Estrogen receptor,cyclic adenosine monophosphate,and protein kinase A are involved in the nongenomic pathway by which estradiol accelerates oviductal oocyte transport in cyclic rats

This investigation examined the role of estrogen receptor (ER) on the stimulatory effect of estradiol (E2) on protein phosphorylation in the oviduct as well as on E2-induced acceleration of oviductal oocyte transport in cyclic rats. Estrous rats were injected with E2 s.c. and with the ER antagonist ICI 182 780 intrabursally (i.b.), and 6 h later, oviducts were excised and protein phosphorylation was determined by Western blot analysis. ICI 182 780 inhibited the E2-induced phosphorylation of some oviductal proteins. Other estrous rats were treated with E2 s.c. and ICI 182 780 i.b. The number of eggs in the oviduct, assessed 24 h later, showed that ICI 182 780 blocked the E2-induced egg transport acceleration. The possible involvement of adenylyl cyclase, protein kinase A (PK-A), protein kinase C (PK-C), or tyrosine kinases on egg transport acceleration induced by E2 was then examined. Selective inhibitors of adenylyl cyclase or PK-A inhibited the E2-induced egg transport acceleration, whereas PK-C or tyrosine kinase inhibitors had no effect. Furthermore, forskolin, an adenylyl cyclase activator, mimicked the effect of E2 on ovum transport and E2 increased the level of cAMP in the oviduct of cycling rats. Finally, we measured PK-A activity in vitro in the presence of E2 or E2-ER complex. Activity of PK-A in the presence of E2 or E2-ER was similar to PK-A alone, showing that E2 or E2-ER did not directly activate PK-A. We conclude that the nongenomic pathway by which E2 accelerates oviductal egg transport in the rat requires absolute participation of ER and cAMP and partial participation of PK-A signaling pathways in the oviduct.     

Acceleration of oviductal transport of oocytes induced by estradiol in cycling rats is mediated by nongenomic stimulation of protein phosphorylation in the oviduct

In order to explore nongenomic actions of estradiol (E2) and progesterone (P4) in the oviduct, we determined the effect of E2 and P4 on oviductal protein phosphorylation. Rats on Day 1 of the cycle (C1) or pregnancy (P1) were treated with E2, P4, or E2 + P4, and 0.5 h or 2.5 h later their oviducts were incubated in medium with 32P-orthophosphate for 2 h. Oviducts were homogenized and proteins were separated by SDS-PAGE. Following autoradiography, protein bands were quantitated by densitometry. The phosphorylation of some proteins was increased by hormonal treatments, exhibiting steroid specificity and different individual time courses. Possible mediation of the E2 effect by mRNA synthesis or protein kinases A (PK-A) or C (PK-C) was then examined. Rats on C1 treated with E2 also received an intrabursal (i.b.) injection of alpha-amanitin (Am), or the PK inhibitors H-89 or GF 109203X, and 0.5 h later their oviducts were incubated as above plus the corresponding inhibitors in the medium. Increased incorporation of 32P into total oviductal protein induced by E2 was unchanged by Am, whereas it was completely suppressed by PK inhibitors. Local administration of H-89 was utilized to determine whether or not E2-induced egg transport acceleration requires protein phosphorylation. Rats on C1 or P1 were treated with E2 s.c. and H-89 i.b. The number and distribution of eggs in the genital tract assessed 24 h later showed that H-89 blocked the E2-induced oviductal egg loss in cyclic rats and had no effect in mated rats. It is concluded that E2 and P4 change the pattern of oviductal protein phosphorylation. Estradiol increases oviductal protein phosphorylation in cyclic rats due to a nongenomic action mediated by PK-A and PK-C. In the absence of mating, this action is essential for its oviductal transport accelerating effect. Mating changes the mechanism of action of E2 in the oviduct by waiving this nongenomic action as a requirement for E2-induced embryo transport acceleration.     

Estrogen and progesterone receptors in the oviduct during egg transport in cyclic and pregnant rats

We investigated the temporal relationships between ovum transport and changes in the concentration of nuclear steroid receptors in the oviduct of cyclic and pregnant rats. A lack of parallelism between estrogen and progesterone fluctuations in plasma and their respective nuclear receptor concentrations in the oviduct predominated during egg transport. In pregnant animals, oviductal egg transport took 24 h longer than in nonpregnant animals. In both conditions, transport was initiated while the action of estrogen and progesterone on the oviduct--measured as nuclear receptor accumulation--was decreasing. Three or four days later, depending on whether the animal was pregnant, the eggs entered the uterus shortly after an increase in the nuclear receptor accumulation of both hormones. Treatment with RU486, a progesterone receptor-blocking agent known to cause premature arrival of eggs in the uterus, advanced estrogen receptor accumulation in the oviduct of pregnant rats. These data suggest that the arrival of eggs in the uterus is timed by a transitory increase in nuclear estrogen receptor in the oviduct that does not necessarily reflect a similar change of circulating estradiol. Moreover, in pregnant rats, the onset of this estrogenic action is delayed by a progesterone receptor-mediated effect that hinders nuclear estrogen receptor accumulation.     

Catechol-o-methyltransferase and methoxyestradiols participate in the intraoviductal nongenomic pathway through which estradiol accelerates egg transport in cycling rats

Estradiol (E(2)) accelerates oviductal egg transport through intraoviductal nongenomic pathways in cyclic rats and through genomic pathways in pregnant rats. This shift in pathways, which we have provisionally designated as intracellular path shifting (IPS), is caused by mating-associated signals and represents a novel and hitherto unrecognized phenomenon. The mechanism underlying IPS is currently under investigation. Using microarray analysis, we identified several genes the expression levels of which changed in the rat oviduct within 6 hours of mating. Among these genes, the mRNA level for the enzyme catechol-O-methyltransferase (COMT), which produces methoxyestradiols from hydroxyestradiols, decreased 6-fold, as confirmed by real-time PCR. O-methylation of 2-hydroxyestradiol was up to 4-fold higher in oviductal protein extracts from cyclic rats than from pregnant rats and was blocked by OR486, which is a selective inhibitor of COMT. The levels in the rat oviduct of mRNA and protein for cytochrome P450 isoforms 1A1 and 1B1, which form hydroxyestradiols, were detected by RT-PCR and Western blotting. We explored whether methoxyestradiols participate in the pathways involved in E(2)-accelerated egg transport. Intrabursal application of OR486 prevented E(2) from accelerating egg transport in cyclic rats but not in pregnant rats, whereas 2-methoxyestradiol (2ME) and 4-methoxyestradiol mimicked the effect of E(2) on egg transport in cyclic rats but not in pregnant rats. The effect of 2ME on egg transport was blocked by intrabursal administration of the protein kinase inhibitor H-89 or the antiestrogen ICI 182780, but not by actinomycin D or OR486. We conclude that in the absence of mating, COMT-mediated formation of methoxyestradiols in the oviduct is essential for the nongenomic pathway through which E(2) accelerates egg transport in the rat oviduct. Yet unidentified mating-associated signals, which act directly on oviductal cells, shut down the E(2) nongenomic signaling pathway upstream and downstream of methoxyestradiols. These findings highlight a physiological role for methoxyestradiols in the female genital tract, thereby confirming the occurrence of and providing a partial explanation for the mechanism underlying IPS.     

Physicochemical characterization of progressive changes in the Xenopus laevis egg envelope following oviductal transport and fertilization

Previous studies have shown that the Xenopus laevis egg envelope exists in three forms with differing ultrastructural, macromolecular, and sperm penetrability properties. The coelomic envelope (CE) is derived from eggs released from the ovary into the body cavity of the female, the vitelline envelope (VE) from eggs which have passed through the oviduct, and the fertilization envelope (FE) from fertilized eggs. In the present study, the physicochemical characteristics of these three envelope types were differentiated. Investigation of envelope solubility, deformability, sulfhydryl reactivity, and hydrophobic dye and ferritin binding capacity demonstrated that profound physicochemical changes occur in envelope conversions CE----VE----FE. The physical strength of the envelopes, as evidenced by deformability studies, ranked FE greater than CE greater than VE. These differences were not accountable by differences in the number of disulfide bonds, although the CE sulfhydryl groups were significantly less accessible than those in the VE or FE. All three envelope forms were hydrophilic in nature, exhibiting little ability to bind 1-anilino-8-naphthalenesulfonic acid. The CE bound greater amounts of ferritin in comparison to the VE and FE, indicating the presence of a basic domain, presumably in the 43-kDa glycoprotein, which is lost upon proteolysis to 41 kDa during the CE----VE conversion. The envelope integrity of all three forms was maintained by both noncovalent and covalent (disulfide) bonds. Measurements of the effect of pH on envelope solubilization indicated the involvement of an ionizable group with pKa of 8.0 in maintaining envelope structure.     

Local distributions of oviductal estradiol, progesterone, prostaglandins, oxytocin and endothelin-1 in the cyclic cow

The cyclic patterns of hormones which regulate the activity of the oviduct in the cow have not been adequately reported. We studied progesterone (P4), estradiol 17 beta (E2), prostaglandin E2 (PGE2), prostaglandin F2 alpha (PGF2 alpha), oxytocin (OT) and endothelin-1 (ET-1) concentrations in the cow oviduct. Reproductive tracts from cyclic Holstein cows in the follicular phase (n = 5), post ovulation phase (n = 5) and luteal phase (n = 5) were collected at a slaughterhouse. Oviducts were separated from the uterus, the lumen vas washed with physiological saline, and the enveloping connective tissues were removed. The fimbria was then separated at first and then the rest was divided into 2 parts of equal length (proximal and distal). After extraction, levels of different hormones in the tissues were measured using double antibody enzyme immunoassays (EIAs). There were no differences in any hormone concentration between the 3 parts of the oviduct at any stage of the estrous cycle. The highest concentration of oviductal P4 was observed during the luteal phase and in the oviduct ipsilateral to the functioning CL. Oviductal OT was unchanged throughout the cycle. The highest E2 concentration was observed during the follicular phase in the oviduct ipsilateral to the dominant follicle. The oviduct ipsilateral to the dominant follicle during the follicular phase and ipsilateral to the ovulation site post ovulation showed higher levels of PGE2, PGF2 alpha and ET-1 than those on the contralateral side or during the luteal phase. The highest PGE2 was observed in the oviduct ipsilateral to the ovulation site during the post ovulation phase. The results suggest that the ovarian products (P4, OT and E2) and the local oviductal products (PGE2, PGF2 alpha, and ET-1) may synergistically control oviductal contraction for optimal embryo transport during the periovulatory period, and provide further evidence for the local delivery of ovarian steroids to the adjacent reproductive tract.     

Effects of estradiol and progesterone on body composition, protein synthesis, and lipoprotein lipase in rats

Prior studies suggest that estradiol and progesterone regulate body composition in growing female rats. Because these studies did not consider the confounding effect of changes in food intake, it remains unclear whether ovarian hormones regulate body composition independently of their effects on food intake. We utilized a pair-feeding paradigm to examine the effects of these hormones on body composition. In addition, skeletal muscle protein fractional synthesis rate and adipose tissue lipoprotein lipase activity were measured to examine pathways of substrate deposition into fat and fat-free tissue. Female Sprague-Dawley rats [pubertal: 7-8 wk old; 190 +/- 0.5 (SE) g] were separated into four groups: 1) sham-operated (S; n = 8), 2) ovariectomized plus placebo (OVX; n = 8), 3) ovariectomized plus estradiol (OVX+E; n = 8), and 4) ovariectomized plus progesterone (OVX+P; n = 8). All ovariectomized groups were pair-fed to the S group. Body composition was measured using total body electrical conductivity. The relative increase in fat-free mass was greater (P < 0.01) in the OVX group (31 +/- 2%) than in the S (17 +/- 2%), OVX+E (18 +/- 2%), and OVX+P (22 +/- 2%) groups. The fractional synthetic rates of gastrocnemius muscle protein paralleled changes in fat-free mass: OVX had a higher (P < 0.05) synthesis rate (21 +/- 3%/day) than S (12 +/- 2%/day), OVX+E (11 +/- 2%/day), and OVX+P (8 +/- 1%/day) groups. Body fat increased in the S group (31 +/- 7%; P < 0.01), whereas the OVX groups lost fat (OVX: -10 +/- 7%; OVX+E: -15 +/- 7%; OVX+P: -13 +/- 7%). No differences in lipoprotein lipase were found. Our results suggest that estradiol and progesterone may regulate the growth of fat and fat-free tissues in female rats. Moreover, ovarian hormones may influence skeletal muscle growth through their effects on skeletal muscle protein synthesis.     

Relationship of estradiol and prostaglandins in sperm retention in the mated rabbit

Estradiol-17β (3 μg), administered 1 hr before mating, significantly increased sperm retention in oviducts, uterus, cervices and vagina of does at 2 hr after mating. Indomethacin itself, administered 1.5 hr before mating, reduced the prostaglandin F₂α concentration in the uterus and reduced the number of sperm recovered from the oviducts, cervices and vagina, but indomethacin did not block the estradiol-induced increase in sperm retention in the reproductive tract. Increased sperm retention due to estradiol is probably not mediated solely by prostaglandin F₂α.     

Effects of stress upon plasma estradiol and progesterone levels and the rate of oviductal embryo transport in the rat.

The possibility that changes in sex steroid levels associated with stress could alter the rate of oviductal embryo transport was investigated in the rat. To this end, the effect of cold-swimming and cold-restraint upon estradiol (E2) and progesterone (P) serum levels and embryo transport were assessed. Swimming in water at 16 degrees C for 10 min two or four times between 16:00 and 22:00 h on day 3 of pregnancy caused a modest acceleration of embryo transport that was not associated with decreased fertility. Restraint at 10 degrees C for 2 h between 13:00 and 15:00 h on the first 4 days of pregnancy did not affect embryo transport. Both stimuli increased corticosterone serum levels. Cold-swimming produced a severe hypothermia as compared to cold-restraint and increased serum E2, decreasing significantly the ratio P/E2. Cold-restraint increased the P/E2 ratio. When rats swam in cold water for 10 min twice and were rewarmed by immersion in water at 38 degrees C during 20 min, embryo transport was accelerated despite that no changes occurred in the blood levels of sex steroids. It is concluded that oviductal embryo transport is minimally affected by stress in the rat and that the effect of acute immersion may be independent of alterations in circulating sex steroid levels.     

The effects of glucose and cyclic GMP on RNA synthesis and nuclear morphology in starved rats.

Feeding rats in diet high in glucose has been demonstrated to inhibit the induction of many enzymes, block the action of glucocorticoids, and, in general, appears to result in decreased cyclic AMP activity. We found that glucose feeding depresses both messenger RNA (mRNA) and non-mRNA synthesis. Electron microscopic examination of the nucleus revealed that glucose feeding decreases the granular component of liver cell nucleoli. It only slightly decreases liver cyclic AMP levels, but produces a sixfold elevation in levels of the cyclic AMP antagonist, cyclic GMP. Administration of bromocyclic GMP, like glucose feeding, depresses mRNA synthesis, but does not simulate the effect of the carbohydrate on nuclear morphology. In addition, glucose feeding halves liver inorganic phosphate and triples ATP levels. Phosphorylation of nuclear proteins, however, remains unaltered. Despite the antagonism between glucose feeding and glucocorticoid activity, the former compound did not change the binding of dexamethasone to liver nuclei.     

Axonal protein synthesis and transport

Recent evidence has challenged our ideas about the nature of axonal protein synthesis and transport. Previous metabolic labeling evidence supported the idea that all axonal proteins were synthesized in the cell body and then transported as formed cytoplasmic structures into the axon. Recent evidence suggests that neither the synthesis nor the transport of axonal proteins is that simple. Though most axonal proteins do appear to be synthesized in the neuronal cell body, a small amount of protein appears to be synthesized intra-axonally in some axons. Though small in amount, intra-axonal protein synthesis may be important functionally in some axons. Recent experiments have also begun to identify the presence of a rich array of transport motors in axons, including many members of the kinesin, dynein and myosin families. Progress is being made in identifying which cargoes are being transported by which of these motors. Finally, recent experiments have addressed an old question about whether axoplasmic proteins are transported as filamentous polymers or as soluble components in axons. The answer is that both mechanism can be used in axons. For example, neurofilament protein can move in its particulate or polymeric state, while tubulin can move in its soluble or unpolymerized state.     

Effects of estradiol on renal cyclic guanosine monophosphate and oxidative stress in spontaneously hypertensive rats

Background: Evidence suggests that estradiol offers protection against the development of cardiovascular and renal pathologies, although the mechanisms involved are still under investigation. The nitric oxide (NO) pathway regulates blood pressure and kidney function, and estradiol is associated with increases in NO bioavailability. We hypothesized that in female spontaneously hypertensive rats (SHRs), estra-diol increases NO bioavailability, activates the NO synthase (NOS) pathway, and suppresses superoxide production compared with rats that underwent ovariectomy (OVX).Objective: The goal of this study was to determine whether estradiol regulates the NO/cyclic guanosine monophosphate (cGMP) pathway and superoxide levels in the kidneys of female SHR.Methods: Three types of SHRs were studied: gonad-intact females, OVX rats, and OVX rats with estra-diol replacement (OVX+E). Renal cortical cGMP levels were measured to assess NO bioavailability. NOS enzymatic activity, NOS protein expression, basal superoxide production, and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity were measured in the renal cortex.Results: Fifty-six SHRs were included in the study (17 intact females, 21 OVX rats, 18 OVX+E rats). Mean (SEM) cGMP levels were significantly lower in the renal cortex of OVX rats (0.03 [0.008] pmol/mg, n = 5) than in intact females (0.1 [0.02] pmol/mg, n = 6; P < 0.05), and estradiol restored cGMP levels to those seen in intact females (0.1 [0.01] pmol/mg, n = 5; P < 0.05). Despite a decrease in cGMP following OVX, renal cortical NOS activity, NOS1 and NOS3 protein expression, and the phosphorylation status of NOS3 were comparable among the 3 groups (n = 7–9 per group). However, mean basal superoxide production in the renal cortex was higher in OVX rats (3.2 [0.3] cpm/mg, n = 12) than in intact females (1.9 [0.3] cpm/mg, n = 8; P < 0.05) and lower in OVX+E rats (1.3 [0.3] cpm/mg, n = 9; P < 0.05). Mean NADPH oxidase activity was comparable in the renal cortex of intact females and OVX rats (81 [4] and 83 [12] cpm/35 μg, respectively [n = 5 per group]). OVX+E rats had significantly lower mean renal cortical NADPH oxidase activity than did rats in the other groups (45 [6] cpm/35 μg, n = 6; P < 0.05), and the decrease in activity was accompanied by a decrease in p22^phox protein expression.Conclusions: In vivo manipulations of estradiol levels influenced renal cortical NO bioavailability, as assessed indirectly by cGMP measurements. The decrease in cGMP following OVX was not due to alterations in the activity or expression of NOS.     

The influence of estradiol and diet on small intestinal glucose transport in ovariectomized rats

Although gender differences exist for intestinal absorption of nutrients and drugs, the possible role estradiol may play in modulating nutrient transport has not been established. Therefore, small intestine glucose transport was measured 1 week after administering estradiol to ovariectomized rats fed diets high in carbohydrate (C) or protein (P). Rats treated with estradiol ate 21% less (P<0.05) and lost body mass (7%; P<0.05) but did not have smaller intestines. Administration of estradiol increased rates of glucose transport, but only when the rats were fed the C diet. These findings indicate that estradiol causes a disconnect between food intake and the dimensions and nutrient transport capacities of the small intestine. Furthermore, the responses to estradiol are influenced by diet composition, are not of the same magnitude for rats and dogs, and can be predicted to affect systemic availability of nutrients and drugs.     

Changes in estrone and estradiol synthesis in fetal and newborn rats

The question was approached whether estradiol synthesis by the rat ovary gained in importance with age relatively to estrone synthesis, which predominated largely in the 20-day-old fetus. Three stages were investigated, i.e. fetal stage 21 days and stages 2 and 7 days after birth. Ovaries were cultured in vitro in the presence of various radioactive androgens, and the conversion percentages into estrone (E1) and estradiol (E2) were determined by double isotopic dilution combined with recrystallization to constant specific activity. Insignificant in the 21-day-old fetus (E1/E2 ratio = 40), estradiol synthesis increased relatively to estrone synthesis in the 2-day-old neonate and still more at the stage of 7 days (E1/E2 ratio = 3). FSH had no effect on estrogen synthesis at the 3 stages investigated.     

Tissue-specific effects of chronic dietary leucine and norleucine supplementation on protein synthesis in rats

Acute administration of leucine and norleucine activates the mammalian target of rapamycin (mTOR) cell-signaling pathway and increases rates of protein synthesis in a number of tissues in fasted rats. Although persistent stimulation of mTOR signaling is thought to increase protein synthetic capacity, little information is available concerning the effects of chronic administration of these agonists on protein synthesis, mTOR signal transduction, or leucine metabolism. Hence, we developed a model of chronic leucine/norleucine supplementation via drinking water and examined the effects of chronic (12 days) supplementation on protein synthesis in adipose tissue, kidney, heart, liver, and skeletal muscle from ad libitum-fed rats. The relative concentration of proteins involved in mTOR signaling and the two initial steps in leucine oxidation were also examined. Leucine or norleucine supplementation was accompanied by increased rates of protein synthesis in adipose tissue, liver, and skeletal muscle, but not in heart or kidney. Supplementation was not associated with increases in the anabolic hormones insulin or insulin-like growth factor I. Chronic supplementation did not cause apparent adaptation in either components of the mTOR cell-signaling pathway that respond to leucine (mTOR, ribosomal protein S6 kinase, and eukaryotic initiation factor 4E-binding protein-1) or the first two steps in leucine metabolism (the mitochondrial isoform of branched-chain amino acid transaminase, branched-chain keto acid dehydrogenase, and branched-chain keto acid dehydrogenase kinase), which may be involved in terminating the signal from leucine. These results suggest that provision of leucine or norleucine supplementation via the drinking water results in stimulation of postprandial protein synthesis in adipose tissue, skeletal muscle, and liver without notable adaptive changes in signaling proteins or metabolic enzymes.     

The role of oligosaccharide in transport of egg yolk riboflavin-binding protein to the egg

The carbohydrate portion of chicken egg yolk riboflavin-binding protein was examined to determine its role in the biological activity of the protein. Yolk RBP was found to contain 5–6 mannose, five galactose, 12 N-acetylglucosamine and four sialic acid residues. Specific modifications of the oligosaccharide moiety were performed which included removal of sialic acid by mild acid hydrolysis, oxidation of galactose oxidase, and removal of N-acetylglucosamine and galactose residues by a mixture of glycosidases from Aspergillus niger. All of the modified proteins retained the ability to bind riboflavin although their capacities were lower than that of native yolk RBP. Circular dichroism of the modified yolk RBP samples showed changes in the near ultraviolet, but molar ellipticities in the far ultraviolet displayed only minor variations indicating no gross structural changes. All samples cross-reacted with RBP-specific antiserum. The plasma half-life of ¹²⁵I-labeled yolk RBP was 62 min. Each of the modified samples was cleared more rapidly from the blood than native yolk RBP. Removal of sialic acid decreased the half-life of yolk RBP by 31%, while the other modifications decreased the half-life by as much as 60%. During a 10-day period following injection of ¹²⁵I-labeled yolk RBP, 5.9% of the labeled protein was recovered from egg yolk. Relative to native yolk RBP, the transport of asialo-yolk RBP was decreased by 82%. The other modifications resulted in even less transport to the egg, the lowest being glycosidase-treated asialo-yolk RBP which was decreased by over 99%. By comparison of samples with similar clearance times, a positive correlation was made between sialic acid and ovarian transport.     

De novo synthesis and release of polypeptides from cyclic and early pregnant porcine oviductal tissue in explant culture

The objective of the present study was to identify and characterize in a limited manner the major de novo oviductal secretory proteins (OSP) synthesized and released by the porcine oviduct. Oviductal tissue was collected on various days of the estrous cycle (EC) and early pregnancy (EP) and cultured in a modified minimal essential medium supplemented with 100 muCi L-[3H]-leucine. Oviductal secretory activity, as measured by the rate of incorporation of 3H-leucine (dpm/mg wet tissue weight) into nondialyzable macromolecules, was greatest (P less than .01) between days 0 and 2 and reached its lowest levels on days 10 to 15. There was no difference between left and right side or pregnancy status. This increased rate of incorporation at proestrus and estrus is temporally associated with elevated levels of estrogen. Incorporation rate for ampulla was greater than for the isthmus. Analysis of oviductal culture medium by one-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis and fluorography revealed three protein bands of relative molecular weight (Mr) 335,000, 115,000, and 85,000, which were associated with proestrus, estrus, and metestrus and were not detectable on other days. All three proteins also incorporated 3H-glucosamine. The 115,000 Mr band was the major 3H-glucosamine-labeled protein. Two protein bands (Mr 60,000 and 20,000) were expressed with increasing progesterone during diestrus. Other de novo synthesized protein bands appear to be present throughout the EC and EP with little modulation by estrogen or progesterone. Thus, this study demonstrates that for the porcine oviduct, the increase in the incorporation rate of 3H-leucine into OSP by both whole oviduct and ampulla and de novo synthesis and secretion of three glycoproteins, Mr 335,000, 115,000, and 85,000, were associated with proestrus and estrus when events such as fertilization and early cleavage stages of embryo development occurred.