Hydrogen Isotope Dilution Research Articles

Colony‐specific differences in decadal longitudinal body composition of a capital‐breeding marine top predator

Abstract Capital breeding animals such as true seals (Phocidae) rely on accumulated body reserves to rear offspring. A mother's body composition at the start of a breeding episode may depend on recent environmental conditions and sets the resources available for the reproductive episode. At two grey seal (Halichoerus grypus) breeding colonies with contrasting demographic characteristics, factors influencing individual variation and temporal trends in the body composition (expressed as the lipid‐to‐protein mass ratio) of females were examined. Maternal reproductive expenditure, and the consequences for mothers and their pups, were investigated. Individual variation in postpartum maternal body composition was considerable. Mean values of 27% (±5%) lipid and 18% (±1%) protein were estimated by hydrogen isotope dilution. No evidence of age effects was detected. Mothers with a high lipid‐to‐protein mass ratio expended a higher proportion of lipid resources, conserved protein and returned with more protein the following year. Average maternal postpartum body composition was similar between the two colonies, but temporal patterns differed at one colony where pup production was decreasing from another where pup production was increasing. Mothers at the declining colony consistently weaned larger pups than mothers at the increasing colony across the range of mother sizes, but measures of maternal body composition did not predict pup weaning masses.

Organochlorine contaminant concentrations in blubber of young Steller sea lion (Eumetopias jubatus) are influenced by region, age, sex, and lipid stores

Contaminant exposure is particularly important for species and populations of conservation concern, such as the Steller sea lion (Eumetopias jubatus). We used blubber samples (n = 120) to determine organochlorine concentrations, including polychlorinated biphenyls (PCBs) and dichloro-diphenyl-trichloroethane (DDTs), and blood samples (n = 107) to estimate total body lipids based on the hydrogen isotope dilution method. We assessed the influence of age, sex, condition, and geographic area on contaminant concentrations in blubber and contaminant body load. The concentration of ΣPCBs was highest in pups (<6 months) from the Aleutian Islands, and the concentrations in males were higher than females in all regions. The ΣPCBs and ΣDDTs concentrations and loads decreased with increasing mass in pups, however, there were no regional or sex differences in contaminant load. Within each of the five age classes, the concentrations of ΣPCBs and ΣDDTs decreased with increasing mass, but overall these OCs increased with age. Further, accounting for the lipid content, a potential proxy for energy balance, in the animal load reduced or removed the regional and sex effects present in age models for contaminants. We propose, that adjusting OCs concentration by the lipid content of the blubber sample alone may not fully account for the variability in OC concentrations associated with differences in condition or energy states between young Steller sea lions.

Sexual Size Dimorphism and Body Condition in the Australasian Gannet.

Sexual size dimorphism is widespread throughout seabird taxa and several drivers leading to its evolution have been hypothesised. While the Australasian Gannet (Morus serrator) has previously been considered nominally monomorphic, recent studies have documented sexual segregation in diet and foraging areas, traits often associated with size dimorphism. The present study investigated the sex differences in body mass and structural size of this species at two colonies (Pope’s Eye, PE; Point Danger, PD) in northern Bass Strait, south-eastern Australia. Females were found to be 3.1% and 7.3% heavier (2.74 ± 0.03, n = 92; 2.67 ± 0.03 kg, n = 43) than males (2.66 ± 0.03, n = 92; 2.48 ± 0.03 kg, n = 43) at PE and PD, respectively. Females were also larger in wing ulna length (0.8% both colonies) but smaller in bill depth (PE: 2.2%; PD: 1.7%) than males. Despite this dimorphism, a discriminant function provided only mild accuracy in determining sex. A similar degree of dimorphism was also found within breeding pairs, however assortative mating was not apparent at either colony (R 2 < 0.04). Using hydrogen isotope dilution, a body condition index was developed from morphometrics to estimate total body fat (TBF) stores, where TBF(%) = 24.43+1.94*(body mass/wing ulna length) – 0.58*tarsus length (r 2 = 0.84, n = 15). This index was used to estimate body composition in all sampled individuals. There was no significant difference in TBF(%) between the sexes for any stage of breeding or in any year of the study at either colony suggesting that, despite a greater body mass, females were not in a better condition than males. While the driving mechanism for sexual dimorphism in this species is currently unknown, studies of other Sulids indicate segregation in foraging behaviour, habitat and diet may be a contributing factor.

MEASURING CHANGES IN JUVENILE GRAY SEAL BODY COMPOSITION

AbstractDetermining body composition in gray seals (Halichoerus grypus) is important when studying their physiology and life history. In this study we investigated the predictability of total body fat (TBF) and protein (TBP) in postweaned gray seal pups from morphometric measurements, blubber thickness using ultrasonographs and bioelectrical impedance analysis (BIA). In postweaned pups, TBF (kg) could be estimated from girth measurements and sex (n= 45, r2= 0.878) using hydrogen isotope dilution methods as the reference. However, TBP could not be reliably estimated from morphometric data. TBF (kg) in yearlings was best predicted from mass (n= 6, r2= 0.776) and TBP (kg) from mass/length (r2= 0.949). Dorsal blubber thickness using B‐mode ultrasound was also a significant predictor of TBF (kg) in postweaned pups (r2= 0.725) but BIA was not. Marked pups were recaptured during their first few months of life (n= 48) and body composition changes investigated. Animals lost mass and TBF after leaving the breeding beach, largely during the first 5–6 mo of life. Postweaned pups were ∼40% TBF and ∼13% TBP whereas yearlings were ∼12% TBF and ∼20% TBP. Pups that survived beyond 6 mo of age then regained mass as protein. Morphometric measurements are a useful field indication of body condition when isotope dilution is impractical.

Food consumption and body composition in mice selected for high wheel-running activity.

The effects of genetic selection for high wheel running (13th generation) and prolonged access (8 weeks) to running wheels on food consumption and body composition were studied in house mice (Mus domesticus). Mice from four replicate lines selected for high wheel-running activity ran over twice as many revolutions per day on activity wheels as did mice from four replicate control lines. At approximately 49 days of age, all mice were placed individually in cages with access to wheels and monitored for 6 days, after which wheels were prevented from rotating for the "sedentary" individuals. During the experiment, five feeding trials were conducted and body mass was measured weekly. After 8 weeks, body composition was measured by hydrogen isotope dilution. Across the five feeding trials, mice in the "active" group (wheels free to rotate) consumed 22.4% more food than mice in the "sedentary" group (wheels locked); mice from the selected lines consumed 8.4% more food than mice from the control lines (average of all trials; body mass-corrected values). In females, but not males, we found a significant interaction between selection and wheel access treatments: within the "active" group the difference in food consumption between selected and control animals was greater than in the "sedentary" group. At the end of the study, mice from the "active" and "sedentary" groups did not differ significantly in body mass; however, mice from the selected lines were approximately 6% smaller in body mass. Estimated lean body mass did not differ significantly either between selected and control lines or between wheel-access groups (P>0.3). Mice from selected lines had lower total body fat compared to mice from control lines (P=0.05; 24.5% reduction; LSMEANS) as did mice from the "active" compared to "sedentary" group (P= 0.03; 29.2% reduction; LSMEANS). Under these conditions, a sufficient explanation for the difference in body mass between the selected and control lines was the difference in fat content.

Evaluating body condition in small mammals

Body condition (energy reserves) can have important fitness consequences. Measuring condition of live animals is typically done by regressing body mass on measures of body size and using the residuals as an index of condition. The validity of this condition index was evaluated by determining whether it reflected measured fat content of five species of small mammals (yellow-pine chipmunks (Tamias amoenus Allen), bushy-tailed wood rats (Neotoma cinerea Ord), deer mice (Peromyscus maniculatus Ord), red-backed voles (Clethrionomys gapperi Vigors), and meadow voles (Microtus pennsylvanicus Ord)). We also determined whether body water could predict fat content, enabling the use of hydrogen-isotope dilution for estimating condition. For all five species, condition estimates weakly predicted fat content and more accurately predicted variation in lean dry mass and water content. The relationship between body water and fat content was inconsistent among the five species, discouraging against the general use of isotope dilution in these animals. Although ecologically important, these indices are best interpreted as explaining variation in all constituents of body composition.

Body composition in mink ( Mustela vison) kits during 21–42 days postpartum using estimates of hydrogen isotope dilution and direct carcass analysis

We compared carcass analysis and hydrogen isotope dilution methods to measure total body water (TBW) and body composition in a small altricial carnivore, the mink. Dilution space ( D) of mink at 21–42 days of age ( n=20), was determined after subcutaneous administration of tritiated water. The same animals were then used to determine TBW and body composition by carcass analysis and to derive predictive empirical relationships between TBW and total body fat, protein and energy. A separate validation set of 27 kits was used to test the accuracy of predicting body composition from TBW. D overestimated TBW by a consistent and predictable 4.1% ( R 2=0.999, P<0.001). Our estimates of fat, protein and energy content, using equations derived from TBW, were not significantly different than those obtained from direct carcass analysis ( P>0.980) in either the initial or validation set of mink. TBW was shown to decrease from 81 to 76% and total body protein to increase from 14 to 19% of LBM of the kits from 21 to 42 days of age. Although a rapidly changing hydration state was apparent in neonates, we conclude that when this is taken into account, accurate estimates of body composition can be obtained from hydrogen isotope dilution.

Variation in milk production and lactation performance in grey seals and consequences for pup growth and weaning characteristics.

Phocid seals are one of the few groups of mammals capable of sustaining the energetic demands of lactation entirely through body nutrient stores while fasting. Lactation performance of the female in turn influences the rate and pattern of pup growth. We examined variation in and patterns of milk composition and production, maternal energy output, and pup growth and energy deposition over the entire lactation period in 18 grey seal mother-pup pairs using hydrogen isotope (3H2O and D2O) dilution. Milk composition was independent of maternal mass and nutrient stores, indicating dependence on other physiological and genetic factors. Heavier females lactated longer (r2=0.653, P<0.001), had higher total milk outputs (r2=0.652, P<0.001), and produced larger pups at weaning (r2=0.417, P=0.005). While fatter females lactated for longer periods of time (r2=0.595, P<0.001), females with a larger lean body mass at parturition produced more milk (r2=0.579, P<0.001). Total milk energy output was the strongest predictor of pup weaning mass, which, along with the pup's efficiency of energy storage, accounted for 91% of the variation in weaning mass. Nevertheless, there was sufficient plasticity in milk composition and energy output that some smaller females produced relatively large pups. Few females appeared to deplete body nutrients to the point where it might limit the duration of lactation.

Fat transfer and energetics during lactation in the hooded seal: the roles of tissue lipoprotein lipase in milk fat secretion and pup blubber deposition.

Hooded seals (Cystophora cristata) lactate for 3.6 days during which females simultaneously fast and transfer large amounts of energy to their pups through fat-rich milk. Pups grow rapidly, principally due to blubber deposition. Lipoprotein lipase (LPL), the primary enzyme responsible for tissue uptake of triglyceride fatty acids, may strongly influence both maternal milk fat secretion and pup blubber deposition. We measured the energetic costs of lactation (using hydrogen isotope dilution, 3H2O), milk composition, prolactin, and LPL activity (post-heparin plasma LPL [PH LPL], blubber, mammary gland and milk; U) in six females. PH LPL and blubber LPL were measured in their pups. Females depleted 216.3 MJ.day-1 of body energy and fat accounted for 59% of maternal mass loss and 90% of postpartum body energy loss, but maternal body composition changed little. Maternal blubber LPL was negligible (0.0-0.2 U), while mammary LPL was elevated (1.8-2.5 U) and was paralleled by changes in prolactin. Estimated total mammary LPL activity was high (up to 20,000 U.animal-1) effectively favoring the mammary gland for lipid uptake. Levels of total blubber LPL in pups increased seven-fold over lactation. Pups with higher PH LPL at birth had greater relative growth rates (P = 0.025). Pups with greater blubber stores and total blubber LPL activity had elevated rates of fat deposition (P = 0.035).

Estimation of total body water in pinnipeds using hydrogen-isotope dilution.

gen-isotope dilution space in pinnipeds and to develop a single Hydrogen-isotope dilution methods have been used widely to predictive equation to estimate TBW in species for which data study the body composition, milk intake, solid-food intake, are lacking. Adopting such an approach will provide a consisand energy metabolism of pinnipeds (Costa 1987; Oftedal and tent basis for comparative studies not only within pinnipeds Iverson 1987) and other vertebrates (Nagy 1987). Indeed, much but in relation to other mammalian taxa. of the current understanding of water balance and of the reproIn the Otariidae (i.e., fur seals and sea lions), only one study ductive and foraging energetics of large mammals comes from has compared hydrogen-isotope dilution space with TBW destudies of this group of species. An important assumption in rived from carcass analysis of the same individuals. Arnould the use of isotope dilution for these purposes is that the tracer et al. (1996) examined the accuracy of both HTO and D2O as mixes completely, and only, with total body water (TBW) of a means of estimating TBW in four pups and five adult female an animal that has been given a known quantity of deuterium Antarctic fur seals (Arctocephalus gazella). HTO dilution space oxide (D2O) or tritiated water (HTO). It has been known for significantly overestimated TBW of pup and adult females, by some time that a small fraction of hydrogen isotope is lost to an average of 1.9% { 1.00% (n A 9). D2O dilution space also rapidly exchangeable hydrogen atoms in organic constituents overestimated TBW of adult females by an average of 1.7% of the body (see, e.g., Ussing 1935). Hevesy and Jacobsen { 1.74% (n A 5), but this difference was not significantly (1940) estimated that 0.5%–2% of body mass is accounted for different than zero. Using these data, Arnould et al. (1996) by rapidly exchanging hydrogen atoms of isotopes in organic derived predictive regression equations to estimate TBW from compounds. For this reason, hydrogen-isotope dilution space either HTO or D2O dilution space. However, in the case of will usually overestimate TBW, and it may be necessary to apply HTO, the slope of the regression did not differ significantly an appropriate correction factor to dilution space. Although from 1.0, and the intercept did not differ from zero. It is relationships between TBW and hydrogen-isotope dilution therefore not clear that the use of this relationship would imspaces have been studied in other mammalian groups (see, prove the estimation of TBW compared to the use of estimates e.g., Nagy and Costa 1980), these relationships in pinnipeds of HTO dilution space. However, given the small sample size, are not well understood. failure to reject the null hypothesis that the slope equals 1.0 The only true validation of isotope-dilution space estimates may have simply reflected the low statistical power of the test. of TBW depends on measurement by isotope dilution followed In contrast to the study in otariids, hydrogen-isotope diluby carcass desiccation of the same individual. This has been tion space has been compared with carcass desiccation in four done in a number of taxa, and the generally close corresponspecies of the Phocidae (i.e., hair seals). Reilly and Fedak (1990) dence between estimated dilution space and TBW measured by administered both D2O and HTO to four grey seals (Halidesiccation indicates that isotope dilution is a valid technique choerus grypus; two pups and two adults) that were subse(Pinson 1952; Nagy and Costa 1980). However, recent practice quently desiccated. Isotope dilution overestimated TBW in all

Measuring the Body Composition of Antarctic Fur Seals (Arctocephalus gazella): Validation of Hydrogen Isotope Dilution

We examined the accuracy of hydrogen isotope dilution (tritium and deuterium) as a means of determining total body water (TBW), and hence body composition, by comparison with whole-carcass chemical analysis in five adult female and four pup Antarctic fur seals. On average, tritium isotope (HTO) dilution overestimated TBW by 1.92% ± 1.00% (n = 9). Deuterium isotope (D₂O) dilution also overestimated TBW (1.71% ± 1.74%, n = 5), but this trend was not significant. The following equations were developed for predicting TBW from hydrogen isotope dilution estimates: TBW (kg) = 0.11 + 0.97 · HTO space (kg) (r² = 0.999, n = 9, P < 0.0001) and TBW (kg) = 2.10 + 0.89 · D₂O space (kg) (r² = 0.980, n = 5, P < 0.001). Whole-carcass chemical analysis showed adult females were composed of 14.5% ± 1.0% total body lipid (TBL), whereas pups were composed of only 9.4% ± 1.0% lipid. Adult females had lower TBW contents (60.1% ± 0.7%) than pups (67.6% ± 1.2%) and, consequently, higher energy contents (adults, 10.59 ± 0.30 MJ · kg⁻¹; pups, 8.24 ± 0.41 MJ · kg⁻¹). Pups had higher ratios of TBW to lean body mass (LBM) (0.747 ± 0.009) than adults (0.707 ± 0.004), which indicates they had not yet reached chemical maturity. The following best-fit predictive equations based on TBW and body mass (BM) were developed for TBL, total body protein (TBP), total body ash (TBA), and total body gross energy (TBGE): TBL (kg) = 0.875 · BM (kg) - 1.187 · TBW (kg) - 0.633 (adults); TBL (%) = 66562 - 0.845 · TBW (%) (pups); TBP (kg) = 0.058 · BM (kg) + 0.209 · TBW (kg) + 0.996 (adults); TBP (kg) = 0.267 · BM (kg) - 0.130 · STBW (kg) + 0.087 (pups); TBA (kg) = 0.071 · TBW (kg) - 0.088 (adults and pups); TBGE (MJ · kg⁻¹) = 34.1 - 0.391 · TBW (%) (adults); TBGE (MJ · kg⁻¹) = 31. 7 - 0.347 · TBW (%) (pups). With the use of these predictive equations, hydrogen isotope dilution is an accurate means of determining body composition in live Antarctic fur seals. Differences in these relationships between adults and pups, and in comparison to those in phocids, emphasize the need for the use of equations derived from appropriate data sets when predicting body composition in pinnipeds.

INDICES OF BODY CONDITION AND BODY COMPOSITION IN FEMALE ANTARCTIC FUR SEALS (ARCTOCEPHALUS GAZELLA)

AbstractAn attempt was made to develop simple, inexpensive, rapid means of determining body composition in Antarctic fur seals (Arctocephalus gazella). Measurements of total body water (TBW) and total body lipid (TBL), obtained by hydrogen isotope dilution, were compared to the results of bioelectrical impedance analysis (BIA) and morphometric indices of body condition in 52 adult females. TBW was weakly correlated with BIA measurements of resistance (v= ‐0.30, P &lt; 0.03). Conductor volume (length2/resistance) was more highly correlated with TBW(r= 0.75, P &lt; 0.0001) and the inclusion of mass into the predictive equation improved the correlation further (r= 0.95, P &lt; 0.0001). A body condition index (mass/length) previously used in pinniped studies was positively correlated to TBL (r= 0.77, P &lt; 0.0001) validating its use as a relative index of condition. However, body mass alone was highly correlated to TBW (r= 0.94, P &lt; 0.0001) and appears to provide a simple, rapid means of estimating body composition in adult females. This technique may also be applicable to juvenile male Antarctic fur seals.

Milk production and milk consumption in polar bears during the ice-free period in western Hudson Bay

Milk yield and milk consumption were measured in polar bears (Ursus maritimus) during the summer ice-free period in western Hudson Bay, a period of severe nutritional restriction. The transfer of milk between adult and dependent offspring was measured for five females with cubs (aged 8 months) and four with yearlings (aged 20 months) by a hydrogen isotope dilution–transfer method. Females with cubs produced significantly more milk energy (10.9 MJ∙day−1) than females with yearlings (2.6 MJ∙day−1). Daily milk production represented a greater proportion of body mass for females with cubs than females with yearlings but milk production was not correlated with maternal mass or litter mass in either group. Milk energy consumption was correlated with body mass in cubs but not in yearlings. Cubs consumed significantly more milk energy (7.8 MJ∙day−1), and displayed a higher relative growth rate, than yearlings (1.5 MJ∙day−1). However, both age groups lost proportionally the same amount of mass during the study period, suggesting that cubs are less able to survive nutritional restrictions and are more dependent on milk for survival than yearlings.

Measurement of the body composition of living gray seals by hydrogen isotope dilution

The body composition of living gray seals (Halichoerus grypus) can be accurately predicted from a two-step model that involves measurement of total body water (TBW) by 2H or 3H dilution and application of predictive relationships between body components and TBW that were derived empirically by slaughter chemical analysis. TBW was overestimated by both 2HHO and 3HHO dilution; mean overestimates were 2.8 +/- 0.9% (SE) with 2H and 4.0 +/- 0.6% with 3H. The relationships for prediction of total body fat (TBF), protein (TBP), gross energy (TBGE), and ash (TBA) were as follows: %TBF = 105.1 - 1.47 (%TBW); %TBP = 0.42 (%TBW) - 4.75; TBGE (MJ) = 40.8 (mass in kg) - 48.5 (TBW in kg) - 0.4; and TBA (kg) = 0.1 - 0.008 (mass in kg) + 0.05 (TBW in kg). These relationships are applicable to gray seals of both sexes over a wide range of age and body conditions, and they predict the body composition of gray seals more accurately than the predictive equations derived from ringed seals (Pusa hispida) (Stirling et al., Can. J. Zool. 53: 1021-1027, 1975) and from the equation of Pace and Rathbun (J. Biol. Chem. 158: 685-691, 1945), which has been reported to be generally applicable to mammals.