Detection of pregnancy in a hibernator based on activity data

Pregnancy and parturition are often difficult to detect in wild-living animals, especially in species that give birth to altricial young in burrows or caves. Current methods to detect pregnancy and birth in hibernating animals often entail disturbances that can affect the animals' reproductive success. We developed a new method to confirm pregnancy and parturition in hibernating brown bears (Ursus arctos) using activity data recorded in dual-axis motion sensors mounted on GPS–GSM neck collars on 30 brown bears during 52 hibernation seasons in Sweden. We adjusted recorded activity to the individual basal activity level for each bear. Pregnant females showed characteristic activity patterns during the period of pregnancy, with significantly higher daily activity levels and frequency of active periods during pregnancy than nonpregnant bears. Pregnant females were active on average 2.20 times more often during the pregnancy period than during the postpartum period, compared with 0.97 times for nonpregnant females. A pregnancy index was defined as the average of the proportion of mean daily activity levels and the proportion of activity events during the pregnancy period compared with the postpartum period. It averaged 2.61?±?1.73 (SD) for pregnant females and 0.94?±?0.24 for nonpregnant females. Using this method, we evaluated a group of adult females that had an unverified reproductive status but were classified as not pregnant because no cubs had been observed after den emergence. The results suggested that 35 % of those females had, in fact, been pregnant.     

The seasonal sensitivity of brown bear denning phenology in response to climatic variability

Background

For brown bears (Ursus arctos), hibernation is a critical part of the annual life cycle because energy savings during hibernation can be crucial for overwintering, and females give birth to cubs at that time. For hibernation to be a useful strategy, timing is critical. However, environmental conditions vary greatly, which might have a negative effect on the functionality of the evolved biological time-keeping. Here, we used a long-term dataset (69 years) on brown bear denning phenology recorded in 12 Russian protected areas and quantified the phenological responses to variation in temperature and snow depth. Previous studies analyzing the relationship between climate and denning behavior did not consider that the brown bear response to variation in climatic factors might vary through a period preceding den entry and exit. We hypothesized that there is a seasonal sensitivity pattern of bear denning phenology in response to variation in climatic conditions, such that the effect of climatic variability will be pronounced only when it occurs close to den exit and entry dates.

Results

We found that brown bears are most sensitive to climatic variations around the observed first den exit and last entry dates, such that an increase/decrease in temperature in the periods closer to the first den exit and last entry dates have a greater influence on the denning dates than in other periods.

Conclusions

Our study shows that climatic factors are modulating brown bear hibernation phenology and provide a further structuring of this modulation. The sensitivity of brown bears to changes in climatic factors during hibernation might affect their ability to cope with global climate change. Therefore, understanding these processes will be essential for informed management of biodiversity in a changing world.

     

Denning in brown bears

1. Hibernation represents an adaptation for coping with unfavorable environmental conditions. For brown bears Ursus arctos, hibernation is a critical period as pronounced temporal reductions in several physiological functions occur.
2. Here, we review the three main aspects of brown bear denning: (1) den chronology, (2) den characteristics, and (3) hibernation physiology in order to identify (a) proximate and ultimate factors of hibernation as well as (b) research gaps and conservation priorities.
3. Den chronology, which varies by sex and reproductive status, depends on environmental factors, such as snow, temperature, food availability, and den altitude. Significant variation in hibernation across latitudes occurs for both den entry and exit.
4. The choice of a den and its surroundings may affect individual fitness, for example, loss of offspring and excessive energy consumption. Den selection is the result of broad‐ and fine‐scale habitat selection, mainly linked to den insulation, remoteness, and availability of food in the surroundings of the den location.
5. Hibernation is a metabolic challenge for the brown bears, in which a series of physiological adaptations in tissues and organs enable survival under nutritional deprivation, maintain high levels of lipids, preserve muscle, and bone and prevent cardiovascular pathologies such as atherosclerosis.

It is important to understand: (a) proximate and ultimate factors in denning behavior and the difference between actual drivers of hibernation (i.e., factors to which bears directly respond) and their correlates; (b) how changes in climatic factors might affect the ability of bears to face global climate change and the human‐mediated changes in food availability; (c) hyperphagia (period in which brown bears accumulate fat reserves), predenning and denning periods, including for those populations in which bears do not hibernate every year; and (d) how to approach the study of bear denning merging insights from different perspectives, that is, physiology, ecology, and behavior.     

Changes in body condition of hibernating bats support the thrifty female hypothesis and predict consequences for populations with white-nose syndrome

White-nose syndrome (WNS) is a new disease of bats that has devastated populations in eastern North America. Infection with the fungus, Geomyces destructans, is thought to increase the time bats spend out of torpor during hibernation, leading to starvation. Little is known about hibernation in healthy, free-ranging bats and more data are needed to help predict consequences of WNS. Trade-offs presumably exist between the energetic benefits and physiological/ecological costs of torpor, leading to the prediction that the relative importance of spring energy reserves should affect an individual''s use of torpor and depletion of energy reserves during winter. Myotis lucifugus mate during fall and winter but females do not become pregnant until after spring emergence. Thus, female reproductive success depends on spring fat reserves while male reproductive success does not. Consequently, females should be “thrifty” in their use of fat compared to males. We measured body condition index (BCI; mass/forearm length) of 432 M. lucifugus in Manitoba, Canada during the winter of 2009/2010. Bats were captured during the fall mating period (n = 200), early hibernation (n = 125), and late hibernation (n = 128). Adult females entered hibernation with greater fat reserves and consumed those reserves more slowly than adult males and young of the year. Consequently, adult females may be more likely than males or young of the year to survive the disruption of energy balance associated with WNS, although surviving females may not have sufficient reserves to support reproduction.     

Weights and hematology of wild black bears during hibernation.

We compared weights and hematological profiles of adult (greater than 3-yr-old) female black bears (Ursus americanus) during hibernation (after 8 January). We handled 28 bears one to four times (total of 47) over 4 yr of varying mast and berry production. Mean weight of lactating bears was greater (P less than 0.0001) than that of non-lactating females. White blood cells (P less than 0.05) and mean corpuscular volume (P = 0.005) also differed between lactating and non-lactating bears. Hemoglobin (P = 0.006) and mean corpuscular hemoglobin concentration (P = 0.02) varied among years; values were lowest during 1975, following decreased precipitation and the occurrence of a second year of mast and berry crop shortages in a three-year period. Significant (P less than 0.05) interaction between reproductive status (lactating versus non-lactating) and study year for hemoglobin, red blood cells, and packed cell volume, and increased mean corpuscular volume, suggested a greater nutritional challenge for lactating females compared to non-lactating females during the 1975 denning season. Our data suggest that hematological characteristics of denning bears may be more sensitive than weights as indicators of annual changes in nutritional status; however, other influential factors, in addition to mast and berry crop production, remain to be examined.     

Black bear movements and habitat use during a critical period for moose calves

In sub-Arctic and north-temperate ecosystems, opportunistic carnivores, such as black bears (Ursus americanus) and brown bears (Ursus arctos), are active on the landscape for a shorter period annually than sympatric gray wolves (Canis lupus). Therefore, bear movement patterns and habitat use might be expected to be more deliberate and of greater consequence, in terms of energy acquisition, than those of predators not undergoing hibernation. Habitat choices concerning feeding, bedding, and denning grounds made by black bears therefore should reflect seasonal abundance and distribution of vegetation and key prey items as these are sites where bears remain and forage for prolonged periods of time. We recorded the movement patterns of 6 GPS-collared black bears from den emergence to onset of moose (Alces alces) parturition in 2003. Over approximately 3 weeks prior to parturition, results from average distance calculations suggest that black bears moved closer to probable moose calving-site habitat. Additionally, the seasonal habitat use by black bears surrounding dens reflected the same trend for areas where cow moose gave birth in spring 2003, with a propensity to use needleleaf forest more than any other habitat.     

Selection of maternity dens by female polar bears in western Hudson Bay,Canada and the effects of human disturbance

We deployed 19 satellite radio collars on adult female polar bears (Ursus maritimus) in western Hudson Bay to study how pregnant females select maternity-den sites and the degree to which human disturbance might influence this process. To eliminate the possibility that being handled while pregnant might influence the final choice of a suitable site, the collars were attached 1 year before the females were expected to select dens. Our results indicated that pregnant females moved inland and selected an area to den shortly after coming ashore in the summer and remained there if not disturbed. We analyzed 24 years of consecutive data (1979–2002) to examine whether disturbance of pregnant females in the denning area in the autumn affected litter size or mass of cubs the following spring. We defined three levels of disturbance (none—bear not seen; low—bear seen but not handled; high—bear seen and handled), and found that disturbance had no effect on either litter size or the mass of male cubs. However, we did find that females handled in the autumn had significantly lighter female cubs than females that were not seen. Our disturbance of pregnant females occurred in August and September, which is only a few months prior to parturition; it is not known what the effects of disturbance might be in late October or early November, closer to the time when cubs are born.     

Hibernating black bears (Ursus americanus) experience skeletal muscle protein balance during winter anorexia

Black bears spend four to seven months every winter confined to their den and anorexic. Despite potential for skeletal muscle atrophy and protein loss, bears appear to retain muscle integrity throughout winter dormancy. Other authors have suggested that bears are capable of net protein anabolism during this time. The present study was performed to test this hypothesis by directly measuring skeletal muscle protein metabolism during the summer, as well as early and late hibernation periods. Muscle biopsies were taken from the vastus lateralis of six free-ranging bears in the summer, and from six others early in hibernation and again in late winter. Protein synthesis and breakdown were measured on biopsies using (14)C-phenylalanine as a tracer. Muscle protein, nitrogen, and nucleic acid content, as well as nitrogen stable isotope enrichment, were also measured. Protein synthesis was greater than breakdown in summer bears, suggesting that they accumulate muscle protein during periods of seasonal food availability. Protein synthesis and breakdown were both lower in winter compared to summer but were equal during both early and late denning, indicating that bears are in protein balance during hibernation. Protein and nitrogen content, nucleic acid, and stable isotope enrichment measurements of the biopsies support this conclusion.     

Influence of pregnancy on locomotor and feeding performances of the skink, Mabuya multifasciata: why do females shift thermal preferences when pregnant?

It has been documented in the many-lined sun skink (Mabuya multifasciata) that pregnant females select lower body temperatures (27.6–30.8 °C) than do nonpregnant females and adult males (29.7–35.7 °C). We therefore used the skink to test the hypothesis that the maximization of reproductive benefits should be achieved in pregnant females by shifting thermal preferences towards the levels optimal for embryonic development but entailing relatively small costs of reproduction. Data on adult males showed that temperatures maximizing swimming stamina (indicative of locomotor endurance) fell within the range of body temperatures selected by nonpregnant females and adult males. Data on swimming stamina and feeding performance of pregnant females, nonpregnant females and adult males measured at 26 and 30 °C showed that: (1) pregnancy impaired locomotor and feeding performances, but such impairments did not persist after parturition; (2) the degree of locomotor impairment during pregnancy was greater at 26 °C than at 30 °C, but the degree of feeding impairment during pregnancy was greater at 30 °C than at 26 °C. Pregnant females of M. multifasciata selecting body temperatures at about 29 °C could not only produce good-quality offspring in a relatively short gestation length but also reduce reproductive costs associated with locomotor and feeding impairments to some extent. Thus, data from M. multifasciata validate the above hypothesis.     

Room without a view—Den excavation in relation to body size in brown bears

Hibernation is an adaptive strategy to survive harsh winter conditions and food shortage. The use of well‐insulated winter dens helps animals minimize energy loss during hibernation. Brown bears (Ursus arctos) commonly use excavated dens for hibernation. Physical attributes of excavated dens are expected to impact the bear's heat retention and energy conservation. The objective of this study was to examine the determinants of cavity size of excavated dens and the impact of physical attributes of excavated dens on energy conservation in hibernating bears, hypothesizing that bears excavate dens in a way to minimize heat loss and optimize energy conservation during hibernation. We predicted that den cavity size would be determined by the bear's body size and that older bears would excavate better‐fitting cavities to minimize heat loss, due to their previous experience. We further predicted that physical attributes of excavated dens would affect the bears’ posthibernation body condition. Our results revealed that bears excavated a den cavity in relation to their body size, regardless of sex, and that older bears tended to excavate better‐fitting den cavities compared to young bears, as we expected. Older bears excavated better‐fitting den cavities, suggesting a potentially experience‐based shift with age in den‐excavation behavior and an optimum cavity size relative to a bear's body size. Our key finding is that insulation of excavated dens provided by wall/rood thickness and bedding materials had a significant positive effect on bears’ posthibernation body condition. We believe that our study provides new insight into how not only the quality of denning habitat, but also the quality of dens may affect hibernating animals, by presenting a potential adaptive aspect of den preparation (age effect on efficiency in den excavation) and effect of den attributes on the posthibernation body condition of brown bears.     

Gestation, thermoregulation, and metabolism in a viviparous snake, Vipera aspis: evidence for fecundity-independent costs

Oxygen consumption of gestating Aspic vipers, Vipera aspis (L.), was strongly dependent on body temperature and mass. Temperature-controlled, mass-independent oxygen consumption did not differ between pregnant and nonpregnant females. Maternal metabolism was not influenced during early gestation by the number of embryos carried but was weakly influenced during late gestation. These results differ from previous investigations that show an increase in mass-independent oxygen consumption in reproductive females relative to nonreproductive females and a positive relationship between metabolism and litter size. These data also conflict with published field data on V. aspis that show a strong metabolic cost associated with reproduction. We propose that, under controlled conditions (i.e., females exposed to precise ambient temperatures), following the mobilisation of resources to create follicles (i.e., vitellogenesis), early gestation per se may not be an energetically expensive period in reproduction. However, under natural conditions, the metabolic rate of reproductive females is strongly increased by a shift in thermal ecology (higher body temperature and longer basking periods), enabling pregnant females to accelerate the process of gestation. Combining both laboratory and field investigation in a viviparous snake, we suggest that reproduction entails discrete changes in the thermal ecology of females to provide optimal temperatures to the embryos, whatever their number. This results in the counterintuitive notion that metabolism may well be largely independent of fecundity during gestation, at least in an ectothermic reptile.     

The timing of hibernation in Tasmanian echidnas: why do they do it when they do?

We investigated the patterns of hibernation and arousals in seven free-ranging echidnas Tachyglossus aculeatus setosus (two male, five female) in Tasmania using implanted temperature data loggers. All echidnas showed a ‘classical’ pattern of mammalian hibernation, with bouts of deep torpor interrupted by periodic arousals to euthermia (mean duration 1.04±0.05 (n=146). Torpor bout length increased as body temperature fell during the hibernation season, and became more variable as temperature rose again. Hibernation started in late summer (February 28±5 days, n=6) and males aroused just before the winter solstice (June 15±3 days, n=3), females that subsequently produced young aroused 40 days later (July 25±3, n=4) while females that did not produce young hibernated for a further two months (arousal Sept 27±5, n=7). We suggest that hibernation in Tasmanian echidnas can be divided into two phases, the first phase, marked by declining minimum body temperatures as ambient temperature falls, appears to be obligatory for all animals, while the second phase is ‘optional’ and is utilised to varying amounts by females. We suggest that early arousal and breeding is the favoured option for females in good condition, and that the ability to completely omit breeding in some years, and hibernate through to spring is an adaptation to an uncertain climate.     

Reproductive performance, population dynamics and anthropometrics of the free-ranging Cayo Santiago rhesus macaques

This report summarizes demographic data collected on the Cayo Santiago colony of rhesus monkeys from 1976-1983 and compares the results with those from 1959-1964 [8,9]. For males and nonpregnant/nonlactating, pregnant, and lactating females mean (+/- 1 SD), body weights, crown-rump lengths, and ponderal indices are tabulated for each age on a large (n = 586) single sampling of this free-ranging population of macaques.     

Ecological drivers of female lion (Panthera leo) reproduction in the Kruger National Park

The role of social cues in the reproduction of social mammals, particularly carnivores, has been thoroughly studied and documented in literature. However, environmental cues such as resources of water, food, and shelter have been identified to a lesser extent. Pregnant lions (Panthera leo) are notoriously secretive during the final stages of pregnancy and postpartum. Behavioral indicators depicted by movement patterns obtained by remote detection of collared female lions in the Kruger National Park were necessary for the monitoring of birth timing. Over the study period, eight plus a potential three parturition incidences of collared females were recorded. Of the variables measured (step length, range size, duration, prey biomass, and rainfall), range size during the month of parturition was the most indicative movement pattern of a successful birth. By backdating the potential birth month of the litters, date of conception was calculated and our results revealed a correlation between the birthing peaks of preferred prey during the month of conception. Birth timing in conjunction with remote sensing and ecological factors were thus identified behaviors associated with denning.     

Factor affecting body mass of young fat dormics (Glis glis) at weaning and by hibernation

This paper describes the factors affecting body mass at weaning and by hibernation of young fat dormice (Glis glis) We studied a free-living population in 1991 and 1992 in a mixed forest where 100 nest-boxes were placed. In the two years, 128 females used the nest-boxes to give birth and rear their litters and more than 900 dormice have been individually marked. Body mass of young at weaning was positively correlated to body mass of the mother in both years and to birth date in 1991, while litter size was negatively correlated to body mass of the young in 1992 only, when birth took place about two weeks later than in 1991. On the contrary, by the time of hibernation, young born later weighed significantly less than those born earlier in the season, and heavier young at weaning remained heavier by hibernation, while maternal body mass and litter size did not significantly affect body mass of young at this stage. Body mass of the mother was negatively related to the date of parturition and early breeding females lost weight during lactation, while females that bred later did not. It seems, therefore, that early breeding carries a cost that only females in good condition can afford. It appears that females may follow two strategies: those in poor body condition will delay pregnancy so that lactation occurs when environmental feeding conditions are at their best; those in good condition will breed earlier, since they can afford to lose weight during lactation. Their young will have more time to grow before hibernation and the mothers themselves will have a longer period of time for restoring their fat reserves.     

Hydrogen sulfide and nitric oxide metabolites in the blood of free-ranging brown bears and their potential roles in hibernation

During winter hibernation, brown bears (Ursus arctos) lie in dens for half a year without eating while their basal metabolism is largely suppressed. To understand the underlying mechanisms of metabolic depression in hibernation, we measured type and content of blood metabolites of two ubiquitous inhibitors of mitochondrial respiration, hydrogen sulfide (H₂S) and nitric oxide (NO), in winter-hibernating and summer-active free-ranging Scandinavian brown bears. We found that levels of sulfide metabolites were overall similar in summer-active and hibernating bears but their composition in the plasma differed significantly, with a decrease in bound sulfane sulfur in hibernation. High levels of unbound free sulfide correlated with high levels of cysteine (Cys) and with low levels of bound sulfane sulfur, indicating that during hibernation H₂S, in addition to being formed enzymatically from the substrate Cys, may also be regenerated from its oxidation products, including thiosulfate and polysulfides. In the absence of any dietary intake, this shift in the mode of H₂S synthesis would help preserve free Cys for synthesis of glutathione (GSH), a major antioxidant found at high levels in the red blood cells of hibernating bears. In contrast, circulating nitrite and erythrocytic S-nitrosation of glyceraldehyde-3-phosphate dehydrogenase, taken as markers of NO metabolism, did not change appreciably. Our findings reveal that remodeling of H₂S metabolism and enhanced intracellular GSH levels are hallmarks of the aerobic metabolic suppression of hibernating bears.     

Capture, anesthesia, and disturbance of free-ranging brown bears (Ursus arctos) during hibernation

We conducted thirteen immobilizations of previously collared hibernating two- to four-year-old brown bears (Ursus arctos) weighing 21-66 kg in central Sweden in winter 2010 and 2011 for comparative physiology research. Here we report, for the first time, an effective protocol for the capture and anesthesia of free-ranging brown bears during hibernation and an assessment of the disturbance the captures caused. Bears were darted in anthill, soil, or uprooted tree dens on eleven occasions, but two bears in rock dens fled and were darted outside the den. We used medetomidine at 0.02-0.06 mg/kg and zolazepam-tiletamine at 0.9-2.8 mg/kg for anesthesia. In addition, ketamine at 1.5 mg/kg was hand-injected intramuscularly in four bears and in six it was included in the dart at 1.1-3.0 mg/kg. Once anesthetized, bears were removed from the dens. In nine bears, arterial blood samples were analyzed immediately with a portable blood gas analyzer. We corrected hypoxemia in seven bears (PaO(2) 57-74 mmHg) with supplemental oxygen. We placed the bears back into the dens and antagonized the effect of medetomidine with atipamezole. Capturing bears in the den significantly increased the risk of den abandonment. One of twelve collared bears that were captured remained at the original den until spring, and eleven, left their dens (mean ± standard deviation) 3.2±3.6 (range 0.5-10.5) days after capture. They used 1.9±0.9 intermediate resting sites, during 6.2±7.8 days before entering a new permanent den. The eleven new permanent dens were located 730±589 m from the original dens. We documented that it was feasible and safe to capture hibernating brown bears, although they behaved differently than black bears. When doing so, researchers should use 25% of the doses used for helicopter darting during the active period and should consider increased energetic costs associated with den abandonment.     

Seasonal and diel movement patterns of brown bears in a population in southeastern Europe

Most animals concentrate their movement into certain hours of the day depending on drivers such as photoperiod, ambient temperature, inter‐ or intraspecific competition, and predation risk. The main activity periods of many mammal species, especially in human‐dominated landscapes, are commonly set at dusk, dawn, and during nighttime hours. Large carnivores, such as brown bears, often display great flexibility in diel movement patterns throughout their range, and even within populations, striking between individual differences in movement have been demonstrated. Here, we evaluated how seasonality and reproductive class affected diel movement patterns of brown bears of the Dinaric‐Pindos and Carpathian bear populations in Serbia. We analyzed the movement distances and general probability of movement of 13 brown bears (8 males and 5 females) equipped with GPS collars and monitored over 1–3 years. Our analyses revealed that movement distances and probability of bear movement differed between seasons (mating versus hyperphagia) and reproductive classes. Adult males, solitary females, and subadult males showed a crepuscular movement pattern. Compared with other reproductive classes, females with offspring were moving significantly less during crepuscular hours and during the night, particularly during the mating season, suggesting temporal niche partitioning among different reproductive classes. Adult males, solitary females, and in particular subadult males traveled greater hourly distances during the mating season in May‐June than the hyperphagia in July–October. Subadult males significantly decreased their movement from the mating season to hyperphagia, whereas females with offspring exhibited an opposite pattern with almost doubling their movement from the mating to hyperphagia season. Our results provide insights into how seasonality and reproductive class drive intrapopulation differences in movement distances and probability of movement in a recovering, to date little studied, brown bear population in southeastern Europe.     

Group Composition Affects Seasonal Birth Timing in Captive Japanese Macaques

We analyzed birth dates recorded during an 18-year period in a group of Japanese macaques housed in the Rome zoo to assess the influence of environmental, physiological, and social factors on birth seasonality. Birth timing differed significantly among years. Birth timing was affected by reproductive condition of females—ones that had given birth in the previous year delivered significantly later than those that had not—but not by their age or dominance rank. We conducted further analyses separately on females that had or had not given birth in the previous year. In both subgroups of females mean birth date was not influenced either by environmental temperature and rainfall during the previous mating season or by group size. On the contrary, among females that had not given birth in the previous year, socionomic sex ratio—ratio of sexually mature males to sexually mature females—is positively correlated with both mean birth date and date of the first birth, but not with date of the last birth. Contrarily, among females that had given birth in the previous year, there is no significant relationship between these variables. We hypothesize that the effects of socionomic sex ratio on birth timing might depend on competition among males for access to fertile females. When the number of males per female was higher, mutual disruption of consort pairs may have led to a delay in the onset of mating.     

Hormonal evidence for induced ovulation in Monodelphis domestica.

Breeding activity in the South American grey opossum, Monodelphis domestica, is stimulated by the presence of males. This study presents the first analysis of changes in concentrations of plasma progesterone during pregnant and nonpregnant cycles. In Expt 1, females were paired with either intact or vasectomized males to stimulate a reproductive cycle, or were isolated from males. Within 4-8 days of pairing, females showed marked changes in the size of the urogenital opening (vulva), which were paralleled by similar changes in body weight (11.4 +/- 2%; mean +/- SEM; n = 13). There was a second increase and decrease in body weight (21.7 +/- 3.3%; n = 13) in pregnant and nonpregnant cycles during the luteal phase of the cycle. Changes in concentrations of plasma progesterone, determined from samples collected three times a week, were similar in pregnant and nonpregnant cycles. Plasma progesterone concentration, which was increased for 15-18 days, showed two distinct peaks. The first peak, 3-4.5 ng ml-1, occurred immediately after the decrease in vulval swelling, and the second peak (1-2 ng ml-1) occurred 8-12 days later. Concentrations of plasma progesterone were low before parturition occurred. Females isolated from males for 35 days showed no changes in body weight or vulval swelling and concentrations of plasma progesterone remained undetectable. In Expt 2, concentrations of luteinizing hormone (LH) in plasma were determined twice a day for the first few days after pairing. In three of five females an LH peak was detected coincident with the maximal swelling of the vulva.