Acrosome reaction in bovine spermatozoa: role of reactive oxygen species and lactate dehydrogenase C4

After capacitation, mammalian spermatozoa accomplish the acrosome reaction (AR), a well-controlled exocytosis process crucial to fertilize mature oocytes that involves several protein kinases such as protein kinase A (PKA), C (PKC), and tyrosine kinase (PTK). Reactive oxygen species (ROS) are involved in both bovine sperm capacitation and AR. Lactate dehydrogenase C4 (LDH-C4) was associated with bovine and mouse sperm capacitation. Our aims were to study the participation of LDH-C4 to contribute with the status redox required for AR and the role of ROS in the regulation of PKA, PKC, and PTK involved in the exocytotic event. Sodium oxamate, an inhibitor of LDH-C4, prevented the AR induced by lysophosphatidylcholine (LPC) or NADH. Hydrogen peroxide promoted and superoxide dismutase (scavenger of superoxide), catalase (scavenger of hydrogen peroxide), diphenyleneiodinum, diphenyliodonium, cibacron blue, and lapachol (inhibitors of NADPH oxidase) prevented the AR, suggesting that ROS and a sperm oxidase are involved in the AR induced by these compounds. Inhibitors of PKA, PKC, and PTK also prevented the AR induced by LPC or NADH, suggesting the involvement of these kinases in the process. These results suggest that LDH-C4 may participate in the regulation of the redox status required to achieve the AR in bovine spermatozoa and that ROS are key elements in the regulation of protein kinases associated with the AR process.     

Acrosome reaction of boar spermatozoa in homologous in vitro fertilization

Ejaculated spermatozoa from four different boars were used to evaluate the acrosome reaction during in vitro fertilization with homologous ovulated oocytes. The acrosome reaction was assessed according to a peroxidase-labeling peanut agglutinin method and a triple-stain technique. An increase in the proportion of living sperm with reacted acrosomes was observed after preincubation and 2 hr of coincubation (P < 0.05). The percentage of true acrosome-reacted sperm remained reasonably constant throughout coincubation. In vitro penetration rates of oocytes varied among boars, but no relationship was found between fertilization rates of oocytes and maximum percentages of acrosome reacted living sperm. © 1993 Wiley-Liss, Inc.     

Acrosome reaction in spermatozoa of the hagfish Eptatretus burgeri (Agnatha)

Fertilization of the hagfish or myxiniformes, a member of the most primitive vertebrate group and an animal of phylogenic interest, is unknown. Here, induction of an acrosome reaction for spermatozoa in the hagfish, Eptatretus burgeri, was successfully achieved by treatment of mature spermatozoa with ionomycin and excess Ca2+. The spermatozoon produced an acrosomal process that elongated from the apex of the long sperm head. The reaction bears resemblance to that of invertebrate spermatozoa rather than that of vertebrate spermatozoa. The result provides insights into the phylogenetical changes that have occurred in this sperm reaction.     

Developmental competence of bovine embryos derived from oocytes collected at various stages of the estrous cycle

The developmental competence of bovine oocytes collected from donors at various stages of the estrous cycle and fertilized in vitro was investigated by comparing the yields of embryos obtained from oocytes isolated from the ovaries of cows slaughtered on estrous cycle Days 7 and 14, 8 and 15, 9 and 16 and on Days 19, 20 and 2. The percentages of oocytes that developed into blastocysts at Day 8 after exposure to spermatozoa were: 11.9 vs 20.0; 13.2 vs 30.5; 20.8 vs 29.8; and 11.7, 4.4 and 16.9, respectively. A significantly higher proportion of oocytes developed into blastocysts following isolation on cycle Days 14 to 16 (24.3 %) than following recovery on Days 7 to 9 (13.0 %; P < 0.05), Days 19 to 20 (6.6 %; P < 0.05) or Day 2 (16.9 %; P < 0.05). Embryo development was also faster in oocytes isolated at the end of the luteal phase (Days 14 to 16). These results demonstrate that the stage of the estrous cycle may influence the developmental potential of oocytes and in vitro embryo production.     

Histological changes in bovine endometrium during the estrous cycle

Endometrial biopsy specimens were obtained from 46 normally cyclic heifers at known stages of their estrous period to show precise characteristic changes. These tissues were embedded in paraffin, sectioned, and stained with hematoxylin and eosin. The following histological changes were observed during the estrous cycle. Metrorrhagia was observed on Days 0 to 1 (estrus = Day 0). Mitoses in glandular epithelium occurred on Day 5. Basal vacuolation in the surface epithelium occurred on Days 5 to 6. Leukocyte invaded the functional layer on Day 7. Stromal mitoses were observed on Days 9 to 12. The results indicate that clincians need to be aware that histological evaluation is important for the diagnosis of endometrial function and that biopsy is useful for this purpose.     

Validation of a mathematical model of the bovine estrous cycle for cows with different estrous cycle characteristics

A recently developed mechanistic mathematical model of the bovine estrous cycle was parameterized to fit empirical data sets collected during one estrous cycle of 31 individual cows, with the main objective to further validate the model. The a priori criteria for validation were (1) the resulting model can simulate the measured data correctly (i.e. goodness of fit), and (2) this is achieved without needing extreme, probably non-physiological parameter values. We used a least squares optimization procedure to identify parameter configurations for the mathematical model to fit the empirical in vivo measurements of follicle and corpus luteum sizes, and the plasma concentrations of progesterone, estradiol, FSH and LH for each cow. The model was capable of accommodating normal variation in estrous cycle characteristics of individual cows. With the parameter sets estimated for the individual cows, the model behavior changed for 21 cows, with improved fit of the simulated output curves for 18 of these 21 cows. Moreover, the number of follicular waves was predicted correctly for 18 of the 25 two-wave and three-wave cows, without extreme parameter value changes. Estimation of specific parameters confirmed results of previous model simulations indicating that parameters involved in luteolytic signaling are very important for regulation of general estrous cycle characteristics, and are likely responsible for differences in estrous cycle characteristics between cows.     

Estrogen and progesterone concentrations in bovine milk during the estrous cycle

Estrogen and progesterone concentrations in milk during the estrous cycle were estimated in 18 normally cycling Holstein dairy cows. The estrogen and progesterone concentrations in milk during the estrous cycle followed the pattern described for them in blood in the corresponding period. During most of the estrous cycle, estrogen concentration remained at approximately 200 pg/ml and reached a proestrous peak of 360 +/- 127 pg/ml on day 19. The progesterone concentration in milk during the estrous cycle increased to a peak on day 13 (45.5 +/- 6.6 ng/ml) and thereafter declined towards estrus. Estrus detection/prediction based on milk progesterone concentrations appears feasible in view of the significant differences in milk progesterone concentrations between the early luteal (post-ovulatory), luteal and rapid follicular growth periods of the estrous cycle.     

Acrosome reaction of stallion spermatozoa evaluated with monoclonal antibody and zona-free hamster eggs

The acrosome of the stallion spermatozoon was visualized by indirect immunofluorescence with monoclonal antibody (18.6) which recognized an integral acrosomal membrane component. Localization was confirmed by electron microscopy using peroxidase labelled antibody. In fresh semen samples (n = 19), 73.9 +/- 9.1% of the spermatozoa from five fertile stallions displayed a uniform bright fluorescence over their acrosome region. In two semen samples from an infertile stallion only 28% and 35% of spermatozoa showed the same pattern of fluorescence. Spermatozoa from fertile stallions incubated for up to 12 hours in TALP medium maintained motility and exhibited a significant progressive loss of acrosomes as detected by immunofluorescence. Alternatively, a similar loss of acrosomes could be induced with calcium ionophore A23187 over a 90 minute incubation. Ultrastructural observations and incubation with zona-free hamster eggs indicated that only with ionophore treatment was immunofluorescent acrosome loss correlated with a physiological acrosome reaction, while prolonged sperm incubation led to degenerative membrane changes. It was concluded that, if carefully validated, immunofluorescent localization of the acrosome of stallion sperm with monoclonal antibody could be used to monitor the acrosome reaction. Furthermore, definitive acrosome visualization would be valuable in assessing semen quality.     

Acrosome reaction in spermatozoa from the amphioxus Branchiostoma belcheri (Cephalochordata, Chordata)

The formation of an acrosomal process at acrosomal exocytosis in spermatozoa of the amphioxus was described in the present report for the first time. A non-reacted acrosome was located in front of the nucleus, where a cup-shaped acrosomal vesicle covered a conical accumulation of subacrosomal material. When naturally spawned spermatozoa were treated with a calcium ionophore, ionomycin, the acrosomal vesicle opened at the apex and an acrosomal process was projected. The process exhibited a filamentous structure. The reaction followed the mode typically seen in marine invertebrates. These observations suggest that the features and function of the acrosome of amphioxus, whose position is on the border between invertebrates and vertebrates, reflect their ecological adaptation and phylogenic position.     

The spermatozoa of ascidians: Acrosome and nuclear envelope

Summary The spermatozoon of Ascidia callosa has a head with a wedge-shaped tip. Between the nuclear envelope and the plasmalemma, at the tip of the head, there are one or two previously undescribed vesicles, 45 to 55 nm in diameter. These vesicles have the characteristics of an acrosome. Their role in the process of fertilization has not been determined. Ultrastructural studies of sperm activation are needed, but claims that the spermatozoa of ascidians do not have an acrosome should be reconsidered.Behind the tip of the sperm there are pores in the nuclear envelope. This part of the envelope also contains a dense band of amorphous material that may have a supportive function. A nearly identical structure, associated with pores has been found in the spermatozoon of Boltenia villosa. An analysis of the nuclear envelope of Ascidia callosa indicates that the same structure has previously been misinterpreted as an acrosome in the spermatozoon of Ascidia nigra.     

Effect of repeated laparoscopic surgery on the bovine estrous cycle

The effects of repeated laparoscopic surgery on the length of the bovine estrous cycle, estrus, ovulation and corpus luteum function were determined after one estrous cycle of normal duration (18 to 24 days). Five, Angus x Hereford cows were subjected to laparoscopy on days 5, 13, 18 and 20 (estrus = day 0) of the subsequent cycle. Blood was collected daily during the cycle in which laparoscopy was performed (surgical cycle) and during the next cycle (postsurgical cycle). Lengths of the surgical and postsurgical cycles (22.3 +/- .5 days and 21.5 +/- .6 days, respectively) did not differ (P>.05) from that of the presurgical cycle (21.8 +/- .2 days). Average concentrations (ng/ml) of LH and progesterone in serum were similar during the surgical and postsurgical cycles (1.2 +/- .1, 2.2 +/- .2 vs 1.3 +/- .2 and 2.3 +/- .1). Progesterone concentrations remained above 1 ng/ml for 17 and 16 days during the surgical and postsurgical cycles, respectively. A pre-ovulatory rise in LH, along with estrus and ovulation was confirmed in all animals. Follicular development, characterized by follicular volume, increased progressively from days 5 to 20, with the largest increase occurring between days 13 and 18. These results indicate that laparoscopy, used at the times and frequency specified, does not alter reproductive function of cyclic cows and can provide information on ovarian activity.     

The apparent in vivo affinity of estradiol binding sites in the uterus varies during the estrous cycle of the rat

An estradiol-exchange assay system was used to determine the amounts of different estradiol receptors in the uterus of rats exhibiting a regular, four day estrous cycle. The total number of all receptors increased from a low at estrus and metestrus to a high at proestrus, before falling sharply between proestrus and estrus. These variations have been explained by alterations in the amounts of estradiol and progesterone in the serum. Similar variations were observed in the classes of free and occupied receptors; the latter varied more dramatically than the former. The changes in the amounts of free and occupied receptors could be explained by variations both in the estradiol levels in the serum and in the total number of receptors in the uterus. However, the amount of occupied receptors did not increase as much from diestrus to proestrus as expected from the increase in the estradiol levels in the serum, indicating that the apparent in vivo affinity of the receptors for estradiol was greater at diestrus than at proestrus. This difference could not be explained in terms of (i) receptor stability during the exchange assay, or (ii) of estradiol retention capacity of the serum, or (iii) of limited uptake of estradiol into the uterine cell. It is concluded that some differences must exist in the function or structure of the receptors at diestrus and proestrus.     

A quantitative method for assessing stages of the rat estrous cycle

The impact of gender and/or hormone variations on a wide variety of neural functions makes the choice between studying males or females (or both) of a given species difficult. Although female rats are widely used experimentally, few studies control for the stage of estrus. More detailed information about how to distinguish the various stages of the estrous cycle is needed. For the present study, vaginal smears were obtained once a day and stained using an adaptation of the Papanicolaou (PAP) procedure. Images are provided of unstained “wet” samples and the corresponding PAP stained smears illustrating the cellular profile for each stage of the cycle as well as post-ovariectomy. The different cell populations across the cycle were quantified and ratios determined to show trends between the predominant and other cell types in each stage of the estrous cycle. Both stained and unstained images and cell quantification data provide valuable guidelines for distinguishing the stages of the estrous cycle.     

A quantitative method for assessing stages of the rat estrous cycle

The impact of gender and/or hormone variations on a wide variety of neural functions makes the choice between studying males or females (or both) of a given species difficult. Although female rats are widely used experimentally, few studies control for the stage of estrus. More detailed information about how to distinguish the various stages of the estrous cycle is needed. For the present study, vaginal smears were obtained once a day and stained using an adaptation of the Papanicolaou (PAP) procedure. Images are provided of unstained “wet” samples and the corresponding PAP stained smears illustrating the cellular profile for each stage of the cycle as well as post-ovariectomy. The different cell populations across the cycle were quantified and ratios determined to show trends between the predominant and other cell types in each stage of the estrous cycle. Both stained and unstained images and cell quantification data provide valuable guidelines for distinguishing the stages of the estrous cycle.     

Expression of superoxide dismutases in the bovine oviduct during the estrous cycle

The superoxide dismutases (SODs) are first-line enzymatic antioxidants that dismute superoxide anion (O(2)(-)) to produce hydrogen peroxide (H(2)O(2)). The primary objective was to characterize, by western blot analysis, the expression of two SODs, the cytosolic (Cu,ZnSOD or SOD1) and the mitochondrial (MnSOD or SOD2) forms in three sections of the oviduct, i.e. isthmus (I), ishtmic-ampullary junction (IA), and ampulla (A), during the estrous cycle. The Cu,ZnSOD and MnSOD proteins were mostly expressed in the ampulla (I相似文献   

Morphological characteristics of the bovine corpus luteum during the estrous cycle and pregnancy

The corpus luteum, one of the biological clocks of the estrous cycle and pregnancy, is known foremost for its production of progesterone that blocks the pituitary release of gonadotropins and prepares the uterus for a pregnancy. The cellular sources of this progesterone are the steroidogenic small and large luteal cells. Other luteal cells that are not steroidogenic, but are believed to have an important role in the function of this gland are the fibroblast, macrophages and endothelial cells. The most prominent luteal cell is the large steroidogenic cell characterized by an abundance of smooth endoplasmic reticulum and densely packed spherical mitochondria that are indicative of its contribution to most of the circulating progesterone believed to be constitutively secreted and not under the control of LH. Other distinguishing features of the large luteal cell are the presence of rough endoplasmic reticulum, prominent Golgi, and secretory granules that are indicative of endocrine cells. This cell undergoes dynamic changes across the estrous cycle and pregnancy, believed to reflect a change in progesterone and protein secretion that will eventually influence a successful pregnancy or another ovulation if pregnancy fails. The morphological characteristics of the bovine luteal cells are the focus of this review.     

Distribution of flehmen reactions of the bull throughout the bovine estrous cycle

A bull's flehmen reactions were recorded daily during 20-minute contact with 3 cows for nine months. During estrous periods (?3 to +1 days) there were 174 observations: 28.2% without flehmen, 21.2% with a single reaction, and 50.6% with repeated reactions. During nonestrous periods there were 624 observations: 49.0% without flehmen, 40.4% with a single reaction, and 10.6% with repeated reactions. Average numbers of all and repeated reactions during estrous periods were 2.11±2.30 and 1.90±2.43 respectively. Corresponding values for nonestrous periods, 0.70±0.93 and 0.29±0.93, were significantly lower (P<0.01).The results suggest that either the estrous pheromone(s) also occur(s) for a limited time or in limited amounts during nonestrus or that flehmen is not a very specific response to the estrus-related odors. In either case, the study of sexual pheromones should encompass other methods as well as flehmen observation.