Preparing Muscles for Diving: Age-Related Changes in Muscle Metabolic Profiles in Harp (Pagophilus groenlandicus) and Hooded (Cystophora cristata) Seals.

In adult marine mammals, muscles can sustain aerobic metabolism during dives in part because they contain large oxygen (O2) stores and metabolic rates are low. However, young pups have significantly lower tissue O2 stores and much higher mass-specific metabolic rates. To investigate how these differences may influence muscle function during dives, we measured the activities of enzymes involved in aerobic and anaerobic metabolic pathways (citrate synthase [CS], ß-hydroxyacyl-coenzyme A dehydrogenase [HOAD], lactate dehydrogenase [LDH]) and the LDH isoform profile in six muscles from 41 harp (Pagophilus groenlandicus) and 30 hooded (Cystophora cristata) seals ranging in age from fetal to adult. All neonatal muscles had significantly higher absolute but lower metabolically scaled CS and HOAD activities than adults (∼ 70% and ∼ 85% lower, respectively). Developmental increases in LDH activity lagged that of aerobic enzymes and were not accompanied by changes in isozyme profile, suggesting that changes in enzyme concentration rather than structure determine activity levels. Biochemical maturation proceeded faster in the major locomotory muscles. In combination, findings suggest that pup muscles are unable to support strenuous aerobic exercise or rely heavily on anaerobic metabolism during early diving activities and that pups' high mass-specific metabolic rates may play a key role in limiting the ability of their muscles to support underwater foraging.

Postnatal development of muscle biochemistry in nursing harbor seal (Phoca vitulina) pups: limitations to diving behavior?

Adult marine mammal muscles rely upon a suite of adaptations for sustained aerobic metabolism in the absence of freely available oxygen (O(2)). Although the importance of these adaptations for supporting aerobic diving patterns of adults is well understood, little is known about postnatal muscle development in young marine mammals. However, the typical pattern of vertebrate muscle development, and reduced tissue O(2) stores and diving ability of young marine mammals suggest that the physiological properties of harbor seal (Phoca vitulina) pup muscle will differ from those of adults. We examined myoglobin (Mb) concentration, and the activities of citrate synthase (CS), beta-hydroxyacyl coA dehydrogenase (HOAD), and lactate dehydrogenase (LDH) in muscle biopsies from harbor seal pups throughout the nursing period, and compared these biochemical parameters to those of adults. Pups had reduced O(2) carrying capacity ([Mb] 28-41% lower than adults) and reduced metabolically scaled catabolic enzyme activities (LDH/RMR 20-58% and CS/RMR 29-89% lower than adults), indicating that harbor seal pup muscles are biochemically immature at birth and weaning. This suggests that pup muscles do not have the ability to support either the aerobic or anaerobic performance of adult seals. This immaturity may contribute to the lower diving capacity and behavior in younger pups. In addition, the trends in myoglobin concentration and enzyme activity seen in this study appear to be developmental and/or exercise-driven responses that together work to produce the hypoxic endurance phenotype seen in adults, rather than allometric effects due to body size.

Development of myoglobin concentration and acid buffering capacity in harp (Pagophilus groenlandicus) and hooded (Cystophora cristata) seals from birth to maturity.

Pinnipeds rely on muscle oxygen stores to help support aerobic diving, therefore muscle maturation may influence the behavioral ecology of young pinnipeds. To investigate the pattern of muscle development, myoglobin concentration ([Mb]) and acid buffering ability (beta) was measured in ten muscles from 23 harp and 40 hooded seals of various ages. Adult [Mb] ranged from 28-97 to 35-104 mg g tissue(-1) in harp and hooded seals, respectively, with values increasing from the cervical, non-swimming muscles to the main swimming muscles of the lumbar region. Neonatal and weaned pup muscles exhibited lower (approximately 30% adult values) and less variable [Mb] across the body than adults. In contrast, adult beta showed little regional variation (60-90 slykes), while high pup values (approximately 75% adult values) indicate significant in utero development. These findings suggest that intra-uterine conditions are sufficiently hypoxic to stimulate prenatal beta development, but that [Mb] development requires additional postnatal signal such as exercise, and/or growth factors. However, because of limited development in both beta and [Mb] during the nursing period, pups are weaned with muscles with lower aerobic and anaerobic capacities than those of adults.

Size and distribution of oxygen stores in harp and hooded seals from birth to maturity.

Pinnipeds rely primarily on oxygen stores in blood and muscles to support aerobic diving; therefore rapid development of body oxygen stores (TBO(2)) is crucial for pups to transition from nursing to independent foraging. Here, we investigate TBO(2) development in 45 harp (Pagophilus groenlandicus) and 46 hooded (Cystophora cristata) seals ranging in age from neonates to adult females. We found that hooded seal adults have the largest TBO(2) stores yet reported (89.5 ml kg(-1)), while harp seal adults have values more similar to other phocids (71.6 ml kg(-1)). In adults, large TBO(2) stores resulted from large blood volume (harp169, hood 194 ml kg(-1)) and high muscle Mb content (harp 86.0, hood 94.8 mg g(-1)). In contrast, pups of both species had significantly lower mass-specific TBO(2 )stores than adults, and stores declined rather than increased during the nursing period. This decline was due to a reduction in mass-specific blood volume and the absence of an increase in the low Mb levels (harp 21.0, hood 31.5 mg g(-1)). Comparisons with other phocid species suggests that the pattern of blood and muscle development in the pre- and post-natal periods varies with terrestrial period, and that muscle maturation rates may influence the length of the postweaning fast. However, final maturation of TBO(2) stores does not take place until after foraging begins.

Panmictic population structure in the hooded seal (Cystophora cristata).

Two putative populations of hooded seals (Cystophora cristata) occur in the North Atlantic. The Greenland Sea population pup and breed on the pack ice near Jan Mayen ('West Ice') while the Northwest Atlantic population is thought to pup in the Davis Strait, in the Gulf of St. Lawrence (the 'Gulf'), and off southern Labrador or northeast Newfoundland (the 'Front'). We used microsatellite profiling of 300 individuals at 13 loci and mitochondrial DNA sequencing of the control region of 123 individuals to test for genetic differentiation between these four breeding herds. We found no significant genetic differences between breeding areas, nor evidence for cryptic nor higher level genetic structure in this species. The Greenland Sea breeding herd was genetically most distant from the Northwest Atlantic breeding areas; however, the differences were statistically nonsignificant. Our data therefore suggest that the world's hooded seals comprise a single panmictic genetic population.

Partitioning of persistent organic pollutants in grey seal (Halichoerus grypus) mother-pup pairs.

Phocid seals have lipid rich milk, which is known to serve as a transfer medium through which persistent organic pollutants (POPs) move from mother to offspring during lactation. However, knowledge on this generational transfer of different POPs and the partitioning of these compounds in maternal and offspring tissues over the course of the lactation are limited. In this study we examined the qualitative and quantitative partitioning of a range of chlorinated POPs in maternal blubber, blood and milk as well as in pup blubber, collected early in the lactation period and late in the lactation period. In the lactating female, the high-chlorinated and hydrophobic compounds were passed less efficiently into the milk than the low-chlorinated compounds and more water-soluble compounds. Significantly, lower maternal blood concentrations than in maternal blubber biopsies suggest a stratification of POP concentrations in the blubber column of lactating female and lower concentrations in the metabolic active inner layers. Over the course of lactation, there was a significant increase in maternal blood and milk concentrations of POPs as opposed to no change in maternal blubber biopsy concentrations. This was most apparent for the hydrophobic and high-chlorinated compounds. The most likely explanation for this is that the metabolic active inner blubber layer, from which the milk lipids are derived from, is in steady state with the circulatory system, while the outer layers are more static and only slowly respond to changes in concentrations elsewhere in the body. The concentrations of the high-chlorinated and hydrophobic compounds were substantially lower in pup blubber than in maternal blubber. This probably relates the combined effect of these compounds stratification in maternal blubber and their slow transfer into the milk. The present study shows that the more hydrophobic and high-chlorinated compounds come to steady state less quickly in the different tissues than the more water-soluble and low-chlorinated compounds in the lactating female and her offspring. This has implications for which matrices to choose when sampling for assessing the toxicological risk of POPs in seals.

Tris (4-chlorophenyl) methane and tris (4-chlorophenyl) methanol in marine mammals from the Estuary and Gulf of St. Lawrence.

Levels of tris (4-chlorophenyl) methanol (TCPM) and its presumed precursor tris (4-chlorophenyl) methane (TCPMe) are reported in marine mammals from the Estuary and Gulf of St. Lawrence, Canada. These compounds were measured in blubber samples of seals and whales using ion trap mass spectrometry (MS/MS) detection. Detectable concentrations of both TCPM and TCPMe were observed in all of the samples analysed. Concentrations of these compounds varied with species ranging from 1.7 to 153 and from 1.3 to 50.6 ng/g lipid wt. for TCPM and TCPMe, respectively. TCPM was from 1.3 to 10 times more concentrated than TCPMe. The highest levels of both TCPM and TCPMe were observed in adult male beluga whales (Delphinapterus leucas) from the St. Lawrence Estuary, while adult female beluga whales from the same area showed levels similar to those in the seals examined. Among the four seal species investigated, TCPM and TCPMe levels were the highest in grey (Halichoerus grypus) and hooded (Cystophora cristata) seals, and lowest in harp seals (Phoca groenlandica). Intermediate levels were found in harbour seals (Phoca vitulina); however, their concentrations might be underestimated considering the younger mean age of these animals. Ratios of both 4,4'-DDE/sigma DDT and TCPM/sigma TCP were very similar between animals from the same species. Strong correlations between sigma TCP and sigma DDT were also observed for each species of mammals, most likely indicating that both sigma TCP and sigma DDT are bioaccumulated in marine mammals. The relationships between sigma DDT and sigma TCP also demonstrate that sigma TCP are less bioaccumulated than sigma DDT by the marine mammal species examined.

Transfer of fatty acids from female seal blubber via milk to pup bladder.

Fatty acids were determined by gas chromatography followed by principal component analysis (PCA) and soft independent modelling of class analogy (SIMCA) in the blubber of 18 female grey seals, Halichoerus grypus, in their milk and in the blubber of their 1-week-old nursing pups. Large individual differences were observed in both blubber and milk content of fatty acids. The fatty acid composition of the milk was systematically different from the composition in maternal blubber, with higher relative amounts of the saturated acids, the monounsaturated with 20 carbon atoms and the n3 polyunsaturated, except 18:3n3. The composition of the fatty acids in the blubber of the pups was different from that of the milk. The same fatty acids that were enriched in the milk were depleted in the blubber of the pups. Therefore the fatty acid composition in the blubber of the pups was similar to that in the adults, although not identical. The results from this investigation imply that the composition of the fatty acids in the blubber of female seals and in the blubber of their pups cannot be determined directly by analysis of fatty acid composition of milk.

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).

Levels and patterns of PCBs and OC pesticides in harbour and grey seals from the St Lawrence Estuary, Canada.

Blubber samples from harbour (Phoca vitulina) and grey seals (Halichoerus grypus) captured in the St Lawrence Estuary were analysed for PCBs and OC pesticides. Concentrations of sigma PCB, sigma DDT, sigma CHLOR and mirex were higher in harbour than in grey seals, while sigma HCH and HCB were similar in the two species. Age vs. concentration plots showed that sigma PCB, sigma DDT and sigma CHLOR concentrations increased with age in males, but plateaued at sexual maturity in females. Concentrations of sigma HCH decreased and mirex increased with age for post-weaning animals, regardless of gender. HCB did not show age-related trends. PCB congener and OC pesticide patterns varied within harbour seals (based on gender and maturity) and between harbour and grey seals. PCB and OC concentrations in harbour seals were lower than those seen in a sample of significantly older beluga whales. Both species remain in the Estuary year-round. PCB and OC pesticide concentrations were higher in resident harbour seals than in either grey or harp seals that visit the Estuary seasonally. PCB and OC pesticide concentrations in harbour seals from the Estuary were generally higher than in harbour seals from other regions of North America. They were comparable to, or higher than those from the NE Atlantic Ocean and the North Sea, and lower than those from the Wadden and Baltic Seas.

Energetics during nursing and early postweaning fasting in hooded seal (Cystophora cristata) pups from the Gulf of St Lawrence, Canada.

In this study we measured growth and milk intake and calculated energy intake and its allocation into metabolism and stored tissue for hooded seal (Cystophora cristata) pups. In addition, we measured mass loss, change in body composition and metabolic rate during the first days of the postweaning fast. The mean body mass of the hooded seal pups (n = 5) at the start of the experiments, when they were new-born, was 24.3 +/- 1.3 kg (SD). They gained an average of 5.9 +/- 1.1. kg.day-1 of which 19% was water, 76% fat and 5% protein. This corresponds to an average daily energy deposition of 179.8 +/- 16.0 MJ. The pups were weaned at an average body mass of 42.5 +/- 1.0 kg 3.1 days after the experiment was initiated. During the first days of the postweaning fast the pups lost an average of 1.3 +/- 0.5 kg of body mass daily, of which 56% was water, 16% fat and 28% protein. During the nursing period the average daily water influx for the pups was 124.6 +/- 25.8 ml.kg-1. The average CO2 .production during this period was 1.10 +/- 0.20 ml.g-1.h-1, which corresponds to a field metabolic rate of 714 +/- 130 kJ. kg-1. day-1, or 5.8 +/- 1.1 times the predicted basal metabolic rate according to Kleiber (1975). During the postweaning fast the average daily water influx was reduced to 16.1 +/- 6.6 ml.kg-1. The average CO2 production in this period was 0.58 +/- 0.17 ml.g-1.h-1 which corresponds to a field metabolic rate of 375 +/- 108 kJ.kg-1.day-1 or 3.2 +/- 0.9 times the predicted basal metabolic rate. Average values for milk composition were 33.5% water, 58.6% fat and 6.2% protein. The pups drank an average of 10.4 +/- 1.8 kg of milk daily, which represents an energy intake of 248.9 +/- 39.1 MJ.day-1. The pups were able to store 73.2 +/- 7.7% of this energy as body tissue.

Energy intake and utilisation by nursing bearded seal (Erignathus barbatus) pups from Svalbard, Norway.

In this study we measure energy intake via milk in nursing bearded seal (Erignathus barbatus) pups and determine how this energy is allocated into metabolism and storage of new tissues. This was accomplished using longitudinal mass gain records and the doubly labelled water technique on nursing pups in combination with cross-sectional data on changes in milk composition from bearded seal mothers. The pups (n = 3) were all less than a week old at the start of the experiments. Pups gained 3.3 +/- 0.4 kg.day-1 of which 50% was fat, 14% protein and 36% water. Average daily water influx for the pups was 69.5 +/- 9.0 ml.kg-1. day-1. Average CO2 production during the study period was 0.99 +/- 0.10 ml.g-1.h-1, which corresponds to a field metabolic rate of 642 +/- 67 kJ.kg-1. day-1, or 6.0 +/- 0.5 times the predicted basal metabolic rate according to Kleiber (1975). The pups drank an average of 7.6 +/- 0.5 kg of milk daily. This corresponds to a daily energy intake of 154 +/- 8 MJ, 47 +/- 14% of which was stored as new body tissue. Despite this high energy intake bearded seal pups do not get as fat as do other nursing phocids. This is in part due to their larger body size but also due to their very active aquatic lifestyle and the lower and more consistent fat content of the milk compared to other phocid species. Bearded seal mothers forage during lactation and may also be involved in teaching their pups to feed independently. All these data suggest that the lactation strategy of bearded seals differs from the phocid norm.

Milk intake, growth and energy consumption in pups of ice-breeding grey seals (Halichoerus grypus) from the Gulf of St. Lawrence, Canada.

In this study we document growth, milk intake and energy consumption in nursing pups of ice-breeding grey seals (Halichoerus grypus). Change in body composition of the pups, change in milk composition as lactation progresses, and mass transfer efficiency between nursing mothers and pups are also measured. Mass transfer efficiency between mother-pup pairs (n = 8) was 42.5 +/- 8.4%. Pups were gaining a daily average of 2.0 +/- 0.7 kg (n = 12), of which 75% was fat, 3% protein and 22% water. The total water influx was measured to be 43.23 +/- 8.07 ml.kg-1.day-1. Average CO2 production was 0.85 +/- 0.20 ml.g-1.h-1, which corresponds to a field metabolic rate of 0.55 +/- 0.13 MJ.kg-1.day-1, or 4.5 +/- 0.9 times the predicted basal metabolic rate based on body size (Kleiber 1975). Water and fat content in the milk changed dramatically as lactation progressed. At day 2 of nursing, fat and water content were 39.5 +/- 1.9% and 47.3 +/- 1.5%, respectively, while the corresponding figures for day 15 were 59.6 +/- 3.6% fat and 28.4 +/- 2.6% water. Protein content of the milk remained relatively stable during the lactation period with a value of 11.0 +/- 0.8% at day 2 and 10.4 +/- 0.3% at day 15. Pups drank an average of 3.5 +/- 0.9 kg of milk daily, corresponding to a milk intake of 1.75 kg per kg body mass gained. The average daily energy intake of pups was 82.58 +/- 19.80 MJ, while the energy built up daily in the tissue averaged 61.72 +/- 22.22 MJ.(ABSTRACT TRUNCATED AT 250 WORDS)

Activity, milk intake and energy consumption in free-living ringed seal (Phoca hispida) pups.

Measurements of growth, activity and energy consumption and estimates of milk intake were made in free-living, nursing ringed seal (Phoca hispida) pups. This was accomplished through the simultaneous use of time-depth recorders and the doubly labelled water technique. The pups spent an average of 52 +/- 7% of their time hauled out on the ice, 37 +/- 5% of the time in the water at the surface, and 11 +/- 5% of the time diving. Average daily mass gain of the pups (n=3) throughout the duration of the study period was 0.35 +/- 0.08 kg. The composition of the mass gain was 76% fat, 6% protein, and 18% water. The total water flux was measured to be 52 +/- 10 ml.kg-1.day-1. Average CO2 production was 0.85 +/- 0.16 ml.g-1.h-1, corresponding to a field metabolic rate of 0.55 +/- 0.10 MJ.kg-1.day-1, or 3.8 +/- 0.6 times the predicted basal metabolic rate based on body size (Kleiber 1975). Average daily milk intake was estimated to be 1379 +/- 390 ml. The field metabolic rate for the different components of seal pup activity budgets were calculated to be FMRhaul out = 1.34 BMR, FMRsurface = 6.44 BMR, and FMRdiving = 5.88 BMR.