In vivo culture of embryos in the immature mouse oviduct

In-vitro culture of mammalian preimplantation embryos is associated with subsequent decreased viability. This phenomenon is more pronounced with the domestic species embryos as culture conditions are at present unable to sustain cleavage of early preimplantation embryos for more than one or two cell divisions. In this study, the immature mouse oviduct is shown to be capable of supporting cleavage and morphological development of rabbit and porcine embryos. The immature mouse oviduct was shown to be comparable to in vitro culture as 76% and 60% of the transferred zygotes developed to the morula stage after 2 and 3 d respectively. The porcine zygotes, however, failed to develop beyond the 4-cell stage in either the immature mouse oviduct or in vitro. Porcine morula showed better tolerance of the oviduct environment and when recovered after 2 d contained an average of 64 cells, which was significantly more than in in vitro cultured morulae (40 cells). Early porcine blastocysts transferred to the mouse oviduct had over a two-fold increase in cell division (104 cells) over comparable blastocysts grown in vitro (57 cells). The immature mouse oviduct is, therefore, a potential surrogate environment for short-term storage of embryos of other species.     

Chicken riboflavin-binding protein. cDNA sequence and homology with milk folate-binding protein

The Rd gene is expressed in the livers and oviducts of laying hens and codes for the riboflavin-binding protein (RfBP) of egg yolk and egg white. A lambda gt11 cDNA library derived from chicken oviduct poly(A)+ RNA was screened with polyclonal rabbit antiserum to chicken RfBP. Positive clones were isolated and rescreened with a mixed oligonucleotide probe corresponding to residues 20-25 of the mature protein. The largest cDNA clone (969 base pairs) was subcloned into plasmid pIBI21, and the nucleotide sequence was determined by the dideoxynucleotide method. This clone contained the entire coding region for RfBP. The published amino acid sequence of the mature protein was confirmed. In addition, the following 17-residue signal peptide was deduced: Met-Leu-Arg-Phe-Ala-Ile-Thr-Leu-Phe-Ala-Val-Ile-Thr-Ser-Ser-Thr-Cys. Unexpectedly, the nucleotide sequence codes for 2 adjacent arginine residues at the carboxyl terminus that are not observed in the mature protein. The amino acid sequence of RfBP is homologous with bovine milk folate-binding protein. Eight of the nine pairs of cysteines involved in disulfide bonds in RfBP are conserved in folate-binding protein, as are all of the tryptophan residues. Sequence identity between homologous regions of these two vitamin-binding proteins is more than 30%.     

Epitheliocytes of fundal glands and the relative volume of the parietal microflora of the stomach and small intestine mucosa in experimental chronic duodenal ulcer

The effect of experimental chronic duodenal ulcers and vagotomy on fundal gland epitheliocytes and membrane microflora has been studied in rats using light microscopy and stereometry on semithin sections. It is shown that in ulcers the relative amount of perietal and zymogen cells increases, while the volume of mucocytes decreases. Vagotomy leads to a decrease in the relative amount of parietal and zymogen cells and increases the relative amount of mucocytes. The relative volume of membrane microflora in gastric fundal and pyloroantral regions, duodenum and jejunum diminishes in ulcers and increases in vagotomy, as compared to the control.     

Use of the rabbit oviduct as a screening tool for the viability of mammalian eggs

The oviducts of both oestrous and pseudopregnant rabbits can be used for the successful culture of mammalian embryos for short periods. This has alowed some selection to be made on the embryos as they are examined on at least two occasions before final transfer. Not only have pregnancy rates been normal, but in some instances they have been higher following a limited period (2–3 days) in the rabbit oviduct. It would appear that these higher pregnancy rates result from a more intensive selection of embryos at the time of transfer rather than from some substance acquired during storage in the oviduct. However, the system is not without disadvantages. There is some loss of embryos (15–30%) in the oviduct and all embryos recovered may not have developed at the normal rate.The rabbit oviduct has been used as a site of xenogenous fertilization. Initial reports indicate that success in that area is lower than when using large animals as the site of fertilization. With more widespread interest in the use of microsurgery in embryos, the rabbit oviduct has been used for the short term storage of agar cylinders and has been found to be unsuitable because of the high rate of degeneration of agar chips. However, the rabbit oviduct is still useful as an experimental tool in the manipulation of embryos from the domestic species.     

Secretory glands of the snail tentacle and their relation to the olfactory organ (Mollusca,Gastropoda)

Summary The epidermis of the posterior tentacles of the terrestrial snail Achatina fulica was examined by histological and histochemial methods. There are seven types of unicellular glands in the tentacle skin: three mucocytes containing either acid mucopolysaccharides or neutral mucopolysaccharides, or both; two mucocytes containing glycoproteins; a lipid gland; and a protein gland. The mucocytes are considerably more abundant along the shaft of the tentacle than at the tip, where the olfactory organ is situated. Conversely, the lipid glands and the protein glands are found almost exclusively in the olfactory organ. With minor exceptions, none of the foregoing cell types is present in the skin of the head or the foot. These observations indicate a high degree of local specificity in secretory products, consistent with a ubiquitous and generous endowment of glands in the molluscan skin. Collar cells, described by previous authors in closely related species, were not observed.     

Regional Distribution of Substance P- and Thyrotrophin-Releasing Hormone-Like Immunoreactivity and Indoleamines in the Rabbit Spinal Cord

The distribution of thyrotrophin-releasing hormone (TRH), substance P, and the indoleamines [5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA)] has been examined in selected regions of the thoracic and lumbar spinal cord of the rabbit using sensitive radioimmunoassays for the first two and HPLC with electrochemical detection for the indoleamines. The levels of TRH- and substance P-like immunoreactivity (TRH-I and SP-I, respectively) were greatest in the ventral and dorsal grey matter, respectively. The level of TRH-I in most thoracic regions was greater than that in equivalent lumbar regions, but the only segmental difference in SP-I was in the ventral grey matter, where the lumbar segment contained more immunoreactivity. 5-HT and 5-HIAA were more evenly distributed than either peptide and showed no segmental variation in levels in equivalent regions, but the ventral grey matter contained significantly higher levels of 5-HT and had a greater 5-HT/5-HIAA ratio than all other regions. The absolute levels and the overall distribution of SP-I, TRH-I, and indoleamines in the thoracolumbar cord of the rabbit was very similar to that previously reported in both rats and humans, and the possible functional role of the peptides and indoleamines in spinal neurones is discussed.     

The motility of rabbit spermatozoa recovered from the female reproductive tract

The motility of rabbit spermatozoa recovered from the vagina, endocervix, uterus, and four regions of the oviduct was assessed visually by phase-contrast microscopy at intervals from one minute to 16 hours after a single mating. The percentage of motile cells in each sample was dependent on the temperature of recovery, ie, 23° vs 37°C, but was not influenced by the temperature of observation. Spermatozoa in the lower isthmus of the oviduct were the most temperature sensitive population to recovery at 23°C. When all manipulations and observations were performed at 37°C, the percentage of spermatozoa with progressive motility varied according to the region sampled and interval after mating. Populations from the vagina, uterus and upper regions of the oviduct usually had a high proportion of progressively motile cells with vigorous flagellar activity. Fewer spermatozoa showed progressive movement on recovery from the endocervix and lower 2 cm of the tubal isthmus and their flagellar activity was generally depressed. The decrease in flagellar beat frequency noted in the latter regions may be a major factor limiting sperm ascent in the female tract. A unique pattern of “activated” motility was seen exclusively in populations taken from the oviducts at 6 to 16 hours after mating. This motility pattern, consisting of alternating episodes of linear progressive and vigorous nonprogressive movement, may be analogous to the activated motility described for capacitated rodent spermatozoa.     

Nucleotide and derived amino acid sequences of a cDNA coding for pre-uteroglobin from the lung of the hare (Lepus capensis).

An almost full-length cDNA coding for pre-uteroglobin from hare lung was cloned and sequenced. The derived amino acid sequence indicated that hare pre-uteroglobin contained 91 amino acids, including a signal peptide of 21 residues. Comparison of the nucleotide sequence of hare pre-uteroglobin cDNA with that previously reported for the rabbit gene indicated five silent point substitutions and six others leading to amino acid changes in the coding region. The untranslated regions of both pre-uteroglobin mRNAs were very similar. The amino acid changes observed are discussed in relation to the different progesterone-binding abilities of both homologous proteins.     

Effects of 19-hydroxy-prostaglandins on oviductal and uterine motility.

The effects of 19-hydroxyprostaglandins (19-OH-PGs) were tested in vivo on the rabbit oviduct and uterus and on the rhesus monkey (Macaca mulatta) uterus. The 19-OH-PGEs suppressed spontaneous oviductal and uterine activity in the rabbit. The qualitative effect on the rabbit oviduct of 19-OH-PGEs was similar to that of PGE2. However, the typical response of the rabbit uterus to PGE2 was an increase in muscle activity. With regard to the rabbit oviduct, 19(R)-OH-PGE2 was as potent as PGE2, but 19(S)-OH-PGE2 was approximately 1/2 as potent as PGE2. Based on the dose of 19-OH-PGEs usually required to cause a minimal suppression and the dose of PGE2 required to cause a minimal stimulation of rabbit uterine activity, 19(R)-OH-PGE2 was twice as potent as PGE2 while 19(S)-OH-PGE2 was 1/2 as potent as PGE2. Stimulatory effects on the rabbit oviduct and uterus were observed following administration of 19-OH-PGFs and PGF2alpha. The potency on the rabbit oviduct of 19(S)-OH-PGF2alpha was about 1/5 to 1/10 that of PGF2alpha; the potency of 19(R)-OH-PGF2alpha was about 1/10 to 1/20 that of PGF2alpha. Both 19-OH-PGFs were approximately 1/5 to 1/10 as potent as PGF2alpha on the rabbit uterus. At the doses tested 19-OH-PGFs were inactive on the monkey uterus. Thus, these compounds are at least 1/5 as active as PGF2alpha. In contrast, 19(R)-OH-PGE2 had approximately the same potency as PGE2 in stimulating monkey uterine activity; but 19(S)-OH-PGE2 was approximately 1/3 as potent as PGE2.     

Identification and localization of plasminogen activator inhibitor-1 within the porcine oviduct

The porcine oviduct synthesizes de novo and secretes a number of proteins into culture medium, many of which are unidentified. The objectives of the present study were to 1) semipurify and identify a M(r) 45 000 secreted protein of the oviduct, 2) examine its synthesis within the three functional segments (infundibulum, ampulla, and isthmus), and 3) evaluate its distribution throughout the oviduct. Oviductal tissue was collected during early pregnancy, divided into functional segments, and subsequently cultured. Medium was collected, and the M(r) 45 000 protein was concentrated by gel-filtration chromatography. The semipurified protein was transferred onto a polyvinylidene fluoride membrane and subjected to N-terminal amino acid analysis. The 26-amino acid sequence was 96% identical to that of pig plasminogen activator inhibitor (PAI)-1. Analysis by 1-dimensional SDS-PAGE and fluorography of rabbit anti-human PAI-1-immunoprecipitated product confirmed PAI-1. Subsequent 2-dimensional SDS-PAGE and fluorographic analyses of media revealed greater PAI-1 synthesis by the isthmus than by the ampulla or infundibulum. PAI-1 was immunolocalized throughout the oviduct and was heavily concentrated in the apical region of epithelial cells. Immunogold electron microscopy localized PAI-1 within putative secretory granules in the epithelial apical region and also associated with cilia in the isthmus. Isthmic PAI expression suggests a crucial role in protecting the preimplantation embryo from proteolytic degradation as well as in regulation of extracellular matrix turnover and remodeling.     

Hastened transport of equine embryos through the oviduct of the mare

The purposes of this experiment were 1) to test the hypothesis that placing rabbit embryos into the mare's uterus would hasten oviduct transport and 2) to determine if placing fluid into the uterus of bred mares on Day 4 and/or Day 5 would subsequently disrupt the mare's pregnancy. The hypothesis that placing rabbit embryos into the mare's uterus would hasten oviduct transport was not supported, since the uterine recovery rate of equine embryos on Day 5 was not significantly higher (P>0.05) for mares receiving rabbit embryos on Day 4 than for mares receiving no uterine infusion on Day 4 (1 10 vs 0 10 , respectively). However, placing fluid into the mare's uterus on Day 4 was apparently responsible for hastened oviduct transport, since mares with media infused into the uterus on Day 4 had a significantly higher (P<0.05) recovery rate of equine embryos on Day 5 than did mares receiving either rabbit embryos or no uterine infusion on Day 4 post ovulation (5 10 vs 1 10 or 0 10 , respectively). The Day-14 pregnancy rate was significantly higher (P<0.05) for mares receiving no uterine infusion on Day 4 or Day 5 than for mares receiving uterine infusion on Day 5 or uterine infusion on both Days 4 and 5 (9 10 vs 4 10 , 2 10 and 0 10 , respectively).     

鸡卵清蛋白基因启动子调控GFP基因在鸡原代输卵管上皮细胞和中国仓鼠卵巢细胞的表达

特异性扩增家鸡卵清蛋白基因上游调控序列 1340bp～ +16 5 5bp片段和第一内含子 +49bp～ +16 5 5bp片段 ,去除pG FP N2载体自身的CMV启动子 ,分别构建了P_2.9koval GFP和P_1.5koval GFP两种表达载体 ,经测序和酶切鉴定表达载体构建正确。采用脂质体转染法分别将这两种载体、pGFP N2 (阳性对照 )质粒及阴性对照转染鸡原代输卵管上皮细胞和中国仓鼠卵巢细胞。用荧光倒置显微镜观测绿色荧光蛋白的表达。结果表明 :两种表达质粒在鸡原代输卵管上皮细胞和中国仓鼠卵巢细胞中都可以表达荧光蛋白。结果既显示卵清蛋白第一内含子对基因的表达起到一定的调控作用 ,也显示卵清蛋白启动子对输卵管上皮细胞和卵巢细胞不存在特异性 ,并且不存在种属差异性。     

Oviductal Function during the Annual Ovarian Cycle of a Wild Avian Species,the Tree Pie Dendrocitta vagabunda

The present investigation was carried out on oviductal activity during the annual ovarian cycle of the Indian tree pie (Dendrocitta vagabunda). The oviductal activity was evaluated by weight, length, histology (gross and quantitative) and peroxidase, acid phosphatase, alkaline phosphatase, sialic acid, glycogen, RNA and protein concentrations of different regions of the oviduct. All these values w ere low during the nonbreeding phase (August to January), increased during the progressive phase (February to March), became maximum during breeding (April to May) and decreased in the regression phase (June to July). The functional change of the oviduct is suggested to be due to fluctuation of ovarian steroid activity in this avian species.     

Oviductal Function during the Annual Ovarian Cycle of a Wild Avian Species, the Tree Pie Dendrocitta vagabunda

The present investigation was carried out on oviductal activity during the annual ovarian cycle of the Indian tree pie ( Dendrocitta vagabunda ). The oviductal activity was evaluated by weight, length, histology (gross and quantitative) and peroxidase, acid phosphatase, alkaline phosphatase, sialic acid, glycogen, RNA and protein concentrations of different regions of the oviduct. All these values w ere low during the nonbreeding phase (August to January), increased during the progressive phase (February to March), became maximum during breeding (April to May) and decreased in the regression phase (June to July). The functional change of the oviduct is suggested to be due to fluctuation of ovarian steroid activity in this avian species.     

Identification of muramyl derivatives in Mollusca Gastropoda tissue

Previous findings have demonstrated the presence of muramic acid and the lack of sialic acid in gastropod glycoconjugates from different tissues. The present study investigated the composition of muramyl derivatives in Mollusca Gastropoda tissue from the foot, mantle and periesophageal ganglia, using HRP-labeled lectins (LTA, UEA I, GSA IB4, GSA II, DBA, SBA, RCA II, WGA, PNA, ConA) and glycosidase digestion (neuraminidase, lysozyme, alpha-L-fucosidase, beta-N-acetylglucosaminidase, alpha-N-acetylgalactosaminidase). Muramyl derivatives from the tissue examined showed some differences related to the composition of the terminal disaccharides. Indeed, foot and mantle mucocytes exhibited muramic acid in a terminal position, linked to (subterminal) N-acetylgalactosamine, whereas in neuron cells muramic acid was present in an internal position and linked to N-acetylglucosamine. Diversities also occurred between foot and mantle mucocytes with respect to the receptor sugar for penultimate N-acetylgalactosamine.     

Inhibition of acrosin by oviductal secretory immunoglobulin A

A major inhibitor of acrosin in rhesus monkey and rabbit oviduct fluid, isolated by isoelectrofocusing in sucrose gradients, displayed a broad peak in the acidic region of the column and was demonstrated to contain secretory IgA specific for acrosin. Its identity was established by immunodiffusion, by the removal of acrosin inhibition with antisera to IgA (α-chain), and by its correct molecular weight during ultracentrifugation. Purified human serum IgA also inhibited rabbit, rhesus monkey, and human acrosins, but neither purified human IgG nor IgM had any inhibitory effect on these acrosins. Neither oviduct fluid secretory IgA nor purified human serum IgA inhibited the activity of bovine pancreatic trypsin. The high specificity of secretory IgA for acrosin and its presence in every rabbit and rhesus monkey oviduct fluid specimen examined suggests a possible regulatory role for this antibody in reproduction.     

The mechanism and control of rabbit oviduct fluid formation

The formation of rabbit oviduct fluid was monitored continuously by using an in situ vascular perfusion technique. Oviduct fluid was secreted linearly for at least 3 h at a mean rate of 20.8 +/- 1.5 microliter/h in estrous does. The rate more than doubled on Day 1 following mating, was similar to the value at estrus on Day 2, and dropped to 8.3 microliter on Day 3. Dibutyryl cyclic adenosine 3',5'-monophosphate (cAMP, 1 mM) added to the vascular medium abolished fluid secretion. The same response was obtained, after a lag period, following the addition of cholera toxin (1 mM), forskolin (1 mM), theophylline (1 mM), phorbol dibutyrate (40 microM), A23187 (2 micrograms/ml), 4-acetamido-4'-isothiocyonatostilbene-2,2'-disulphonic acid (SITS, 1 mM), and bumetanide (10 microM) to the vascular medium. N-ethylmaleimide (1 mM), which inhibits adenylate cyclase, stimulated oviduct fluid formation. The transmural potential difference (p.d.) across the oviduct was 5.46 +/- 1.01 mV. This was increased after cAMP addition to 8.7 +/- 1.22 mV. The p.d. in oviducts taken 3 days post-ovulation was 7.6 +/- 1.75 mV, and was increased by cAMP to 12.7 +/- 0.53 mV. Exposure to cholera toxin and forskolin almost doubled the cAMP content of the oviduct. The undirectional flux of chloride ions from the vascular compartment into the lumen was reduced by about 75% after the addition of cAMP, SITS, and bumetanide. A tentative model to account for the formation and regulation of rabbit oviduct fluid in terms of ion fluxes and cAMP and calcium ion concentrations is presented.     

Progesterone binding in rabbit oviduct and uterus.

Progesterone binding of high affinity with a dissociation constant of 10(-9) M was identified in cytosol of rabbit oviduct and uterus. Macromolecules with sedimentation coefficients of 7-8 S and 4-5 S were present. Progesterone receptor concentration was two to fivefold lower in the oviduct when compared with the uterus. The receptor concentration declined steadily from 3 hr until 144 hr after mating in the uterus; however, the decline in oviductal receptor was not significant until the sixth day of pregnancy. Progesterone receptor concentration in rabbit oviduct and uterus in estrus and early pregnancy was greater than estradiol receptor levels.     

Synthesis and secretion of sulphated glycoproteins by rabbit oviduct explants in vitro

The ability of rabbit oviduct explants to incorporate radiolabelled precursors into specific secretory products was investigated. Ampullary and isthmic oviduct segments were cultured in the presence of [3H]glucosamine or [35S]sodium sulphate. Medium samples were analysed for the presence of secreted, labelled macromolecules. Explants incorporated the [3H]glucosamine and secreted labelled glycoproteins in vitro. SDS gel electrophoresis and subsequent fluorographic analysis of culture medium demonstrated a differential secretion of glycoproteins between the ampulla and the isthmus. Although ampullary tissue secreted a greater amount of labelled glycoproteins during the sampling period, the major secretory constituent of Mr approximately 66,000 was common to both oviduct segments. Tissue incubated with [35S]sodium sulphate also secreted a labelled glycoprotein or subunit of Mr approximately 66,000. The results indicate that rabbit oviduct explants are capable of synthesis and secretion of specific sulphated glycoproteins in vitro and that there is a difference in the type and amount of secretion produced between the two oviduct segments.