GREAT/LGR8 is the only receptor for insulin-like 3 peptide

During male development testes descend from their embryonic intraabdominal position into the scrotum. Two genes, encoding the insulin-like 3 peptide (INSL3) and the GREAT/LGR8 G protein-coupled receptor, control the differentiation of gubernaculum, the caudal genitoinguinal ligament critical for testicular descent. It was established that the INSL3 peptide activates GREAT/LGR8 receptor in vitro. Mutations of Insl3 or Great cause cryptorchidism (undescended testes) in mice. Overexpression of the transgenic Insl3 causes male-like gubernaculum differentiation, ovarian descent into lower abdominal position, and reduced fertility in females. To address the question whether Great deletion complements the mutant female phenotype caused by the Insl3 overexpression, we have produced Insl3 transgenic mice deficient for Great. Such females had a wild-type phenotype, demonstrating that Great was the only cognate receptor for Insl3 in vivo. We have established that pancreatic HIT cells, transfected with the INSL3 cDNA, produce functionally active peptide. Analysis of five INSL3 mutant variants detected in cryptorchid patients showed that P49S substitution renders functionally compromised peptide. Therefore, mutations in INSL3 might contribute to the etiology of cryptorchidism. We have also showed that synthetic insulin-like peptides (INSL4 and INSL6) were unable to activate LGR7 or GREAT/LGR8.     

Cryptorchidism: an indicator of testicular dysgenesis?

Cryptorchidism is a common ailment of new-born boys, affecting 1–9% of full term boys at birth. Cryptorchidism has been associated with an increased risk of testicular cancer and reduced fertility. Aetiology of cryptorchidism remains obscure in most cases. Familial occurrence suggests a heritable susceptibility to cryptorchidism; however, seasonal variation in the incidence of cryptorchidism suggests that environmental factors also contribute. Testicular descent is characterised by androgen-dependent regression of cranial suspensory ligament and androgen + insulin-like hormone 3 (Insl3)-dependent gubernacular outgrowth. Even though hormonal defects are rarely detected in patients, both hypo-and hypergonadotropic hormonal patterns have been associated with cryptorchidism. Moreover, cryptorchid boys have significantly reduced serum androgen bioactivity at 3 months of age when normal boys have a strong surge of reproductive hormones. Defects in Insl3 action cause cryptorchidism in male mice, and over-expression in female mice causes ovarian descent. Defects in leucine-rich repeat-containing G-protein-coupled receptor 8/G-protein-coupled receptor affecting testis descent (LGR8/GREAT), the receptor for Insl3, manifest the same phenotype as Insl3 knockout mutants. Even though mutations found in Insl3 and LGR8/GREAT genes are not a common cause of cryptorchidism in patients, it remains to be resolved whether low Insl3 levels during development are associated with cryptorchidism. Cryptorchidism may reflect foetal testicular dysgenesis that may later manifest as subfertility or testicular cancer.This work was supported by the Turku University Central Hospital, the Academy of Finland and the European Commission (contracts BMH4-CT96-0314, QLK4-CT1999-01422, QLK4-CT2001-00269 and QLK4-CT2002-00603).     

Defining the LGR8 residues involved in binding insulin-like peptide 3

The peptide hormone insulin-like peptide 3 (INSL3) is essential for testicular descent and has been implicated in the control of adult fertility in both sexes. The human INSL3 receptor leucine-rich repeat-containing G protein-coupled receptor 8 (LGR8) binds INSL3 and relaxin with high affinity, whereas the relaxin receptor LGR7 only binds relaxin. LGR7 and LGR8 bind their ligands within the 10 leucine-rich repeats (LRRs) that comprise the majority of their ectodomains. To define the primary INSL3 binding site in LGR8, its LRRs were first modeled on the crystal structure of the Nogo receptor (NgR) and the most likely binding surface identified. Multiple sequence alignment of this surface revealed the presence of seven of the nine residues implicated in relaxin binding to LGR7. Replacement of these residues with alanine caused reduced [(125)I]INSL3 binding, and a specific peptide/receptor interaction point was revealed using competition binding assays with mutant INSL3 peptides. This point was used to crudely dock the solution structure of INSL3 onto the LRR model of LGR8, allowing the prediction of the INSL3 Trp-B27 binding site. This prediction was then validated using mutant INSL3 peptide competition binding assays on LGR8 mutants. Our results indicated that LGR8 Asp-227 was crucial for binding INSL3 Arg-B16, whereas LGR8 Phe-131 and Gln-133 were involved in INSL3 Trp-B27 binding. From these two defined interactions, we predicted the complete INSL3/LGR8 primary binding site, including interactions between INSL3 His-B12 and LGR8 Trp-177, INSL3 Val-B19 and LGR8 Ile-179, and INSL3 Arg-B20 with LGR8 Asp-181 and Glu-229.     

Male mice that do not express group VIA phospholipase A2 produce spermatozoa with impaired motility and have greatly reduced fertility

The Group VIA Phospholipase A(2) (iPLA(2)beta) is the first recognized cytosolic Ca(2+)-independent PLA(2) and has been proposed to participate in arachidonic acid (20:4) incorporation into glycerophosphocholine lipids, cell proliferation, exocytosis, apoptosis, and other processes. To study iPLA(2)beta functions, we disrupted its gene by homologous recombination to generate mice that do not express iPLA(2)beta. Heterozygous iPLA(2)beta(+/-) breeding pairs yield a Mendelian 1:2:1 ratio of iPLA(2)beta(+/+), iPLA(2)beta(+/-), and iPLA(2)beta(-/-) pups and a 1:1 male:female gender distribution of iPLA(2)beta(-/-) pups. Several tissues of wild-type mice express iPLA(2)beta mRNA, immunoreactive protein, and activity, and testes express the highest levels. Testes or other tissues of iPLA(2)beta(-/-) mice express no iPLA(2)beta mRNA or protein, but iPLA(2)beta(-/-) testes are not deficient in 20:4-containing glycerophosphocholine lipids, indicating that iPLA(2)beta does not play an obligatory role in formation of such lipids in that tissue. Spermatozoa from iPLA(2)beta(-/-) mice have reduced motility and impaired ability to fertilize mouse oocytes in vitro and in vivo, and inhibiting iPLA(2)beta with a bromoenol lactone suicide substrate reduces motility of wild-type spermatozoa in a time- and concentration-dependent manner. Mating iPLA(2)beta(-/-) male mice with iPLA(2)beta(+/+), iPLA(2)beta(+/-), or iPLA(2)beta(-/-) female mice yields only about 7% of the number of pups produced by mating pairs with an iPLA(2)beta(+/+) or iPLA(2)beta(+/-) male, but iPLA(2)beta(-/-) female mice have nearly normal fertility. These findings indicate that iPLA(2)beta plays an important functional role in spermatozoa, suggest a target for developing male contraceptive drugs, and complement reports that disruption of the Group IVA PLA(2) (cPLA(2)alpha) gene impairs female reproductive ability.     

Do sires and juvenile male mice (C57BL/6) contribute to the rearing of the offspring?

Copulation and/or cohabitation with a pregnant female facilitate paternal behavior in male mice. However, their contribution to the rearing of the offspring is still not well understood. Our aims were to investigate the behavior of sires toward own or alien pups; the immediate consequences of the presence of fathers on the offspring and the behavior of the mother; and whether the exposure of juvenile males to newborn siblings, in an overlapping litters context, facilitates paternal behavior in C57BL/6 mice. We found that sires behaved paternally toward alien pups at both postpartum days 3 and 7; did not affect the behavior of the mother (e.g., licking and grooming, retrieval behavior, time in the nest, and crouching postures); and reduced the time offspring stayed alone in the nest. The exposure to newborn siblings did not promote paternal behavior in juvenile males. Therefore, sires are more paternal than usually described in the literature for laboratory mice, suggesting a facultative role in the rearing of the offspring. However, juvenile male mice, in contrast to juvenile females, could be adapted to leave the nest earlier without major contribution to the offspring.     

Characterization of novel splice variants of LGR7 and LGR8 reveals that receptor signaling is mediated by their unique low density lipoprotein class A modules

The relaxin and insulin-like peptide 3 receptors, LGR7 and LGR8, respectively, are unique members of the leucine-rich repeat-containing G-protein-coupled receptor (LGR) family, because they possess an N-terminal motif with homology to the low density lipoprotein class A (LDLa) modules. By characterizing several LGR7 and LGR8 splice variants, we have revealed that the LDLa module directs ligand-activated cAMP signaling. The LGR8-short variant encodes an LGR8 receptor lacking the LDLa module, whereas LGR7-truncate, LGR7-truncate-2, and LGR7-truncate-3 all encode truncated secreted proteins retaining the LGR7 LDLa module. LGR8-short and an engineered LGR7 variant missing its LDLa module, LGR7-short, bound to their respective ligands with high affinity but lost their ability to signal via stimulation of intracellular cAMP accumulation. Conversely, secreted LGR7-truncate protein with the LDLa module was able to block relaxin-induced LGR7 cAMP signaling and did so without compromising the ability of LGR7 to bind to relaxin or be expressed on the cell membrane. Although the LDLa module of LGR7 was N-glycosylated at position Asn-14, an LGR7 N14Q mutant retained relaxin binding affinity and cAMP signaling, implying that glycosylation is not essential for optimal LDLa function. Using real-time PCR, the expression of mouse LGR7-truncate was detected to be high in, and specific to, the uterus of pregnant mice. The differential expression and evolutionary conservation of LGR7-truncate further suggests that it may also play an important role in vivo. This study highlights the essential role of the LDLa module in LGR7 and LGR8 function and introduces a novel model of GPCR regulation.     

Adamts-1 is essential for the development and function of the urogenital system

Successful ovulation and implantation processes play a crucial role in female fertility. Adamts-1, a matrix metalloproteinase with disintegrin and thrombospondin motifs, has been suggested to be regulated by the progesterone receptor in the hormonal pathway leading to ovulation. With the primary aim of investigating the role of Adamts-1 in female fertility, we generated Adamts-1 null mice. Forty-five percent of the newborn Adamts-1 null mice die, with death most likely caused by a kidney malformation that becomes apparent at birth. Surviving female null mice were subfertile, whereas males reproduced normally. Ovulation in null females was impaired because of mature oocytes remaining trapped in ovarian follicles. No uterine phenotype was apparent in Adamts-1 null animals. Embryo implantation occurred normally, the uteri were capable of undergoing decidualization, and no morphological changes were observed. These results demonstrate that a functional Adamts-1 is required for normal ovulation to occur, and hence the Adamts-1 gene plays an important role in female fertility, primarily during the tissue remodeling process of ovulation.     

Moderate caloric restriction initiated in rodents during adulthood sustains function of the female reproductive axis into advanced chronological age

Age-related ovarian failure in women heralds the transition into postmenopausal life, which is characterized by a loss of fertility and increased risk for cardiovascular disease, osteoporosis and cognitive dysfunction. Unfortunately, there are no options available for delaying loss of ovarian function with age in humans. Rodent studies have shown that caloric restriction (CR) can extend female fertile lifespan; however, much of this work initiated CR at weaning, which causes stunted adolescent growth and a delayed onset of sexual maturation. Herein we tested in mice if CR initiated in adulthood could delay reproductive aging. After 4 months of CR, the ovarian follicle reserve was doubled compared to ad libitum (AL)-fed age-matched controls, which in mating trials exhibited a loss of fertility by 15.5 months of age. In CR females returned to AL feeding at 15.5 months of age, approximately one-half remained fertile for 6 additional months and one-third continued to deliver offspring through 23 months of age. Notably, fecundity of CR-then-AL-fed females and postnatal offspring survival rates were dramatically improved compared with aging AL-fed controls. For example, between 10 and 23 months of age, only 22% of the 54 offspring delivered by AL-fed females survived. In contrast, over 73% of the 94 pups delivered by 15.5- to 23-month-old CR-then-AL-fed mice survived without any overt complications. These data indicate that in mice adult-onset CR maintains function of the female reproductive axis into advanced age and dramatically improves postnatal survival of offspring delivered by aged females.     

Vertical transmission of Neospora caninum in BALB/c mice determined by polymerase chain reaction detection.

Vertical transmission of Neospora caninum was evaluated in BALB/c mice using an N. caninum-specific polymerase chain reaction (PCR) assay as a means of detecting parasite transmission to offspring. BALB/c mice were infected with the NC-1 isolate of N. caninum during pregnancy (days 8-15 gestation). Transmission of parasite, detected by PCR, was determined in 2- to 23-day-old offspring. When dams were infected on days 13-15 of gestation, transfer of parasites was detected in only a proportion of the litter. Infection between days 8 and 12 of gestation resulted in a high frequency of parasite transmission; every offspring from all litters was infected. The tissue locations of parasites in pups of different ages were determined. In young pups (2- to 4-days-old), the predominant sites of infection were the lungs and the brain. In older pups (7- and 23-days-old) the predominant site of infection was the brain. This study shows that PCR may be useful for evaluation of candidate vaccines against horizontal N. caninum infection, vertical transmission, or both.     

Direct production of gene-targeted mice from ES cells by nuclear transfer and gene transmission to their progeny.

In order to evaluate the usefulness of a cloning technique to produce gene-manipulated mice for the field of laboratory animal science, we produced mice cloned from gene-targeted embryonic stem (ES) cells and examined the vertical transmission of a targeted gene to their progeny. Of 1257 eggs constructed by nuclear transfer using M-phase ES donor cells targeted with an oviduct-specific glycoprotein (OGP) gene, 990 formed a pseudo-pronucleus and a polar body after activation. Of 504 cloned embryos transferred into recipients, 20 live cloned pups (2%) were recovered by Caesarean section at 19.5 days of gestation. Fourteen of these cloned mice were studied. Genotyping of the OGP locus and 20 microsatellite loci showed that they were genetically identical to the OGP gene-targeted TT2 cells. Eight cloned pups grew into adults, of which 7 were male and 1 was female (missing the Y chromosome). Mating experiments using the cloned mice were carried out. Of 89 F1 mice produced from the mating of cloned and C57BL/6J mice, 50 had the targeted OGP gene heterozygously. Thirty-seven F2 mice from 4 pairs of the OGP-/+ mice were composed of 9 OGP-/-, 18 OGP-/+, and 10 OGP+/+. Moreover, 26 offspring of one pair of the cloned mice were composed of 10 OGP-/-, 12 OGP-/+, and 4 OGP+/+. These offspring were fertile and transmitted the mutant OGP gene to the next generation. Comparison of these results with those of germline chimeric mice indicates that gene-targeted mice can be produced at least one generation earlier by nuclear transfer than by the conventional methods. In addition, the targeted OGP gene was constantly transmitted to the progeny of the gene-targeted mice. Cloning techniques are potentially a more efficient way to generate gene-manipulated mice for laboratory animal science, although such techniques include many unresolved problems, such as low production efficiency, and selection of a cell source for gene manipulation among others.     

Haploinsufficiency of the follicle-stimulating hormone receptor accelerates oocyte loss inducing early reproductive senescence and biological aging in mice

Female mice that are null for the FSH-receptor (FSH-R) gene are estrogen deficient, acyclic, and sterile. However, the heterozygous (+/-) mice initially have reduced fertility and stop breeding by 7-9 mo. The purpose of this study was to understand the basis of reduced fertility in mice with haploinsufficiency of the FSH-R. Heterozygous females were compared to +/+ females at 3, 7, and 12 mo of age. By 7 mo most of the +/- females were acyclic and <50% delivered pups. The wild-type females were normal in these respects. None of the 1-yr-old +/- females gave viable offspring (73% in +/+). Many degenerative changes, including atresia and apoptosis, and profound loss of oocytes, were apparent in +/- mice by 7 mo. The 1-yr-old +/- ovary had very few follicles and consisted mostly of fibroid tissue and cysts. Our data support the hypothesis that reproductive deficits in +/- FSH-R mice occur because of accelerated oocyte loss due to increased cell death in the ovary. These events contribute to early reproductive senescence and biological aging in mice. Thus FSH-R status is an important determinant of ovarian aging and all phenomena that arise from subsequent estrogen deficiency and other aberrations.     

Absence of p53 and FasL has sexually dimorphic effects on both development and reproduction

Reproduction and development are highly dependent on apoptosis to balance the proliferation that necessarily occurs during these processes. How the absence of two apoptotic factors in mice would affect reproduction and development was examined. Given previous reports of increased neural tube defects in p53-/- female fetuses, decreased fertility in gld female mice, and altered spermatogenesis in both p53 and gld male mice, the possibility that these phenotypes might be enhanced by the elimination of a second apoptotic factor was investigated. The reproductive vigor and the health of offspring were monitored during the production of the new double-deficient strain (FasL-/-p53-/-) for any changes from the reported phenotypes. Thus, any unusual phenotypes that could lead to new models for studying mechanisms of health and disease would be identified. Double-deficient male offspring appeared healthy and occurred at expected frequencies. Additionally, spermatogenesis and male fertility were unaffected by the gene deficiencies. On the other hand, FasL+/+p53-/- and FasL-/-p53-/- female mice were susceptible to increased malformations and post-natal death. These abnormalities were consistent with previous reports of neural tube defects in p53-/- female mice. Fertility rates were also significantly decreased in p53-/- female mice that lived to be adults, an observation not previously reported. Finally, the absence of both FasL and p53 led to dystocia in pregnant female mice, suggesting that the two genes play complementary roles in parturition. Therefore, although male mouse development and reproduction remained unaffected by p53 and FasL deficiencies, female mouse development was adversely affected by the absence of p53, and no live litters were born to female mice with the combined absence of both FasL and p53. In this report, we suggest a potential mechanism involving corpora luteal regression to explain this defect in parturition in FasL-/-p53-/- female mice.     

Analysis of LGR4 Receptor Distribution in Human and Mouse Tissues

LGR4 is an R-spondin receptor with strong positive effect on Wnt signaling. It plays a critical role in development as its ablation in the mouse led to total embryonic/neonatal lethality with profound defects in multiple organs. Haplotype insufficiency of LGR4 in human was associated with several diseases, including increased risk of squamous cell carcinoma of the skin, reduced birth weights, electrolyte imbalance, and decreased levels of testosterone, which are similar to the phenotypes of LGR4-hypomorphic mice. Tissue distribution of LGR4 was extensively analyzed in the mouse using gene-trap reporter enzyme alleles. However, its expression pattern in human tissues remained largely unknown. We have developed LGR4-specific monoclonal antibodies and used them to examine the expression of LGR4 in selected adult human and mouse tissues by immunohistochemical analysis. Intense LGR4-like immunoreactivity was observed in the epidermis and hair follicle of the skin, pancreatic islet cells, and epithelial cells in both the male and female reproductive organs. Of particular interest is that LGR4 is highly expressed in germ cells and pancreatic islet cells, which have important implications given the role of R-spondin-LGR4 signaling in the survival of adult stem cells. In addition, the majority of colon tumors showed elevated levels of LGR4 receptor. Overall, the expression pattern of LGR4 in human tissues mapped by this IHC analysis is similar to that in the mouse as revealed from gene trap alleles. Importantly, the pattern lends strong support to the important role of LGR4 in the development and maintenance of skin, kidney, reproductive systems, and other organs.     

Marker genes identify three somatic cell types in the fetal mouse ovary

The two main functions of the ovary are the production of oocytes, which allows the continuation of the species, and secretion of female sex hormones, which control many aspects of female development and physiology. Normal development of the ovaries during embryogenesis is critical for their function and the health of the individual in later life. Although the adult ovary has been investigated in great detail, we are only starting to understand the cellular and molecular biology of early ovarian development. Here we show that the adult stem cell marker Lgr5 is expressed in the cortical region of the fetal ovary and this expression is mutually exclusive to FOXL2. Strikingly, a third somatic cell population can be identified, marked by the expression of NR2F2, which is expressed in LGR5- and FOXL2 double-negative ovarian somatic cells. Together, these three marker genes label distinct ovarian somatic cell types. Using lineage tracing in mice, we show that Lgr5-positive cells give rise to adult cortical granulosa cells, which form the follicles of the definitive reserve. Moreover, LGR5 is required for correct timing of germ cell differentiation as evidenced by a delay of entry into meiosis in Lgr5 loss-of-function mutants, demonstrating a key role for LGR5 in the differentiation of pre-granulosa cells, which ensure the differentiation of oogonia, the formation of the definitive follicle reserve, and long-term female fertility.     

The reproductive capacity of male mice protected against the superlethal action of gamma radiation by the administration of a mixture of Archangelica officinalis and Ledum palustre extracts]

Thirty-day old albino mongrel male mice were exposed to gamma radiation (LD90/30) after preventive single injection of a mixture of extracts from Archangelica officinalis and Ledum palustre. One month after irradiation, the survivors were mated to nonirradiated females. Healthy offspring were obtained from 11 out 12 experimental males. The number of mouse pups was 10.2 +/- 0.6 and 7.4 +/- 0.7 in the experimental and nonirradiated groups respectively. The number of both sexes in the posterity of nonirradiated parents was equal, whereas in offspring of experimental groups, the number of female pups was 2.3 times larger than that of males. The experimental posterity was found to be resistant to supralethal radiation doses.     

Genetic control of fertility and embryonic waste in the mouse: A rolefor angiotensinogen

The purpose of this study was to evaluate the impact of angiotensinogen gene (Agt) deficiency on reproductive fitness in a rodent model. Mice with 0 (Agt(-/-)), 1 (Agt(-/+)), and 2 (Agt(+/+)) copies of Agt were bred according to the following schemes: 1) Agt(-/-) x Agt(-/-), 2) Agt(-/+) x Agt(-/+), 3) Agt(+/+) x Agt(+/+), and 4) Agt(+/+) female symbol x Agt(-/+) male symbol. There were 4 breeding pairs per scheme. Breedings were time mated. Mice and litters were weighed daily. Southern blotting was used for genotyping. We found that Agt(-/-) breeding pairs had fewer litters (2 [range 1-2] vs. 4 [range 3-5]; P = 0.01), fewer pups per litter (4 [range 1-7] vs. 6 [range 1-10]; P = 0.006), and longer interpregnancy intervals (43 days [range 31-44] vs. 35.5 days [range 22-58]; P = 0.04) compared to wild-type controls. The ratio of postcoital plugs to subsequent litters was 4.0 and 1.2 for Agt(-/-) and Agt(+/+) breedings, respectively (P = 0.03). Median maternal weights during all trimesters of pregnancy were significantly lower for Agt-deficient mice compared to wild-type controls. Among Agt(-/+) x Agt(-/+) breedings, the proportions of Agt(+/+) (n = 17), Agt(-/+) (n = 38), and Agt(-/-) (n = 4) offspring differed significantly from the expected 1:2:1 Mendelian inheritance pattern (P = 0.03). Neonatal survival among the offspring derived from the Agt(-/-) x Agt(-/-) breeding scheme was significantly reduced (P = 0. 001). We conclude that Agt deficiency is associated with an in utero lethal effect, decreased fertility, and impaired neonatal survival.     

Pyrroloquinoline quinone improves growth and reproductive performance in mice fed chemically defined diets

Growth, reproductive performance, and indices of collagen maturation and expression were investigated in Balb/c mice fed chemically defined, amino acid-based diets with or without the addition 6 micro Mpyrroloquinoline quinone (PQQ)/kg diet. The diets were fed to virgin mice for 8 weeks before breeding. At weaning, the pups from successful pregnancies were fed the same diet as their respective dams. Reproductive performance was compromised in mice fed diets devoid of PQQ, and their offspring grew at slower rates than offspring from mice fed diets supplemented with PQQ. Successful mating (confirmed vaginal plugs) was not affected by the presence or absence of PQQ; however, pup viability (number of pups at parturition/number of pups at Day 4 of lactation) was decreased in PQQ-deprived mice. Conception (percentage of females giving live births) and fertility (percentage of births) were also decreased in PQQ-deprived mice. The slower rates of growth in offspring from PQQ-deprived mice were associated with decreased steady-state mRNA levels for Type I procollagen alpha(1)-chains in skin and lungs from neonatal mice. Values for lysyl oxidase accumulation as protein in PQQ-deficient mice also tended to be lower than corresponding values from PQQ-supplemented or -replete mice. Skin collagen solubility was increased in PQQ-deprived mice. These results indicate that PQQ supplementation can improve reproductive performance, growth, and may modulate indices of neonatal extracellular matrix production and maturation in mice fed chemically defined, but otherwise nutritionally complete diets.     

Targeted disruption of one of the importin α family members leads to female functional incompetence in delivery

Importin?α mediates the nuclear import of proteins through nuclear pore complexes in eukaryotic cells, and is common to all eukaryotes. Previous reports identified at least six importin α family genes in mice. Although these isoforms show differential binding to various import cargoes in vitro, the in vivo physiological roles of these mammalian importin α isoforms remain unknown. Here, we generated and examined importin α5 knockout (impα5(-/-)) mice. These mice developed normally, and showed no gross histological abnormalities in most major organs. However, the ovary and uterus of impα5(-/-) female mice exhibited hypoplasia. Furthermore, we found that impα5(-/-) female mice had a 50% decrease in serum progesterone levels and a 57% decrease in progesterone receptor mRNA levels in the ovary. Additionally, impα5(-/-) uteruses that were treated with exogenous gonadotropins displayed hypertrophy, similarly to progesterone receptor-deficient mice. Although these mutant female mice could become pregnant, the total number of pups was significantly decreased, and some of the pups were dead at birth. These results suggest that importin α5 has essential roles in the mammalian female reproductive organs.     

Mania‐ and anxiety‐like behavior and impaired maternal care in female diacylglycerol kinase eta and iota double knockout mice

Genome‐wide association studies linked diacylglycerol kinase eta and iota to mood disorders, including bipolar disorder and schizophrenia, and both genes are expressed throughout the brain. Here, we generated and behaviorally characterized female mice lacking Dgkh alone, Dgki alone, and double Dgkh/Dgki‐knockout (dKO) mice. We found that fewer than 30% of newborn pups raised by dKO females survived to weaning, while over 85% of pups survived to weaning when raised by wild‐type (WT) females. Poor survival under the care of dKO mothers was unrelated to pup genotype. Moreover, pups from dKO dams survived when fostered by WT dams, suggesting the poor survival rate of dKO‐raised litters was related to impaired maternal care by dKO dams. Nest building was similar between WT and dKO dams; however, some dKO females failed to retrieve any pups in a retrieval assay. Pups raised by dKO dams had smaller or absent milk spots and reduced weight, indicative of impaired nursing. Unlike WT females, postpartum dKO females showed erratic, panicked responses to cage disturbances. Virgin dKO females showed behavioral signs of anxiety and mania, which were not seen in mice lacking either Dgkh or Dgki alone. Our research indicates that combined deletion of Dgkh and Dgki impairs maternal behavior in the early postpartum period, and suggests female dKO mice model symptoms of mania and anxiety.