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Simple Summary Corpora lutea (CL) are temporary endocrine structures that secrete progesterone, which is essential for maintaining a healthy pregnancy. A variety of regulatory factors come into play in modulating the functional lifespan of CL, with luteotropic and luteolytic effects. Many aspects of luteal phase physiology have been clarified, yet many others have not yet been determined, including the molecular and/or cellular mechanisms that maintain the CL from the beginning of luteolysis during early CL development. This paper summarizes our current knowledge of the endocrine and cellular mechanisms involved in multifactorial CL lifespan regulation, using the pseudopregnant rabbit model. Abstract Our research group studied the biological regulatory mechanisms of the corpora lutea (CL), paying particular attention to the pseudopregnant rabbit model, which has the advantage that the relative luteal age following ovulation is induced by the gonadotrophin-releasing hormone (GnRH). CL are temporary endocrine structures that secrete progesterone, which is essential for maintaining a healthy pregnancy. It is now clear that, besides the classical regulatory mechanism exerted by prostaglandin E2 (luteotropic) and prostaglandin F2α (luteolytic), a considerable number of other effectors assist in the regulation of CL. The aim of this paper is to summarize our current knowledge of the multifactorial mechanisms regulating CL lifespan in rabbits. Given the essential role of CL in reproductive success, a deeper understanding of the regulatory mechanisms will provide us with valuable insights on various reproductive issues that hinder fertility in this and other mammalian species, allowing to overcome the challenges for new and more efficient breeding strategies.

Resveratrol is one of the most investigated natural polyphenolic compounds and is contained in more than 70 types of plants and in red wine. The widespread interest in this polyphenol derives from its antioxidant, anti-inflammatory and anti-aging properties. Several studies have established that resveratrol regulates animal reproduction. However, the mechanisms of action and the potential therapeutic effects are still unclear. This review aims to clarify the role of resveratrol in the male and female reproductive functions, with a focus on animals of veterinary interest. In the female, resveratrol has been considered a phytoestrogen due to its capacity to modulate ovarian function and steroidogenesis via sirtuins, SIRT1, in particular. Resveratrol has also been used to enhance aged oocyte quality, and as a gametes cryo-protectant with mainly antioxidant, and anti-apoptotic effects. In the male, resveratrol enhanced testes function and spermatogenesis through activation of AMPK pathway. Furthermore, resveratrol has been supplemented to semen extenders improving the preservation of sperm quality. In conclusion, resveratrol has potentially beneficial effects for ameliorating ovarian and testes function. However, due to unclear data, further studies are necessary to consolidate these findings.

Freeze-drying (FD) has been exhaustively tried in several mammalian species as an alternative technique to sperm cryopreservation, but few studies have been done in rabbits (Oryctolagus cuniculus). The main objective of this study was to compare the protective effect of various antioxidants added to EDTA medium on structural and functional components of FD rabbit spermatozoa and on their status of global DNA methylation. FD media used were composed of basic FD medium (10 mM Tris-HCl buffer and 50 mM NaCl) supplemented with either 50 mM EDTA alone (EDTA) or added with 105 µM of rosmarinic acid (RA, EDTA-RA) or 10 µM of melatonin (MLT, EDTA-MLT). The effect of each medium on the preservation of FD spermatozoon structure was evaluated with light and scanning electron microscopy (SEM). Global DNA methylation was quantified in all FD sperm samples as well as in fresh spermatozoa. Morphologically, fracture points were evidenced in the neck, mid and principal piece of the spermatozoon tail. No differences in spermatozoon fracture points were evidenced among FD treatments: intact spermatozoa were the largest (p < .01) category, whereas the most frequent (p < .01) injury was the neck fracture, resulting in tailless heads. At SEM, the head of spermatozoa showed a well-conserved shape and intact membrane in all treatments. DNA methylation status was the same in all FD treatments. In conclusion, supplementation of EDTA, EDTA-RA and EDTA-MLT during FD preserved rabbit sperm morphological integrity and methylation status as well. Therefore, the difficulty of getting viable offspring using FD semen is likely unrelated to the impact of the lyophilization process on DNA methylation and morphology of lyophilized spermatozoa.

To investigate the ovulatory mechanisms triggered by raw semen (RS) in rabbits, we examined the expression of nerve growth factor (NGF)—a supposed ovulation-inducing factor (OIF)—and cognate receptors in anterior pituitary, ovary, and cervix as well as plasma NGF and luteinizing hormone (LH) concentrations. Six does/group were sham-inseminated with sterile saline (PBS), naturally mated (NM), inseminated with RS alone or after lumbar anesthesia (ARS), or treatment with COX inhibitors (CIRS). Immunohistochemistry revealed positive signals for NGF and receptors in all tissues. RT-PCR confirmed the presence of the target transcripts in the same tissues, except NTRK1 in the cervix. Circulating NGF concentrations rose 3- to 6-fold (P < 0.01) 15 min after semen deposition into the genital tract of NM, RS, and ARS rabbits and remained sustained thereafter. Circulating NGF was 4-fold lower (P < 0.01) in CIRS than in RS does indicating that NGF is mainly synthesized by the uterus. A concomitant rise of LH and NGF concentrations was found in 83.3%, 50.0%, and 16.7% of NM, RS, and CIRS does, respectively, but not in ARS (despite high NGF circulating levels). Seminal plasma NGF concentration was 151.9 ± 9.25 μg/mL. The ovulatory responses were 0%, 83.3%, 66.7%, 16.7%, and 0% in PBS, NM, RS, ARS, and CIRS groups, respectively. Present data confirm that, although RS may induce ovulation via endocrine mechanisms through binding to NGF receptors in the ovary, a novel OIF-mediated neural mechanism facilitates ovulation in rabbits.

[EN] To evaluate how rearing programmes could affect resources allocation and reproductive performance of primiparous rabbit females, a total of 118 rabbit females were used to evaluate the effects of five rearing feeding programmes on their performance from 1st to 2nd parturition: CAL, fed ad libitum C diet (11.0 MJ digestible energy (DE), 114 g digestible protein (DP) and 358 g NDF/kg dry matter (DM) until 1st parturition; CR, fed ad libitum with C diet until 12 weeks of age and then C diet restricted (140 g/day) until 1st parturition; F, fed ad libitum with F diet (8.7 MJ DE, 88 g DP and 476 NDF/kg DM) until 1st parturition; FC, fed with F diet ad libitum until 16 weeks of age, and C diet ad libitum until 1st parturition; FCF, fed with F diet ad libitum until 16 weeks of age, then C diet ad libitum until 20 weeks and then F diet ad libitum until 1st parturition. From 1st parturition, C diet was ad libitum offered to all the experimental groups until 2nd parturition. CAL females presented lower feed intake than females of F, FC and FCF groups in the 1st week of lactation (on av. ¿16.6%; P, The authors thank the Spanish Ministry of Education and Science (Project AGL2006-07596) for the economic support to conduct this study.

Kisspeptin (KiSS) and its related receptors (KiSS1R) have a critical role in the reproduction of mammals. The KiSS/KiSS1R system is expressed in numerous reproductive organs including the ovary. Here, we studied the expression of the KiSS/KiSS1R system and its functional role in rabbit corpora lutea (CL) at days 4 (early-), 9 (mid-), and 13 (late-stage) of pseudopregnancy. In vitro progesterone, prostaglandin (PG) F2α (PGF2α) and E2 (PGE2) productions and prostaglandin-endoperoxide synthase 1 (PTGS1) and 2 (PTGS2) activities were evaluated. Immune reactivity (IR) for KiSS and KiSS1R were detected in luteal cells at nuclear and cytoplasmic level at all luteal stage for KiSS and only at early- and mid-stage for KiSS1R; IR decreased from early- to later stages of pseudopregnancy. The KiSS-10 augmented progesterone and PGE2 and diminished PGF2α secretions by early- and mid-CL; KiSS-10 reduced PTGS2 activity at early- and mid-stages, but did not affect PTGS1 at any luteal stages. The antagonist KiSS-234 counteracted all KiSS-10 effects. This study shows that the KiSS/KiSS1R system is expressed in CL of pseudopregnant rabbits and exerts a luteotropic action by down-regulating PTGS2, which decreases PGF2α and increases PGE2 and progesterone.

The reproductive cycle of an invasive alien Italian grey squirrel population was studied to understand its adaptation and limit its spread, in order to conserve the autochthonous red squirrel. Female and male genital traits were evaluated throughout the reproductive cycle, including the ovary, uterus, testicle, epididymis, seminiferous tubule morphometry, and germinative epithelium histology. Moreover, individual female fecundity was determined by counting uterine scars. Ovary width and uterus weight, length, and width reached their highest values in the luteal and pregnancy phases. On conducting a histological evaluation of the testicular germinal epithelium, four morphotypes related to the different reproductive phases of the male squirrels were identified: immature, pubertal, spermatogenesis, and regressive. Testicle and epididymis weights and seminiferous tubule diameters reached their largest values during spermatogenesis. Uterine scar analysis showed that 69% of the females had given birth to one or two litters, while 31% had no uterine scars. Litters were larger in the first breeding period than in the second, annual fecundity was 4.52 &plusmn, 1.88 uterine scars/female. Umbrian grey squirrels have adapted to their non-native range, showing two annual mating periods at times similar to those in their native range, and high reproductive success.

This study investigates the effects of linseed (rich in ɑ-linolenic acid (ALA)) and fish oil (rich in eicosapentaenoic (EPA) and docosahexaenoic acid (DHA)) supplementation on the insulin resistance of pregnant rabbits. Two months before insemination, the rabbits (15 animals/group) were fed different diets: commercial standard (group C), supplemented with 10% extruded linseed (group L), and 3% fish oil (group FO). The L group does showed both the highest feed intake before AI (P < 0.01) and the highest body weight (BW) throughout pregnancy (P < 0.001). The L does yielded less milk than the C does (P < 0.001); however, no differences were observed in either weight or size of litter at weaning. Regardless of diet, insulin concentrations and HOMA-IR values were higher during the first half of pregnancy (P < 0.001). Nevertheless, the L does showed higher mean insulin concentrations than FO rabbits (P < 0.01) and the lowest glucose clearance (P < 0.01) during pregnancy. On the other hand, pregnant FO rabbits showed the lowest glucose concentrations (P < 0.05) and the lowest Homeostasis model assessment values for insulin resistance (HOMA-IR, P < 0.05) as well as a faster restoration of baseline glucose levels following glucose load (P < 0.001). Before and during pregnancy, the BW of the rabbits was positively related to fasting sample- and tolerance test-derived indices of insulin resistance (P < 0.05) suggesting that a high pre-pregnancy BW predisposes to gestational insulin resistance. Linseed supplementation increased BW and predisposed to insulin resistance during pregnancy; whereas, fish oil improved insulin sensitivity without significant changes in BW.

[EN] This study examined the effects of feed restriction at different stages of rabbit pregnancy on body condition and productive performance. Just after insemination, pregnant primiparous New Zealand White does were assigned to 4 groups (10/group): the control group (C) was fed with 130 g/d of commercial feed while the others received 90 g/d from day 0 to 9 (R1), from day 9 to 18 (R2) or from day 19 to 28 (R3) of pregnancy and 130 g/d the remaining periods. A 3-point scale for loin and rump was used to calculate the aggregate body condition score (BCS), while perirenal fat weight (PFW) was estimated by ultrasound measurement of its thickness. The C does showed a positive balance of the pregnancy (0-26 d) for both body weight (P, This work is part of the PhD programme in “Animal health, livestock production and food safety” XXVIII cycle, financed by the University of Perugia, Italy.

A total of 190 rabbit females were used to evaluate five feeding programmes from 9 weeks of age to the first parturition: CAL, fed ad libitum with a control diet (C: 11.0 MJ digestible energy (DE) and 114 g digestible protein (DP)/kg dry matter (DM)) until first parturition; CR, fed ad libitum with C diet until 12 weeks of age and then C diet restricted (140 g/day) until first parturition; F, fed ad libitum with a low-energy, high-fibre diet (F: 8.7 MJ DE and 88 g DP/kg DM) until first parturition; FC, fed with F diet ad libitum until 16 weeks of age, and C diet ad libitum until first parturition; FCF, fed with F diet ad libitum until 16 weeks of age, then C diet ad libitum until 20 weeks and then F diet ad libitum until first parturition. The rabbits were artificially inseminated at 18 weeks of age. CAL group had a higher mortality rate compared with the other groups between 9 and 12 weeks of age (34% v. 3%; P,0.05) and during the last 3 weeks of first pregnancy (14% v. 3%; P,0.05). The CAL and FC females presented higher BW and perirenal fat thickness (PFT) than CR females at 11 days of pregnancy (10.41 kg and 10.6 mm; P,0.05), with F females showing medium values. The type of feeding procedure did not affect the fertility rate of young females at first artificial insemination. Differences in BW disappeared at parturition, when only CAL females presented a greater PFT than CR and FC females (10.3mm; P,0.05). In comparison with FCF, CAL females had smaller and thinner live born litters (22.5 kits and 2139 g, respectively; P,0.05), with CR, F and FC females showing medium values. The low number of kits born alive for CAL females was because of their lesser total number of kits born (21.7 kits; P,0.05) and the greater mortality of their litters at birth (113.9%; P,0.05) compared with FCF females. Non-esterified fatty acid was higher in the blood of females fed C diet (CAL and CR) than in others at partum day (on average 10.15 mmol/l; P,0.05). In conclusion, the ad libitum use of diets for lactating rabbit does throughout the rearing period could lead young rabbit females to present a higher risk of early death and smaller litter size at first parturition. Feed restriction or earlier use of suitably fibrous diets led females to achieve the critical BW and fat mass at first mating to ensure reproduction., The authors thank the Spanish Ministry of Education and Science (Project AGL2006-07596) for the economic support to conduct this study.

To study the role of endothelial factors in luteal function, the dynamic profiles of genes for endothelin 1 (EDN1), its receptor subtypes, EDNRA and EDNRB, and angiotensin converting enzyme (ACE) were examined in corpora lutea (CL) obtained from rabbits on Days 4 and 9 of pseudopregnancy after prostaglandin (PG) F2alpha analogue (alfaprostol) treatment. The cell type distribution of EDN1 in the ovaries and its mechanisms of actions in vitro and in vivo were also studied. Positive immunostaining for EDN1 was localized in the luteal and endothelial cells, in granulosa cells of the follicles, and in the ovarian epithelium. The basal mRNA levels for EDNRA, EDNRB, and ACE were lower (P

The objective of the present study was to investigate in rabbit corpora lutea (CL), at both the cellular and molecular level, intraluteal cyclooxygenase (COX)-1, COX-2 and prostaglandin (PG) E2-9-ketoreductase (PGE2-9-K) enzymatic activities as well asin vitroPGE2 and PGF2α synthesis following PGF2α treatment at either early- (day-4) or mid-luteal (day-9) stage of pseudopregnancy. By immunohistochemistry, positive staining for COX-2 was localized in luteal and endothelial cells of stromal arteries at both the stages. In CL of both stages, basal COX-2 mRNA levels were poorly expressed, but rose (P< 0.01) 4- to 10-fold 1.5–6 h after treatment and then gradually decreased within 24 h. Compared to mid-stage, day-4 CL had lower (P< 0.01) COX-2 and PGE2-9-K basal activities, and PGF2α synthesis rate, but higher (P< 0.01) PGE2 production. Independent of luteal stage, PGF2α treatment did not affect COX-1 activity. In day-4 CL, PGF2α induced an increase (P< 0.01) in both COX-2 activity and PGF2α synthesis, whereas that of PGE2 remained unchanged. In day-9 CL, PGF2α up-regulated (P< 0.01) both COX-2 and PGE-9-K activities, and PGF2α production, but decreased (P< 0.01) PGE2 synthesis. All changes in gene expression and enzymatic activities occurred within 1.5 h after PGF2α challenge and were more marked in day-9 CL. Our data suggest that PGF2α directs intraluteal PG biosynthesis in mature CL, by affecting the CL biosynthetic machinery to increase the PGF2α synthesis in an auto-amplifying manner, with the activation of COX-2 and PGE-9-K; this may partly explain their differentially, age-dependent, luteolytic capacity to exogenous PGF2α in rabbits.

This study examined the effects of food restriction during rabbit pregnancy on hormones and metabolites involved in energy homeostasis and metabolic programming. Pregnant does were assigned to four groups: the control group was fed a standard ration while the others received a restricted amount of food (30% restriction) during early (0–9 days), mid (9–18 days), and late (19–28 days) pregnancy. The pregnancy induced a coordinated range of adaptations to fulfil energy requirements of both mother and foetus, such as hyperleptinaemia and hyperinsulinaemia, reduced insulin sensitivity, increased cortisol and non-esterified fatty acid. Food restriction altered leptin, insulin, T3, non-esterified fatty acids and glucose concentrations depending on the gestational phase in which it was applied. Collectively, present data confirm that the endocrinology of pregnancy and the adaptive responses to energy deficit make the rabbit an ideal model for studying nutritional-related disorders and foetal programming of metabolic disease.

[EN] Sexual receptivity of rabbit does at insemination greatly influences fertility and is generally induced by hormones or techniques known as “biostimulation”. Searching for more sustainable farming systems, an original alternative would be to utilise the genetic pathway to increase the does’receptivity. The purpose of the present study was to identify genetic and non-genetic factors that influence rabbit doe sexual receptivity, in the context of a divergent selection experiment over 1 generation. The experiment spanned 2 generations: the founder generation (G0) consisting of 140 rabbit does, and the G1 generation comprising 2 divergently selected lines (L and H lines) with 70 does each and 2 successive batches from each generation. The selection rate of the G0 females to form the G1 lines was 24/140. The selection tests consisted of 16 to 18 successive receptivity tests at the rate of 3 tests per week. On the basis of 4716 tests from 275 females, the average receptivity was 56.6±48.2%. A batch effect and a test operator effect were revealed. The contribution of females to the total variance was 20.0%, whereas that of bucks was only 1.1%. Throughout the experiment, 18.2% of does expressed a low receptivity (< 34%), 50.7% a medium one and 33.1% a high one (>66%). Some does were frequently receptive, whereas others were rarely receptive. The repeatability of sexual receptivity was approximately 20%. The results confirmed the high variability of sexual receptivity of non-lactating rabbit does maintained without any biostimulation or hormonal treatment. A lack of selection response on receptivity was observed. Accordingly, the heritability of receptivity was estimated at 0.01±0.02 from an animal model and at 0.02±0.03 from a sire and dam model. The heritability of the average receptivity of a doe was calculated as 0.04. In agreement with the low estimated heritability, the heritability determined was no different from zero. Nevertheless, the occurrence of pseudopregnancies due to uncontrolled ovulations and the presence of corpora lutea, as assessed by progesterone titrations, could have interfered with receptivity. Further studies would be necessary to confirm the low heritability of female rabbit receptivity., The authors are grateful to the Department of Animal Genetics of the National Institute of Agronomical Research for its financial contribution, to P. Aymard, J.M. Bonnemère, Annette Debrusse, H. Girotto and D. Labatut, technicians at the PECTOUL Experimental Rabbit Farm at INRA in Toulouse, to J. Ruesche, the IT team from SAGA for their collaboration and to G. Wagman for her careful linguistic revision.

The aim of this study was to better understand the role of the endothelin-1 (ET-1) system in the process of controlling the corpora lutea (CL) life span in rabbits. ET-1 (10 microg iv) administration at d 9 and 12 of pseudopregnancy induced a functional luteolysis within 24 h of injection, but it was ineffective at both d 4 and 6. Pretreatments with Bosentan, a dual ET(A)/ET(B) receptor antagonist, or cyclooxygenase (COX) inhibitor blocked the luteolytic action of ET-1 but not that induced by prostaglandin F2alpha (PGF2alpha). In CL cultured in vitro, ET-1 increased (P/= 0.01) both PGF(2alpha) production and luteal nitric oxide synthase activity but decreased (Por = 0.01) progesterone release. Addition of ET(A) receptor antagonist BQ123 or COX inhibitor blocked the ET-1 luteolytic effects. Positive staining for ET-1 receptors was localized in ovarian blood vessels, granulosa cells of large follicles, and luteal cells. Immunoblot analysis of ET-1 receptor protein revealed a strong band of approximately 48 kDa in d-9 CL. Up to d 6 of pseudopregnancy, ET-1 mRNA abundance in CL was poorly expressed but then increased (Por = 0.01) at d 9 and 13. ET(A)-receptor transcript increased (Por = 0.01) at d 6, remained at the same level up to d 13, and then declined to the lowest (Por = 0.01) levels at d 22. ET(B)-receptor mRNA increased (Por = 0.01) throughout the late-luteal stage from d 13 up to d 18. Our data suggest that the luteolytic action of ET-1 may be a result of PGF2alpha synthesis from both luteal and accessory cells, via the COX pathways.

The in vivo chronic and in vitro acute effects of di(2-ethylhexyl) phthalate (DEHP) on the reproductive function of peroxisome proliferator-activated receptor gamma (PPARG) were studied in rabbit corpora lutea (CL) at early stage (Day 4), midstage (Day 9), and late stage (Day 13) of pseudopregnancy. The rabbits were in vivo treated with DEHP for 15 days before induction of pseudopregnancy. Immunohistochemistry provided evidence for the presence of PPARG, prostaglandin endoperoxide synthase 1 (PTGS1), PTGS2, prostaglandin E2-9-ketoreductase (PGE2-9-K), and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) in all the luteal cells during pseudopregnancy. DEHP decreased progesterone plasma levels and CL production in all the luteal stages and PPARG protein and gene expressions in early and mid-CL. DEHP in vivo treatment reduced PTGS2 protein expression at the late stage and that of PGE2-9-K at all the stages, whereas PTGS1 and 3beta-HSD were not affected. In in vitro cultured CL, DEHP alone, the PPARG antagonist T0070907 alone, or DEHP plus T0070907 diminished progesterone production and 3beta-HSD activity and increased PGF2alpha and PTGS2 in early and mid-CL, whereas DEHP plus the PPARG agonist 15d-PGJ2 did not affect these hormones and enzymes. All the in vitro treatments did not affect PGE2 secretion as well as PTGS1 and PGE2-9-K enzymatic activities in all the luteal stages. These results provided evidence that DEHP favors functional luteolysis of pseudopregnant rabbit CL, with a mechanism that seems to involve PPARG expression down-regulation, an increase of PTGS2 activity and prostaglandin F2alpha secretion, 3beta-HSD down-regulation, and decrease in progesterone.

Dopamine (DA) receptor (DR) type 1 (D1R) has been found to be expressed in luteal cells of various species, but the intrinsic role of the DA/DRs system on corpora lutea (CL) function is still unclear. Experiments were devised to characterize the expression of DR types and the presence of DA, as well as the in vitro effects of DA on hormone productions by CL in pseudopregnant rabbits. Immunoreactivity and gene expression for D1R decreased while that for D3R increased in luteal and blood vessel cells from early to late pseudopregnant stages. DA immunopositivity was evidenced only in luteal cells. The DA and D1R agonist increased in vitro release of progesterone and prostaglandin E2 (PGE2) by early CL, whereas the DA and D3R agonist decreased progesterone and increased PGF2α in vitro release by mid- and late CL. These results provide evidence that the DA/DR system exerts a dual modulatory function in the lifespan of CL: the DA/D1R is luteotropic while the DA/D3R is luteolytic. The present data shed new light on the physiological mechanisms regulating luteal activity that might improve our ability to optimize reproductive efficiency in mammal species, including humans.

The expression of peroxisome proliferator-activated receptor gamma (PPARgamma) and its role in corpora lutea (CL) function were studied in pseudopregnant rabbits. Corpora lutea were collected at an early stage (Day 4), midstage (Day 9), and late stage (Day 13) of pseudopregnancy. Immunohistochemistry found evidence for the presence of PPARgamma in the perinuclear cytoplasm and nucleus of all the luteal cells; immunoreactivity decreased from the early to the late stage, with immunonegativity of the nuclei of late stage CL. PPARgamma mRNA transcript was expressed in all the luteal stages with the lowest level in the late stage. In CL cultured in vitro, the PPARgamma agonist (15-deoxy delta12,14 prostaglandin J2 [15d-PGJ2], 200 nM) increased and the antagonist (T0070907, 50 nM) decreased progesterone secretion at early and midluteal stages, whereas 15d-PGJ2 reduced and T0070907 increased PGF2alpha at the same stages. Prostaglandin-endoperoxide synthase 2 (PTGS2) activity was reduced by 15d-PGJ2 and increased by T0070907 in CL of early and midluteal stages. Conversely, 15d-PGJ2 increased and T0070907 reduced 3betahydroxysteroid dehydrogenase (3beta-HSD) activity in early and midluteal stage CL. PGE2 in vitro secretion as well as PTGS1 and 20alpha-HSD enzymatic activities were not affected by 15dPGJ2 and T0070907 in any CL types. These results indicate that PPARgamma plays a luteotropic role in pseudopregnant rabbits, through PTGS2 down-regulation and 3beta-HSD up-regulation, with a consequent PGF2alpha decrease and progesterone increase.

[EN] Primiparous does express low mating acceptance during lactation and different oestrus induction methods are employed to improve sexual receptivity. The aim of this work was to study the effect of a biostimulation, a 48 h doe-litter separation (DLS), in primiparous lactating rabbits on mating acceptance, external characteristics of the vulva and plasma concentrations of estradiol, testosterone, progesterone and cortisol. Seventy primiparous does from local population were divided on day 1 after kindling into 2 equal groups: control (C, n=35), where does always had free access to the nest-box, and DLS (n=35), where free suckling was adopted, except from day 9 to day 11 post-partum (PP). Litter size was equalised to 6-7 kits. Before mating, blood samples were obtained from 10 does per group on 9th and 11th d post-partum (dpp). The results showed that the DLS treatment improved the mating acceptance on 11th dpp (88.5 vs. 54.2%; P0.05). In contrast, cortisol plasma levels were not affected by the DLS treatment (-12.4 and -14.4% for control and DLS group; P>0.05). At day 11 PP, higher plasma E2 and P4 levels were described in DLS in comparison to C, although no significant difference was found, while plasma testosterone concentrations tended to be higher in DLS (165.5±27.1 vs. 114.9±31.9 pg/mL, for DLS and C, respectively; P=0.075). In summary, the 48 h doe-litter-separation acted as an efficient oestrus inductor in primiparous lactating rabbits. Insemination of biostimulated does must occur after the 1st suckling episode following the separation. This biostimulation method did not modify cortisol secretion of the rabbit doe., This work was supported by the National Research Program (Algeria). The authors wish to thank Dr. Bellazzoug of Algiers Hospital for the hormone assays.

The expression of gonadotropin-releasing hormone (GNRH) receptor (GNRHR) and the direct role of GNRH1 on corpora lutea function were studied in Mediterranean buffalo during diestrus. Immunohistochemistry evidenced at early, mid, and late luteal stages the presence of GNRHR only in large luteal cells and GNRH1 in both small and large luteal cells. Real-time PCR revealed GNRHR and GNRH1 mRNA at the three luteal stages, with lowest values in late corpora lutea. In vitro corpora lutea progesterone production was greater in mid stages and lesser in late luteal phases, whereas prostaglandin F2 alpha (PGF2alpha) increased from early to late stages, and PGE2 was greater in the earlier-luteal phase. Cyclooxygenase 1 (prostaglandin-endoperoxide synthase 1; PTGS1) activity did not change during diestrus, whereas PTGS2 increased from early to late stages, and PGE2-9-ketoreductase (PGE2-9-K) was greater in late corpora lutea. PTGS1 activity was greater than PTGS2 in early corpora lutea and lesser in late luteal phase. In corpora lutea cultured in vitro, the GNRH1 analog (buserelin) reduced progesterone secretion and increased PGF2alpha secretion as well as PTGS2 and PGE2-9-K activities at mid and late stages. PGE2 release and PTGS1 activity were increased by buserelin only in late corpora lutea. These results suggest that GNRH is expressed in all luteal cells of buffalo, whereas GNRHR is only expressed in large luteal phase. Additionally, GNRH directly down-regulates corpora lutea progesterone release, with the concomitant increases of PGF2alpha production and PTGS2 and PGE2-9-K enzymatic activities.

Food deprivation affects female reproduction. The goal of the present study was to elucidate in the rabbit model the effects of acute energy restriction on ovarian function (follicle development, atresia rate and in vitro oocyte maturation) and embryonic development and gene expression of some candidate genes. Serum metabolic parameters (non-esterified fatty acids (NEFA), triglycerides, glucose, insulin and leptin concentrations) and endocrine markers (oestradiol-17β and progesterone concentrations) were also studied. A control group of nulliparous does fed ad libitum and a 72-h fasted group were used. At the end of the nutritional treatment, the ovaries of half of the animals were retrieved while the other animals were re-fed and artificially inseminated to recover embryos at 84 h after insemination, during the luteal phase. At the end of fasting, increased serum NEFA and decreased leptin concentrations were observed in the fasted group, but no differences appeared in serum steroid concentrations, follicle population and atresia rate or nuclear and cytoplasmic oocyte maturation. In the luteal phase, insulin concentrations increased notably in the fasted group. The number of recovered embryos per female and the speed of embryo development were reduced in the food-deprived group. Acute fasting altered both metabolic and endocrine markers and embryo development, but follicle and oocyte development and embryo gene expression were not affected.

International audience; The present study sought to assess whether the receptors for adrenocorticotropic hormone (ACTH), MC2R, and for glucocorticoid (GR) are expressed in corpora lutea (CL) of pseudopregnant rabbits and whether ACTH and cortisol exert any direct action on luteal function. By immunohistochemistry, positive reaction for MC2R and GR was detectable within luteal cells of CL. The MC2R mRNA levels were five-fold less abundant in day 9 than in day 4 CL (P

[EN] The effect of 2 different techniques of mother-litter separation on rabbit doe performance was analysed. In the first experiment, executed in a commercial rabbitry, multiparous lactating hybrid does were successively artificially inseminated (620 Al) while in the second, a flock of 100 nulliparous was followed (524 Al). Does Nere submitted to one of the following treatments: control no treatment; 24 hours of mother-litter separation, 3 days befare Al, with 2 different procedures: closing the nestbox (Experiment 1) and change of caga (Experiment 2). AH the does were inseminated 11 days after kindling. Mother-litter separation did not significantly affect reproductiva results while, as expected, sexual receptivity and physiological status strongly influenced it. Probably longar stimuli are necessary to induce oestrus, particularly when does are in lactation. Nulliparous and multiparous non-lactating does showed a significant positiva reaction to biostimulation (Experiment 2 - gain of fertillty rata: 12.9% and 14.0%, respectively)., [FR] Deux techniques différentes de séparation de la mere et de sa portée ont été étudiées. Dans la premiare expérience menée dans un élevage industrie/, des lapínes hybrídes plurípares a/laítantes ont été inséminées artificiellement (620 JA) tandis que dans la deuxieme expérimentation un groupe de 100 lapines ont été contr61ées jusqu'a obtenir 524 JA. Les traitements suivants ont été appliqués : groupe controle sans traitement, 24 heures de séparation mere-portée trois iours avant /'/A selon deux méthodes - fermeture de la boíte a níd (expérience 1) - changement de cage (expérience 2). Toutes les lapines ont été inséminées 11 jours apres la mise bas. La séparation mere-portée n'a pas significatívement influencé les performances de reproductíon tandis que la réceptivité sexual/e et le statut physiologique ont été fortement ínfluencés. De plus· longues stimulatíons sont sans doute nécessaires pour induire l'oestrus, particulierement quand les feme/les sont allaitantes. Les lapines non allaitantes nullípares ou pluripares montrent une réaction positiva significativa a la biostimulation (expérience 2: augmentation du taux de fertilité: 12,9 et 14,0 %, respectivement)

In this issue of "W0RD RABBIT SCIENCE", five papers deal with the so called "biostimulation" methods which are intended to increase the reproductive performance of rabbit without resort to exogenous hormones. This is not just a fortunate coincidence, but rather the evidence of the role undertaken by the IRRG (Intemational Rabbit Reproduction Group) on rabbit research. lt is my responsibility to briefly illustrate here the main objectives and the concerted actions on which our work-group is engaged.

[EN] In this short synthesis the authors triad to drive sorne conclusions from 4 studies on breeding rabbit does biostimulation, conducted within the lnternational Rabbit Reproduction Group (IRRG). The aim of these biostimulations was the improvement of does' fertility after artificial insemination (Al). Ali experiments were conducted according a 42 days reproduction rhythm. The trial of flushing, i.e. momentary improvement of the energy balance of does 4 days before Al, failed to improve fertility. The main reason was the low palatability of the experimental diet : the daily energy intake was lower than with control diet. Transfer of does to another caga, 2 days befare Al, can improve fertility of adults does, but in opposition with literatura, was inefficient in nulliparous does. However this technique is difficult to manage in larga commercial rabbitries. Efficiency of biostimulation through separation of mother from her litter for a short period depends of the timing and duration of the stimulus. A 36 to 48 hours separatlon just befare Al may improve fertility by 7 to 30 points ; but kits' weaning weight is reduced by 5% to 10%. Finally, further experiments are needed to know the practica! interest of a long term utilisation of such biostimulations, and to know the physiological mechanism., [FR] Dans ce bref artícle de synthiJse, les auteurs tentent de tirar les enseígnements de 4 études portant sur la "bíostimulatíon" de la reproduction chez la lapine, études conduites dans le cadre du Groupe lntemational sur la Reproduction (IRRG). Les différentes biostímulatlons étudiées avaient toutes pour objectif de favoriser la fertílité de tapines inséminées artificiellement dans le cadre d'une conduite en bandas selon un rythme a 42 jours. L'essai de f/ushing par distributlon d'un aliment plus concentré n'a pas donné les résultats espérés en raison d'une sous-consommation relativa de /'aliment expérimental initialement destiné a stimuler l'ingestion énergétique des femelles. Un simple changement de caga des tapines pendant les 2 jours précédant /'/A permet d'améliorer significativement la fertílité (+14 points), sauf pour les femelles nullipares. Toutefois, cette technique est difficile a app/iquer dans des élevages commerciaux. Enfin, la biostimulation de la reproduction par séparation de la mere et de sa portée donne des résu/tats dépendants de la durée et du moment de la séparation par rapport a l'IA. Une séparation de 36 a 48 heures juste avant l'IA permet d'accroitre la fertílité des tapines de 7 a 30 points, mais elle réduit d'environ 5% a 10% le poids des lapereaux au sevrage. De nouvelles expérimentations sont nécessaires pour validar /'efficacité de ces biostimulations a long terma et é/ucider le mécanisme de leur action.

[EN] The effect of environmental temperature and age on plasma hormonal concentrations of testosterone (Tl, dihydrotestosterone (DHT}, cortisol (C), triiodothyronine (T3) and thyroxine (T4) were evaluated in 40 male Grimaud rabbits. Twenty animals were reared at 20ºC (TNT = Thermic Neutrality Temperature) and the other 20 at 27ºC (HST = Heat Stress Temperature). Far both treatments relative humidity levels averaged 77 % and the photoperiod was 8D:16L with a light intensity of about 40 lux. The animals were fed ad libitum with a commercial pelleted feed. Blood samples were collected from each ammal by intracardiac puncture at 71 and 85 days of age. No interaction effect between temperature and age was found. With regard to effect of temperature, the HST animals hada lower (P, [FR] Cette étude réalisée sur 40 lapins Grimaud males a pour but d'évaluer l'effet de la température ambiante et de l'age sur les concentrations _plasmat1ques des hormones swvantes : Testostérone (T), Dihydrotestostérone, (DHT), Cortisol {C), Triiodothyronine (T3), Thyroxine (T4). Vingt males ont été maintenus a 2óºC (TNT = Temeerature de Neutralité Thermique) et les vingt autres a 27 C (HST = Température de Stress de Chaleur). Pour les deux lots /'humidite relative moyenne était de 77 %,'la photopériode était de 8D:16L avec une intensité lumineuse d'environ 40 lux. Les animaux ont été nourris ad libitum avec un a/iment granulé du commerce. Sur chaque animal, des prélevements de sang ont été effectués par ponction intracardiaque a l'age de 71 et 85 jours. Aucun effet d'interaction entre l'age et la température n'a été observé. Pour ce qui concerne l'effet de la température, on a enregistré pour les animaux HST un poids vif final (2325 vs 2555 g ), un gain de poids journalier (32.7 vs 38.1 g) et une consommation (106 vs 126 g/jour) inférieurs (P, Research supported by National Research Council of Italy, Special project RAISA, Sub project n. 3; Paper nº 2245

CB1 is a member of the G-protein-linked receptor superfamily that is present in the central nervous system as well as in certain peripheral neuronal and non-neuronal tissues. Recently, the presence of CB1 was found in the ductal system of the major salivary glands of laboratory animals, but no data are available for domestic mammals. Thus, in the present study, we examined the presence and distribution of CB1 in the major salivary glands of dogs using immunohistochemical techniques. CB1 was found in the parotid and mandibular glands of adult dogs; positive immunoreaction was localized to the cells of the striated ducts, with a peculiar localization on or near the apical membrane. This particular localization may be explained by the characteristics of this receptor as membrane-associated. The acinar structures were completely negative for CB1. We conclude that CB1 is involved in the control of dog salivary secretion via endogenous substances, likely endocannabinoids. The localization of CB1 highlights that endocannabinoids promote qualitative and/or quantitative changes of the primary saliva in the ductal system.

[EN] The aim of this study was to examine sexual receptivity and productivity of does injected 48 h before insemination with 8 or 25 IU of eCG (groups 8 and 25, respectively) in comparison with a control group (no injection, group 0). One hundred twenty four does were inseminated during 11 consecutive cycles (reproduction rythm: 35 d; inseminations: 4 d post partum and weaning: 28 d post partum). In comparison with the control group, eCG signifi cantly improves the receptivity of does (60.3 vs 80.5 and 79.8%; P

[EN] In order to study the metabolic adaptation in response to 48 h transient doe-litter separation (DLS) in young rabbits (5 rabbits/d group) between postnatal 9 and 11 d, plasma concentrations of thyroid hormones T3 and T4, insulin, leptin, glucose, triglycerides (TG), and free fatty acids (FFA) were examined before (6-8 d), during (9-11 d), and after separation (12-16 d). T3 concentrations in newborn control rabbits gradually increased from 0.6 ng/mL at postnatal 6 d to 1.0 ng/mL at postnatal 16 d, whereas those of T4 remained fairly constant (25 ng/mL) up to postnatal 14 d, when T4 gradually declined to 8 ng/mL. T3 values of DLS newborn rabbits did not differ from those of controls at postnatal 10 and 11 d, but were lower (P, This work was partially supported by a grant from the Italian “Ministero Istruzione Università e Ricerca” (Progetto Ricerca Interesse Nazionale – Prot. 2001072484 and 2003074002) and by generous donations from the BC Red-water Trust (Perugia, Italy). Procedures conformed to animal care guidelines and were approved by the Bioethical Committee of the University of Perugia in accordance with the current laws of Italy. The authors gratefully acknowledge the revision of the English text by Sheila Beatty.

The expression of type I GNRH receptor (GNRHR-I) and the direct role of GNRH-I on corpora lutea (CL) function were studied in the pseudopregnant rabbit model. Immunohistochemistry evidenced GNRHR-I and GNRH-I in luteal cells at early (day 4 pseudopregnancy)-, mid (day 9)-, and late (day 13)-luteal stages. Real-time RT-PCR and western blotting revealed GNRHR-I mRNA and protein at the three luteal stages. Buserelin in vivo treatment at days 9 and 13 decreased plasma progesterone levels for 48 and 24 h respectively. In in vitro cultured CL, buserelin reduced progesterone secretion, increased prostaglandin F2α (PGF2α) secretion and cyclo-oxygenase-2 (COX-2) and nitric oxide synthase (NOS) activities at days 9 and 13, and decreased PGE2 at day 13. Co-incubation with antagonists for GNRH-I (antide), inositol 1,4,5-trisphosphate (IP3, 2-amino-ethoxydiphenylborate), and diacylglycerol (DAG, 1-hexadecyl-2-acetyl glycerol) or inhibitors for phospholipase C (PLC, compound 48/80), and protein kinase C (PKC, staurosporine) counteracted the buserelin effects. Buserelin co-incubated with COX inhibitor (acetylsalicylic acid) increased progesterone and decreased PGF2α and NOS activity at days 9 and 13, whereas co-incubation with NOS inhibitor (N-nitro-l-arginine methyl ester) increased progesterone at the same luteal stages. These results suggest that GNRHR-I is constitutively expressed in rabbit CL independently of luteal stage, whereas GNRH-I down-regulates directly CL progesterone production via PGF2α at mid- and late-luteal stages of pseudopregnancy, utilizing its cognate type I receptor with a post-receptorial mechanism that involves PLC, IP3, DAG, PKC, COX-2, and NOS.

The aims of this study were to compare the hormones and metabolites serum levels and the reproductive performances of nulliparous (n=100) and primiparous pregnant does submitted to artificial insemination (AI) 11 days post-partum. On the day of AI, all the does were weighed and the sexual receptivity was evaluated. The kits were weaned at 26 day. Blood samples were collect by punc- ture of the marginal ear vein from one day before AI until few days before the kindling and assayed for hormones and metabolites. The higher sexual receptivity and the fertility in nulliparous than in primiparous does confirmed the negative effect of lactation. Nulliparous does showed higher blood con- centration of leptine than primiparous, and in both the groups such level lowered during pregnancy, probably reflecting the reduction of the fat reserve. The insuline level increased during pregnancy in either groups as a consequence of the growing of the foetuses. In nulliparous does the cortisol, NEFA and T3 concentrations were higher than primiparous does. The glucose levels were similar in both the groups probably due to the homeostatic mechanisms controlling the glycemia. Hormonal and metabo- lite analyses represent a good tool for understanding the physiological mechanisms required to meet higher reproductive performance.

[EN] Restriction of young rabbit females during rearing is a widespread management technique that could have negative consequences on their welfare and behaviour. In the present work, a total of 24 young rabbit females aged 9 wk were used to evaluate 3 rearing feeding programmes until first parturition: CAL, fed ad libitum with a control diet [C: 11.0 MJ digestible energy (DE) and 114 g digestible protein (DP) per kg dry matter (DM)]; CR, receiving the C diet restricted (140 g/d) from 12 wk of age; and F, fed ad libitum with a low energy/high fibre diet [F: 8.7 MJ DE and 88 g DP per kg DM]. F females presented lower body weight than CAL and CR females at week 18 (–0.4 kg and –0.2 kg; P, The authors thank the Spanish Ministry of Education and Science (Projects AGL2006-07596 and AGL2014-53405-C2-1-P) for the economic support to conduct this study.

This study compared reproductive performance and behaviour of does raised in a group-housing system and in a regular cage system. The group-housing pen was divided into different functional areas for suckling, resting, and eating and special hiding areas for kits when they had left the nest-boxes and does to favour the species specific behavioural traits. Does had access to their nest-box by means of an individual Electronic Nest-box Recognition System (ENRS) activated by a coded transponder placed in their eartags. Eight does were housed in each pen. Natural mating (NM, with a buck in the group) or artificial inseminations (AI) were applied. Litter size, kit mortality and kit weight at 14 d of age were similar for group-housing and cages when NM were applied. With a natural reproduction rhythm group-housing led to an increase of +38% of litters. However, from a management point of view, a cycled production system with AI is preferred. With AI and group-housing, a lower kindling rate and a lower kit weight at weaning were found. The lower kindling rate was partly caused by pseudo-pregnancies that were found in 23% (P < 0.01) of the does in the group-housing system against 0% in the control group. Sixteen to 20% of the does in the group-housing system had skin injuries, which is an indicator for aggression among does. Most of the injuries were seen on the body and most of them were superficial bites. Based on the results of this study, it can be concluded that group-housing of rabbit does seems possible, but more research is needed to solve the problems of the decreased kindling rate and occurrence of pseudo-pregnancies, the lower weight at weaning and aggressiveness among does.

In this study, we have examined the presence and the distribution of receptors for leptin (Ob-R) in the oviduct of rabbits, and the effects of leptin on the release of prostaglandin (PG) F2α and PGE2 and on the activity of nitric oxide (NO) synthase (NOS) by oviducts cultured in vitro. Rabbits were killed during the follicular phase and the oviducts were incubated in vitro with leptin, PGF2α, PGE2, NO donor and inhibitors of NOS and cyclo-oxigenase (COX). Using immunohistochemistry, Ob-R-like positive reaction was observed only in the cytoplasm of secretory cells, having stronger intensity in the infundibulum and ampulla tracts than in the isthmus. Both leptin and NO donor inhibited PGE2 release, whereas they enhanced PGF2α release; NOS inhibitor alone or with leptin increased PGE2 and decreased PGF2α production; NOS activity was enhanced by leptin, while PGs did not affect this enzyme. This study suggests that the oviduct could be a potential target for endocrine regulation by leptin, whose circulating levels may act as a metabolic signal modulating oviductal PG release through mediation of the NOS/NO system.

We studied leptin involvement in rabbit corpora lutea (CL) activity, and its post-transcriptional signalling pathway. The expression of leptin receptor (Ob-R) in rabbit ovary at day 9 of pseudopregnancy was evaluated by immunohistochemistry and Western blot analysis. The specificity of the Ob-R receptor antibodies was characterised by immunoprecipitation and competition with blocking peptide. Day 9 CL were incubated in vitro with leptin alone or with inhibitors of PLC (phospholipase C), PLD (phospholipase D), AC (adenylate cyclase), JAK (janus kinase), MAPK (mitogen-activated protein kinase) and both cAMP- and cGMP-specific PDE (phosphodiesterase). Prostaglandin F2α(PGF2α), PGE2 and progesterone levels were measured in the culture medium, while NOS (nitric oxide synthase) and cAMP- and cGMP- specific PDE activities were measured in CL tissue. Positive staining for Ob-R was found within the cytoplasm of large luteal cells of CL as well as in granulosa cells of follicles and oocytes. Immunoblots detected a band of about 99 kDa size in Ob-R immunoprecipitates from CL homogenates. This band was not detectable after pre-incubation of the primary antibody with the immunising leptin peptide. Leptin increased PGF2αand cAMP-specific PDE, decreased basal progesterone and did not affect PGE2 and NOS levels. Leptin used the JAK pathway in increasing PGF2α, and MAPK and cAMP-specific PDE in decreasing progesterone. This study supports a permissive luteolytic role for leptin in rabbit CL.

The gene expressions for macrophage chemoattractant protein-1 (MCP-1), interleukin (IL)-1β, IL-2 and p53 were examined by semi-quantitative RT-PCR in corpora lutea (CL) of rabbits during spontaneous luteolysis at days 13, 15, 18 and 22 of pseudopregnancy. In the same luteal tissue, total activity of nitric oxide (NO) synthase (NOS) and genes for both endothelial (eNOS) and inducible (iNOS) isoforms were also analysed. From day 13 to 15, MCP-1 and IL-1βmRNA levels rose (P≤ 0.01) almost 2-fold, and the transcript for p53 almost 8-fold, but then all dropped (P≤ 0.05) from day 18 onward. IL-2 mRNA abundance was higher (P≤ 0.01) on day 13 and then gradually declined. During luteolysis, eNOS mRNA decreased 40% (P≤ 0.05) by day 15, but thereafter remained unchanged, while iNOS mRNA was barely detectable and did not show any clear age-related pattern throughout the late luteal stages. Total NOS activity progressively increased (P≤ 0.01) from day 13 to 18 of pseudopregnancy and then dropped to the lowest (P≤ 0.01) levels on day 22. Luteal progesterone content also declined during CL regression from 411 to 17 pg/mg found on days 13 and 22 respectively, in parallel with the decrease in blood progesterone concentrations. These data further support a physiological role of NO as modulator of luteal demise in rabbits. Locally, luteal cytokines may be involved in the up-regulation of NOS activity, while downstream NO may inhibit steroroidogenesis and induce expression of p53 gene after removal of the protective action of progesterone.

An experiment was performed to assess the metabolic adaptation and hormonal regulation in young female rabbits during long-term food restriction and subsequent compensatory growth during rearing. Feeding level was either ad libitum (AL, no. = 52) or restricted (R, no. = 52). From 6 to 12 weeks of age, food intake ofR was kept at a constant level. This resulted in an increase in relative restriction as compared with AL to 0-54of AL intake at 12 weeks of age (restriction period). Thereafter food intake gradually increased to 0-95 of AL at 17 weeks of age (recovery period). During the last 5 days before insemination at 17-5 weeks of age, all animals were fed to appetite. Blood samples were taken weekly from 6 to 17 weeks of age from 11 animals in each group. Growth rate of R was reduced during the restricted period (29 (s.d. 2) v. 44 (s.d. 5) g/day for R and AL, respectively; P 0-05), but was higher in the recovery period (30 (s.d. 3) v. 27 (s.d. 4) g/day, respectively; P

Total activity of nitric oxide synthase (NOS) and the gene expression of both endothelial NOS (eNOS) and inducible NOS (iNOS) isoforms in corpora lutea of pseudopregnant rabbits were examined during prostaglandin F(2alpha) (PGF(2alpha))-induced luteolysis. Corpora lutea were collected at 0, 6, 12, 24 and 48 h after an injection of PGF(2alpha) at day 9 of pseudopregnancy. At 12 h after PGF(2alpha) administration, luteal mRNA encoding eNOS decreased (P

[EN] The aim of this study was to verify the effectiveness of post partum pharmacological oestrus synchronisation with alfaprostol or eCG in improving rabbit doe reproductive performances and to evaluate the incidence of pseudopregnancy in this species. One hundred and eighty commercial hybrid rabbit does, multiparous and lactating, were randomly divided into three groups (n=60) and treated on day 8 post partum (pp) as follows: Group A, 1 mL s.c. vitaminic solution with 200 mg alfaprostol, a synthetic analogue of PGF2a; Group B, 1 mL sc vitaminic solution with 20 I.U eCG; Group C (Control) 1 mL sc vitaminic solution. On day 11 pp (3 days after treatment), all the rabbits were artificially inseminated (AI), and injected with 0.8 mg of buserelin i.m. to induce ovulation. Concurrently, blood samples were taken for progesterone (P4) analysis by RIA with the following schedule: day 8 pp (before treatment), day 11 pp (before AI and the induction of ovulation) and day 16 pp (5 days after AI). Fertility was not influenced by hormonal treatments (71.7% and 71.2% respectively) compared to control (66.1%). Conversely, both PGF2a and eCG hormonal synchronisation treatments increased (P1 ng/mL). On day 5 after insemination 95.5% had P4 values, which attest to the presence of functional corpora lutea (CL) (P4>2 ng/mL). The results of this study show that as long as rabbitry is properly managed zootechnically and sanitarily, drugs such as alfaprostol and eCG, while not ameliorating fertility rate when used for post partum oestrus sychronisation, can increase litter size.

Total activity of nitric oxide (NO) synthase (NOS) and expression of both endothelial (eNOS) and inucible (iNOS) isoform were examined in corpora lutea (CL) of rabbits across pseudopregnancy by quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis, Western blot and immunohistochemistry. CL were collected at early- (day 4), mid- (day 9) and late- (day 13) luteal phase of pseudopregnancy. The PCR product of rabbit luteal eNOS was cloned and its direct sequence exhibited 90% homology with those of other species. The steady state mRNA levels encoding eNOS remained fairly constant throughout both early- and mid- luteal stages of pseudopregnancy but dropped almost to half (P£ 0.05) by day 13. By contrast, luteal eNOS proteins increased two-fold (P£ 0.05) from early- to late-luteal phase. Independently of CL age, iNOS mRNA was very poorly expressed while protein levels gradually declined from early- to late-luteal stage. Intense eNOS-like immunoreactivity was detected in large luteal cells, while iNOS staining was targeted to a few, isolated cells, probably macrophages. Basal NOS activity was greater in day 4 CL (18930 ± 535 d.p.m./mg wet tissue) than in both day 9 and day 13 CL (10460 ± 394 and 15430 ± 485 d.p.m., respectively). These data are the first to characterize in rabbit CL the temporal expression patterns of NOS isoforms across different luteal stages of pseudopregnancy and, collectively, suggest the existence of an expressional control for this constitutive isoform, which might have a physiological role in regulating CL function during development.

[EN] Using data from three different experiments, the effects of oestrus synchronisation by doe-litter separation (DLS), applied 48 hours befare artificial insemination from day 9 to day 11 post partum, on productive performance of free nursing rabbit does and their litters in relation to parity and number of successive DLS treatments were evaluated. lnitially, 200 New Zealand White does of different parity were homogeneously and definitively distributed in two groups (DLS and Control). On the whole, DLS improved fertility (+23.9%, Ps0.001 ), but reduced the growth of young rabbits from day 9 to weaning (-1.7 g/d, Ps0.01) and their weaning weight (-38 g, Ps0.01) at day 35. DLS also increased by 19.2%, 32.5% and 35.0% (Ps0.01) the fertility rate of does from parity 1 to 3, respectively. but was ineffective on does of parity, [FR] En utilisant des données de trois expériences différentes, nous avons évalué J'effet de la synchronisation de l'oestrus par séparation mere-jeune (SMJ), réalisée 48 heures avant insémination artificielle, du 9eme (J9) au 11 eme jour (J11) apres la mise bas, sur la productivité de lapines en allaitement libre et de leurs portées, en fonction de la parité et du nombre de SMJ successives. Deux cents lapines Néo Zélandais Blanc, de parité variable, ont été réparties de fa9on homogene et définitive en deux groupes (SMJ et témoin). Globalement, SMJ a amélioré Ja fertilité (+23.9%, P, This work was supported by a grant from "Ministero per l'Istruzione, l'Università e la Ricerca"

Studies were conducted to characterize receptors for prostaglandin (PG) F(2alpha) (PGF(2alpha)) and PGE(2), and the signalling pathways regulating total nitric oxide synthase activity and progesterone production in rabbit corpora lutea (CL) of different luteal stages. CL were obtained at days 4, 9 and 13 of pseudopregnancy and cultured in vitro for 2 h with PGF(2alpha) or PGE(2) and with activators and inhibitors of G protein (Gp), phospholipase C (PLC), protein kinase C (PKC), adenylate cyclase (AC) and protein kinase A (PKA). High affinity PGF(2alpha) receptor (K(d)=1.9+/-0.6 nM mean+/-s.e.m. ) concentrations increased (P< or =0.01) four- to five-fold from early to mid- and late-luteal phases (50.6+/-8.5, 188.3+/-36.1 and 231.4+/-38.8 fmol/mg protein respectively). By contrast, PGE(2) receptor (K(d)=1.6+/-0.5 nM) concentrations decreased (P< or =0.01) from day 4 to day 9 and 13 (27.5+/-7.7, 12.4+/-2.4 and 16.5+/-3.0 fmol/mg protein respectively). The Gp-dependent AC/PKA pathway was triggered only on day 4 CL, mimicking the PGE(2) treatment and increasing progesterone production. In both day 9 and day 13 CL, the Gp-activated PLC/PKC pathway evoked a luteolytic effect similar to that induced by PGF(2alpha). The time-dependent selective resistance to PGF(2alpha) and PGE(2) by rabbit CL is mediated by factors other than a lack of luteal receptor-ligand interactions.