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The early developing embryo faces a continuously changing microenvironment to supports its growth. In the European roe deer, this environment accompanies embryonic diapause, a period of up to 4 months in which fertilization and subsequent implantation are decoupled. Diapause is characterised by a deceleration of embryonic growth. In most ruminants such as cattle and sheep, interferon tau (IFNt) plays a major role in maternal recognition of pregnancy. Uniquely to ruminants, the roe deer embryo does not secrete IFNt. The roe deer was used as a model species to gain insights into the changing uterine environment devoid of IFNt that supports prolonged decelerated embryo development, resumption of developmental velocity, and subsequent implantation. Uterine fluid samples from 188 female does were collected during regular huntings between September and January, and 4 developmental stages-blastocysts at early, mid, and late diapause and elongated embryos (16, 57, 97, and 18 does per developmental stage, respectively)-were defined. The developmental stages were assigned based on morphological characteristics of the embryo and the embryonic genomic DNA content. For the analysis of amino acids (AA), all 188 uterine fluid samples were subjected to targeted liquid chromatography-tandem mass spectrometry. Almost all AA increased over the course of embryo development. Although most AA showed developmental stage-specific concentration peaks, serine, glycine, alanine, glutamate, and glutamine were most abundantly present irrespective of the developmental progression. For the analysis of the protein abundances in the uterine fluid in a selected subset of samples (n=5 per developmental stage), holistic liquid chromatography-tandem mass spectrometry identified and quantified a total of 819 proteins with a false discovery rate of

Implantation is essential for the establishment of a successful pregnancy, and the preimplantation period plays a significant role in ensuring implantation occurs in a timely and coordinated manner. This requires effective maternal–embryonic signalling, established during the preimplantation period, to synchronise development. Although multiple factors have been identified as present during this time, the exact molecular mechanisms involved are unknown. Polyamines are small cationic molecules that are ubiquitously expressed from prokaryotes to eukaryotes. Despite being first identified over 300 years ago, their essential roles in cell proliferation and growth, including cancer, have only been recently recognised, with new technologies and interest resulting in rapid expansion of the polyamine field. This review provides a summary of our current understanding of polyamine synthesis, regulation and function with a focus on recent developments demonstrating the requirements for polyamines during the establishment of pregnancy up to the implantation stage, in particular the role of polyamines in the control of embryonic diapause and the identification of an alternative pathway for their synthesis in sheep pregnancy. This, along with other novel discoveries, provides new insights into the control of the peri-implantation period in mammals and highlights the complexities that exist in regulating this critical period of pregnancy.

Embryonic developmental arrest, known as diapause, has been reported in more than 130 species. However, its mechanisms are still not completely understood. In the roe deer, the only known ungulate that exhibits this phenomenon, diapause lasts for approximately 5 months, starting after the rut period in mid-July to early August and ending with embryo elongation and implantation in December/January. Little is known regarding oocyte characteristics during this period. Here, we analysed the roe deer oocyte transcriptome as a model to understand diapause effects on oocyte features during embryonic developmental arrest and reactivation. During regular hunting, immature oocytes were obtained by ovary slicing from diapause and nondiapause stages, and classified according to morphological characteristics. Only oocytes with >2 layers of compact cumulus cells and cytoplasm from 30 hunted females were used for analyses. Immature oocytes were denuded and snap frozen. Additional oocytes were cultured in maturation medium for 20-24h. Matured oocytes with a present polar body were snap frozen. Two pools of 10 immature and mature oocytes for both diapause and nondiapause stages were included (at least 4 donors/pool). Oocyte pools were processed using the Smart-seq 2 single-cell protocol (Illumina Inc., San Diego, CA, USA) for full-length cDNA and library preparation. We performed RNA-seq on an Illumina sequencer. The obtained Fastq files were clipped and analysed with a locally installed version of the Galaxy platform. Sequences were mapped against the roe deer transcriptome (unpublished data) and annotated against human and bovine transcripts. Differentially expressed genes (DEG; false discovery rate

In the tammar wallaby, Macropus eugenii, the blastocyst normally remains in embryonic diapause for 11 months without cell division or apoptosis occurring. Progesterone regulates reactivation by inducing active secretion from the endometrium, but the molecular cross-talk between the endometrium and blastocyst is unknown. This process may involve the phospholipid paf. Paf is an embryotrophin that acts as a trophic/survival factor in the early embryo, partly by inactivating (via the PI3K/Akt pathway) the expression of p53, a cell cycle arrest factor (1,2). In vitro, paf production from the tammar endometrium increases after diapause (3). This study examined the expression of the paf receptor (pafr) and p53 in the tammar endometrium and embryo at entry into, during and reactivation from diapause. Both pafr and p53 mRNA were expressed in the endometrium at all stages. However there was no quantitative change in pafr expression. In the endometrium, pafr protein is present on the membrane of the glandular epithelium at all stages examined, but p53 was not expressed in the endometrial nuclei at any stage and hence does not appear to be active. Both pafr and p53 mRNA were also expressed in the embryo from the early cleavage stages, during diapause and in the reactivated blastocyst. Pafr protein was present in the embryo both before and after diapause, but levels were greatly reduced during diapause, indicating it may be necessary for active growth. Unexpectedly, the expression of p53 in the embryo does not appear to depend on the presence or absence of pafr. p53 was expressed in the nuclei of the cleavage stage embryonic cells before diapause, but not during or after diapause. These results suggest that paf and pafr may participate in the molecular control of embryonic diapause in the tammar independent of p53. (1) Jin XL et al. (2009) Biology of Reproduction 80: 286–294.(2) O’Neill C (2005) Human Reproduction Update 11(3): 215–228.(3) Kojima T et al. (1993) Reproduction Fertility Development 5: 15–25.

Embryonic diapause, a suspension of cell division and growth at the blastocyst stage, is widespread amongst mammals, but is especially common in the kangaroos and wallabies. In the tammar, Macropus eugenii, the sequence of endocrine events leading to embryonic diapause and reactivation are well defined[1]. The blastocyst can remain in diapause for up to 11 months without cell division, measurable metabolism or apoptosis occurring [2]. The ovarian hormones, especially progesterone, exert their effects on the blastocyst by alterations in the endometrial secretions [3], but the molecular cross-talk between the endometrium and blastocyst is unknown. There is increasing evidence for the involvement of leukaemia inhibitory factor (LIF)but the epidermal growth factor (EGF) family of growth factors are also likely to be involved.This study examined the expression of EGF and HB-EGF as well as their receptors, ERBB1 and ERBB4, in the tammar endometrium and blastocyst at entry into, and reactivation from, diapause. The genes for these factors were highly conserved in the tammar with orthologues in human and mouse. Quantitative RT-PCR of all four factors in the endometrium showed that expression changed with stage. Although expression levels of both receptors did not change between diapause and reactivation, both HB-EGF and EGF levels increased at reactivation from diapause and levels of HB-EGF decreased at entry into diapause. All factors were immunopositive in the endometrium. Studies underway will determine whether the cellular location and quantity of these factors change with entry into or exit from diapause, and define the molecular interactions occurring between the blastocyst and endometrium. These results are consistent with a role for the EGF family of growth factors in the control of embryonic diapause in tammars.

Embryonic diapause is widespread amongst mammals, but is especially common in the kangaroos and wallabies. In the tammar, Macropus eugenii, the sequence of endocrine events leading to embryonic diapause and reactivation are well defined and the blastocyst can remain in diapause for up to 11 months without cell division or apoptosis occurring (Renfree and Shaw 2000). The ovarian hormones exert their effects on the blastocyst by alterations in the endometrial secretions, but the molecular cross-talk between the endometrium and blastocyst is unknown. One possible regulator of diapause is the phospholipid PAF, an embryotrophin that acts as a trophic/survival factor for the early embryo (O'Neill 2005) partly by inactivating the expression of p53, a cell cycle inhibitor, via the PI3-K pathway. PAF is released from the tammar endometrium around the time of reactivation from diapause (Kojima et al. 1993). This study examined the expression of PAF-R and p53 in the tammar endometrium at entry into, and reactivation from, diapause. PAF-R and p53 were highly conserved with orthologueues in human and mouse. PAF-R and p53 expression was assessed by RT–PCR and both genes were expressed in the endometrium at all stages examined. Quantitative PCR (QPCR) studies performed for PAF-R in the endometrium show that levels of PAF-R vary depending on the stage examined and appear to be increasing at entry into diapause and decreasing at exit from diapause. Immunohistochemical (IHC) studies are in progress to determine the cellular location of PAF-R in the endometrium and confirm the QPCR results. QPCR and IHC studies are in progress to determine if there is any change in levels of expression or cellular location of p53 between the stages examined and how this relates to PAF-R availability. These results suggest that the control of diapause in the tammar involves interactions between multiple factors. (1) Renfree MB, Shaw G (2000) Diapause. Annu Rev Physiol 62, 353–375 (2) O'Neill C (2005) The role of Paf in embryo physiology. Human Reproduction Update 11, 215–228 (3) Kojima T et. al. (1993) Production and secretion of progesterone in vitro and presence of PAF in early pregnancy of the marsupial, Macropus eugenii. Reproduction Fertility Development 5, 15–25.

Marsupials give birth to an undeveloped altricial young after a relatively short gestation period, but have a long and sophisticated lactation with the young usually developing in a pouch. Their viviparous mode of reproduction trades placentation for lactation, exchanging the umbilical cord for the teat. The special adaptations that marsupials have developed provide us with unique insights into the evolution of all mammalian reproduction. Marsupials hold many mammalian reproductive ‘records’, for example they have the shortest known gestation but the longest embryonic diapause, the smallest neonate but the longest sperm. They have contributed to our knowledge of many mammalian reproductive events including embryonic diapause and development, birth behaviour, sex determination, sexual differentiation, lactation and seasonal breeding. Because marsupials have been genetically isolated from eutherian mammals for over 125 million years, sequencing of the genome of two marsupial species has made comparative genomic biology an exciting and important new area of investigation. This review will show how the study of marsupials has widened our understanding of mammalian reproduction and development, highlighting some mechanisms that are so fundamental that they are shared by all today’s marsupial and eutherian mammals.

Macropus dorsalis was capable of breeding throughout the year. The period of gestation was 33 - 36 days. Birth was usually followed by an oestrus and mating, and a subsequent lactation-controlled embryonic diapause. The period between loss of pouch-young and birth was 29 - 30 days. Pouch life was 192 - 225 days and weaning occurred 81 - 159 days after permanent pouch emergence. Sexual maturity, defined as age at first fertility, occurred at 11.3 months in females and 15.7 months in males. Linear mixed-effects models were used to describe polynomial growth equations for age determination of pouch young using both head and pes length. The relationship between error in age prediction and each body measurement was also defined. Head measurements provided the most accurate estimates of the age of pouch young.

Reproduction of the purple-necked rock-wallaby, Petrogale purpureicollis, was studied in captivity. The length of the oestrous cycle was 36–38 days followed by a gestation period of 33–35 days. Birth was usually followed by an oestrus and mating, and a subsequent lactation-controlled embryonic diapause. The interval between loss of pouch young and birth was 30–36 days. Pouch life was 178–197 days and weaning occurred 92–171 days after permanent emergence from the pouch. The youngest age at which sexual maturity was reached was 21.8 months for males and 18 months for females. Linear mixed-effects models were used to describe polynomial growth equations for age determination of pouch young using both head and pes length. The relationship between error in age prediction and each body measurement was also defined. Head measurements provided the most accurate estimates of the age of pouch young.

Reproduction in the northern bettong, Bettongia tropica, was studied in captivity. B. tropica is capable of breeding throughout the year, and mating behaviour is similar to that reported for other Bettongia species. The length of the oestrous cycle was 21–23 days, and the period of gestation was 20–23 days. Birth was usually followed by an oestrus and mating, and a subsequent lactation-controlled embryonic diapause. The interval between loss of pouch young and birth was 19–20 days. Permanent emergence from the pouch occurred at 102–112 days, and young at foot were weaned at 166–185 days of age. Linear mixed-effects models were used to describe polynomial growth equations for age determination of pouch young using both head and pes length. The relationship between error in age prediction and each body measurement was also defined. Pes measurements provided the most accurate estimates of the age of pouch young.

Reproduction in the northern bettong, Bettongia tropica, was studied in captivity. B. tropica is capable of breeding throughout the year, and mating behaviour is similar to that reported for other Bettongia species. The length of the oestrous cycle was 21–23 days, and the period of gestation was 20–23 days. Birth was usually followed by an oestrus and mating, and a subsequent lactation-controlled embryonic diapause. The interval between loss of pouch young and birth was 19–20 days. Permanent emergence from the pouch occurred at 102–112 days, and young at foot were weaned at 166–185 days of age. Linear mixed-effects models were used to describe polynomial growth equations for age determination of pouch young using both head and pes length. The relationship between error in age prediction and each body measurement was also defined. Pes measurements provided the most accurate estimates of the age of pouch young.

Reproduction in the Proserpine rock-wallaby, Petrogale persephone, was studied in captivity. Sexual maturity, defined as age at first fertility, was attained at 20.5 months in females whereas males were not mature until 24.8 months. P. persephone is capable of breeding throughout the year. The length of the oestrous cycle was 33–38 days, while the period of gestation was 30–34 days. Birth was usually followed by an oestrus and mating, and a subsequent lactation-controlled embryonic diapause. The mean interval between loss of a pouch young and birth was 31.5 days. Pouch life was 203–215 days and young at foot were weaned 105–139 days after permanent emergence from the pouch. Linear mixed-effects models were used to describe polynomial growth equations for age determination of pouch young using both head and pes length. The relationship between error in age prediction and each body measurement was defined. Head measurements provided the most accurate estimates of the age of pouch young.

It has been shown that changes to the frequency of sucking by the pouch young do not affect the time of reactivation of the quiescent corpus luteum and diapause embryo in Bettongia penicillata; these observations led to the suggestion that the corpus luteum may have an inherent maximum duration of quiescence. The aim of the present study was to investigate the effect of the age of the corpus luteum on the timing of its reactivation. Ovulation fails to occur post partum in female B. penicillata isolated from males, and the introduction of a male B. penicillata induces oestrus in females suckling a young in the pouch. Oestrus was induced from Day 23 of lactation to Day 76, in different females, and the corpus luteum became quiescent. All parturitions occurred between Day 95 and Day 103 of lactation (average, Day 98.8), at the time expected if the corpus luteum had been formed post partum. Duration of quiescence of the corpus luteum ranged from 60 days to 10 days, compared with 83.9 +/- 0.43 days in females that mated post partum. The results show that the age of the corpus luteum does not affect the timing of its redevelopment and release of the embryo from diapause near the end of the period of the young being suckled within the pouch.

Petrogale assimilis has a typical life history and reproductive ecology for a macropodid of its size. Both sexes are capable of reproducing continuously; gestation is about the same length as the oestrous cycle (approximately one month); a single young is born and, a post-partum oestrus and embryonic diapause probably occurs. The sex ratio of young is unbiased. Pouch young remain permanently attached to the teat until 110 - 143 days (n=11). Permanent exit from the pouch occurs at 180 - 231 days (mean=201 days, n=25), and weaning occurs between 267 - 387 days (n=5). Sexual maturity occurs at a minimum age of 17.5 months in females and 23 months in males.

Ruminant and non-ruminant members of the Artiodactyla appear to use different mechanisms for the maternal recognition of pregnancy. In ruminants, the trophoblast of the developing blastocyst produces a characteristic Type I interferon of the interferon-tau family; this acts by blocking endometrial synthesis of the oxytocin receptor. A similar mechanism applies to domestic ruminants (sheep, cow, goat) and deer. In roe deer, which have embryonic diapause and are monoestrus, there is no requirement for a pre-implantation blastocyst signal to ensure maternal recognition of pregnancy. This paper compares the mechanisms leading to the maternal recognition of pregnancy in ruminants with that suggested to occur in a non-ruminant (the domestic pig).

The metabolism of tammar wallaby (Macropus eugenii) blastocysts was analysed by means of quantitative fluorescence microscopy during embryonic diapause and 2, 3, 4, 5, 8 and 10 days after reactivation to determine nutrient preferences during metabolic reactivation of the blastocyst. The surface area of quiescent blastocysts was 0.16 +/- 0.02 mm2 (mean +/- s.e.m.), and increased to 0.44 +/- 0.04 mm2 (P < 0.05) by Day 8 after removal of the sucking stimulus of the pouch young (RPY). Day-10 blastocysts, analysed over two successive breeding seasons, were significantly different in size from each other (Group A, 1992: 4.44 +/- 1.47 mm2; Group B, 1993: 18.87 +/- 4.62 mm2; P < 0.01), and both groups were significantly different in size from diapausing blastocysts (P < 0.01). There was no significant difference in carbohydrate uptake or production by blastocysts during the first five days after RPY. Glucose uptake by blastocysts recovered 8 days after RPY (61.9 +/- 30.0 pmol embryo-1 h-1) was significantly greater than that by Day-0 blastocysts (17.9 +/- 5.5 pmol embryo-1 h-1) and glucose uptake by both groups of Day-10 blastocysts (Group A, 174.0 +/- 28.4 pmol embryo-1 h-1; Group B, 616.0 +/- 239.0 pmol embryo-1 h-1) was significantly different from that by Day-0 blastocysts (P < 0.01). Pyruvate uptake by Day-10 blastocysts (Group A, 46.0 +/- 32.2 pmol embryo-1 h-1; Group B, 250.0 +/- 136.0 pmol embryo-1 h-1; P < 0.01) increased significantly compared with that by Day-0 blastocysts (6.4 +/- 1.6 pmol embryo-1 h-1; P < 0.01). Lactate production by Day-10 blastocysts (Group A, 186.7 +/- 30.3 pmol embryo-1 h-1; Group B, 285 +/- 129 pmol embryo-1 h-1; P > 0.01) was also significantly different from that by quiescent blastocysts (41.20 +/- 9.6 pmol embryo-1 h-1). There was a linear relationship between surface area and glucose uptake and surface area and pyruvate uptake (r2 = 0.965 and r2 = 0.971 respectively). Despite increases in carbohydrate uptake, there was a proportional decrease in lactate production indicating an increase in oxidative metabolism during reactivation. This suggests that there may be a metabolic switch at, or around, Day 5 after RPY.

This first account of the morphology of the female reproductive tract of the Australian sea lion, Neophoca cinerea, is based on examination of 15 specimens. The morphology of the female reproductive tract is similar in most respects to that in other pinniped species; only features peculiar to the species are described. The Y-shaped, bicornuate uterus is for the most part septate, but has a common uterine canal that is relatively much longer than that in other otariids. The uterine mucosa of newborns is slightly hypertrophied and regresses considerably by 3 weeks postpartum. During the embryonic diapause the uterine mucosa has serrated luminal epithelium, coiled uterine glands, and tall luminal and glandular epithelia. The mucosa of the adult vagina decreases in thickness from 100-mu-m at about 2 weeks postpartum to 20-mu-m 14.5 weeks postpartum, indicative of the regressive phase typical of the delay period in other pinnipeds. The urethral meatus lacks urinary papillae, unlike most other pinnipeds. A relatively large clitoris is present, and an os clitoridis was collected from one old individual. A monoestrous cycle seemed to occur in four animals, but a polyoestrous cycle is suspected in at least one individual. Ovulation occurs from alternate ovaries in successive pregnancies. Corpora albicantia are retained for at least three breeding seasons and probably longer. Implantation occurs in the midsection of the uterine cornu, ipsilateral to the ovary that released the egg. The maximum period of embryonic diapause cannot be ascertained from the available data, although a delay of about 8-9 months is possible if the breeding cycle (pregnancy cycle) lasts about 18 months.

The patterns of breeding and reproduction in Cercartetus concinnus were investigated through the histology of specimens in Australian museums, field observations in north-western Victoria, and published accounts. Histology of the reproductive tracts of museum specimens of female C. nanus, C. lepidus, Acrobates pygmaeus and Distoechurus pennatus was also undertaken to elaborate on reproduction in these species. Young of C. concinnus are born in most months, and some reproductively active males are probably present at all times of the year. Embryonic diapause occurs during the unilaminar blastocyst stage of embryonic development; the blastocysts expand slowly during diapause. The presence of embryos in the uteri of lactating female D. pennatus and C. lepidus suggests that diapause is also probable in these species. The gross anatomies of the female reproductive tracts of the five small possum species are described. Cercartetus spp. have a vaginal system characterised by voluminous culs-de-sac, and only vestigial remains of a median septum. This contrasts with the condition in the two acrobatids, in which the vaginal culs-de-sac are smaller and divided by a prominent septum. The male reproductive anatomy of C. concinnus is similar to that of C. lepidus. The prostate is carrot-shaped and its internal structure is unlike that described for other marsupial groups.

Contrary to the results of previous hybridization studies on Australian crickets, diapause has been shown to occur in eggs from the T. oceanicus 9 x T. comrnodus 6 cross. The reciprocal cross develops without delay. Diapause occurs at the same embryonic stage as in T, commodus and is accompanied by a similar drop in respiratory rate. In the light of our observations, the conclusion that diapause is absent from hybrids between diapausing and non-diapausing crickets in Japan and North America may be premature.

Sterile offspring were produced by crosses between strains of Teleogryllus commodus (Walker) from northern Australia (Queensland) and southern Australia (Victoria, Adelaide). If the northern and southern populations are not connected by a series of freely interbreeding populations, they must be recognized as distinct species. Embryonic diapause occurred in none of the eggs of the Queensland strain, and in different proportions of the eggs of the two southern strains, when incubated at 28�C. After several generations of laboratory rearing the proportions of eggs that entered diapause at 28�C decreased in both the southern strains.

The female reproductive tract is described and compared with that of other Macropodidae. It is shown that the characters considered to be diagnostic of the Potoroinae are more variable than hitherto supposed and occur also in the Macropodinae. For the greater part of the oestrous cycle the lateral vaginae are probably occluded and the pseudovaginal canal closes within a day of parturition, so that the contents of the vaginal smear are derived mainly from the posterior vaginal sinus. B. lesueur is polyoestrous and monovular. The oestrous cycle has a modal length of 23 days and gestation modal length of 21 days, post-partum oestrus following about 1 day later. The reproductive cycle is delayed during lactation and embryonic diapause occurs. The young leaves the pouch at 115 days and the delayed embryo is born a few days later. Experimental removal of the pouch young results in resumption of development and birth 20 days after removal. The young weighs 0.317 g at birth and it reaches an adult weight at about 280 days, when sexual maturity is attained. Animals less than 1 yr old can be distinguished by the presence of the sectorial premolar 3 in the upper jaw, which is shed at 1 yr. The full adult dentition is attained by 18 months. In close confinement females bred regularly from February to September but about 70% spontaneously lost their young by the age of 3 weeks. In large pens young were carried through lactation to independence. Reproduction in Bettongia is compared to that of Potorous, a closely related rat-kangaroo, and a general hypothesis is developed to reconcile several disparate observations of marsupial reproduction. It is postulated that most marsupials have a period of obligatory diapause interposed in the development of the embryo, and this is followed by a short and relatively constant period of embryogenesis. The diapause may be short, as in Perameles, or long as in Potorous, and, in the special conditions of the Macropodidae may be greatly extended during lactation. This latter phenomenon may be secondarily advantageous to the species ecologically but it is emphasized that its primary adaptive significance is likely to be found in the need to synchronize embryonic and uterine development for intra-uterine nourishment of the embryo.

A method is presented for splitting open fixed blastocysts and mounting the entire vesicle on a flat surface. A description is given of the red kangaroo blastocyst during lactation-controlled embryonic diapause. The protoderm is unilaminar and contains about 85 cells. No mitotic figures are seen, and there is no growth or differentiation. The marsupial blastocyst is compared with blastocysts of eutherian mammals during diapause.

The reproductive system of the female grey kangaroo consists as in other marsupiais, of two ovaries, two uteri, two lateral vaginae, and a median vaginal canal. The changes which occur during the oestrous cycle or pregnancy can be divided into three phases: (1) a proliferative phase characterized by maturation and rupture of an ovarian follicle and its subsequent transformation into a new corpus luteum, and by cell division in the uteri; (2) a luteal phase, when the corpus luteum is fully formed and the luteal cells attain their maximum size, and the uterine gland cells are tall columnar, with basally situated nuclei; and (3) a post-luteal phase when the secretory cells of the corpus luteum and uteri degenerate. Repair begins in the uteri before the end of pregnancy and this is supplemented by post-partum mitotic activity, but post-partum oestrus does not occur. The uterine glands are small, and ovarian activity is inhibited during the initial period of lactation. A small proportion of females with pouch young over 100 days old return to oestrus and mate. Mitotic activity then ceases in the uteri, in the newly formed corpus luteum, and the blastocyst until the pouch young is lost or approaches the end of pouch life, when mitotic activity is resumed in the corpus luteum and blastocyst. The occurrence of embryonic diapause has been confirmed in both wild and captive animals.

The reproductive tracts of seven Cercartetus concinnus with pouch young were examined. One female had recently given birth, four were pregnant, and two were in anoestrus. The post-partum female was near oestrus. The ovaries contained corpora lutea and large follicles, and luteal regression was occurring in the uteri. Ovaries of pregnant females contained old corpora lutea and corpora lutea of pregnancy. Blastocysts recovered from the uteri increased in size with age of the pouch young. Corpora lutea of pregnancy out-numbered blastocysts in many females, and blastocysts often out-numbered teats. Ovaries of anoestrous females contained only old corpora lutea. During birth the young pass through a cleft in the connective tissue between the posterior end of the vaginal culs-de-sac and the posterior vaginal sinus. This cleft, the pseudovaginal canal, closes after parturition. The observation that embryos of C. concinnus do not enter diapause contrasts with conclusions previously drawn from circumstantial evidence that the pigmy possum resembled several species of Macropodidae in undergoing embryonic diapause.

The effects of temperature on the termination of diapause in eggs of Acheta commodu8 were investigated over the range of -16� 5 to + 12� SoC; the effects on the entry into diapause were also measured by exposing pre-diapause eggs to temperatures of -10�0 to +12�SoC.