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Pregnancy failure in female prairie deermice related to parity and social environment
Axim. Behav ., 1969,17, 1 04-108
PREGNANCY FAILURE IN FEMALE PRAIRIE DEERMICE RELATED TO PARITY AND SOCIAL ENVIRONMENT BY C .
RICHARD TERMAN
Laboratory of Endocrinology and Population Ecology Biology Department, College of William and Mary, Williamsburg, Virginia
Confined laboratory populations of prairie deermice even though supplied with excess food and water do not continue to increase indefinitely ; rather growth is controlled at a numerical level which is variable under constant conditions of the physical environment (Terman 1963, 1965) . Control of population growth is achieved in the above conditions either by complete cessation of reproduction or by failure of young to survive . The experiment herein reported is related to the first of these two mechanisms of controlthat of failure of reproductive function . In our previous population experiments, inhibition of reproduction appears to be due to stimuli influencing the females on at least two different levels . First, the two or three females producing litters in each population stopped reproducing even though copulation was observed in some cases . The second level of reproductive control involved an inhibition of reproductive maturation and function of animals born into the populations and as a consequence 94 per cent of the females at least ninety days of age did not produce young (Terman 1965) . Further, the reproductive organs of both males and females born into the populations were inhibited in development and were significantly lighter in weight than control animals of similar age kept as bisexual pairs (Terman 1968) . Some recent findings which may have significance to the inhibition of reproductive function in the populations mentioned above are those related to the phenomenon known as `pregnancy block' or the `Bruce effect' . These studies have shown that when recently inseminated female house mice, Mus musculus, are exposed to a strange male, to shavings soiled by strange males or the urine of a strange male within a period of 4 days post-insemination, the pregnancies of the females are interrupted due to failure of implantation and the females return to normal cycle within a few days . Females made anosmic by having their olfactory tracts severed do not have their pregnancies blocked by exposure to bedding soiled by strange males and consequently olfactory stimuli released by males
have been suggested as probable stimuli for pregnancy failure (Bruce 1960, 1961 ; Bruce & Parrott 1960 ; Parkes & Bruce 1962 ; Dominic 1966) . While experiments with deermice have not been as extensive as with Mus, pregnancy block has been demonstrated and appears to have many characteristics similar to the phenomenon in Mus (Bronson & Eleftheriou 1963 ; Bronson, Eleftheriou & Garick 1964 ; Eleftheriou, Bronson & Zarrow 1962) . The purpose of the present experiment was to examine the relationship between parity and social environment in the occurrence of pregnancy block in prairie deermice since both variables are continually part of the population context. Methods Animals The animals used in these experiments were prairie deermice (Peromyscus maniculatus bairdii) whose ancestors had been part of a laboratory colony for 13 years. Sib matings, however, were prevented and wild caught animals were added to the colony once per year when possible during the past 5 yr. Both nulliparous females and females which had produced young were used in this experiment . The nulliparous females approximated 70 days of age and the parous females 157 days of age . All females had been weaned at 21 days of age and maintained with sibs of the same sex until mating at an age older than 60 days . Procedure Vaginal smears were begun for the nulliparous females on the day following pairing . Young of parous females were removed on the day of birth and vaginal smears were begun on the same day. All smears were obtained by rinsing out the vagina with a small quantity of water twice per day, at 8-9 a .m . and 4-5 p .m . during five consecutive days or until insemination . A female was regarded as inseminated when a smear revealed at least twenty sperm per field of 100 magnifications . Each inseminated female was randomly 104
TERMAN : PREGNANCY FAILURE IN DEERMICE
assigned to one of four treatments . Beginning on the afternoon of the day following insemination, each female was kept in her home cage for a period of 48 hr, either alone, with her stud male, with a strange male, or with her stud male plus a strange male . Following the treatment period, all males were removed and the females remained in their home cages for seven additional days at which time they were sacrificed and examined for embryos . There were thirty parous and thirty nulliparous females exposed to each post-insemination treatment . All animals in this experiment were kept in a room 15 ft by 15 ft which also contained a breeding colony of deermice . The room was ventilated by a window fan and its only illumination was by two 40 W fluorescent tubes between 7 .30 a .m . and 7 .30 p .m . daily. Results Figure 1 presents the percentage of females pregnant in all treatments . A female was considered pregnant provided at least one embryo was viable .
1 05
Comparing parous versus nulliparous females In all four treatment situations the parous females exhibited a significantly higher percentage of pregnancy than the nulliparous females (0 .005>P) . Further, the difference between the parous and the nulliparous in all treatments was essentially the same . Comparing treatments for parous and nulliparous females Figure 1 also presents the results of statistical comparisons of the pregnancy proportions of both parous and nulliparous females . The probability of the calculated Chi square values for comparisons between treatments are given in the horizontal columns headed by `P' which indicates the treatment with which comparisons were made . For both nulliparous and parous females, there was an increasingly higher percentage of pregnancy in the following treatment order : female plus strange male, female plus stud and strange male, female alone, and female plus stud male. PAROUS ®
NONPAROUS
90
83 H z
a z 0
63
LL)
a
47
a
N
? a S a A
7
a A
Fig . 1 . Percentage of parous and nulliparous females pregnant at 10 days post-insemination . Each female was exposed to one of the following situations during 24-72 hr post insemination : female alone, female with stud male, female with strange male, female with stud and strange male simultaneously .
1 06
ANIMAL BEHAVIOUR, 17,
When the female alone situation was compared with the female plus stud male treatment for both nulliparous and parous females, higher but not significantly higher pregnancy percentages were recorded in the latter situation . When both treatments (females alone and females plus their stud males) were compared with the female plus strange male treatment, significantly smaller pregnancy proportions were recorded in the strange male situation in three of the four comparisons (0 . 05>P). Thus, pregnancy block occurred under these circumstances . When the female plus stud male treatment was compared with the situation in which both types of males were with the female, a higher proportion of pregnant females occurred in the former (0 . 02>P). Comparison of the female plus stud male and strange male situation with the the female plus strange male only situation revealed a higher proportion of pregnancies in the former, but this difference was also not significant at the P>0 . 05 level . These two comparisons therefore indicate that the presence of the stud male during the time the strange male was present increased the pregnancy success of the female, but the presence of the strange male decreased the pregnancy success exhibited when the female was with the stud male only.
1
Combined comparisons Figure 2 shows the pregnancy proportion when treatment groups were combined irrespective of parity . `P' indicates the treatment situation with which comparisons were made and the probabilities of the calculated Chi square values for the comparisons are recorded in the horizontal column to the right of each `P'. As noted previously, pregnancy success was significantly greater among females kept alone and those kept with their stud males when compared with the female plus strange male situation (0 . 025>P and 0 .0005 >P respectively) . Presence of the stud male while the strange male was present significantly increased the pregnancy success compared to the treatment when the female was alone with the strange male (0 .05>P) . Further the presence of the strange male decreased the pregnancy success of females maintained with the stud male only. This decrease was significant at 0 . 1 >P . Discussion The percentage of pregnancies was significantly smaller among nulliparous females than parous females in all treatments of the experimental design . This difference was relatively constant irrespective of the treatment . Confounded with the parity difference, however, was an age
80 fz
a z
68
0 W
62
crv a W °
co a
43
Fz W 0
P
NS
.I)
.0005)
a: P
W (L
NS
P
2
a S
2
a S a A
.025) .05)
2 a
A
Fig . 2 . Percentage of females pregnant at 10 days post-insemination when the data were combined irrespective of parity .
TERMAN : PREGNANCY FAILURE IN DEERMICE
difference, since the parous females were also older. Chipman & Fox (1966) have recently shown that six month-old house mice (M. musculus) females are more resistant to pregnancy blocking than two-month-old females . Age as well as parity may, therefore, be an important variable producing the differences observed in the present study . However, to my knowledge, the age variable has not been tested with deermice . One of the most interesting findings of the present experiment was the influence of the stud male on the stimulation of pregnancy block by the strange male . When the females were exposed to the strange male with the stud male present, the percentage of pregnancies was increased compared to the situation in which the stud male was not present during this exposure . However, the presence of the strange male in addition to the stud male decreased the percentage of pregnancies noted when stud males only were present with the females . Pregnancy success of the females when both males were present was almost exactly intermediate to the treatment situations in which the females were with one type of male or the other . These results vary somewhat from those mentioned without detailed desciption by Parkes & Bruce in their 1961 review ; namely that the presence of the stud male with the female during exposure to alien males largely eliminated pregnancy blocking . These differences, however, may reflect only differences in technique and the species of mice studied . The above findings raise the interesting question of whether the presence of the stud male prevented the strange male from releasing the pheromones which function to block the pregnancies, or whether the stud male influenced the response of the female to the pheromone . Both possibilities are relevant to the experimental study of population dynamics as well as to basic questions relating behaviour to reproductive physiology . Certainly, the behavioural definition of the stud male versus the strange male is difficult to establish within the population context . Clearly established by these data, however, is the fact that the occurrence of `pregnancy block', a pheromone induced phenomenon, may he influenced by social factors per se . Prior evidence suggesting such an influence but in a different context was provided by Bruce (1963), who exposed groups composed of differing numbers of female Mus to a single male and found that the females were protected against pregnancy blocking in direct proportion to the
107
number in the group at the time of exposure . The relationship between these two kinds of `social factors' to pregnancy block or to each other is not clear . Summary The occurrence of `pregnancy block' was noted in nulliparous and recently parous female prairie deermice when exposed to one of the following situations during a period from 24 to 72 hr subsequent to insemination : (1) stud male removed and female left alone, (2) stud male left with female, (3) stud male removed and strange male placed with female and (4) strange male placed with female and stud male . All females were killed at 10 days post-insemination and examined for embryos . There was a significantly greater frequency of pregnancy among parous females than nulliparous, although age differences may have been an important confounding variable . Further, there were decreasing pregnancy proportions in both parous and nulliparous females in the treatment sequences of (2) stud male left with female, (1) stud male removed and female left alone, (4) strange male placed with female and stud male and (3) stud male removed and strange male placed with female . Acknowledgments This study was partially supported by Public Health Service Grant NIH-MH-08289 from the National Institutes of Health . REFERENCES Bronson, F . H . & Eleftheriou, B . E . (1963) . Influence of strange males on implantation in the deermouse . Gen . comp . Endocr., 3, 515-518 . Bronson, F . H ., Eleftheriou, B . E . & Garick, E . I . (1964) . Effects of intra- and inter-specific social stimulation on implantation in deermice. J. Reprod. Fert ., 8, 23-27 . Bruce, H. M . (1960) . A block to pregnancy in the mouse caused by the proximity of strange males . J. Reprod. Fert., 1, 96-103 . Bruce, H . M . (1961) . An olfactory block to pregnancy in mice. Part 1 : Characteristics of the block . Proc . I Vth Int. Cong. An . Reprod., 159-162 . Bruce, H. M . (1963) . Olfactory block to pregnancy among grouped mice . J. Reprod. Fert ., 6, 451-460 . Bruce, H . M . & Parrott, D . M . V . (1960) . Role of Olfactory sense in pregnancy block by strange males . Science, N . Y., 131, 1526 . Chipman, R . K . & Fox, K . A . (1966) . Factors in pregnancy blocking, age and reproductive background of females : numbers of strange males . J . Reprod. Fert ., 12, 399-403 . Dominic, C . J . (1966) . Observations on the reproductive pheromones of mice . 1 . Source . J. Reprod . Fert ., 11, 407-414.
108
ANIMAL BEHAVIOUR, 17,
Eleftheriou, B . E ., Bronson, F. H . & Zarrow, M . X . (1962) . Interaction of olfactory and other environmental stimuli on implantation in the deermice. Science, N.Y., 137, 764 . Parkes, A . S . & Bruce, H. M . (1961) . Olfactory stimuli in mammalian reproduction . Science, N.Y., 134, 1049-1054. Parkes, A . S . & Bruce, H . M . (1962) . Pregnancy block in female mice placed in boxes soiled by males . J. Reprod. Fert., 4, 303-308 . Terman, C. R. (1963) . The influence of differential early social experience upon spatial distribution within
1
population of prairie deermice. Anim . Behav ., 11, 246-262 . Terman, C . R. (1965) . A study of population growth and control exhibited in the laboratory by prairie deermice . Ecology, 46, 890-895 . Terman, C . R . (1968) . Weights of the reproductive organs, adrenal glands and other selected organs of prairie deermice from laboratory populations with a long duration of asymptote . Unpublished ms .-submitted for publication . (Received 16 June 1967 ; revised 12 March 1968 ; Ms . number: A669)
PREGNANCY FAILURE IN FEMALE PRAIRIE DEERMICE RELATED TO PARITY AND SOCIAL ENVIRONMENT BY C .
RICHARD TERMAN
Laboratory of Endocrinology and Population Ecology Biology Department, College of William and Mary, Williamsburg, Virginia
Confined laboratory populations of prairie deermice even though supplied with excess food and water do not continue to increase indefinitely ; rather growth is controlled at a numerical level which is variable under constant conditions of the physical environment (Terman 1963, 1965) . Control of population growth is achieved in the above conditions either by complete cessation of reproduction or by failure of young to survive . The experiment herein reported is related to the first of these two mechanisms of controlthat of failure of reproductive function . In our previous population experiments, inhibition of reproduction appears to be due to stimuli influencing the females on at least two different levels . First, the two or three females producing litters in each population stopped reproducing even though copulation was observed in some cases . The second level of reproductive control involved an inhibition of reproductive maturation and function of animals born into the populations and as a consequence 94 per cent of the females at least ninety days of age did not produce young (Terman 1965) . Further, the reproductive organs of both males and females born into the populations were inhibited in development and were significantly lighter in weight than control animals of similar age kept as bisexual pairs (Terman 1968) . Some recent findings which may have significance to the inhibition of reproductive function in the populations mentioned above are those related to the phenomenon known as `pregnancy block' or the `Bruce effect' . These studies have shown that when recently inseminated female house mice, Mus musculus, are exposed to a strange male, to shavings soiled by strange males or the urine of a strange male within a period of 4 days post-insemination, the pregnancies of the females are interrupted due to failure of implantation and the females return to normal cycle within a few days . Females made anosmic by having their olfactory tracts severed do not have their pregnancies blocked by exposure to bedding soiled by strange males and consequently olfactory stimuli released by males
have been suggested as probable stimuli for pregnancy failure (Bruce 1960, 1961 ; Bruce & Parrott 1960 ; Parkes & Bruce 1962 ; Dominic 1966) . While experiments with deermice have not been as extensive as with Mus, pregnancy block has been demonstrated and appears to have many characteristics similar to the phenomenon in Mus (Bronson & Eleftheriou 1963 ; Bronson, Eleftheriou & Garick 1964 ; Eleftheriou, Bronson & Zarrow 1962) . The purpose of the present experiment was to examine the relationship between parity and social environment in the occurrence of pregnancy block in prairie deermice since both variables are continually part of the population context. Methods Animals The animals used in these experiments were prairie deermice (Peromyscus maniculatus bairdii) whose ancestors had been part of a laboratory colony for 13 years. Sib matings, however, were prevented and wild caught animals were added to the colony once per year when possible during the past 5 yr. Both nulliparous females and females which had produced young were used in this experiment . The nulliparous females approximated 70 days of age and the parous females 157 days of age . All females had been weaned at 21 days of age and maintained with sibs of the same sex until mating at an age older than 60 days . Procedure Vaginal smears were begun for the nulliparous females on the day following pairing . Young of parous females were removed on the day of birth and vaginal smears were begun on the same day. All smears were obtained by rinsing out the vagina with a small quantity of water twice per day, at 8-9 a .m . and 4-5 p .m . during five consecutive days or until insemination . A female was regarded as inseminated when a smear revealed at least twenty sperm per field of 100 magnifications . Each inseminated female was randomly 104
TERMAN : PREGNANCY FAILURE IN DEERMICE
assigned to one of four treatments . Beginning on the afternoon of the day following insemination, each female was kept in her home cage for a period of 48 hr, either alone, with her stud male, with a strange male, or with her stud male plus a strange male . Following the treatment period, all males were removed and the females remained in their home cages for seven additional days at which time they were sacrificed and examined for embryos . There were thirty parous and thirty nulliparous females exposed to each post-insemination treatment . All animals in this experiment were kept in a room 15 ft by 15 ft which also contained a breeding colony of deermice . The room was ventilated by a window fan and its only illumination was by two 40 W fluorescent tubes between 7 .30 a .m . and 7 .30 p .m . daily. Results Figure 1 presents the percentage of females pregnant in all treatments . A female was considered pregnant provided at least one embryo was viable .
1 05
Comparing parous versus nulliparous females In all four treatment situations the parous females exhibited a significantly higher percentage of pregnancy than the nulliparous females (0 .005>P) . Further, the difference between the parous and the nulliparous in all treatments was essentially the same . Comparing treatments for parous and nulliparous females Figure 1 also presents the results of statistical comparisons of the pregnancy proportions of both parous and nulliparous females . The probability of the calculated Chi square values for comparisons between treatments are given in the horizontal columns headed by `P' which indicates the treatment with which comparisons were made . For both nulliparous and parous females, there was an increasingly higher percentage of pregnancy in the following treatment order : female plus strange male, female plus stud and strange male, female alone, and female plus stud male. PAROUS ®
NONPAROUS
90
83 H z
a z 0
63
LL)
a
47
a
N
? a S a A
7
a A
Fig . 1 . Percentage of parous and nulliparous females pregnant at 10 days post-insemination . Each female was exposed to one of the following situations during 24-72 hr post insemination : female alone, female with stud male, female with strange male, female with stud and strange male simultaneously .
1 06
ANIMAL BEHAVIOUR, 17,
When the female alone situation was compared with the female plus stud male treatment for both nulliparous and parous females, higher but not significantly higher pregnancy percentages were recorded in the latter situation . When both treatments (females alone and females plus their stud males) were compared with the female plus strange male treatment, significantly smaller pregnancy proportions were recorded in the strange male situation in three of the four comparisons (0 . 05>P). Thus, pregnancy block occurred under these circumstances . When the female plus stud male treatment was compared with the situation in which both types of males were with the female, a higher proportion of pregnant females occurred in the former (0 . 02>P). Comparison of the female plus stud male and strange male situation with the the female plus strange male only situation revealed a higher proportion of pregnancies in the former, but this difference was also not significant at the P>0 . 05 level . These two comparisons therefore indicate that the presence of the stud male during the time the strange male was present increased the pregnancy success of the female, but the presence of the strange male decreased the pregnancy success exhibited when the female was with the stud male only.
1
Combined comparisons Figure 2 shows the pregnancy proportion when treatment groups were combined irrespective of parity . `P' indicates the treatment situation with which comparisons were made and the probabilities of the calculated Chi square values for the comparisons are recorded in the horizontal column to the right of each `P'. As noted previously, pregnancy success was significantly greater among females kept alone and those kept with their stud males when compared with the female plus strange male situation (0 . 025>P and 0 .0005 >P respectively) . Presence of the stud male while the strange male was present significantly increased the pregnancy success compared to the treatment when the female was alone with the strange male (0 .05>P) . Further the presence of the strange male decreased the pregnancy success of females maintained with the stud male only. This decrease was significant at 0 . 1 >P . Discussion The percentage of pregnancies was significantly smaller among nulliparous females than parous females in all treatments of the experimental design . This difference was relatively constant irrespective of the treatment . Confounded with the parity difference, however, was an age
80 fz
a z
68
0 W
62
crv a W °
co a
43
Fz W 0
P
NS
.I)
.0005)
a: P
W (L
NS
P
2
a S
2
a S a A
.025) .05)
2 a
A
Fig . 2 . Percentage of females pregnant at 10 days post-insemination when the data were combined irrespective of parity .
TERMAN : PREGNANCY FAILURE IN DEERMICE
difference, since the parous females were also older. Chipman & Fox (1966) have recently shown that six month-old house mice (M. musculus) females are more resistant to pregnancy blocking than two-month-old females . Age as well as parity may, therefore, be an important variable producing the differences observed in the present study . However, to my knowledge, the age variable has not been tested with deermice . One of the most interesting findings of the present experiment was the influence of the stud male on the stimulation of pregnancy block by the strange male . When the females were exposed to the strange male with the stud male present, the percentage of pregnancies was increased compared to the situation in which the stud male was not present during this exposure . However, the presence of the strange male in addition to the stud male decreased the percentage of pregnancies noted when stud males only were present with the females . Pregnancy success of the females when both males were present was almost exactly intermediate to the treatment situations in which the females were with one type of male or the other . These results vary somewhat from those mentioned without detailed desciption by Parkes & Bruce in their 1961 review ; namely that the presence of the stud male with the female during exposure to alien males largely eliminated pregnancy blocking . These differences, however, may reflect only differences in technique and the species of mice studied . The above findings raise the interesting question of whether the presence of the stud male prevented the strange male from releasing the pheromones which function to block the pregnancies, or whether the stud male influenced the response of the female to the pheromone . Both possibilities are relevant to the experimental study of population dynamics as well as to basic questions relating behaviour to reproductive physiology . Certainly, the behavioural definition of the stud male versus the strange male is difficult to establish within the population context . Clearly established by these data, however, is the fact that the occurrence of `pregnancy block', a pheromone induced phenomenon, may he influenced by social factors per se . Prior evidence suggesting such an influence but in a different context was provided by Bruce (1963), who exposed groups composed of differing numbers of female Mus to a single male and found that the females were protected against pregnancy blocking in direct proportion to the
107
number in the group at the time of exposure . The relationship between these two kinds of `social factors' to pregnancy block or to each other is not clear . Summary The occurrence of `pregnancy block' was noted in nulliparous and recently parous female prairie deermice when exposed to one of the following situations during a period from 24 to 72 hr subsequent to insemination : (1) stud male removed and female left alone, (2) stud male left with female, (3) stud male removed and strange male placed with female and (4) strange male placed with female and stud male . All females were killed at 10 days post-insemination and examined for embryos . There was a significantly greater frequency of pregnancy among parous females than nulliparous, although age differences may have been an important confounding variable . Further, there were decreasing pregnancy proportions in both parous and nulliparous females in the treatment sequences of (2) stud male left with female, (1) stud male removed and female left alone, (4) strange male placed with female and stud male and (3) stud male removed and strange male placed with female . Acknowledgments This study was partially supported by Public Health Service Grant NIH-MH-08289 from the National Institutes of Health . REFERENCES Bronson, F . H . & Eleftheriou, B . E . (1963) . Influence of strange males on implantation in the deermouse . Gen . comp . Endocr., 3, 515-518 . Bronson, F . H ., Eleftheriou, B . E . & Garick, E . I . (1964) . Effects of intra- and inter-specific social stimulation on implantation in deermice. J. Reprod. Fert ., 8, 23-27 . Bruce, H. M . (1960) . A block to pregnancy in the mouse caused by the proximity of strange males . J. Reprod. Fert., 1, 96-103 . Bruce, H . M . (1961) . An olfactory block to pregnancy in mice. Part 1 : Characteristics of the block . Proc . I Vth Int. Cong. An . Reprod., 159-162 . Bruce, H. M . (1963) . Olfactory block to pregnancy among grouped mice . J. Reprod. Fert ., 6, 451-460 . Bruce, H . M . & Parrott, D . M . V . (1960) . Role of Olfactory sense in pregnancy block by strange males . Science, N . Y., 131, 1526 . Chipman, R . K . & Fox, K . A . (1966) . Factors in pregnancy blocking, age and reproductive background of females : numbers of strange males . J . Reprod. Fert ., 12, 399-403 . Dominic, C . J . (1966) . Observations on the reproductive pheromones of mice . 1 . Source . J. Reprod . Fert ., 11, 407-414.
108
ANIMAL BEHAVIOUR, 17,
Eleftheriou, B . E ., Bronson, F. H . & Zarrow, M . X . (1962) . Interaction of olfactory and other environmental stimuli on implantation in the deermice. Science, N.Y., 137, 764 . Parkes, A . S . & Bruce, H. M . (1961) . Olfactory stimuli in mammalian reproduction . Science, N.Y., 134, 1049-1054. Parkes, A . S . & Bruce, H . M . (1962) . Pregnancy block in female mice placed in boxes soiled by males . J. Reprod. Fert., 4, 303-308 . Terman, C. R. (1963) . The influence of differential early social experience upon spatial distribution within
1
population of prairie deermice. Anim . Behav ., 11, 246-262 . Terman, C . R. (1965) . A study of population growth and control exhibited in the laboratory by prairie deermice . Ecology, 46, 890-895 . Terman, C . R . (1968) . Weights of the reproductive organs, adrenal glands and other selected organs of prairie deermice from laboratory populations with a long duration of asymptote . Unpublished ms .-submitted for publication . (Received 16 June 1967 ; revised 12 March 1968 ; Ms . number: A669)