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J Appl Physiol 94: 677-684, 2003. First published November 1, 2002; doi:10.1152/japplphysiol.00469.2002
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Vol. 94, Issue 2, 677-684, February 2003

Differential metabolic capacity of mice selected for magnitude of swim stress-induced analgesia

Iwona B. Łapo1, Marek Konarzewski2, and Bogdan Sadowski1,3

1 Institute for Genetics and Animal Breeding, Polish Academy of Sciences, 05-552 Wólka Kosowska; 2 Institute of Biology, University of Białystok, 15-950 Białystok; and 3 Department of Experimental Pathology, Medical Academy of Warsaw, 00-325 Warsaw, Poland


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Maximum oxygen consumption (VO2) elicited by swimming in 20°C water or by exposure to -2.5°C in helium-oxygen (Helox) atmosphere is higher in mice selected for low (LA) than for high (HA) stress-induced analgesia (SIA) produced by swimming. However, this line difference is greater with respect to swim- than to cold-elicited VO2. To study the relationship between the analgesic and thermogenic mechanisms, we acclimated HA and LA mice to 5°C or to daily swimming at 20 or 32°C. Next, the acclimated mice were exposed to a Helox test at -2.5°C and to a swim test at 20°C to compare VO2 and hypothermia (Delta T). Cold acclimation raised VO2 and decreased Delta T. These effects were similar in both lines in the Helox test but were smaller in the HA than in the LA line in the swim test. HA and LA mice acclimated to 20 or 32°C swims increased VO2 and decreased Delta T elicited by swimming, but only HA mice acclimated to 20°C swims increased VO2 and decreased Delta T in the Helox test. We conclude that the between-line difference in swim VO2 results from a stronger modulation of thermogenic capacities of HA mice by a swim stress-related mechanism, resulting in SIA. We suggest that the predisposition to SIA observed in laboratory as well as wild animals may significantly affect both the results of laboratory measurements of VO2 and the interpretation of its intra- and interspecific variation.

maximum oxygen consumption; thermogenic capacity; swim stress


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

MEASUREMENTS OF SUMMIT METABOLIC capacity of animals are particularly important in physiological and evolutionary ecology (3, 9, 13, 17-19, 34). The most often analyzed measure of metabolic capacity is maximum oxygen consumption (VO2), reflecting the ability to mobilize energetic resources and convert them to heat or mechanical work. Such mobilization often takes place in threatening situations (such as fleeing from a predator or a sudden heat loss in a cold environment), as well as when chasing a prey or during competition for a mate. Thus a high VO2 is potentially of great importance for animals' survival and reproductive success (45).

To date, research on the physiology of VO2 was mainly focused on mechanisms underlying heat production or mechanical work (2, 5, 7, 17, 18). However, in our earlier study (33), we demonstrated that VO2 differs in mice selectively bred for high and for low stress-induced analgesia (SIA), which is a transient decrease of pain sensitivity frequently observed in animals faced with both artificial and natural threatening stimuli (25, 50). We estimated that genetically determined predisposition of mice to SIA accounts for no less than 10% of variability of their VO2 (33). The relationship between VO2 and SIA was strong enough to suggest the existence of a negative genetic correlation. This means that VO2 not only depends on animals' metabolic capacity, but is also affected by stress-related mechanisms, resulting in a suppression of pain sensitivity.

SIA has been reported not only in laboratory rodents, but also in wild animals (see Refs. 1, 25) and is elicited by various emergencies, such as intraspecific agonistic competitions or predator-prey interactions (26, 27, 38, 51). Because such emergencies are also likely to elicit a rapid increase in the rate of metabolism, the relationship between VO2 and SIA may have important consequences for the animals' survival.

It is intriguing, however, that, in laboratory mouse lines divergently selected for high and low swim SIA, we observed a negative relationship between VO2 and SIA magnitude. The animals were subjected to forced swimming, which imposed, in addition to muscular exercise and hypothermic challenge, an important emotional load, resulting from the affection of an emergency condition. The high analgesia (HA) mouse line, manifesting a considerably elevated nociceptive threshold after completion of the swim, displayed significantly lower maximum metabolic rates during swimming (VO2 swim) and greater postswim hypothermia (Delta Tswim), compared with the low analgesia (LA) line, in which the analgesic effect of swim stress was hardly detectable (33). Importantly, not only swimming, but also 15-min exposure to ambient cold in Helox (79% helium-21% oxygen) atmosphere, causing a rapid decrease in core temperature, activated the analgesic mechanism in HA mice (47) and resulted in between-line differences in the level of maximum VO2 (VO2 Helox) and post-Helox hypothermia (Delta THelox) (33, 47). Thus it is likely that SIA affects animals' thermogenic capacity. Because VO2 swim is lower than VO2 Helox, whereas Delta Tswim is higher than Delta THelox (33), the effect of SIA may be more pronounced during swimming due to an emotional load. This is supported by a higher swim-elicited analgesia than that elicited by exposure to ambient cold in Helox (47).

In the concurrent study on HA mice, we showed that acclimation to cold did not affect predisposition to SIA, whereas acclimation to repeated swimming significantly attenuated SIA (34a). Here we have taken advantage of these findings. To study the mechanisms of the interaction between an emotional load of swimming and metabolic capacity, we compared swim-elicited and Helox-elicited VO2 and Delta T in mice of both lines acclimated to ambient cold (which enhanced their thermogenic capacities, without affecting predisposition to SIA) or repeated swimming (which attenuated SIA). To further evaluate the effect of the emergency condition of swimming on the animals' thermogenic capacity, we compared VO2 and Delta T in subgroups of naive mice forced to swim and merely immersed in water.


    MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Animals

Subjects in all experiments were Swiss-Webster mice of both sexes, selectively bred for over 40 generations toward high and low swim SIA. The selection was carried out in the Institute for Genetics and Animal Breeding, Polish Academy of Sciences, Jastrzebiec, Poland, as described elsewhere (41). Briefly, outbred mice were exposed to 3-min swimming in 20°C water and, 2 min after completion of the swim, were measured for the latency of a nociceptive reflex on a hot plate at 56°C. Males and females displaying the longest (50-60 s) and the shortest (<10 s) postswim latencies of the hindpaw flick or lick response were chosen as progenitors of the HA and the LA line, respectively. A similar procedure was repeated in each offspring generation, and only subjects displaying the longest and the shortest postswim hot plate latencies were mated to maintain, respectively, the HA and the LA line.

The animals, except for the cold-acclimated and their control subgroups (see below), were housed in same-sex and same-family groups of four to five per cage (290 × 210 × 100 mm) at 23°C. In all experiments, mice were maintained on 12:12-h light-dark cycle and had unlimited access to murine chow and water.

Acclimation to Cold

At the age of 3 wk, HA and LA mice of the 43rd generation were randomly assigned to two groups. The experimental group was acclimated during the subsequent 6 wk to 5°C, and the unacclimated group was maintained at 23°C. Because small rodents tend to huddle together to improve their heat economy at low ambient temperatures (10), each mouse was confined to a separate cage. This procedure necessarily affected the social status of the animals; therefore, it was applied both to acclimated and unacclimated groups.

Acclimation to Repeated Swimming

Seven-week-old HA and LA mice of the 44th generation were randomly assigned to three groups. The first group was exposed for 14 days to a 3-min swim at 20°C, a standard condition consistently causing a pronounced between-line difference in Delta Tswim. The second group was given the same daily swims at 32°C, which were found earlier to produce only a little Delta Tswim in the HA line (40). Animals swam individually in a plastic container filled with tap water up to 300 mm above the floor. The control group did not swim but was kept in the same room to experience a comparable amount of environmental disturbance.

Sequence of Trials

Mice of the 43rd generation acclimated to cold and mice of the 44th generation acclimated to swimming, as well as their unacclimated controls, were given a Helox test (15-min exposure to -2.5°C in Helox atmosphere) and 3 days later a swim test (swimming in 20°C water for 5 min). The above temporal and temperature parameters were found earlier to produce maximum metabolic rates during cold exposure in Helox and during swimming (33). Additionally, part of naive HA and LA mice of the 47th generation either were given a swim test or were merely immersed in 20°C water. All mice were exposed to the Helox test, the swim test, and the immersion test at ~9-10 wk of age. Their average body mass was 35 g (males) and 29 g (females).

Measurements of VO2

To measure VO2 Helox, we placed the mouse for 15 min in a 350-ml metabolic chamber vented with Helox gas mixture and submerged in temperature-controlled (-2.5 ± 0.2°C) glycol-based coolant. The use of Helox atmosphere was justified by a fourfold higher heat conductivity of helium compared with nitrogen (44), which ensures rapid removal of heat from the mouse body.

To measure VO2 swim, we used a vertically positioned cylindrical Plexiglas metabolic chamber (250 mm high, 115-mm diameter), vented with atmospheric air. The chamber was partly filled with water, leaving the air volume of 560 ml above the water level. The temperature of water within the chamber was maintained at 20 ± 0.2°C. Each mouse was placed just above the water level on a movable platform and allowed 10 min for adaptation. The platform was then abruptly submerged to force the animal to swim.

The same metabolic chamber was used to measure VO2 during the immersion (VO2 imm). After 10-min adaptation, the platform was lowered to 50 mm below the surface of water. The mouse remained partially submerged with its head above the water surface but was not forced to swim. As soon as the mouse completed the swim or immersion trial, the platform was raised, and the mouse was removed from the metabolic chamber.

Because metabolic measurements during swimming require 5 min to minimize measurement error (33), mice were exposed to swim as well as to immersion for 5 min, instead of 3 min applied in the selection protocol (41).

For all metabolic measurements, we used a positive-pressure, open-circuit respirometry system. Outside atmospheric air or Helox was pushed through a column of Drierite to remove water vapor and then was passed through a mass flow controller (Sierra Instruments, Monterey, CA, or ERG-1000, Warsaw, Poland) at the rate of 700 ml/min. Before reaching the metabolic chamber, the gas stream was forced through a copper coil, also submerged in the same water bath to equalize the temperature. Depending on the type of measurement, we simultaneously used one (VO2 Helox) or two metabolic chambers (VO2 swim or VO2 imm), monitored by separate channels of the measurement setup. The gas stream from the metabolic chamber was directed to a computer-controlled channel multiplexer, which is a part of a Sable Systems TR-1 oxygen analyzer setup (Henderson, NV). The analyzed gas stream was scrubbed of CO2 (Carboabsorb AS, BDH Laboratory Supplies), redried (Drierite), then subsampled at the rate of 75 ml/min with a subsampler, and passed through the sensor of an S-3A/I Applied Electrochemistry (Pittsburgh, PA) analyzer. The electrical signal from the analyzer was filtered through a baselining system and then interfaced to an analog-to-digital converter and computer that averaged readings every 0.5 s. Maximum metabolic rate was defined as the highest VO2 averaged over 2 min of 5-min swimming and immersion (VO2 swim and VO2 imm, respectively), and the highest VO2 averaged over 2 min of the last 5 min of 15-min Helox exposure (VO2 Helox). Whenever two chambers were simultaneously used, the VO2 data were corrected for a subtle difference in gas flow through either measurement system.

Metabolic data were analyzed with Sable System DATACAN V software. We calculated VO2 rates using Equation 4a of Withers (52) and attempted to correct instantaneous values of the highest VO2 for the chamber washout time by applying a Z transformation (46).

Measurement of Core Temperature and Body Mass

Colonic temperature was measured to the nearest 0.1°C with a thermocouple thermometer (BAT-12, Physitemp Instruments, Clifton, NJ) twice, i.e., before and after each test. The second measurement was made immediately after cold exposure in Helox and 2 min after completion of the swim or immersion, i.e., as the temperature attained the lowest level (47, 48). The difference between pre- and postexposure core temperatures was taken as the magnitude of Delta THelox, Delta Tswim, or Delta Timm.

Before each trial, the animals were weighed to the nearest 0.1 g. All trials were conducted between 0800 and 1900.

Statistics

The general linear models of analysis of covariance (ANCOVA) were used, taking body mass as covariate (49). First, the effects of line and treatment (acclimation to cold or swim, or immersion vs. swimming) as fixed factors were tested by using a cross-nested three-way ANCOVA model. Family affiliation nested within line × treatment interaction was a random factor controlling for the possible effect of the animal's relatedness. Next, a two-way nested ANCOVA model (treatment as a fixed factor and family nested within treatment as a random factor) was used to test the effects of treatment on VO2 and Delta T within each selected line. A similar model (line as a fixed factor and family nested within line as a random factor) was used to test the effects of the selected line on VO2 and Delta T within a given treatment. It is important to note that the differentiation of the number of families for the particular tests has affected the number of degrees of freedom in the analyses. Sex was omitted in all comparisons, because preliminary analyses did not detect any sex-related differences. Detailed comparisons between subgroups were made with planned contrasts. Body mass was a significant covariate in all reported ANCOVAs (P < 0.05).


    RESULTS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Acclimation to Cold

Helox test. As shown in Fig. 1, VO2 Helox in cold-acclimated mice was significantly higher, whereas Delta THelox was lower than in unacclimated controls (nested three-way ANCOVAs; Table 1). However, neither VO2 Helox nor Delta THelox differed between the lines.


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  		Fig. 1.   Effects of cold acclimation (5°C) on oxygen consumption (VO2) elicited by exposure to 79% helium-21% oxygen (Helox) (VO2 Helox; A) and post-Helox hypothermia (Delta THelox; B) in the high-analgesia (HA) and in the low-analgesia (LA) line. Data are presented as least squares means from analysis of covariance within each line with body mass as covariate ± SE. * Different from unacclimated controls, P < 0.05.


                              
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  		Table 1.   Nested ANCOVAs of VO2 and Delta T in mice acclimated to cold

Swim test. VO2 swim significantly differed between the lines and increased due to cold acclimation (nested three-way ANCOVA; Table 1; Fig. 2A). A significant line × cold acclimation interaction indicates that this increase was less conspicuous in the cold-acclimated HA mice. Nevertheless, it was highly statistically significant within each line (F1,32 = 34.87, P < 0.0001 and F1,32 = 131.92, P < 0.0001, separate nested two-way ANCOVAs for the HA and the LA line, respectively).


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  		Fig. 2.   Effects of cold acclimation (5°C) on maximum metabolic rates during swimming (VO2 swim; A) and postswim hypothermia (Delta Tswim; B) in HA and in LA mice. Data are presented as least squares means from analysis of covariance within each line with body mass as covariate ± SE. Different from * unacclimated controls and dagger  the respective subgroup of the other line, P < 0.05.

Compared with the LA line, lower VO2 swim in cold-acclimated HA mice was accompanied by higher Delta Tswim (nested three-way ANCOVA; Table 1; Fig. 2B). However, the Delta Tswim decrease caused by cold acclimation was significant in the LA but not in the HA line (F1,32 = 7.80, P < 0.01 and F1,32 = 0.21, P = 0.6503, respectively, separate nested two-way ANCOVAs within lines; Fig. 2B).

Acclimation to Repeated Swimming

Helox test. VO2 Helox was higher in LA than in HA mice and was markedly augmented by the acclimation to repeated swimming (nested three-way ANCOVA; Table 2; Fig. 3A). A significant line × repeated swimming interaction suggests that the effect of swim acclimation on VO2 Helox differed between the lines. This is supported by a significant difference between swim-acclimated and unacclimated HA mice (F2,58 = 12.22, P < 0.001), whereas no such difference was seen in the LA line (F2,58 = 0.24, P = 0.7866, separate nested two-way ANCOVAs within each line). An increase in VO2 Helox was significant in HA mice acclimated to repeated swims in 20°C (P < 0.001, planned contrasts), but not in 32°C water (Fig. 3A).

                              
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  		Table 2.   Nested ANCOVAs of VO2 and Delta T in mice acclimated to repeated swimming



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  		Fig. 3.   Effects of repeated swimming in 20 and 32°C water on VO2 Helox (A) and Delta THelox (B) in HA and LA mice. Data are presented as least squares means from analysis of covariance within each line with body mass as covariate ± SE. Different from * unacclimated controls and dagger  the respective subgroup of the other line, P < 0.05.

The between-line difference in Delta THelox was inversely related to the difference in VO2 Helox. HA mice displayed significantly higher Delta THelox than LA mice (nested three-way ANCOVA; Table 2; Fig. 3B). The change in Delta THelox due to swim acclimation was line dependent, as suggested by a significant line × repeated swimming interaction and confirmed by within-line comparisons. Thus the repeated swimming caused an attenuation of Delta THelox only in HA mice (F2,58 = 6.10, P < 0.01, nested two-way ANCOVA) and only if it was performed in 20°C water (P < 0.01, planned contrasts). No attenuation of Delta THelox due to swim acclimation was observed in the LA line.

Swim test. VO2 swim was lower in the HA than in the LA line and significantly rose after repeated swimming. The increase in VO2 swim was equal in both lines, as shown by nonsignificant line × repeated swimming interaction (nested three-way ANCOVA; Table 2; Fig. 4A), and did not depend on the repeated swim temperature.


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  		Fig. 4.   Effects of repeated swimming in 20 and 32°C water on VO2 swim (A) and Delta Tswim (B) in the HA and in the LA line. Data are presented as least squares means from analysis of covariance within each line with body mass as covariate ± SE. Different from * unacclimated controls and dagger  the respective subgroup of the other line, P < 0.05.

Although Delta Tswim in HA mice significantly decreased after repeated swimming, it was still considerably higher than in the LA line (nested three-way ANCOVA; Table 2; Fig. 4B). A nonsignificant line × repeated swimming interaction indicates that the attenuation of Delta Tswim did not differ between the mouse lines. No difference was seen between the effectiveness of 20 and 32°C swimming in the HA line. In LA mice, repeated swimming in 20°C water caused a slightly, but significantly, greater attenuation of Delta Tswim, compared with swimming at 32°C (P < 0.01, planned contrasts).

Immersion in Water vs. Swimming

VO2 was higher in naive mice merely immersed for 5 min in 20°C water than in their swimming counterparts (nested three-way ANCOVA; Table 3; Fig. 5A). The difference between VO2 imm and VO2 swim was greater in HA than in LA mice, as indicated by a significant effect of line and a significant line × water immersion interaction. Nevertheless, HA mice manifested lower VO2 imm as well as VO2 swim than LA mice (F1,20 = 5.54, P < 0.03 and F1,19 = 87.54, P < 0.0001, separate nested two-way ANCOVAs within immersed and swimming mice, respectively; Fig. 5A).

                              
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  		Table 3.   Nested ANCOVAs of VO2 and Delta T in mice immersed in water



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  		Fig. 5.   Effects of immersion in 20°C water for 5 min on VO2 (A) and Delta T (B) in HA and LA mice. Data are presented as least squares means from analysis of covariance within each line with body mass as covariate ± SE. Different from * swimming subgroups and dagger  the respective subgroup of the other line, P < 0.05.

VO2 was inversely related to the magnitude of Delta T. Accordingly, the immersion elicited far lower Delta T than swimming (nested three-way ANCOVA; Table 3; Fig. 5B). This difference was greater in HA than in LA mice, as shown by a significant effect of line and a significant line × water immersion interaction. Both Delta Timm and Delta Tswim in HA mice were significantly higher than those in LA mice (F1,20 = 39.93, P < 0.0001 and F1,19 = 485.83, P < 0.0001, separate two-way ANCOVAs within immersed and swimming groups, respectively; Fig. 5B).


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Acclimation to ambient cold and to daily swims increased metabolic rates in HA and LA mice when exposed both to ambient cold in Helox and to swimming. The increase in VO2 did not differ between the lines when it was measured under the acclimation conditions, i.e., when cold-acclimated mice were subject to Helox exposure and swim-acclimated animals were given the swim test. However, the increase in VO2 did differ between the lines, when it was measured under conditions of the other procedure, i.e., when cold-acclimated mice were given the swim test and swim-acclimated mice were exposed to ambient cold in Helox. This suggests that the two acclimation procedures affected different physiological mechanisms controlling the animals' thermal homeostasis.

Acclimation to Cold

A long-term exposure of animals to low ambient temperatures elicits an increase in metabolic rate to balance the heat loss and maintain body temperature in a cold environment (8). Accordingly, the observed increase in VO2 Helox (Fig. 1A), together with the decrease in Delta THelox (Fig. 1B), appears to reflect an increased thermogenic capacity of cold-acclimated mice. Cold acclimation also elevated metabolic rates during swimming in 20°C water (Fig. 2A). The elevated heat production in acclimated rodents is mainly linked to nonshivering thermogenesis and the functioning of the brown adipose tissue (8, 24). Brown adipose tissue is likely to produce heat during swimming, because the capacity of nonshivering thermogenesis, elicited by norepinephrine injection, significantly increases after acclimation to repeated swimming at water temperature causing hypothermia (personal communication). It is, however, noteworthy that the cold-acclimated HA mice displayed a considerably smaller increase in VO2 swim than LA mice did (Fig. 2A). Clearly, a mere acclimation to cold, without repeated swim experience, was not sufficient to counteract the hypothermic challenge of swimming in the HA line.

Acclimation to Repeated Swimming

One might assume that the increase in VO2 swim in HA and LA mice after repeated swims (Fig. 4A), similar to rats exposed to repeated treadmill exercise (7, 43), was due to enhanced muscle capacity resulting from physical training. This assumption is plausible because, in the muscles of rats exposed to repeated swimming (16), the activity of respiratory enzymes rose just like after repeated treadmill exercise (14, 15, 23). Because the improvement of muscle capacity did not depend on the temperature of swimming (16), our data showing that VO2 swim rose to the same extent in mice given repeated swims at 20°C as at 32°C seem to be consistent with this observation.

However, it should be taken into account that repeated swimming is less efficient than treadmill runs in improving the capacity of exercising muscles (Ref. 16 vs. Refs. 14, 15). Furthermore, the treadmill training is known to increase the exercise metabolism without influencing the metabolic rate in ambient cold (7). In the present study, daily swimming in 20°C water elevated the thermogenic capacities of HA mice, not only under conditions of swim stress, but also during exposure to cold Helox atmosphere (Fig. 3). We assume that this last effect was due to thermal acclimation of HA mice to a repetitive marked hypothermia that accompanied each daily swim at 20°C. Such interpretation is supported by our most recent data showing that daily swimming in 20°C water for 3 wk raised the capacity of nonshivering thermogenesis more in HA and LA mice (personal communication). Nevertheless, we claim that the elevation of swim metabolism in HA mice exposed to repeated swims resulted from an acclimation to the emergency component of the swim stress, rather than to the swim-produced hypothermia. This is supported by a considerable attenuation of their SIA after acclimation to repeated swimming (34a). In consequence, thermoregulatory processes in swim-acclimated HA mice became less susceptible to the stressful condition of swimming, and thus the increase in VO2 swim and decrease in Delta Tswim in HA mice were similar to those seen in the LA line (Fig. 4).

Metabolic Capacity and Swim Stress-related Mechanisms

A comparison of metabolic consequences of cold acclimation and repeated swimming suggests that the thermogenic capacity of the two selected lines is essentially similar, but during swimming it is damped more in HA than in LA mice. The impairment of thermogenesis is most likely to be emotional in nature, swim stress-related mechanism, resulting in SIA. This is in agreement with our results showing that swim-elicited SIA of HA mice was attenuated by acclimation to repeated swimming, but not by acclimation to cold (34a). Furthermore, this is congruent with high emotional sensitivity of HA mice, as revealed by their high magnitude of acoustic startle response and a suppression of open-field behavior (4). We hypothesize, therefore, that a differential modulation of thermogenic capacities of HA and LA mice by an extrathermal stress factor may account for the between-line difference in VO2 swim. An important argument for this hypothesis is that HA mice remaining merely immersed manifested only slightly lower VO2 (Fig. 5A) and slightly higher Delta T (Fig. 5B) than LA mice. This effect was surprisingly similar to that observed during the exposure to ambient cold in Helox. Accordingly, the magnitude of analgesia assessed after immersing HA mice was far lower than after swimming (34a). Thus not a mere contact with cold water, but also the condition in which the mouse can or cannot sense ground under its feet, appears essential for the swim thermogenesis.

The phenomenon of the unexpectedly low metabolic rates that accompany changes in behavior was observed in many swimming rodents (2, 37). We recently demonstrated (42) that the emotional component of the swim stress can account for differential swimming behavior of the selected mouse lines. In contrast to LA mice, HA mice remain relatively inactive when swimming and, after initial excitation, perform only a minimum amount of movements, enough to keep the nose over the surface of the water. At certain swim parameters, the immobility of HA mice is reversed by antidepressant treatment, not effective after administration of naltrexone, an opioid receptor antagonist (42). These observations, together with the upregulation of multiple opioid systems in HA mice (30-32, 35, 39), suggest that the differential thermogenic capacities of HA and LA mice during swimming can, at least in part, depend on the difference in swim stress-induced adaptive mechanisms mediated by endogenous opioids. Thus the line differentiation in swim behavior might also explain why swim metabolism in HA mice is lower than that in LA mice (Figs. 2A and 4A), whereas generally higher VO2 Helox, not involving swimming behavior, differs only little, and inconsistently (Figs. 1A and 3A; see also Ref. 33), between the lines.

Significance of the Relationship Between Metabolic Capacity and SIA

Significant population differences of the levels of VO2 (6, 17, 22), as well as of SIA (28, 29), indicate that both high metabolic capacity and high SIA may be a target of natural selection. For example, Hayes and O'Connor (20) reported better winter survival of Peromyscus maniculatus individuals characterized by above-average VO2. Furthermore, Marek and Szacki (36) found that wild house mice (Mus musculus) and field mice (Apodemus agrarius) are characterized by twofold higher SIA elicited by swimming than outbred laboratory Swiss mice, whereas SIA magnitudes in both wild species are similar to those observed in HA mice. For this reason, a reduction of VO2 due to SIA reported here is puzzling, because one can expect that an increase, not a decrease, of metabolic capacity would be beneficial under stressful circumstances inducing analgesia.

Undoubtedly, an increase in metabolic rate and high SIA are the components of the integrated behavioral and physiological response to stressful situations. For example, island populations of deer mice, compared with mainland conspecifics, display higher opioid-mediated analgesia and reduced level of locomotor activity in response to stressors (28). Moreover, a passive defensive behavior or freezing commonly observed in animals confronting dangerous situations is often associated with a reduction in heart and respiratory rates as well as Delta T (e.g., Refs. 11, 12). It is possible that a suppression of locomotory activity and a transient decrease in metabolic rate of animals faced with a threatening stimulus are primarily defensive responses. Under some circumstances, such as rapid heat loss elicited by cold exposure, a decrease in pain sensitivity may be maladaptive because of the modulation of thermoregulatory mechanisms. The reduction of VO2 due to predisposition to SIA reported in our study most likely results from a significant, negative genetic correlation between these traits (33) and, therefore, cannot be attributed to genetic drift inherent to the process of artificial selection (21), which would question its relevance to natural populations. Clearly, VO2 magnitude is determined not only by metabolic and thermoregulatory processes, but also by psychophysiological phenomena. Thus our study calls attention to the possible, confounding effect of SIA on the results of measurements of metabolic rates. Moreover, this illustrates the need to take into account the relationship between metabolic capacity and SIA in inter- and intraspecific comparisons.


    ACKNOWLEDGEMENTS

The authors thank J. S. Mogil and two anonymous reviewers for criticism and comments. We also thank I. Gasiorkiewicz, A. Koszewnik, M. Lewoc, B. Lewonczuk, and B. Sobolewska for technical assistance.


    FOOTNOTES

This study was supported by Polish Committee for Scientific Research Grants Nr 6PO4C01616 and Nr 6PO4C01818 (to B. Sadowski).

Address for reprint requests and other correspondence: I. B. Łapo, Institute for Genetics and Animal Breeding, Polish Academy of Sciences, 05-552 Wólka Kosowska, Poland (E-mail: i.lapo{at}ighz.pl).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

First published November 1, 2002;10.1152/japplphysiol.00469.2002

Received 28 May 2002; accepted in final form 11 October 2002.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

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J APPL PHYSIOL 94(2):677-684
8750-7587/03 $5.00 Copyright © 2003 the American Physiological Society



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