The effects of space flight during gestation on rat uterine smooth muscle

Pregnant rats were flown on a NASA space shuttle during days 11-20 of gestation and the effects of pregnancy maintenance and uterine myometrial smooth muscle were evaluated. There was no effect of space flight on fetal mass in rats examined at recovery on day 20 of gestation (G-20). Rats allowed to go to term delivered vaginally at days 22-23, but space flight significantly decreased pup mass at birth. Space flight did not alter myometrial smooth muscle volume at G-20 and postpartum when compared to flight delayed synchronous controls, but flight animals showed a 37% decrease in myometrial smooth muscle volume between G-20 and postpartum. In contrast, smooth muscle volume of the myometrium of controls, postpartum, was similar to G-20. We conclude that myometrial smooth muscle, which hypertrophies during space flight similar to controls, reacclimates acutely to earth's gravity between G-20 and parturition with a dramatic reduction in volume.     

Phagocytosis of cytoplasmic blebs derived from smooth muscle cells by fibroblast-like cells and macrophages in the post-partum rat uterus

Numerous blebs were observed in contact with smooth muscle cells (SMC) by light microscopy in the myometrium of the rat uterus after parturition. Electron-microscopically the cell surface of SMC showed bulbous protrusions, which often lacked a basement membrane and were less electron-dense than the surrounding cytoplasm or sometimes nearly electron-lucent. Many bulbous protrusions were separated from SMC and became the isolated structures which we called cytoplasmic blebs. These bulbous protrusions and cytoplasmic blebs were often found to be phagocytosed by fibroblast-like cells and macrophages. A series of these tissue changes in the uterine myometrium after delivery, possibly due to hypoxic conditions, contribute to a rapid involution of SMC which have enlarged during pregnancy.     

Expression and regulation of SLC39A family zinc transporters in the developing mouse intestine

Several ZIP genes (SLC39A family of metal transporters) play roles in zinc homeostasis. Herein, the temporal and spatial patterns of expression of the mouse ZIP1, 3, 4, and 5 genes in the developing intestine and the effects of maternal dietary zinc deficiency on these patterns of expression were examined. ZIP1 and ZIP3 genes, conserved members of the ZIP subfamily II, were found to be coexpressed during development. Expression of these genes was detected on day 14 of gestation in smooth muscle and the pseudostratified endoderm. By 5 days post-partum, prominent expression became restricted to muscle and connective stroma. In contrast, expression of ZIP4 and ZIP5 genes, members of the ZIP subfamily called LIV-1, coincided with epithelial morphogenesis. ZIP5 expression was detected on d16 of gestation and localized to the basolateral membranes of the single-layered epithelium. ZIP4 expression was detected on d18 of gestation and localized to the apical membrane of villus epithelial cells. When dams were fed a zinc-deficient diet beginning at parturition, ZIP4 expression in the nursing neonate was greatly induced. In contrast, neonatal ZIP5 expression remained unchanged, but this protein was removed from the basolateral membrane of the enterocyte. These responses to dietary zinc deficiency mimic those found in the adult intestine. These studies reveal cell-type-specific expression of ZIP genes during development of the intestine, and suggest that the mouse intestine can elicit an adaptive response to dietary zinc availability at birth.     

Ultrastructure and localization of substance P and met-enkephalin immunoreactivity in the human fetal gastric antrum

Summary The ultrastructural localization and relations of substance P- and met-enkephalin-labeled neuronal structures were examined in the wall of the human gastric antrum during early fetal life. By 14–16 weeks of gestation, clearly discernable neural plexuses and a well developed external muscle coat were present. In the submucous coat, neural plexuses varied from immature forms consisting of 1–4 neurites partially enveloped by Schwann cell processes to more mature plexuses where neurons were completely enclosed by Schwann cell processes. Neuronal profiles with substance P- and met-enkephalin-like immunoreactivities were observed in the submucous plexus. In the myenteric plexus met-enkephalin-like immunoreactivity was seen within cell bodies and neurites. By contrast, although substance P-like immunoreactivity was observed in neurites in the myenteric plexus, no substance P-labeled somata could be identified. Unlabeled terminals were seen in contact with both unlabeled dendrites and met-enkephalinergic neurons. An increase in electron density was observed at the sites of contact. These structures probably represent early stages in the development of synaptic specializations. In addition, met-enkephalin-labeled varicosities were seen in apposition to smooth muscle cells of the circular muscle coat. This suggests that antral smooth muscle cells are directly innervated by met-enkephalin neurons.     

Qualitative and quantitative differences in protein synthesis comparing fetal lamb ductus arteriosus endothelium and smooth muscle with cells from adjacent vascular sites

During late gestation, intimal cushions form in the ductus arteriosus (DA) and these cause the vessel to close when it constricts in the postnatal period. The formation of intimal cushions suggests highly specialized functions of DA endothelial and smooth muscle cells. To investigate these properties, we established, from fetal lambs on Day 138 of a 148-day term gestation, primary cell cultures of DA endothelium and smooth muscle and compared them to cells derived from the adjacent pulmonary artery and aorta. Purity of the endothelial cell cultures from each vascular site was assessed by the contact inhibited "cobblestone" monolayer phenotype, by positive immunofluorescence for factor VIII and by angiotensin converting enzyme activity. Purity of smooth muscle cell cultures at each vascular site was assessed by the "hills and valleys" phenotype and by positive immunofluorescence with a smooth muscle actin specific monoclonal antibody. Endothelial and smooth muscle cells had different growth curves, ultrastructural features, and protein profiles on single and two-dimensional SDS-polyacrylamide gel electrophoresis (PAGE), but vascular sites were similar. To further determine whether differences related to DA origin were indeed present, endothelial and smooth muscle cells from all three vascular sites were incubated with the radiolabeled amino acids [14C]leucine, [14C]proline, and [14C]valine and the proteins in both the cells and the conditioned medium were analyzed by autoradiography after SDS-PAGE. A dense band corresponding to a 42-kDa protein was observed in valine-labeled DA endothelial cells and conditioned medium and a 52-kDa protein was observed in the conditioned medium of leucine-labeled DA smooth muscle cells only. Further isolation and characterization of these endothelial and smooth muscle proteins will be necessary to determine whether they are related to the mechanism of intimal cushion formation in the late gestation DA or are present abnormally in association with the intimal proliferation observed in pulmonary and systemic vascular disease.     

Ultrastructural analysis of the transdifferentiation of smooth muscle to skeletal muscle in the murine esophagus

The ultrastructure of the mouse esophagus at the level of the diaphragm was studied from embryo day 17 to adult. The transdifferentiation of smooth muscle into skeletal muscle was categorized into seven ultrastructural stages: during phase I normal smooth muscle myogenesis was observed. In phase II subpopulations of cells changed into aggregates of myoblast-like cells. At the center of these cell aggregates, phase III cells appeared that contained condensed myofilaments. Dense bodies and dense bands appeared enlarged by the accumulation of thin filaments. In phase IV the condensed myofilaments organized into sarcomere pretemplate structures. The dense bodies and dense bands formed rudimentary Z-lines. In phase V the sarcomere templates appeared as more defined structures and began to align. An elaborate perinuclear region appeared. During phase VI, skeletal muscle sarcomeres were apparent and myofilaments were arranged in a typical hexagonal array. Phase VII skeletal muscle fibers were unique with sarcomeric bifurcations and anastomoses between adjacent myofibrils. Non-contractile organelles were less organized in these cells than in skeletal muscles such as rectus and vastus lateralis muscles. During the transdifferentiation process, other cell types remained unchanged, except the number of interstitial cells of Cajal became reduced. Immunocytochemical studies with antibodies against smooth and skeletal muscle myosin were also performed during the process of transdifferentiation. An osmium tetroxide/potassium ferricyanide en bloc mordant enabled the use of ultrathin Unicryl sections for immunocytochemistry. Cells exhibited smooth muscle myosin-like immunoreactivity from the smooth muscle stage through the condensed myofilament stage. Cells were immunopositive for skeletal muscle myosin before the formation of sarcomere templates, during the condensed stage, and after development of mature skeletal muscle cells. We also observed a hybrid muscle cell with properties of both smooth and skeletal muscle cells.     

Expression of actin isoforms in developing rat intestinal epithelium

A minimum of six very similar but distinct actin isoforms are encoded by the mammalian genome. Developmental regulation of these genes results in a tissue-specific distribution of the isoforms in the adult. Using a panel of actin specific monoclonal antibodies (MAb), we recently reported the expression of two unique actin isoforms in adult rat intestinal brush border. In this report, we examine the developmental expression of these and other actin isoforms in rat intestinal epithelial cells. Isoforms containing the HUC 1-1 and/or C4 epitopes are present by day 15 of gestation and are continuously expressed throughout adult life. Unexpectedly, the gamma-enteric smooth muscle isoactin, defined by the B4 epitope, is transiently expressed in these non-muscle cells late in gestation. The alpha-vascular smooth muscle isoform, however, is not expressed in intestinal epithelial cells during development and, as previously reported, both smooth muscle isoforms are absent in epithelial cells of adult intestine. In addition, we demonstrate that although multiple isoforms are expressed simultaneously in these cells, they are not uniformly distributed at the subcellular level, suggesting that the cell recognizes the actin isoforms as functionally distinct entities.     

Serum 17,20 beta-dihydroxy-4-pregnen-3-one Levels in pregnant and non-pregnant female rockfish, Sebastes schlegeli, viviparous teleost, and its production by post-ovulatory follicles

Changes in serum 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17,20 beta-DP) levels around the gestation time of normal pregnant and experimentally non-pregnant females were investigated in the black rockfish, Sebastes schlegeli, a viviparous teleost. The serum 17,20 beta-DP in both females showed similar changes and levels, increasing from the early to late gestation periods and declining just before parturition in pregnant females and egg-release in non-pregnant females, respectively. These results suggest that the maintenance of high serum levels of 17,20 beta-DP after oocyte maturation is not correlated with gestation or parturition, but occurs spontaneously in this species. The decline of 17,20 beta-DP levels prior to egg-release in non-pregnant females tended to occur one week earlier than those prior to parturition in pregnant females, suggesting that both a decline in 17,20 beta-DP levels in mothers and some response from embryos are needed for a smooth parturition. The post-ovulatory follicles were maintained throughout the gestation period and produced a considerable amount of 17,20 beta-DP in vitro (3.44-6.96 pg/ml/mg tissue), but little estradiol-17beta (0.92-1.66 pg/ml/mg tissue). The production of 17,20 beta-DP tended to be enhanced by the addition of a precursor steroid, pregnenolone, in the pre-, early and mid-gestation periods. These results strongly suggest that the follicle cells in black rockfish have the ability to synthesize 17,20 beta-DP during the post-ovulatory period, and high serum 17,20 beta-DP during gestation is supplied by the post-ovulatory follicles, which in Sebastes are considered to be functionally homologous to the mammalian corpus luteum.     

An electron microscopic study of the post-partum involution of the rat uterus. With a note on apparent crinophagy of collagen

During pregnancy the increase in mass of the rat uterus has been accounted for mainly by hypertrophy rather than hyperplasia of the smooth muscle cells and by an increase in collagen content. Following parturition the rat uterus regains its non-pregnant weight within little more than a week. Autophagy is a likely mechanism for the reduction in size of the smooth muscle cells. However, many previous investigations have failed to demonstrate an extensive lysosomal apparatus in uterine smooth muscle cells, a morphologically detectable prerequisite for augmented autophagy. The superfluous collagen has been assumed to be digested by macrophages. In the present electron microscopical study the involution of the rat uterus was followed at various time points post-partum. The results indicated that three pathways were involved namely, autophagocytosis, heterophagocytosis and crinophagy. Autophagocytosis was noted in smooth muscle cells--apparently a means to reduce their size. In some instances extensive autophagy appeared to lead to cell death. Macrophages and also to some extent fibroblasts have the ability to endocytose collagen and cell debris including split off portions of smooth muscle cell cytoplasm. The evidence obtained suggested that at the time of parturition the (pro)collagen synthesized but not yet secreted was retained within the fibroblasts and degraded by means of crinophagy.     

Periodic organization of the contractile apparatus in smooth muscle revealed by the motion of dense bodies in single cells

To study the organization of the contractile apparatus in smooth muscle and its behavior during shortening, the movement of dense bodies in contracting saponin skinned, isolated cells was analyzed from digital images collected at fixed time intervals. These cells were optically lucent so that punctate structures, identified immunocytochemically as dense bodies, were visible in them with the phase contrast microscope. Methods were adapted and developed to track the bodies and to study their relative motion. Analysis of their tracks or trajectories indicated that the bodies did not move passively as cells shortened and that nearby bodies often had similar patterns of motion. Analysis of the relative motion of the bodies indicated that some bodies were structurally linked to one another or constrained so that the distance between them remained relatively constant during contraction. Such bodies tended to fall into laterally oriented, semirigid groups found at approximately 6-microns intervals along the cell axis. Other dense bodies moved rapidly toward one another axially during contraction. Such bodies were often members of separate semirigid groups. This suggests that the semirigid groups of dense bodies in smooth muscle cells may provide a framework for the attachment of the contractile structures to the cytoskeleton and the cell surface and indicates that smooth muscle may be more well-ordered than previously thought. The methods described here for the analysis of the motion of intracellular structures should be directly applicable to the study of motion in other cell types.     

Calponin and SM22 as differentiation markers of smooth muscle: spatiotemporal distribution during avian embryonic development

Abstract. Calponin and SM 22 are two proteins related in sequence that are particularly abundant in smooth muscle cells. Here, the distribution patterns of calponin and SM 22 were compared with that of other smooth muscle contractile and cytoskeletal components in the avian embryo using immunofluorescence microscopy and immunoblotting. Like myosin-light-chain kinase and heavy caldesmon, both calponin and SM 22 were more or less exclusively found in smooth muscle cells, during embryonic development and in the adult. Labelling of other cell types including striated muscle was not observed. In contrast, tropomyosin, smooth muscle α-actin, filamin and desmin could also be detected in many other cell types in addition to smooth muscles, at least during part of embryonic life. Calponin and SM 22 appeared almost synchronously during the differentiation of all smooth muscle cell populations, though with a slight time difference in the case of the aorta. The appearance of calponin, SM22 and heavy caldesmon was generally delayed in relation to desmin, tropomyosin, smooth muscle α-actin, myosin-light-chain kinase and filamin and a marked increase in abundance of these proteins was observed in the late embryo and in the adult. From these observations we can conclude that both calponin and SM 22 belong to a group of late differentiation determinants in smooth muscle and may constitute convenient and reliable markers to follow the differentiation of most, if not all, smooth muscle cell populations.     

High iNOS mRNA and protein levels during early third trimester suggest a role for NO in prelabor cervical ripening in the bovine

Nitric oxide (NO) plays a key role in the processes leading to cervical softening prior to labor. Inducible nitric oxide synthase (iNOS) contributes most to the increased production of NO during labor, as demonstrated in the rat cervix, or at term pregnancy in women. Changes in expression of iNOS during late gestation have not yet been studied longitudinally in any species, because repeatedly taking biopsies could not be performed. iNOS mRNA (n = 6) and protein expression (n = 3) in serial cervical biopsies of pregnant pluriparous cows taken around days 225, 250, and 275 of pregnancy and within 1.5 hr after calving (d225, d250, d275 and parturition biopsies, respectively) were measured using quantitative RT-PCR and Western blotting. iNOS mRNA expression decreased from the d225 biopsy onwards, differences being significant between the d250 and d275 (P < 0.05) and between the d275 and parturition biopsies (P < 0.05). iNOS protein expression decreased from d225 to d250 onwards. Immunohistochemical analysis of biopsies showed, besides positive staining in endothelium and epithelium, which remained unchanged at different time points, that iNOS expressing cells in the connective tissue cells of early biopsies were predominantly spindle shaped (mostly smooth muscle cells and some fibroblasts). In the parturition biopsies, iNOS reactivity was mainly found in mononuclear leucocytes. These results lead us to suggest that iNOS from spindle shaped cells is involved in prepartum cervical ripening, while iNOS in mononuclear inflammatory cells may be important for local tissue repair mechanisms during postpartum cervical involution.     

Immunohistochemical demonstration of cyclooxygenase-2 (COX-2) and prostaglandin receptors EP2 and FP expression in the bovine intercaruncular uterine wall around term

During parturition, uterine-derived prostaglandins (PG) play an outstanding role regarding the functional elimination of the corpus luteum and the promotion of uterine contraction. The rate-limiting enzyme cyclooxygenase-2 (COX-2), highly regulated in a cell-type and localization specific manner throughout pregnancy, is involved in uterine prostanoid production. Prostaglandins exert their effects via G-protein-coupled receptors. Distribution and cellular localization of these receptors are decisive factors for prostaglandin-mediated actions. Since both COX-2 and PG receptors have only been assessed during pregnancy in the cow, these parameters were localized immunohistochemically near term to evaluate their specific role at parturition. Thus, during two periods, segments of the intercaruncular uterine wall were collected from cows at slaughter being eight and nine months pregnant, from cattle during caesarean section, and after spontaneous calving.

Results reveal that COX-2 was mainly localized in the cytoplasm of surface epithelial cells with a high expression in animals with induced parturition. The enzyme could also be found in lower concentrations within the glandular epithelium without any effect of gestational time or labour. In contrast to relaxant prostaglandin E receptor type 2 (EP2), not showing any change in all tissue layers observed, contractile prostaglandin F₂ receptor (FP) was modulated during the peripartal period revealing a peak expression in animals with induced parturition. FP was localized in surface and glandular epithelial cells as well as in endometrial stroma and myometrial smooth muscle cells.

Our study indicates that labour and induction of parturition may have an effect on amounts of immunohistochemically detectable COX-2 and FP. EP2 remains rather unchanged during the peripartal period. COX-2 and FP thus contribute via changes in amount and distribution to mechanisms associated with parturition.     

Modifications of Erectile Tissue Components in the Penis during the Fetal Period

Background

The penile erectile tissue has a complex microscopic anatomy with important functions in the mechanism of penile erection. The knowledge of such structures is necessary for understanding the normal physiology of the adult penis. Therefore, it is important to know the changes of these penile structures during fetal development. This study aims to analyze the development of the main components of the erectile tissue, such as collagen, smooth muscle fibers and elastic system fibers, in human fetuses.

Methodology/Principal Findings

We studied the penises of 56 human fetuses aged 13 to 36 weeks post-conception (WPC). We used histochemical and immunohistochemical staining, as well as morphometric techniques to analyze the collagen, smooth muscle fibers and elastic system fibers in the corpus cavernosum and in the corpus spongiosum. These elements were identified and quantified as percentage by using the Image J software (NIH, Bethesda, USA). From 13 to 36 WPC, in the corpus cavernosum, the amount of collagen, smooth muscle fibers and elastic system fibers varied from 19.88% to 36.60%, from 4.39% to 29.76% and from 1.91% to 8.92%, respectively. In the corpus spongiosum, the amount of collagen, smooth muscle fibers and elastic system fibers varied from 34.65% to 45.89%, from 0.60% to 11.90% and from 3.22% to 11.93%, respectively.

Conclusions

We found strong correlation between the elements analyzed with fetal age, both in corpus cavernosum and corpus spongiosum. The growth rate of these elements was more intense during the second trimester (13 to 24 WPC) of gestation, both in corpus cavernosum and in corpus spongiosum. There is greater proportional amount of collagen in the corpus spongiosum than in corpus cavernosum during all fetal period. In the corpus spongiosum, there is about four times more collagen than smooth muscle fibers and elastic system fibers, during all fetal period studied.     

Delayed parturition: constraint or coping mechanism in a viviparous gekkonid?

The extended gestation of fully developed embryos in viviparous reptiles can be the consequence of a lack of appropriate environmental conditions and/or an adaptive contribution to increased offspring viability. I examined the cause and effects of extended gestation in a gekkonid lizard Hoplodactylus maculatus , which delays parturition of developed embryos for up to 9 months from autumn into a second spring. Comparisons of microhabitat temperatures showed that thermal opportunities were equally conducive to birth in autumn and spring. The responsiveness of females to a hormonal stimulant for parturition also did not differ between these seasons. Multiple offspring traits were measured, including body size, locomotory ability and lipid reserves, and were found not to increase with extended gestation. Incorporating these results with previously published evidence for microhabitat thermal variation, I discuss whether delayed parturition may be an adaptive mechanism maintained by the selective pressures of an uncertain thermal environment on components of reproduction, including vitellogenesis, maternal birthing cues and neonatal survival.     

Differential expression of Kv4 pore-forming and KChIP auxiliary subunits in rat uterus during pregnancy

Regulation of voltage-gated K(+) (K(v)) channel expression may be involved in controlling contractility of uterine smooth muscle cells during pregnancy. Functional expression of these channels is not only controlled by the levels of pore-forming subunits, but requires their association with auxiliary subunits. Specifically, rapidly inactivating K(v) current is prominent in myometrial cells and may be carried by complexes consisting of Kv4 pore-forming and KChIP auxiliary subunits. To determine the molecular identity of the channel complexes and their changes during pregnancy, we examined the expression and localization of these subunits in rat uterus. RT-PCR analysis revealed that rat uterus expressed all three Kv4 pore-forming subunits and KChIP2 and -4 auxiliary subunits. The expression of mRNAs for these subunits was dynamically and region selectively regulated during pregnancy. In the corpus, Kv4.2 mRNA level increased before parturition, whereas the expression of Kv4.1 and Kv4.3 mRNAs decreased during pregnancy. A marked increase in KChIP2 mRNA level was also seen at late gestation. In the cervix, the expression of all three pore-forming and two auxiliary subunit mRNAs increased at late gestation. Immunoprecipitation followed by immunoblot analysis indicated that Kv4.2-KChIP2 complexes were significant in uterus at late pregnancy. Kv4.2- and KChIP2-immunoreactive proteins were present in both circular and longitudinal myometrial cells. Finally, Kv4.2 and KChIP2 mRNA levels were similarly elevated in pregnant and nonpregnant corpora of one side-conceived rats. These results suggest that diffusible factors coordinate the pregnancy-associated changes in molecular compositions of myometrial Kv4-KChIP channel complexes.     

Dense-cored membranous structures in smooth muscle cells of the vas deferens of the rat

Summary Smooth muscle cells from rat vas deferens were studied by electron microscopy. Vesicular and tubular membranous structures containing an electron-opaque material were found in the smooth muscle cells. Similar structures were also found in a subfraction (F3) of microsomes of vas deferens smooth muscle which was shown to be rich in both plasma membrane and putative endoplasmic reticulum markers. Treatment of the tissues with calcium-free Krebs solution containing EGTA prior to fixation eliminated almost completely the presence of these dense-cored membranous structures (DMS), whereas incubation of the subcellular membrane fraction with EGTA solution had no effect on the appearance of the DMS. Plasma membrane infoldings were found in the smooth muscle cells extending well into their interior. Horseradish peroxidase penetrates vesicles in a location similar to that of DMS in smooth muscle cells, suggesting that some of the DMS may be connected to the extracellular space. We conclude that the dense-core material within the DMS is calcium dependent. We also suggest that some of the DMS represent infoldings of the plasma membrane extending into the cell's interior.     

The ovary of the gestating South American plains vizcacha (Lagostomus maximus): suppressed apoptosis and corpora lutea persistence

The South American plains vizcacha, Lagostomus maximus, displays an exceptional ovulation rate of up to 800 eggs per cycle, the highest rate recorded for a mammal. Massive polyovulation arises from the overexpression of the apoptosis-inhibiting BCL2 gene leading to a suppression of apoptotic pathways responsible for follicular atresia in mammals. We analyzed the ovarian histology, ovarian apoptosis, and apoptosis-related protein expression with special emphasis in corpora lutea throughout the 5-mo-long gestation period, at parturition day and early postpartum, in L. maximus. Corpora lutea were abundant throughout gestation with no sign of structural regression even at the end of gestation. Both immunohistochemistry and Western blot analysis showed strong signals for apoptosis-inhibiting BCL2 protein, whereas the proapoptotic BAX protein was just detected in isolated luteal cells in gestating females and postpartum females. Apoptosis-associated DNA fragmentation detected by TUNEL was very scarce and occasional and correlated with BAX detection in luteal cells. Marked expression of progesterone and alpha-estrogen receptors in luteal cells was found at early, mid-, and late gestation as well as at parturition day and early postpartum samples. Additionally, serum level of progesterone increased markedly to reach maximal values at late gestation and decreasing at parturition to levels found at early gestation, suggesting that corpora lutea remained functional throughout gestation. These results point out that the unusual ovarian environment of L. maximus in which germ cell demise is abolished through antiapoptotic BCL2 gene overexpression also preserves structural integrity and functionality of corpora lutea during the whole gestation. Overexpression of antiapoptotic BCL2 gene may represent a strategy for an essential need of ovary and corpora lutea in order to maintain pregnancy until term.     

Factors Affecting Date of Implantation,Parturition, and Den Entry Estimated from Activity and Body Temperature in Free-Ranging Brown Bears

Knowledge of factors influencing the timing of reproduction is important for animal conservation and management. Brown bears (Ursus arctos) are able to vary the birth date of their cubs in response to their fat stores, but little information is available about the timing of implantation and parturition in free-ranging brown bears. Body temperature and activity of pregnant brown bears is higher during the gestation period than during the rest of hibernation and drops at parturition. We compared mean daily body temperature and activity levels of pregnant and nonpregnant females during preimplantation, gestation, and lactation. Additionally we tested whether age, litter size, primiparity, environmental conditions, and the start of hibernation influence the timing of parturition. The mean date of implantation was 1 December (SD = 12), the mean date of parturition was 26 January (SD = 12), and the mean duration of the gestation period was 56 days (SD = 2). The body temperature of pregnant females was higher during the gestation and lactation periods than that of nonpregnant bears. The body temperature of pregnant females decreased during the gestation period. Activity recordings were also used to determine the date of parturition. The parturition dates calculated with activity and body temperature data did not differ significantly and were the same in 50% of the females. Older females started hibernation earlier. The start of hibernation was earlier during years with favorable environmental conditions. Dates of parturition were later during years with good environmental conditions which was unexpected. We suggest that free-ranging pregnant brown bears in areas with high levels of human activities at the beginning of the denning period, as in our study area, might prioritize investing energy in early denning than in early parturition during years with favorable environmental conditions, as a strategy to prevent disturbances caused by human.