Comparisons between warm and cold water swim stress in mice

The following experiments evaluated the effects of warm- or cold-water swim stress on tail-flick latencies (TFL) in mice. To first determine the appropriete control group, the TFL's of dry-vs-dunked mice were compared. Dry mice had significantly shorter TFL's than dunked mice, implying that the dampness of the mouse's tail contributed to the increase in the TFL. Therefore, dunked mice were used as the relevant control for the swum mice. Cold water swimming (2°C) produced a significant increase in the TFL; this was not blocked by the opiate antagonist naloxone (3 mg/kg sc) or potentiated by the enkephalinase inhibitor thiorphan (100 mg/kg sc). Warm water swimming (32°C) up to 3 min produced an inconsistent effect on TFL's, implying that the effects were at the threshold of detectability. Naloxone attenuated and thiorphan modestly potentiated the effects of warm water swimming on TFL's. This suggests that warm water swim stress-induced increases in mouse TFL's may involve opioid pathways, whereas cold water swim stress-induced changes in mice TFL's appear not to be opioid mediated.     

The effect of psychoemotional state on Lewis lung carcinoma metastasis

The effect of a depression-like status formed by chronic stress on development of Lewis lung carcinoma metastases in C57Bl/6J mice was investigated. Two types of acute stress (restraint and social stress) were used for comparison. The depression-like status was induced by eight-week exposure to repeated but unpredictable stressors (chronic mild stress model) and was assessed in the forced swim test. Tumor cells were inoculated an hour after the onset of social stressor or immediately after physical or chronic stressor impacts. The number of metastases was counted 17 days after the inoculation. The results indicate that chronic mild stress provokes the development of a depression-like state in mice and causes a twofold increase in the number of metastases in the lungs, while both types of acute stress have no such effects. Thus, a depression-like psychoemotional status of animals enhances the metastasis of Lewis lung carcinoma.     

Work in artificial cold environments

The physiological characteristics of work in cold stores, as a typical artificial cold environment, are reviewed mainly from our various field and experimental studies. There are about 4,000 cold stores in Japan, and 85% of them are kept at temperatures below -20 degrees C. Although the duration of cold exposure per stay in a cold store was very short, forklift workers entered the cold stores very frequently. Cold stress and the decrease in workers' performance were the same as for continuous exposure to cold. Since the peripheral skin temperature of subjects at night is higher than that in the afternoon, they are less likely to feel cold or pain sensation at night. However, there was a marked decrease in rectal temperature and in manual performance. There is an increased risk of both hypothermia and accidents for those who work at night. The cold store workers, however, had adapted to cold through daily repeated cold exposures.     

Physical dependence on physiologically released endogenous opiates

Mice which had been submitted to a chronic schedule of warm water swimming exhibited a naloxone precipiated withdrawal behaviour which was remarkably similar to that produced in mice following chronic morphine treatment. These results are consistent with the activation of endogenous opiates during swim stress in mice and present the possibility that opiate receptors are activated in a manner analogous to the repeated application of exogenous opiates, producing both tolerance and withdrawal-like behaviour.     

Hypothermia-induced tyrosine phosphorylation of SIRPα in the brain

Signal regulatory protein α (SIRPα) is a neuronal membrane protein that undergoes tyrosine phosphorylation in the brain of mice in response to forced swim (FS) stress in cold water, and this response is implicated in regulation of depression-like behavior in the FS test. We now show that subjection of mice to the FS in warm (37 °C) water does not induce the tyrosine phosphorylation of SIRPα in the brain. The rectal temperature (T(rec) ) of mice was reduced to 27° to 30 °C by performance of the FS for 10 min in cold water, whereas it was not affected by the same treatment in warm water. The level of tyrosine phosphorylation of SIRPα in the brain was increased by administration of ethanol or picrotoxin, starvation, or cooling after anesthesia, all of which also induced hypothermia. Furthermore, the tyrosine phosphorylation of SIRPα in cultured hippocampal neurons was induced by lowering the temperature of the culture medium. CD47, a ligand of SIRPα, as well as Src family kinases or SH2 domain-containing protein phosphatase 2 (Shp2), might be important for the basal and the hypothermia-induced tyrosine phosphorylation of SIRPα. Hypothermia is therefore likely an important determinant of both the behavioral immobility and tyrosine phosphorylation of SIRPα observed in the FS test.     

Interaction of genotype and parental environment in the adaptation of wild House mice Mus musculus to cold

Wild House mice, Mus musculus, were bred in two laboratory environments, one warm (controls) and one cold (Eskimo). At the seventh generation, mice of both stocks were cross-fostered at birth in both environments. In the warm environment, differences in both genotype and nest environment influenced growth: (1) Eskimo reared by Eskimo females were the heaviest of the four classes of fostered young; and (2) control foster parentage retarded growth. There was, however, no good evidence of differences in the reproductive performance of the four classes of fostered mice. In the cold environment, the effects of both genetical differences and of fostering were greater. Both the superior growth of Eskimo reared by Eskimo and the retarding effect of control foster parentage were more marked. Moreover, adult males with control foster parents had less fat than had those with Eskimo foster parents. Reproductive performance was also affected: (1) the young of the pairs with Eskimo genotype were heavier than the young of control pairs; (2) the litters of mice with Eskimo foster parents were larger than those of mice with control foster parents, and their young were heavier. Differences among the young of fostered mice represent a grandmother effect. Evidently, selection in a cold environment had led, not only to adaptive genetical changes in the ability to respond directly to cold, but also to changes in parental performance; and the latter enhanced the fitness, in the cold environment, of their offspring and grandoffspring.     

Learning and cross drug effects: Thermic effects of pentobarbital and amphetamine

The effects of environmental cues explicitly paired or unpaired with pentobarbital on the thermic effects of pentobarbital and amphetamine were investigated. Rats received 19 injections of pentobarbital in a distinctive environment and were subsequently tested for the thermic effects of pentobarbital and amphetamine in the distinctive environment, another environment previously associated only with saline, or in the colony room not previously associated with injections. Rats tested in the context of the environmental cues previously associated with pentobarbital were tolerant to the hypothermic effect of pentobarbital, but rats tested in the environment previously associated only with saline or in the colony room were not tolerant. Pentobarbital-experienced rats administered amphetamine in the context of the usual pentobarbital cues exhibited an exaggerated hyperthermic reaction compared to previously drug-naive rats administered amphetamine. Pentobarbital-experienced rats injected with amphetamine in the homeroom exhibited a smaller hyperthermic response than previously drug-naive rats administered amphetamine in the home room. These results demonstrate that an animal's response to a drug can be affected by cues paired and unpaired with drug administration.     

Sudden Failure of Swimming in Cold Water

To investigate the effect of cold water on swimming four men who declared themselves good swimmers were immersed fully clothed on separate days in water at 23·7° and 4·7° C. The time that they were able to swim in the cold water was much shorter than in the warm. The two shortest swims ended after 1·5 and 7·6 minutes, before rectal temperature fell, when the men suddenly floundered after developing respiratory distress with breathing rates of 56–60/min. The other cold swims, by the two fattest men, ended less abruptly with signs of general and peripheral hypothermia.It is concluded that swimming in cold water was stopped partly by respiratory reflexes in the thin men and hypothermia in the fat, and partly by the cold water''s high viscosity. The longer swimming times of the fat men are attributed largely to their greater buoyancy enabling them to keep their heads above water during the early hyperventilation.The findings explain some reports of sudden death in cold water. It is clearly highly dangerous to attempt to swim short distances to shore without a life-jacket in water near 0° C.     

Tolerance and cross tolerance with morphine resulting from physiological release of endogenous opiates

Mice which had been exposed to a chronic schedule of warm water swimming showed the development of a significant tolerance to the antinociceptive response (tail-flick latency) and a significant, two-fold increase in the ED50 of morphine (tail-flick latency and abdominal constriction response). These results suggest the involvement of endogenous opiates during swim stress in mice and are consistent with the hypothesis that during chronic stress the opiate receptors are activated in a manner analogous to the repeated application of exogenous opiates producing tolerance, morphine cross tolerance and (as previously reported) withdrawal-like behaviour.     

Effects of cold on nest-building by wild and domestic mice, Mus musculus L.

Nest-building by male and virgin female wild and laboratory mice kept at a temperature of 2^o C was compared with that of controls kept at 23^o C. The amount of cotton wool pulled into the cage was recorded over 24 hours. Nest quality was also assessed. The tests of nest-building were conducted in both cold and warm environments. Cotton pulling was usually at a lower rate in the cold environment, but there was no corresponding decrement in final nest quality. Previous experience in the cold, compared with absence of such experience, resulted in higher scores by wild mice tested in either environment, and by laboratory mice tested in the cold environment. Wild mice that built high quality nests used less cotton wool in the cold than in the warm environment. The tendency for wild mice to pull more cotton wool and build better nests than laboratory mice was more pronounced among cold-treated animals. Differences in body weight did not account for the differences between wild and laboratory mice.
In further experiments nest-building was observed over 16 days. In the cold environment there was an initial depression of nest-building by both wild and laboratory mice, followed by a steady improvement over 6 days.
Both males and virgin females sometimes made well constructed nests even in the warm environment. In the cold environment the effect of previous cold-exposure on wild mice was the rapid construction of a good nest.     

Metabolic rate and oxidative stress in insects exposed to low temperature thermal fluctuations

Fluctuating temperatures are a predominant feature of the natural environment but their effects on ectotherm physiology are not well-understood. The warm periods of fluctuating thermal regimes (FTRs) provide opportunities for repair leading to increased survival, but there are also indications of negative effects of warm exposure. In this study, we examined respiration and oxidative stress in adult Alphitobius diaperinus exposed to FTRs and to constant low temperatures. We hypothesized that cold exposure will cause oxidative stress and that FTRs would reduce the amount of chill injuries, via activation of the antioxidant system. We measured V˙CO2, activities of super oxide dismutase (SOD), amounts of total (GSHt) and oxidized glutathione (GSSG) during cold and warm periods of FTRs. Increased severity of cold exposure caused a decrease in the glutathione pool. SOD levels increased during the recovery period in the more severe FTR. The antioxidant response was sufficient to counter the reactive oxygen species production, as the GSH:GSSG ratio increased. We conclude that cold stress causes oxidative damage in these beetles, and that a warm recovery period activates the antioxidant system allowing repair of cold-induced damage, leading to the increased survival previously noted in beetles exposed to fluctuating versus constant temperatures.     

Cold water swim stress inhibits the nociceptive responses to intrathecally administered somatostatin,but not substance P

The effects of cold water swim stress (CWSS) on the nociceptive responses to i.t. administered substance P (SP) and somatostatin (SST) were examined. Male ICR mice, weighing about 30 g, were forced to swim in water at 20°C for 3 min. In unstressed mice, i.t. injection of SP (0.1 nmol) and SST (1 nmol), respectively, produced nociceptive-related behaviors. Although CWSS had no effect on the intensity of the SP-induced nociceptive responses, CWSS significantly reduced the intensity of the SST-induced nociceptive responses. The effect of CWSS on the SST-induced nociceptive responses was blocked by naloxone (5 mg/kg, s.c.) and naltrindole (1 mg/kg, s.c.), a selective δ-opioid receptor antagonist, but not by β-funaltrexamine (20 mg/kg, s.c.), a selective μ-opioid receptor antagonist. These results indicate that CWSS may selectively reduce the SST-induced nociceptive responses primarily through δ-opioid receptors.     

Is stress-induced analgesia a measure of stress severity?

When comparing magnitudes of "behavioural despair" (duration of immobility) and stress-induced analgesia in the tail suspension test and cold water swim test with SHR and NMRI male mice. The results might depend on saline injection prior the test and on the fact that exposure to cold water in swim test was sufficient to alter the response patterns. The findings show that the main parameters are closely related to each other. Stress-induced analgesia seems to be a measure of stress as the stress becomes stronger analgesia changes in linear dependence, whereas duration of immobility has an invert U-shaped function.     

The effect of benzodiazepine on the increase of oxygen consumption in cold exposed rats

1. Oxygen consumption and rectal temperature of warm and cold acclimated rats were measured after chronic and acute injections of saline or benzodiazepine (diazepam). 2. Benzodiazepine has blocked the increase in oxygen consumption of warm acclimated rats on exposure to cold. 3. After cold acclimation, the benzodiazepine did not affect the increase in oxygen consumption. 4. Benzodiazepine caused a slight hypothermia when injected chronically, but did not affect rectal temperature over a short period of time.     

Changes in pain reactions and 3H-naloxone binding to opiate receptors of the hypothalamus and midbrain in rats after repeated swimming in cold water

The dynamics of nociceptive reactions and character of 3H-naloxone binding to hypothalamus and midbrain synaptic membranes were studied in rats subjected to repeated cold swim stress (3 min. daily during 3, 5 and 15 days). It was shown that an increase of latencies of background nociceptive reactions (hot-plate and tail-flick tests) was accompanied by an ambiguous changes of kinetic parameters of 3H-naloxone binding in the studied brain structures. The results suggest that an increase of antinociceptive systems tone under repeated cold swim stress may be caused by a dynamic transformation of opiate u-receptor apparatus in various brain structures.     

Involvement of microtubules in the tolerance of cardiomyocytes to cold ischemia-reperfusion

Before transplantation, the heart graft is preserved by the use of cold storage in order to limit ischemia-reperfusion stress. However, sustained exposure to low temperature may induce myocardial ultrastructural damage, particularly microtubules (MT) disruption. Previous data suggested that tubulin-binding agents are able to attenuate cold-induced cytoskeleton alterations. Thus, the aim of the present work was to study the influence of docetaxel (DX, a tubulin-binding taxane) on the effects of deep hypothermia (4°C) and of simulated cold ischemia-reperfusion on the MT network and oxidative stress of cardiomyocyte (CM) in monolayer cultures prepared from newborn rat ventricles. The MT network was explored by immunocytochemistry and Western-blotting, the cell stress by tetrazolium dye assay (MTT) and lactate dehydrogenase (LDH) release, and the superoxide production by the dihydroethidium probe (DHE). The MT assembly remained stable after 4 and 8 h of hypothermia. Tubulin acetylation was promoted in CM subjected to 4-h hypothermia. Low temperature reduced the mitochondrial function and increased the basal LDH release. The cold ischemia during 4 and 8 h preserved MT network. Docetaxel promoted MT polymerization and tubulin acetylation in basal and in cold conditions. This drug decreased the release of LDH induced by cold ischemia. Moreover, hypothermia (4 h) significantly raised the anion superoxide production. Docetaxel decreased this oxidative stress in the control CM and in CM submitted to 4 h of hypothermia. These data demonstrated that stabilizing MT with DX exerted a protective effect on CM subjected to hypothermia and to cold ischemia-reperfusion. Tubulin-ligands should be thus considered to improve the tolerance of the heart graft toward stressing conservative conditions.     

Effects of pyridostigmine bromide on human thermoregulation during cold water immersion

This study examined the effects of an oral 30-mg dose of pyridostigmine bromide (PYR) on thermoregulatory and physiological responses of men undergoing cold stress. Six men were immersed in cold water (20 degrees C) for up to 180 min on two occasions, once each 2 h after ingestion of PYR and 2 h after ingestion of a placebo. With PRY, erythrocyte cholinesterase inhibition was 33 +/- 12% (SD) 110 min postingestion (10 min preimmersion) and 30 +/- 7% at termination of exposure (mean 117 min). Percent cholinesterase inhibition was significantly related to lean body mass (r = -0.91, P less than 0.01). Abdominal discomfort caused termination in three of six PYR experiments but in none of the control experiments (mean exposure time 142 min). During immersion, metabolic rate, ventilatory volume, and respiratory rate increased significantly (P less than 0.05) over preimmersion levels and metabolic rate increased with duration of immersion (P less than 0.01) in both treatment but did not differ between conditions. PYR had no significant effect on rectal temperature, mean body temperature, thermal sensations, heart rate, plasma cortisol, or change in plasma volume. It was concluded that a 30-mg dose of PYR does not increase an individual's susceptibility to hypothermia during cold water immersion; however, in combination with cold stress, PYR may result in marked abdominal cramping and limit cold tolerance.     

In a Model of Batten Disease,Palmitoyl Protein Thioesterase-1 Deficiency Is Associated with Brown Adipose Tissue and Thermoregulation Abnormalities

Infantile neuronal ceroid lipofuscinosis (INCL) is a fatal neurodegenerative disorder caused by a deficiency of palmitoyl-protein thioesterase-1 (PPT1). We have previously shown that children with INCL have increased risk of hypothermia during anesthesia and that PPT1-deficiency in mice is associated with disruption of adaptive energy metabolism, downregulation of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), and mitochondrial dysfunction. Here we hypothesized that Ppt1-knockout mice, a well-studied model of INCL that shows many of the neurologic manifestations of the disease, would recapitulate the thermoregulation impairment observed in children with INCL. We also hypothesized that when exposed to cold, Ppt1-knockout mice would be unable to maintain body temperature as in mice thermogenesis requires upregulation of Pgc-1α and uncoupling protein 1 (Ucp-1) in brown adipose tissue. We found that the Ppt1-KO mice had lower basal body temperature as they aged and developed hypothermia during cold exposure. Surprisingly, this inability to maintain body temperature during cold exposure in Ppt1-KO mice was associated with an adequate upregulation of Pgc-1α and Ucp-1 but with lower levels of sympathetic neurotransmitters in brown adipose tissue. In addition, during baseline conditions, brown adipose tissue of Ppt1-KO mice had less vacuolization (lipid droplets) compared to wild-type animals. After cold stress, wild-type animals had significant decreases whereas Ppt1-KO had insignificant changes in lipid droplets compared with baseline measurements, thus suggesting that Ppt1-KO had less lipolysis in response to cold stress. These results uncover a previously unknown phenotype associated with PPT1 deficiency, that of altered thermoregulation, which is associated with impaired lipolysis and neurotransmitter release to brown adipose tissue during cold exposure. These findings suggest that INCL should be added to the list of neurodegenerative diseases that are linked to alterations in peripheral metabolic processes. In addition, extrapolating these findings clinically, impaired thermoregulation and hypothermia are potential risks in patients with INCL.     

House mice bred for many generations in two environments

Wild house mice, Mus musculus L., were trapped, and their descendants reared, in permanently mated pairs, for a number of generations in two laboratory environments, at about 21°C (controls) and -3°C, respectively. All mice had sawdust and cottonwool for bedding; but the nests of those at -3°C were colder than those in the warm, and fluctuated greatly in temperature.
Reproductive performance was inferior in the cold environment: more pairs were barren, and the fecund pairs reared fewer young than the controls. Yet litters at birth were usually larger in the cold, and the young at three weeks were always heavier. Over ten generations nestling mortality declined at -3°C.
From generation 1 on, adult mice at -3°C were heavier than the controls, but there was no corresponding increase in body length. Tails were much shorter relative to body length in the first generations in the cold, but returned to control proportions by generation 10. Most of the structural changes in the cold accord with the "rules" of Bergmann and Allen.
The incidence of abnormal sixth lumbar vertebrae was low in all generations at both temperatures.
After nine generations, some mice were transferred from the cold to the warm environment, and bred for a further three generations. There they outstripped the controls both in reproductive performance and in growth. They also had more fat, and a heavier and longer small intestine; but the heart, stomach and kidneys were lighter than those of the controls. Adrenal weights at 21°C declined over the generations, but those of the mice at -3°C did not.
The secular changes observed, especially those in the cold environment, are attributed principally to differential selection of genotypes, not to inbreeding; but maternal effects may also have been involved.