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Parathionis an organophosphorate pesticide amply used in agriculture. Many alterations induced by organophosphorate pesticides have been described, such as: cytogenetic alterations in germinal cells, oligozoospermia and teratozoospermia in the mouse. The effect of Parathion¤, both pure (PP) and commercial (PC), on mouse interstitial cell testosterone production was evaluated in vivo and in vitro. Male mice were intraperitoneally injected with a single dose of 1/3 LD50 of Parathion¤, both PP and PC. The animals were sacrificed at 1, 8 and 40 days post injection to evaluate the impact of disrupting testosterone production on spermatogonia, spermatocytes and elongated spermatids. The plasma testosterone was assayed by standard radioimmunoanalysis. The same method was used to assay testosterone in the culture medium of interstitial cells obtained from the control and Parathiontreated animals at the same time intervals. Sperm count, sperm teratozoospermia and tubular blockage were analyzed for an appraisal of spermatogenesis. Increase in the teratozoospermia and tubular blockage was detected in the PP and PC group at 8 and 40 days post injection. Plasma testosterone levels drop significantly at 8 days and recovered slowly at 40 days only in PP animals as detected in vivo, implying interference of testicular steroidogenesis due to the toxicant. Recuperation of normality occurs at long time intervals. In conclusion, Parathiondisturbs the synthesis of testosterone in mice affecting qualitatively the spermatogenesis

Most studies in alpaca reproductive biology have been focused on female physiology. Only recent research is being conducted in order to increase the knowledge on males. Semen characteristics during breeding periods will contribute to understanding the poor fertility rates in alpaca. Ten adult male alpacas were distributed randomly into two groups and submitted alternatively to two regimens of semen collection of 12 days duration (day 1, initial day of semen collection). Semen samples were collected using an artificial vagina and a receptive, non-pregnant female. With regimen 1, males were maintained with females except for the days of sexual rest (6 and 7). Semen was collected on days 1, 5, 8 and 12. With regimen 2, males were exposed to females for daily semen collection only, before and after sexual rest. Mating duration, color and volume of ejaculates, spermatozoa concentration and morphology were evaluated. No statistical differences for the variables were found between regimens that were used for semen collection. With respect to influence of day, however, the total numbers of spermatozoa ejaculated on days 1 and 5 of semen collection were statistically different (p0.05). Azoospermic samples increased on days 5 and 12 of semen collection. Partial recovery in spermatozoa concentration and number of spermatozoa ejaculated were observed after sexual rest. Although normal spermatozoa percentage was less on day 1 (p0.05) as compared with values found in the following ejaculates (days 5 and 12), the total number of normal spermatozoa was greater. These results support the conclusion that when male alpaca have a daily ejaculation during five consecutive days, they might copulate without having enough spermatozoa for fertilization towards the end of the mating period.

Pregnancies at high-altitudes are influenced by hypoxia and oxidative stress and frequently affected by IUGR. However, a common thought is that early pregnant women visiting altitude have no major complications for gestation development, since IUGR is developed during the second half of pregnancy. Thus, using a well-characterized sheep-model, we aimed to determine whether long-and/or short-term exposure to high-altitude may affect maternal steroidogenesis and therefore embryo-fetal growth from conception. The second aim was to differentiate the relative role of hypoxia and oxidative stress by assessing the effects of supplementation with antioxidant agents during this early-pregnancy stage, which were previously found to be useful to prevent IUGR. The results indicate that both long- and shortterm exposure to high-altitude causes disturbances in maternal ovarian steroidogenesis and negatively affects embryo-fetal growth already during the very early stages of gestation, with the consequences being even worsened in newcomers to high-altitude. The supply of antioxidant during this period only showed discrete effects for preventing IUGR. In conclusion, the present study gives a warning for clinicians about the risks for early-pregnant women when visiting high-altitude regions and suggests the need for further studies on the effects of the length of exposure and on the interaction of the exposure with the pregnancy stage. © 2015 Parraguez et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Increasing the knowledge of the semen characteristics in the alpaca will contribute to understanding one of the many factors that affect the poor fertility rate in this species. Ten adult male alpacas, 2.6-10 years of age, average weight 64.7 +/- 4.7 kg were used. The animals were distributed randomly into two groups of five each and submitted alternatively to two semen collections, using an artificial vagina and sexually receptive females. For the first semen collection the animals had a sexual rest period of about 90 and 45 days before the second. Duration of semen collection, color and volume of ejaculate were recorded, and sperm concentration and morphology (light microscopy) were evaluated. Descriptive statistical analyses were used for each variable, considering all samples obtained (n= 19). An analysis of variance for animal groups and opportunity of collection were used for quantitative variables. Most frequent color was opalescent white (84.2%). There were no statistical differences among male groups or between semen collections. The average values and standard deviations for the quantitative variables were: 12.3 +/- 7.2 min for semen collection time, 1.8 +/- 0.8 ml for ejaculate volume, (17.6 +/- 26.1) x 10(6) sperm/ml for sperm concentration and 34.0 +/- 52.2 x 10(6) for total number of sperm per ejaculate. The percentage of normal spermatozoa was 51.0 +/- 12.4%. From the total abnormalities, that of mid piece segment (14.4%) was the most frequent. These results indicate that male alpaca have poor semen quality, when compared with other domestic species. Nevertheless, for the evaluation of male alpaca as breeders it would be necessary to create a protocol for the selection of them, where phenotypic, behavioral and seminogram aspects are considered. The values reported herein define the characteristics of the alpaca semen that could be considered as the initial base of the seminal analysis to select male alpacas before mating.

This study evaluated ovarian activity in late gestation and post-partum in guanacos in captivity. Follicular dynamics was monitored every second day from 40 days before and other 40 after delivery by transrectal sonography and by plasma steroids concentrations. Seven out of eight (87.5%) of gestating females presented ovarian follicular activity under progesterone levels >3 nmol/l with maximum follicular size of 8.42 ± 0.83 mm from days 23 to 1 before delivery. After delivery, all females have follicular wave development from day 0 to 38, with larger follicular size and longer follicular wave phases and interwave interval when compared with pre-partum data. During post-partum period, there was a close relationship between follicle size and estradiol-17β concentration, with r = 0.69 at the beginning of growth phase and r = 0.86 in association with the largest dominant follicle. Plasma estradiol-17β concentration varied from 11.92 to 198.55 pmol/l. Plasma estrone sulfate, free estrone and progesterone returned to baseline concentrations during peripartal period and remained basal thereafter. The results described follicular activity during late gestation and early post-partum period. These findings provide relevant information to understand physiological changes occurring during this reproductive key period in seasonal breeders with long gestation duration as New and Old World camelids.

The purpose of the present study was to investigate the pattern of distribution of cytokeratins, vimentin and muscular actin in the testis of vicuna (Vicugna vicugna) and llama (Lama glama) two species of camelids native of the Andean high plateau of South America. Testicular biopsies of four vicunas and five llamas were used. Animals were healthy breeders. The tissues were processed by standard immunohistochemistry with antipancytokeratinAE1/AE3, antikeratin 18 (K 18), CAM 5.2 (antikeratin 5, 18, and 19), antivimentin, and smooth-muscle-specific antiactin antibodies to track the cytoskeletal pattern of testicular cells. Using AE1/AE3 antibody the immunostaining was found in the epithelial lining of tubuli recti and rete testis. The reaction was relatively stronger in the apical cytoplasm of epithelial cells. The testicular cells of the two species showed no reaction to K 18 and CAM 5.2 antibodies. Antivimentin antibody stained the basal cytoplasm of the Sertoli cells, the Leydig cells, and the epithelial lining of tubuli recti and rete testis. In the last two structures the immunostain was relatively more intense in the basal cytoplasm of epithelial cells. Antiactin antibody stained the peritubular cells and the muscle cells of the lamina propria oftubuli recti and rete testis. The presence in these species of only some keratins found in man, its coexpression with vimentin in epithelial lining of tubuli recti and rete testis and the peritubule organization, so different from other ungulates may reflect a differential adaptation of the cytoskeleton to particular reproductive strategies.

Plasma progesterone concentration during pregnancy and the early postpartum period was measured by radioimmunoassay in five alpacas (Lama pacos), of the Huacaya breed, whose pregnancy length had a mean of 344.8 +/- 4.4 days. Concentration of progesterone increased (P0.001) from low premating values (0.11 +/- 0.06 nmol/l) to greater values at 30 days of pregnancy (8.05 +/- 1.13 nmol/l) and remained high up to 2 months of pregnancy. A slight transitory decline was observed between 3 to 7 months. Concentration of plasma progesterone dropped markedly during the 72 h before parturition especially, at the day of parturition.

At high altitude, hypoxia and/or oxidative stress may compromise fertility. This study tested the relative effect of short- or long-term exposure to high-altitude hypobaric hypoxia and oxidative stress in sheep on preovulatory follicle dynamics and gonadotrophin secretion. Thus, growth dynamics, stereidogenic function and competence to ovulate of preovulatory follicles, as well as FSH and LH availability throughout the entire oestrous cycle, were compared among sheep native from low and high altitude, and sheep newcomers to high altitude. The results indicates that short-term exposure in sheep newcomers to high altitude has a deleterious effect on both the ovarian function (affecting preovulatory follicular development) and the pituitary function (diminishing plasma LH availability). On the other hand, there were no detected differences in the preovulatory follicular development in sheep adapted to high altitude for generations and, conversely, LH secretion was increased, which suggests an adaptive mechanism. The treatment with antioxidant agents during a relative short period for the time of folliculogenesis (approximately 1 month and a half) changed substantially the development of preovulatory follicles in short-term exposed sheep to similar patterns than in sheep native and living to both high and low altitude. These results highlight the role of oxidative stress in the detriment of the reproductive function in individuals recently exposed to high-altitude hypoxic environment. © 2014 Blackwell Verlag GmbH.

Sheep pregnancy in high-altitude environments frequently involves hypoxia and oxidative stress and causes intrauterine growth retardation. The adverse effects of altitude on fetal growth can be prevented by the administration of antioxidant vitamins, but the mechanisms responsible are not well known. The maintenance of a viable pregnancy depends largely on adequate placental steroidogenesis, especially in the last two-thirds of pregnancy. Thus, in the present study we evaluated the effect of antioxidant vitamins (C and E) on plasma concentrations of progesterone and 17β-oestradiol during the last two-thirds of high-altitude pregnancies in ewes both native and naïve to the high-altitude environment. In addition, pregnancy outcomes were evaluated by determining the bodyweight of newborn lambs. Sex steroid patterns differed between ewes with and without vitamin supplementation. Concentrations of plasma progesterone and 17β-oestradiol were significantly higher in the supplemented groups from approximately 40 days before parturition until near term. Newborn weights were significantly lower in animals not adapted to the higher altitude, and vitamin supplementation prevented this decrease. In conclusion, the administration of antioxidant vitamins in the present study enhanced placental steroidogenesis, thus favouring fetal development in pregnancies developing at high altitudes.

The objective of the present study was to describe ovarian activity in 11 adult non-mated guanacos, evaluated every second day for 40 days by transrectal ultrasonography and by plasma estradiol-17beta and progesterone concentrations. An inverse relationship (r = -0.29, P0.001) was observed between the diameter of the largest ovarian follicle and the total number of follicles indicating that follicular growth in guanacos occurs in waves. The mean duration of follicular wave was 15.1 +/- 4.2 days with a range from 9 to 26. The follicular growth phase was 7.0 +/- 2.4 days, the static phase 3.0 +/- 1.2 days, the regression phase 5.2 +/- 2.1 days and the inter-wave interval was 12.6 +/- 5.6 days. The maximum follicular diameter in each follicular wave was 10.2 +/- 2.1 mm with a range from 7.2 to 16.1 mm. Inter-wave intervals of longer duration were associated with a larger maximum follicle diameter (P0.001). Follicular activity alternated between ovaries in 93% of the waves with 48% of dominant follicles occurring in the left and 52% in the right ovary without differences (P0.05). Plasma estradiol-17beta concentrations showed a wave-like pattern, varying from 20.0 to 92.1 pmol/L. Plasma progesterone concentrations remained below 1 nmol/L without any ultrasonic evidence of ovulation during the study. These results in guanacos suggest a follicular wave pattern more similar to the llama (Lama glama) than previously described in other South American and Old World camelid species.

Fetal weight and the placenta of sheep at high altitude (HA) are affected by hypoxia. Placental changes (an increase in placental size and vascularization) are greater in ewes from populations that have lived for several generations at HA than in those exposed during just 1 gestation. This study investigated placental expression of vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS), 2 molecules involved in placental angiogenesis that could be upregulated by hypoxia. Two groups of ewes were maintained at HA (3589 m) during pregnancy: HA-native ewes (group HH) and ewes native to lowlands but moved to HA immediately after the diagnosis of pregnancy (group LH). A control group (LL) was kept at sea level. Near term, placentomes were removed, weighed, and processed for immunohistochemical detection of VEGF and eNOS, as well as for vascular area measurement. Placental weight was significantly higher in the HH group than in the LH and LL groups; between the latter 2 groups there was no significant difference. The placental area occupied by vasculature was significantly greater in both the HA groups than in the LH group; the number of placentomes was greatest in the LL group. The density of VEGF and eNOS in the placentome tissue was significantly greater in both HA groups than in the LL group. Although the density of VEGF was significantly lower in the HH group than in the LH group, no differences were observed in eNOS density between the HH and LH animals. These results demonstrate that chronic hypoxia upregulates the expression of placental VEGF and eNOS, suggesting an important role of these molecules in the placental response to HA hypoxia. In addition, an attenuated response to hypoxia in VEGF synthesis may be part of the long-term process of adaptation to HA.

Plasma concentrations of progesterone (P4), estradiol-17beta (E2), estrone (E1) and estrone sulfate (E1S) were measured during gestation in eight guanacos kept in captivity. Gestational length was 346.1+/-9.8 days. P4 plasma concentrations increased after ovulation and remained elevated until parturition. However, during the last 4 weeks of gestation, a gradual decrease from 4.17x1.17(+/-1)nmol/L to 2.02x1.95(+/-1)nmol/L on day 5 before parturition was observed, followed by a more abrupt final decline to baseline concentrations which were reached on the day after parturition. Mean E2 plasma concentrations started to increase during the eighth month of gestation, and were significantly elevated up to maximum concentrations of 484.7x1.21(+/-1)pmol/L during the last 2 months of pregnancy. Concentrations returned to baseline during the last 2 days of gestation. An increase of E1S concentrations (p

The aim of the study was to investigate the histological characteristics and steroid concentrations in follicular fluid of different populations of follicles at different stages of development, during pregnancy and the oestrous cycle in cows. Follicles from ovaries collected at a slaughterhouse were allocated into three size categories (small, 2–5.9 mm; medium, 6–13.9 mm; and large, 14–20 mm) in pregnant and non-pregnant cows. Slices were stained with HE and PAS for histological analysis. Follicular fluid was pooled according to size and pregnancy status and estradiol, testosterone and progesterone concentrations in follicular fluid were determined by RIA. Characteristics of healthy follicles did not differ, regardless of follicle size or pregnancy status. Total histological atresia was significantly higher in pregnant cows than in non-pregnant cows (p < 0.05). Estradiol increased and testosterone decreased significantly, while follicles increased in size, in both non-pregnant and pregnant cows (p < 0.05). Nonpregnant cows had the highest estradiol values in follicles of all sizes. Medium and large follicles from pregnant cows showed the lowest testosterone concentration (p < 0.05). Progesterone levels increased with follicle size only in non-pregnant animals. In large follicles, progesterone concentration was significantly higher in non-pregnant cows than in pregnant cows (p < 0.05). Considering steroid concentration and histological findings, most large follicles might be atretic during pregnancy in cattle.

South American camelids are induced ovulators and require a stimulus to trigger the LH surge responsible for the ovulation. Seminal plasma (SP) of fertile alpacas (Lama pacos) was tested using a bioassay of pituitary cells to study the effect of seminal plasma on LH release. Plates containing rat pituitary cells (2 x 10(5) cells/90-95% viability) were cultured adding: (A) whole SP (WSP) treated with charcoal-dextran, or 1:2 or 1:4 proportions diluted in culture medium (DMEM/HEPES + antibiotics), or (B) 1:2 SP + anti-GnRH rabbit serum (inhibitory potency 10(-5) M), or (C) 1:2 SP + anti-GnRH + 100 nM synthetic GnRH (buserelin acetate) or (D) 100 nM, 50 nM, 10 nM, and 1 nM synthetic GnRH. Concentration (ng/ml) of LH secreted (Sec) and contained (Con) was analyzed using RIA 125I and the percentage of Sec and Con in each experiment was determined. The results of LH Sec for the cells treated with 50, 10, and 1 nM GnRH were 39, 13, and 1.5%, respectively (r2 = 98.41%, r = 0.9920) but cells treated with 100 nM GnRH secreted 10% of LH. With WSP, 1:2, or 1:4 SP the LH Sec was of 44.5% (3.25 ng/ml), 27% (1.9 ng/ml), and 18% (1.2 ng/ml), respectively. The exposure of cells to 1:2 SP + anti-GnRH, or to 1:2 SP + anti-GnRH/100 nM GnRH produced 31% (2.20 ng/ml) and 30% (1.8 ng/ml) of LH Sec, respectively. These results suggest that the SP of alpacas could have some factor(s) different from GnRH that would contribute to the mechanisms of LH secretion and to the induced ovulation in the female alpaca.

Background At high altitudes, hypoxia, oxidative stress or both compromise sheep fertility. In the present work, we tested the relative effect of short- or long-term exposure to high altitude hypobaric hypoxia and oxidative stress on corpora luteal structure and function.Methods The growth dynamics of the corpora lutea during the estrous cycle were studied daily by ultrasonography in cycling sheep that were either native or naïve to high-altitude conditions and that were supplemented or not supplemented with antioxidant vitamins. Arterial and venous blood samples were simultaneously drawn for determination of gases and oxidative stress biomarkers and progesterone measurement. On day five after ovulation in the next cycle, the ovaries were removed for immunodetection of luteal HIF-1alpha and VEGF and IGF-I and to detect IGF-II gene expression.Results The results showed that both short- and long-term exposure to high-altitude conditions decreased luteal growth and IGF-I and IGF-II gene expression but increased HIF-1 alpha and VEGF immunoexpression. The level of plasma progesterone was also increased at a high altitude, although an association with increased corpus luteum vascularization was only found in sheep native to a high-altitude location. Administration of antioxidant vitamins resulted in a limited effect, which was restricted to decreased expression of oxidative stress biomarkers and luteal HIF-1alpha and VEGF immunoexpression.Conclusions Exposure of the sheep to high-altitude hypobaric hypoxia for short or long time periods affects the development and function of the corpus luteum. Moreover, the observed association of oxidative stress with hypoxia and the absence of any significant effect of antioxidant vitamins on most anatomical and functional corpus luteum traits suggests that the effects of high altitude on this ovarian structure are mainly mediated by hypoxia. Thus, these findings may help explain the decrease in sheep fertility at a high altitude. © 2013 Parraguez et al.; licensee BioMed Central Ltd.

The present study evaluated the hypothesis that the effects of hypoxia on sheep pregnancies at high altitude (HA) are mediated by oxidative stress and that antioxidant vitamins may prevent these effects. Both HA native and newcomer ewes were maintained at an altitude of 3589 m during mating and pregnancy. Control low altitude (LA) native ewes were maintained at sea level. Half of each group received daily oral supplements of vitamins C (500 mg) and E (350 IU) during mating and gestation. Near term, maternal plasma vitamin levels and oxidative stress biomarkers were measured. At delivery, lambs were weighed and measured, and placentas were recovered for macroscopic and microscopic evaluation. Vitamin concentrations in supplemented ewes were two- or threefold greater than in non-supplemented ewes. Plasma carbonyls and malondialdehyde in non-supplemented ewes were consistent with a state of oxidative stress, which was prevented by vitamin supplementation. Vitamin supplementation increased lamb birthweight and cotyledon number in both HA native and newcomer ewes, although placental weight and cotyledon surface were diminished. Placentas from vitamin-supplemented HA ewes were similar to those from ewes at sea level, making these placental traits (weight, number and diameter of cotyledons) similar to those from ewes at sea level. Vitamin supplementation had no effect on LA pregnancies. In conclusion, supplementation with vitamins C and E during pregnancy at HA prevents oxidative stress, improving pregnancy outcomes.