but increased significantly during cold stimulation in both age groups. Under anesthesia and myorelaxation before and during cold stimulation aged mice produced about 20% more heat than adult mice. Because in these experiments all sources of facultative thermogenesis, except nonshivering, were suppressed by anesthesia and myorelaxation, and because brown adipose tissue is the major source of nonshivering thermoproduction, we concluded that aged mice housed at room temperature have an increased thermogenesis in brown adipose tissue. This conclusion was also supported by the finding that the concentration of uncoupling protein measured in the mitochondria of brown adipose tissue after single cold exposure was significantly higher in aged than in adult mice. Therefore, we propose that the lower, cold-induced, heat production typically observed in nonanesthetized aged mice may reflect reduced thermogenic capacity of skeletal muscles. While aged mice have less brown adipose tissue than adult animals, the remaining brown adipose tissue may compensate by increasing the concentration of uncoupling protein.     

A longitudinal study of tolerance to cold stress among C57BL/6J mice

C57BL/6J male mice of different ages were movement-restricted and exposed to 10 degrees C for 3-hr periods every other week while colonic temperature was measured. A longitudinal trend in cold tolerance related to age and to initial colonic temperature was demonstrated. Adaptative thermoregulatory changes during cold exposure occurred during the first two tests. These were similar for all age groups except 30-month-old mice. There was no adaptation of colonic temperature during cold exposure among aged mice with repeated testing; however, their baseline colonic temperatures prior to testing increased after the first two tests. This finding suggests that old animals adjust to repeated cold stress differently than do younger mice. Specifically, younger animals are capable of adjusting their thermoregulatory response during cold stress with no change in baseline (pre-stress) temperature. Old animals do not modify the responses emitted during the stress; however, they do adapt by raising their baseline temperatures. Repeated cold exposure started later in life increased mortality among old animals but did not affect maximum lifespan.     

Metabolic heat production and cold tolerance during cold stress of different intensity of adult and aged male C57BL/6J mice

Adult and aged male C57BL/6J mice were subjected to a 3-h cold stress test at either 24°C, 18°C, 12°C, or 6°C. Body mass was measured before the test, and colonic temperature, O₂ consumption, and CO₂ production were measured during the test. The slopes of colonic temperature over time of test and the mean metabolic heat production were calculated for each animal. While adult mice had a relatively small reduction in colonic temperature during the test at all four ambient temperatures, in the aged mice ambient temperatures resulted in steeper reductions of colonic temperature. In adult mice, an increase in metabolic heat production was proportional to ambient cold. The thermogenic response of aged mice at 24°C and at 18°C was similar to adult mice, suggesting that the ability of aged mice to respond to cold by increasing heat production does not diminish with age. However, in aged mice metabolic heat production at 12°C and 6°C was significantly below that of adult mice, which indicated a reduced capacity for thermogenesis.     

Physiological and behavioral correlates of lifespan in aged C57BL/6J mice

Physiological and behavioral measurements were made in a cohort of 29-month-old male C57BL/6J mice to determine whether any correlated significantly with lifespan. Significant linear relationships with lifespan were found among the physiological measures, including hematocrit and hemoglobin levels and collagen denaturation rate; however, body weight failed to be a significant predictor of survival. Among the behavioral variables studied, significant quadratic relationships with lifespan were found for exploratory activity and passive avoidance learning, while performance on a rotorod and a tightwire showed no significant relationships with lifespan. Through the use of multiple regression techniques, about one-third of the variance in lifespan could be explained by a combination of physiological variables, and about two-fifths could be explained by a combination of behavioral variables.     

Oxygen consumption in adult and aged C57BL/6J mice during acute treadmill exercise of different intensity

Submaximal and maximal oxygen consumption was determined in untrained adult and aged male C57BL/6J mice during treadmill running. Each of 12-month-old (ADULT) and 24-month-old (AGED) male mice was tested on a motor-driven treadmill once at different speeds. VO₂ was measured before, during, and after exercise by means of indirect calorimetry in metabolic treadmill chambers. The resting VO₂ averaged 3064.67 ± 87.71 mL/kg/h for ADULT mice and 2472.95 ± 69.41 mL/kg/h for AGED mice. During exercise, VO₂ increased linearly with work intensity (running speed): ADULT mice-from 5908.06 ± 422.35 mL/kg/h at 3 m/min to 10861.99 ± 174.03 mL/kg/h at 25 m/min; AGED mice-from 5217.25 ± 263.26 mL/kg/h at 3 m/min to 7817.32 ± 290.28 mL/kg/h at 20 m/min. Further increase of the running speed resulted in a decline of VO₂ in ADULT and refusal to run in AGED mice. The results of this study demonstrated that in untrained C57BL/6J mice VO₂max and maximal exercise capacity declined with age. At the same absolute and relative workloads, VO₂ was lower in AGED mice.     

Hybrid C57BL/6J x FVB/NJ mice drink more alcohol than do C57BL/6J mice

BACKGROUND: From several recent strain surveys (28 strains: Bachmanov et al., personal communication; 22 strains: Finn et al., unpublished), and from data in >100 other published studies of 24-hr two-bottle ethanol preference, it is known that male C57BL/6 (B6) mice self-administer about 10-14 g/kg/day and that female B6 mice self-administer about 12-18 g/kg/day. No strain has been found to consume more ethanol than B6. In one of our laboratories (Texas), we noted a markedly greater intake of ethanol in an F1 hybrid of B6 and FVB/NJ (FVB) mice. METHODS: To confirm and extend this finding, we repeated the study at another site (Portland) using concentrations up to 30% ethanol and also tested B6xFVB F1 mice in restricted access drinking procedures that produce high levels of alcohol intake. RESULTS: At both sites, we found that B6xFVB F1 mice self-administered high levels of ethanol during two-bottle preference tests (females averaging from 20 to 35 g/kg/day, males 7-25 g/kg/day, depending on concentration). F1 hybrids of both sexes drank significantly more 20% ethanol than both the B6 and FVB strains. Female F1 hybrids also drank more 30% ethanol. In the restricted access tests, ethanol consumption in the F1 hybrids was equivalent to that in B6 mice. CONCLUSIONS: These data show that this new genetic model has some significant advantages when compared to existing inbred strains, and could be used to explore the genetic basis of high ethanol drinking in mice.     

Heat loss during cold exposure in ADULT and AGED C57BL/6J mice

Metabolic heat production (MHP), colonic temperature (Tco), and nonevaporative (dry) heat loss were measured in ADULT and AGED C57BL/6J male mice during cold exposure. Dry heat loss was assessed as a differential temperature (Td) between incoming and outgoing air through the chamber for indirect calorimetry. The average Td during cold exposure normalized to surface area for ADULT mice was significantly higher than that for the AGED animals (0.0618 ± 0.0003°C/cm² and 0.0553 ± 0.0005°C/cm², respectively). Linear regression analysis showed that at the same Tco AGED mice showed lower values of Td normalized to surface area, indicating that at the same body temperature they were losing less heat than ADULT animals. It was concluded that age-related decline in cold tolerance in mice is not due to a lack of ability to reduce heat loss during cold exposure. On the contrary, AGED animals had lower heat loss in comparison with ADULT. We suggest that augmentation of heat conservation mechanisms is an adaptive response to diminishing cold-induced heat production.     

Hormonal,metabolic and morphologic studies of aged C57BL/6J obese mice

Genetically obese mice (C57BL/6J-ob/ob), fed ad libitum, demonstrated a precipitous increase in the spontaneous death rate after 50 weeks. The first signs of morbidity were a ruffled hair coat and a progressive motor ataxia. Necropsy revealed that obese mice had pale and fatty livers, urolithiasis and grossly distended bladders. Microscopically, the hepatocellular changes observed in all aged obese mice included: a loss of orientation of hepatocytes, an enormous variability in the size of both hepatocytes and their nuclei, and an extensive deposition of both large and small lipid droplets, confirmed by an increase content of triacyl3lycerols. A subacute-to-chronic, multifocal, necrotizing hepatitis was also present. Kidneys from aged obese mice contained hypertrophied glomeruli and increased PAS-stained material. Tubular dilation with compaction of the tubular cells was also seen. There were no significant alterations in the microanatomy or mineralization of femurs from obese mice, yet there was a significant increase in plasma alkaline phosphatase activity. In obese mice at 62–63 weeks of age, hyperglycemia was present even in spite of hyperinsulinemia. Pituitary immunoreactive ACTH and its molar ration to pituitary immunoreactive β-endorphin were also increased in obese mice at this age. Even though the etiology of the decreased lifespan of genetically obese mice remains uncertain, the possibility is discussed that an overall defect in the central nervous system may be involved.     

Effect of intermittent feeding on thermoregulatory abilities of young and aged C57BL/6J mice

At 6 mth of age, male C57BL/6J mice were subjected to an intermittent schedule of feeding (every-other-day) or continued to be maintained on an ad libitum diet (24% protein). This regimen of dietary restriction resulted in increased mean and maximum lifespan (11%) compared to the survival of ad libitum-fed mice. At 8 and 26 mth of age, different groups of mice were tested for cold tolerance during 3 h exposure to 10 degrees C. Aged mice (26 mth) fed ad libitum compared to young mice (8 mth) on the same diet had significantly lower baseline colonic temperatures prior to cold exposure, and impaired cold tolerance, as measured by the rate of decline in colonic temperature during cold exposure. Dietary restriction by intermittent feeding had no significant effect on cold tolerance in young mice or baseline colonic temperature in young or aged mice. However, the cold tolerance of aged mice subjected to 20 mth of intermittent feeding was markedly improved over that of aged cohorts maintained on an ad libitum diet and in fact was indistinguishable from that of young counterparts. Thus, it appeared that this regimen of dietary restriction when implemented in young adults prevented the age-related decline in cold tolerance observed among ad libitum fed mice of this strain.     

Development of an alcohol deprivation and escalation effect in C57BL/6J mice

BACKGROUND: Relapse-like drinking has been studied through the expression of the alcohol deprivation effect (ADE), which is measured by a pronounced increase in ethanol preference and consumption after imposed abstinence. No studies have characterized the ADE in C57BL/6J (B6) mice. The present study examined the effects of length and number of deprivations on the expression of the ADE in B6 mice. METHODS: Adult male B6 mice received 24-hour continuous access to ethanol and water for 6 weeks (baseline). Experiment 1 determined the ADE in mice receiving weekly access to 15% ethanol (i.e., exposed 1 day a week and deprived during the other 6 days) for a total of 10 weeks. Experiments 2 and 3 determined the ADE after a single 2-week deprivation period in mice receiving a single concentration of 15% ethanol or multiple concentrations of 7.5, 15, and 30% ethanol, respectively, followed by weekly access to their respective ethanol solutions for 10 weeks. Experiment 4 determined the ADE after a single 2-week deprivation period, followed by daily access to 15% ethanol. Mice never deprived of ethanol (i.e., continuous access) were used as age-matched drinking controls. RESULTS: The ADE was observed after the initial 6-day deprivation period and was profoundly enhanced (i.e., escalation of the ADE) following weekly reexposure to 15% ethanol. Compared with a single concentration of 15% ethanol, concurrent access to multiple ethanol concentrations resulted in a near 2-fold increase in baseline ethanol consumption. Regardless of having access to single or multiple concentrations of ethanol, the ADE was not observed immediately after a 2-week deprivation period. The ADE was observed (although to a lesser magnitude and duration) following weekly reexposure to single or multiple concentrations of ethanol. Alternatively, following a 2-week deprivation period, mice receiving daily access to 15% ethanol showed a significant decrease in ethanol intake and preference (i.e., negative ADE). CONCLUSIONS: Short-term deprivations followed by repeated intermittent (weekly) reexposure to ethanol produces a robust ADE in B6 mice. Increasing the initial deprivation length to 2 weeks produces various opposing effects, including erasure of an initial ADE, diminished expression and magnitude of the ADE following weekly exposure, and complete reversal of the ADE following daily exposure to ethanol.     

链脲菌素剂量对C57BL/6J小鼠糖尿病诱导效应的影响

目的 观察不同剂量链脲菌素(STZ)对C57BL/6J小鼠糖尿病诱导效应的影响,探讨其量效关系及最佳剂量范围.方法 将C57BL/6J小鼠按数字随机法分为9个STZ剂量组(A～I组,STZ分别为30、60、80、100、120、150、180、210、240 ms/kg体重),每组15只,腹腔注射;1个对照组,10只,腹腔注射等体积缓冲液.观察各组血糖、体重、血胰岛素和45 d生存率的变化,分析其与STZ剂量的关系.同时取A、C、G及对照组小鼠胰腺、肾脏组织做病理学检杏,并行免疫组化观察胰腺胰岛素及肾脏CD<,68>的表达.结果 C～G组较对照组血糖增高、体重及血胰岛素含量较对照组下降非常显著(P<0.05),且STZ剂量与血糖呈正相关(r=0.984,P<0.05),与血胰岛素含量呈负相关(r=-0.994,P<0.05).C～G组成模率达86.7%～100%,显著高于A、B组的0和40%(P<0.05);45 d生存率为46.7%～73.3%,显著高于H、I组的13.3%和0(P<0.05).A组胰腺、肾脏组织未见明显破坏;C组及G组出现典型的胰岛萎缩变形,胰岛素分泌颗粒减少,肾小球系膜外基质沉着及球周臣噬细胞浸润.结论 C57BL/6J小鼠腹腔注射STZ以80～180 mg/kg体重的剂量制模率高、生存率高,且靶器官损伤典型;该剂量与血糖呈正相关,与血胰岛素含量呈负相关.     

Schedule-induced ethanol self-administration in DBA/2J and C57BL/6J mice

BACKGROUND: The purpose of these experiments was to provide an initial investigation into ethanol self-administration elicited in the schedule-induced polydipsia (SIP) paradigm. METHODS: Mature male mice were food deprived to between 80 and 85% of their baseline weight and received 20 daily 1 hr SIP test sessions in which a food pellet (20 mg) was delivered on a fixed-time 60 sec schedule. In different groups, the acquisition of drinking 5% (v/v) ethanol solution (experiment 1) or water (experiment 2) was recorded along with other behaviors that occurred in the test chambers. RESULTS: Results indicated that C57BL/6J mice drank significantly more ethanol than DBA/2J mice and that C57 mice achieved blood alcohol concentrations as high as 300 mg/dl. Blood alcohol concentrations were consistently correlated with g/kg ethanol intake. The groups did not differ in consumption of water. SIP test sessions using higher concentrations of ethanol (10-20% v/v, experiment 1) or sucrose solutions (0.1-2% w/v, experiment 2) then were performed. Group differences in ethanol consumption were maintained at all ethanol concentrations. Although DBAs drank more of a low concentration of sucrose (0.1%), when expressed as g/kg, sucrose intake was equivalent in the two strains at all concentrations. Analysis of the time course of drinking clearly showed that this behavior was adjunctive in nature. CONCLUSION: These results demonstrate the effectiveness of this procedure in inducing ethanol self-administration and its utility for investigating the genetic bases of vulnerability toward excessive ethanol consumption.     

Body temperature of C57BL/6J mice with age

On the basis of a study of rectal temperature in a group of 180, C57BL/6J male mice, ranging in age from 3 months to 30 months, the following conclusions were drawn: 1) The positive correlation between body weight and body temperature typical for rodents was found only for young adults of the C57BL/6J strain; 2) body temperature of male C57BL/6J did not appear to decline until about 23.5 months, after which there was a significant negative correlation (r = -0.53) between age and temperature.     

Experimental scoliosis in melatonin-deficient C57BL/6J mice without pinealectomy

The etiology of idiopathic scoliosis is unknown. Scoliosis with many characteristics closely resembling those seen in idiopathic scoliosis has been produced in young chickens and bipedal rats after pinealectomy. In this study, we induced experimental scoliosis in C57BL/6J mice without pinealectomy and melatonin treatment suppressed the development of scoliosis. A total of 100 mice were divided into four groups: 20 quadrupedal mice served as controls; 30 mice underwent resection of two forelegs and tail at 3 wk of age (bipedal mice); the remaining 20 quadrupedal and 30 bipedal mice received intraperitoneal melatonin (8 mg/kg BW) at 19:00 hr daily. Before killing, blood samples were collected in the middle of dark cycle and melatonin levels were measured by radioimmunoassay. Spine X-ray and helical 3D-CT were examined after killing at 5 months of age. The bipedal mice without a tail were able to walk with standing posture, whereas the quadrupedal mice did not walk with standing posture. In C57BL/6J mice, the serum melatonin was reduced to nearly zero; however, the normal level was restored in both bipedal and quadrupedal mice after the injection of melatonin. Scoliosis with rib humps developed in 29 of 30 bipedal and in five quadrupedal mice. None of mice with melatonin treatment developed scoliosis. The results suggest that melatonin deficiency in bipedal mice appears to play crucial role for development of scoliosis. Also the restoration of melatonin levels prevents the development of scoliosis.     

Calcium and calmodulin changes with ageing in C57BL/6J mice

Male C57BL/6J mice ranging in age from 50 to 1186 days were used to measure total calcium and calmodulin concentrations. The increase in calcium between 0 and 1,000 days of age was 260% for kidney, followed by brain (189%), heart (173.5%), lung (106.5%) and liver (78.5%). Calcium in femur declined by 28.2%. The calmodulin content of liver increased with ageing. Both liver and kidney calmodulin concentrations declined early in life followed by ageing-related increases. Brain, lung and heart calmodulin concentrations did not change significantly with ageing. We conclude that changes in calcium homeostasis are not reflected in calmodulin changes. The loss of calcium in bone is consistent with the occurrence of osteoporosis in ageing C57 mice.     

Iron accumulation and lipid peroxidation in aging C57BL/6J mice

Total iron concentrations in organs from C57BL/6J male mice increased with age. In animals ranging 45 to 900 days of age liver iron increased by 216%, heart by 66%, kidney by 54% and brain by 27%. Two separate phases of iron accumulation were found in brain, kidney and liver. Between 45-355 days of age brain iron increased by 33% and after 355 days there was no change. For both kidney and liver no change was found until after 355 days of age when liver increased by 140% and kidney by 44%. The kidney and liver results suggest a true aging phenomenon. Liver peroxidation potential as measured by the thiobarbituric acid test tended to increase with age but the differences were not significant. The addition of metal ion chelators greatly reduced lipid peroxide values for all organs but no significant age-related trend was evident. We conclude that the large aging-related increases in tissue iron are not correlated with increased concentrations of lipid peroxides.     

Aging and uterine growth during implantation in C57BL/6J mice

Uterine growth during implantation was compared in C57BL/6J mice aged 3–7 mo vs. 11–12 mo. All mice had given birth to at least one previous litter. Older mice had smaller uteri during early gestation (3 to 10 days post coitum) as measured by DNA content, although wet weight and protein were generally similar in both age groups. However, there were no age effects on uterine growth during implantation, as measured by the increments of DNA, as well as protein and wet weight per implantation site (decidual swelling) on days 6–10. These data are discussed in terms of the major increase in fetal death and resorptions in older mice subsequently observed by day 12–13. Larger age-related impairment of the artificially-induced decidual response is also considered. We conclude that impairments of the artificially induced decidual response do not predict the extent of decidual responses during pregnancy in aging mice.     

Temporal decrease of body temperature in middle-aged C57BL/6J mice

Colonic temperature of C57BL/6J mice of both sexes between 12 and 19 months was measured at normal ambient temperature in four independent studies. One study was cross-sectional and the others were longitudinal. In all samples, colonic temperature at about 14 months was always lower than temperature at earlier ages. Furthermore, temperature always returned to earlier levels within 2 weeks; however, it remained unstable up to the age of 18 months after which it returned to the pre-14-month level until finally it fell late in life (after 23 months). Additionally, parametric studies showed that there were differences in the reliability of temperature probes made by different manufacturers to measure absolute temperature. These differences, however, did not affect relative differences among temperatures. Further parametric studies showed that colonic temperature in restrained mice rises 1.2 degrees C within the first 5 min after restraint indicating that duration of restraint is an important factor in studies such as these.