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Several seminal plasma components, besides NGF, are implicated as ovulation-inducing factors in mammals. This study investigated the IL1B and its receptor IL1R1 in the testis (T), male accessory glands, prostate (P) and seminal vesicles (SV), and uterus (U) of adult rabbits using immunohistochemistry (IHC) and quantitative reverse transcription PCR (RT-qPCR). We also assessed the presence of IL1B in seminal plasma through Western blotting (WB) and examined the interaction between IL1B and NGF in vitro by measuring their production with enzyme-linked immunosorbent assay (ELISA) in the presence of NGF and IL1B alone or with their respective receptor antagonists. IHC revealed IL1B system expression in all reproductive organs studied, with IL1B and IL1R1 localized to the germinative epithelium of the T and the epithelial cells of the accessory glands and U. IL1B gene transcript levels were significantly higher ( p < 0.01) in the P and SV compared to the T, while IL1R1 levels were significantly higher ( p < 0.001) in the P compared to the other tissues, while IL1R1 levels were three times higher ( p < 0.001) in the P. WB confirmed the presence of IL1B in seminal plasma with a 30-35 kDa band. The in vitro study demonstrated that IL1B increased ( p < 0.05) basal NGF production in the U, whereas NGF had no effect on IL1B production. These findings provide evidence of the expression of the IL1B/IL1R1 system in both male and female rabbit reproductive tracts and suggest that IL1B in seminal plasma may influence uterine endocrine activity. The results propose a potential role for IL1B in ovulation, in conjunction with NGF, supporting that ovulation may involve inflammatory-like processes.

We study and characterize the inclusion relations of global classes in the general weight matrix framework in terms of growth relations for the defining weight matrices. We consider the Roumieu and Beurling cases, and as a particular case, we also treat the classical weight function and weight sequence cases. Moreover, we construct a weight sequence which is oscillating around any weight sequence which satisfies some minimal conditions and, in particular, around the critical weight sequence ( p ! ) 1 / 2 , related with the non-triviality of the classes. Finally, we also obtain comparison results both on classes defined by weight functions that can be defined by weight sequences and conversely., (© The Author(s) 2024.)

The use of contrast-enhanced ultrasound (CEUS) has been widely reported for reproductive imaging in humans and animals. This review aims to analyze the utility of CEUS in characterizing canine reproductive physiology and pathologies. In September 2022, a search for articles about CEUS in canine testicles, prostate, uterus, placenta, and mammary glands was conducted on PubMed and Scopus from 1990 to 2022, showing 36 total results. CEUS differentiated testicular abnormalities and neoplastic lesions, but it could not characterize tumors. In prostatic diseases, CEUS in dogs was widely studied in animal models for prostatic cancer treatment. In veterinary medicine, this diagnostic tool could distinguish prostatic adenocarcinomas. In ovaries, CEUS differentiated the follicular phases. In CEH-pyometra syndrome, it showed a different enhancement between endometrium and cysts, and highlighted angiogenesis. CEUS was shown to be safe in pregnant dogs and was able to assess normal and abnormal fetal-maternal blood flow and placental dysfunction. In normal mammary glands, CEUS showed vascularization only in diestrus, with differences between mammary glands. CEUS was not specific for neoplastic versus non-neoplastic masses and for benign tumors, except for complex carcinomas and neoplastic vascularization. Works on CEUS showed its usefulness in a wide spectrum of pathologies of this non-invasive, reliable diagnostic procedure.

In this retrospective study, we assessed the accuracy of different blood flow imaging in diagnosing testicular tumor types in dogs. We recruited 27 dogs with leydigomas (14), seminomas (eight), sertoliomas (six), and mixed cells (five) confirmed histopathologically. In intact dogs, Pampiniform plexus and marginal arteries were scanned through pulsed Doppler. Blood flow and presence of intralesional/perilesional arteries were assessed by color and power Doppler, B-flow, and contrast-enhanced ultrasound. Tumor types did not differ by B-Mode ultrasonography characters. Pampiniform and testicular arteries of sertoliomas had higher ( p < 0.05) pulsatility and resistive indexes. The proportion of leydigomas with a perilesional and/or perilesional/intralesional blood flow pattern detected by color and pulsed Doppler and B-flow was higher ( p < 0.05) than that of the other tumors counted together. This resulted in a sensitivity of 81.8%, 83.3%, and 85.7%, a specificity of 76.5%, 56.3%, and 73.7%, and a correct classification rate of 78.6%, 67.9%, and 78.8%, respectively. While contrast enhanced ultrasound was highly effective in detecting all tumors, qualitative and quantitative parameters did not contribute to their differential diagnosis. In conclusion, results indicate that different testicular tumor types of dogs have subtly different vascular patterns, a condition that could help in identifying leydigomas.

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.

We prove that the Hermite functions are an absolute Schauder basis for many global weighted spaces of ultradifferentiable functions in the matrix weighted setting and we determine also the corresponding coefficient spaces, thus extending the previous work by Langenbruch. As a consequence, we give very general conditions for these spaces to be nuclear. In particular, we obtain the corresponding results for spaces defined by weight functions., (© The Author(s) 2020.)

Stimuli that lead to the release of gonadotropin-releasing hormone (GnRH) and pituitary gonadotropins and, consequently, ovulation in mammals fall into two broad categories. In the first, high plasma oestrogen concentrations induce the events that trigger ovulation, a characteristic of spontaneous ovulators. In the second, nerve stimuli occurring during mating reach the hypothalamus and trigger the release of GnRH and ovulation with a neuroendocrine reflex that characterizes induced ovulators.In this review, we will give an overview of the distribution of NGF and its expression in the different tissues of the male accessory sex glands, the main sites of NGF production. Next, we will highlight the role of NGF in sperm function and its potential cryopreserving role in artificial insemination techniques. Finally, we will evaluate the functions of NGF in ovulation, particularly in induced ovulators. Overall, the information obtained so far indicates that NGF is widely distributed in organs that regulate the reproductive activity, in both males and females. In spontaneous ovulators, NGF exerts mainly a luteotrophic action, while, in induced ovulators it is the main ovulation-inducing factor. A better understanding of the role of NGF in reproduction would be of great interest, since it could help finding innovative therapeutic aids to improve mammalian fertility., (© 2021. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

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 male and female reproductive functions, with a focus on animals of veterinary interest. In females, resveratrol has been considered as 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 males, resveratrol enhances testes function and spermatogenesis through activation of the 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.

The objectives of this study were to evaluate in horse testes the expression of kisspeptin (KiSS) and GnRH1 neuropeptides and their cognate receptors, KiSS1R and GnRH1R, as well as their action on testosterone, GnRH1, prostaglandin F2α (PGF2α), and PGE2 synthesis and cyclooxygenase 1 (COX1) and COX2 activity by Leydig cells in vitro. Testes were obtained from 9 sexually mature horses by surgical castration. Immunohistochemistry, evidenced the presence of KiSS, KiSS1R, GnRH, and GnRH1R in Leydig cells, whereas germinal and Sertoli cells were positive only for GnRH1. Transcripts for both neuropeptides and their cognate receptors were revealed in isolated Leydig cells by RT-PCR. Isolated and purified Leydig cells were in vitro cultured with agonists and antagonists of KiSS (KiSS-10 and KiSS-234, respectively) and GnRH1 (buserelin and antide, respectively). KiSS-10 and buserelin increased (P < 0.01) COX1 activity and testosterone and PGF2α basal secretion, while decreased (P < 0.01) that of PGE2. KiSS-10 and buserelin did not affect COX2 activity. GnRH1 basal production was increased (P < 0.01) by KiSS-10, but not by buserelin. Antide counteracted the KiSS and GnRH1 effects, whereas KiSS-234 influence only those of KiSS. Summarizing, the KiSS/GnRH1 system is present in horse Leydig cells and modulates their endocrine activity. In particular, the endocrine effects of KiSS are mediated by GnRH1, so suggesting that hypothalamic-like interaction between KiSS and GnRH1 occurs also in Leydig cells., (Copyright © 2020. Published by Elsevier Inc.)

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 ± 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.

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., (© 2019 Blackwell Verlag GmbH.)

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<0.05). During 1st lactation, the perirenal fat thickness change in CAL females was not different from 0 (+0.02 mm), while in the other four groups it increased (on av. +0.44 mm; P<0.05). Plasma of females fed with F diet during rearing (F, FC and FCF) had lower non-esterified fatty acids content than those exclusively fed with C diet (-0.088 and -0.072 mmol/l compared to CAL and CR, respectively; P<0.05). FCF litters had higher weight than F litters at day 21 of lactation (+247 g; P<0.05), but FCF litter had significantly lower weight than FC litters at weaning (+170 g; P<0.05). CR females had the shortest average interval between the 1st and 2nd parturition (49 days) and FCF females the longest (+ 9 days compared to CR; P<0.05). At 2nd parturition, liveborn litters of F females were larger and heavier than litters of FCF females (+2.22 kits and +138 g; P<0.05), probably due to the lower mortality at birth of F litters (-16.5 percentage points; P<0.05). In conclusion, rearing females on fibrous diets seems to increase the ability of primiparous rabbit females to obtain resources, especially at the onset of lactation.

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.

In the present study, we evaluated the dynamic changes of intra-ovarian blood flow, by real-time colour-coded and pulsed Doppler ultrasonography, as well as the immunopresence of prostaglandin F2α (PGF2α) receptor (FP) and peripheral plasma progesterone concentrations in pseudopregnant rabbit after PGF2α treatments at either early- (4 days) and mid-luteal (9 days) stages. During the pre-treatment observation interval of one hour, the ovarian blood flows showed a fluctuating pattern. Independently of luteal stage, PGF2α administration caused a fourfold decline in the blood flow within 40 min that was followed 50 min later by a reactive hyperaemia that lasted several hours, while the resistive index showed an opposite trend. Twenty-four hour later, the blood flow was one half that measured before PGF2α injection. At day 4 of pseudopregnancy, PGF2α did not affect peripheral plasma progesterone concentrations, but at day 9, it caused functional luteolysis as progesterone levels declined 6 hr later to reach basal values after 24 hr. The changes in the ovarian blood flows of pseudopregnant rabbits receiving PGF2α were accompanied by simultaneous changes in the resistance index. This biphasic response in the blood flow and vascular resistances likely reflects reactive hyperaemia following vasoconstriction. By immunohistochemistry, strong positive immune reaction for FP was detected in the cytoplasm of endothelial cells of ovarian arteries, veins and capillaries. In conclusion, these results suggest that PGF2α could acutely regulate the ovarian blood flow of pseudopregnant rabbits, even if there is no evidence of a blood flow reduction anticipating luteolysis., (© 2018 Blackwell Verlag GmbH.)

We evaluated the potential usefulness of CEUS to assess fetal-maternal circulation during pregnancy in dogs. Nine bitches were examined at 23, 30, and 45 days of gestation using an ultrasound machine (LOGIQ E9) and SonoVue ® contrast media as echo-signal enhancer. Qualitative and quantitative evaluation of contrast enhancement patterns of uterine artery and utero/placental vessels were performed on recorded images. Independently of the gestational periods, the qualitative evaluation showed the initial wash-in phase from the first appearance of the uterine artery to the rapid distribution in embryonic vesicles or placenta to the progressive washout, whilst there was no enhancement of either embryos or fetuses in any bitch. Independent of gestational age, parameters derived from quantitative analysis of time intensity-curves of contrast enhancement (peak intensity, time to peak, rise time, washout) did not vary between proximal placenta, distal placenta, and uterine artery. With the progression of gestation, AUC values did not change in both proximal and distal placenta, but in the uterine artery it was lower (P ≤ 0.05) at day 30 than at day 23 (464.8 ± 16.1 vs.596.4 ± 28.1, respectively). In conclusion, CEUS appears to safely permit evaluation of the maternal and fetal vessels in the first two third of gestation, without any clinically relevant adverse effects. Further studies in a larger number of bitches in different stages of pregnancy are needed to establish standard parameters for normal pregnancies that can be used to detect abnormalities of pregnancy., (Copyright © 2018 Elsevier Inc. All rights reserved.)

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., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

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.

Gall-bladder diseases are common in dogs and two-dimensional ultrasonography is a current standard method for diagnosis and treatment planning. However, findings from this modality can be nonspecific. The aim of this retrospective, case series study was to describe conventional and contrast-enhanced ultrasound (using SonoVue ® ) findings in a group of dogs with histologically confirmed gall bladder disease. A total of 65 dogs were included. Branchlike, heterogeneous, and homogeneous contrast enhancement of echogenic intraluminal mass-forming lesions was a contrast-enhanced ultrasound characteristic of polypoid lesions due to cystic mucosal hyperplasia of the gallbladder and/or tumor, which had different wash-in and washout characteristics. In dogs with mobile or immobile biliary sludge or mucocele, the echogenic intraluminal masses remained unenhanced. A double rim mark or enhancement defect in the gallbladder wall was a characteristic of edema or necrosis/rupture of the wall, respectively. Conventional ultrasonography correctly identified biliary sludge or mucocele in 36/37 dogs, cholecystitis/edema in 44/47 dogs, necrosis/rupture in 19/25 dogs, and gallbladder neoplasia in three of three dogs with these pathologies. It falsely identified biliary sludge or mucocele in eight of 28 dogs, cholecystitis/edema in three of 15 dogs, necrosis/rupture in 13/37 dogs, and gall-bladder neoplasia in 20/59 dogs that did not have these pathologies. Contrast-enhanced ultrasound correctly identified cholecystitis/edema in 42/47 dogs, but falsely identified cholecystitis/edema in three of 18 dogs. It correctly identified necrosis/rupture, benign polypoid lesions, and gallbladder neoplasia in all dogs with no false-positive results. Findings supported contrast-enhanced ultrasound as a complement to conventional ultrasonography for dogs with suspected gallbladder pathologies such as edema, necrosis, and rupture., (© 2018 American College of Veterinary Radiology.)

We explored the expression and cell type distribution of cannabinoid receptors type 1 (CB1) and cannabinoid receptors type 2 (CB2) in the mandibular glands of pigs in relation to different physical forms of the diet. Thirty-two crossbred growing pigs (ages 5-6 weeks) were randomly allotted to four experimental groups (eight pigs/group) and fed four different physical types of the same diet for 4 weeks: finely ground pellet (FP), coarsely ground meal (CM), coarsely ground pellet (CP) and coarsely ground extruded (CE) with dMEAN of 0.46, 0.88, 0.84 and 0.66 mm respectively. At the end of the feeding trial, the pigs were euthanized and the mandibular gland was collected after dissection. By immunohistochemistry, positive signals for CB1 were found in the cytoplasm of duct epithelial cells of pigs fed CP, FP and CE diets and in the serous cells of mixed acini in pigs fed the coarser CM diet. Positive signals for CB2 were detected in duct epithelial cells and in neurons of ganglia close to major secretory ducts of all pigs. The differential expression and localization of these receptors in response to variable chewing activity due to the type of diet suggest that endocannabinoids may influence the functional activity of the mandibular gland by modifying qualitative and/or quantitative aspects of salivary secretion., (© 2017 Blackwell Verlag GmbH.)

Pregnancy and lactation represent critical periods of canine reproductive life and different hormones are required to maintain homeostasis and the correct energy requirements. The aim of this study was to evaluate leptin, insulin, triiodotyronine (T3), thyroxin (T4) and cortisol serum concentrations to better describe the canine maternal energy metabolism. Twelve Bloodhound bitches that had at least one litter and were considered to be healthy submitted blood samples every 15 days starting from day 0 (ovulation) throughout pregnancy up to the end of lactation (day 120). We found that leptin concentrations increased from day 0 to reach maximum value at day 45 (P < 0.001), decreased at day 60 (P < 0.05) and rose again at day 105 and day 120 (P < 0.05 and P < 0.01 respectively). Insulin concentrations significantly increased during pregnancy peaking at day 60 (P < 0.05). After parturition dropped and reached progressively lower values similar to those at day 0 at the end of lactation. During pregnancy Body Weight was in low correlation with leptin (r = 0.32, P < 0.01) and there was no correlation with insulin (r = 0.13, P > 0.1); moreover, no correlation was found during lactation. T3 concentrations showed a U-shaped trend with a significant reduction at day 75 (P < 0.05). T4 concentrations fluctuated without any significant change (ranging from 27.0 ± 1.6 nmol/l to 32.1 ± 1.9 nmol/l). Cortisol concentrations ranged within the reference values (minimal value at day 30 (49.8 ± 6.3 nm/L) and maximal value at day 45 (72.5 ± 16.1 nm/L)). Our study states that canine pregnancy and lactation evoke many hormonal changes necessary to mother and fetuses. In particular, this is the first report regarding the pattern of leptin during whole pregnancy and lactation. The present work lays the groundwork for further studies on endocrine homeostatic mechanisms using the dog as a model for human medicine., (Copyright © 2017. Published by Elsevier Inc.)

Background: Cardiac output (Q˙) is a key parameter in the assessment of cardiac function, its measurement being crucial for the diagnosis, treatment, and prognostic evaluation of all heart diseases. Until recently, Q˙ determination at peak exercise has been possible through invasive methods, so that normal values were obtained in studies based on small populations., Methods: Nowadays, peak Q˙ can be measured noninvasively by means of the inert gas rebreathing (IGR) technique. The present study was undertaken to provide reference values for peak Q˙ in the normal general population and to obtain a formula able to estimate peak exercise Q˙ from measured peak oxygen uptake (V˙o 2 )., Results: We studied 500 normal subjects (age, 44.9 ± 1.5 years; range, 18-77 years; 260 men, 240 women) who underwent a maximal cardiopulmonary exercise test with peak Q˙ measurement by IGR. In the overall study sample, peak Q˙ was 13.2 ± 3.5 L/min (men, 15.3 ± 3.3 L/min; women, 11.0 ± 2.0 L/min; P < .001) and peak V˙o 2 was 95% ± 18% of the maximum predicted value (men, 95% ± 19%; women, 95% ± 18%). Peak V˙o 2 and peak Q˙ progressively decreased with age (R 2 , 0.082; P < .001; and R 2 , 0.144; P < .001, respectively). The V˙o 2 -derived formula to measure Q˙ at peak exercise was (4.4 × peak V˙o 2 ) + 4.3 in the overall study cohort, (4.3 × peak V˙o 2 ) + 4.5 in men, and (4.9 × peak V˙o 2 ) + 3.6 in women., Conclusions: The simultaneous measurement of Q˙ and V˙o 2 at peak exercise in a large sample of healthy subjects provided an equation to predict peak Q˙ from peak V˙o 2 values., (Copyright © 2017 American College of Chest Physicians. Published by Elsevier Inc. All rights reserved.)

Gall bladder necrosis and rupture are life-threatening conditions in dogs requiring surgical intervention and early diagnosis is essential. Human patients with suspected gall bladder necrosis/rupture are commonly evaluated with contrast-enhanced ultrasonography (CEUS), however this procedure has not been described in dogs with suspected gall bladder necrosis/rupture. In a prospective diagnostic cohort study, CEUS (using SonoVue contrast medium) was performed in 93 dogs with gallbladder lesions identified by abdominal conventional ultrasonography. Necrosis/rupture was identified by CEUS as a focal lack of enhancement of the gallbladder wall. Dogs with positive CEUS finding for necrosis/rupture (complete lack of regional wall enhancement) underwent immediate surgery as did dogs with other biliary disorders requiring surgery. Dogs with negative CEUS findings or those not requiring surgery were managed medically. In cases undergoing surgery, necrosis/rupture was confirmed intraoperatively (and via histopathology). Absence of necrosis/rupture was confirmed either intraoperatively (via histopathology) or was assumed to be absent by complete recovery with medical management. Forty-nine dogs underwent surgery and cholecystectomy: 24 had necrosis/rupture. CEUS was more accurate (100% sensitive and specific) in diagnosing gallbladder wall necrosis/rupture than conventional ultrasonography (75% sensitive and 81% specific) (P < 0.03). In conclusion, CEUS provides accurate characterization of gallbladder wall integrity that can impact decisions regarding clinical management, either surgical or medical., (© 2016 American College of Veterinary Radiology.)

The aim of the present study was to evaluate: (1) the presence of nerve growth factor (NGF), neurotrophic tyrosine kinase receptor 1 (NTRK1), and nerve growth factor receptor (NGFR) in the rabbit uterus; and (2) the in vitro effects of NGF on PGF2α and PGE2 synthesis and on the PGE2-9-ketoreductase (PGE2-9-K) activity by the rabbit uterus. Nerve growth factor, NTRK1, and NGFR were immunolocalized in the luminal and glandular epithelium and stroma cells of the endometrium. reverse transcriptase polymerase chain reaction indicated the presence of messenger RNA for NGF, NTRK1, and NGFR in the uterus. Nerve growth factor increased (P < 0.01) in vitro secretions of PGF2α and PGE2 but coincubation with either NTRK1 or oxide nitric synthase (NOS) inhibitors reduced (P < 0.01) PGF2α production and blocked (P < 0.01) PGE2 secretion. Prostaglandins releases were lower (P < 0.01) than control when uterine samples were treated with NGF plus cyclooxygenase inhibitor. However, addition of NGFR inhibitor reduced (P < 0.01) PGF2α secretion less efficiently than NTRK1 or NOS inhibitors but had no effect on PGE2 yield. Nerve growth factor increased (P < 0.01) the activity of PGE2-9-K, whereas coincubation with NTRK1 or NOS inhibitors abolished (P < 0.01) this increase in PGE2-9-K activity. However, cotreatment with either cyclooxygenase or NGFR inhibitors had no effect on PGE2-9-K activity. This is the first study to document the distribution of NGF/NTRK1 and NGFR systems and their effects on prostaglandin synthesis in the rabbit uterus. NGF/NTRK1 increases PGF2α and PGE2 productions by upregulating NOS and PGE2-9-K activities, whereas NGF/NGFR augments only PGF2α secretion, through an intracellular mechanism that is still unknown., (Copyright © 2016 Elsevier Inc. All rights reserved.)

We evaluated the diagnostic accuracy of the contrast-enhanced ultrasonography (CEUS), using a second-generation microbubble contrast agent, in differentiating the different types of adrenal mass lesions in 24 dogs. At B-mode ultrasound, 9 lesions involved the right adrenal gland, 14 the left, and 1 was bilateral. Each dog received a bolus of the contrast agent into the cephalic vein, immediately followed by a 5-mL saline flush. The first contrast enhancement of each adrenal lesion was evaluated qualitatively to assess the degree of enhancement and its distribution during the wash-in and wash-out phases, as well as the presence of non-vascularized areas and specific vascular patterns. Pathological diagnoses were determined in all dogs by histopathology or by cytology. Combining enhancement degree and vascularity resulted in the best predictive model, allowing CEUS to differentiate adrenocortical adenoma (n=10), adenocarcinoma (n=7), and pheochromocytoma (n=7) with an accuracy of 91.7% (P < 0.001). Combining enhancement degree and vascularity, CEUS can discriminate malignant versus benign adrenal lesions with a sensitivity of 100.0%, a specificity of 80.0%, and an accuracy of 91.7% (P < 0.001). In conclusion, results of this study confirm that CEUS is useful for differentiating between the different types of adrenal tumors in dogs.

Experiments were devised to characterize the expression of nerve growth factor, beta polypeptide (NGF), and its cognate receptors neurotrophic tyrosine kinase receptor type 1 (NTRK1) and nerve growth factor receptor (NGFR) in rabbit male sex organs, as well as the concentrations of NGF in both seminal and blood plasma of sexually mature male rabbits. Immunoreactivity and gene expression for NGF and cognate receptors were detected in testis, prostate gland and seminal vesicle. The highest levels of NGF and NTRK1 transcripts were found in the prostate, while intermediate expressions were found in the testis. NGFR transcripts were expressed at the same levels in both testis and prostate and were more abundant than in seminal vesicles. The widespread distribution of NGF in all prostate glandular cells, together with its relative high mRNA abundance, confirms that the prostate of rabbits is the main source of this neurotrophin. In conclusion, the present data suggest that the NGF system is involved in the testicular development and spermatogenesis of rabbits and that NGF may act as a potential ovulation-inducing factor being abundantly present in the seminal plasma., (© 2015 Blackwell Verlag GmbH.)

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., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Leptin is a polypeptide secreted by adipocytes which binds to a specific receptor (Ob-R) that is expressed in various tissues. The wide distribution of the Ob-R suggests that leptin might exert diverse biological functions, not only by regulating energy metabolism and appetite, but also by acting as a mitogen in many cell types, including keratinocytes. In this study, the presence and localization of Ob-R was investigated in the skin of the dog using RT-PCR and immunohistochemical techniques. RT-PCR revealed the presence of Ob-R m-RNA in the skin specimens collected from the dorsal region of two smooth coat breed dogs. Through immunohistochemistry performed on the skin of five dogs, the expression of the receptor was observed in the basal layer of the epidermis, in the hair follicles as well as in the apocrine sweat and sebaceous glands. No staining for Ob-R was detected in the suprabasal epidermis layers. Strong positive signals were observed in many cells of the outer root sheath of hair follicles in growing and in regressive phases. The identification of Ob-R in the above targets suggests that leptin may play a role in the regulation of cyclic renewal of the epidermis and skin appendages in dog. This study represents an important contribution to understand the complex mechanisms that are involved in the skin biology in this species., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

Cardiopulmonary exercise testing (CPET) is a procedure widely used in daily clinical activity to investigate cardiac and pulmonary disorders. Peak oxygen consumption (VO2 peak) is the most validated and clinically accepted parameter used to report aerobic capacity in healthy individuals and in different clinical settings. However, peak VO2 is influenced by several factors, whose variability is nowadays particularly evident due to the extensive use of CPET even in very young and very old subgroups of patients. Thus, its diagnostic and prognostic significance may be improved by the use of % of predicted VO2. At present, many sets of normal values are available, making the identification of the most proper max VO2 predicted value a challenging problem. In fact, normal value sets have been obtained from studies whose accuracy was reduced by important limitations, such as small sample size, low grade of heterogeneity of the population enrolled, poor rigorousness of methods, or difficulty in interpreting results. Accordingly, the aim of the present review is threefold: (A) to report some illustrative cases to show how the choice of the normal value set can influence the report of CPET; (B) to describe the most known and used reference value sets, highlighting the main characteristics of sample population, the most important methodological aspects, and the major limitations of the studies; (C) to suggest which equation should be used, if any, and to underline its weakness.

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.

This is a review of original data concerning new extra- and intracellular regulators of rabbit ovarian functions. Effects of some hormones including leptin, ghrelin, oxytocin, arginine-vasotocin, endothelin (ET-1), gonadotropin releasing hormone (GnRH), adrenocorticotropic hormone (ACTH), growth factors such as insulin-like growth factor-I (IGF-I), epidermal growth factor (EGF), nuclear peroxisome proliferator-activated receptor gamma (PPARγ), pharmacological regulators of some protein kinases such as protein kinase A (PKA), mitogen-activated protein (MAP) kinase, cell division cycle protein 2 homolog (CDC2 kinase, CDK), tyrosine kinases), and plant molecules (resveratrol, rapamycin) on the functions of ovarian cells (proliferation, apoptosis, secretory activity, expression of some protein kinases) and reproductive end points (blood level of reproductive hormones, ovarian morphology, number of ovulations, embryo yield and quality, number and viability of offspring), and their possible interrelationships and practical application in rabbit breeding are reviewed., (Copyright © 2014 Elsevier B.V. All rights reserved.)

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.

In brainstem slices of male rats, we examined in single neurons of the medial vestibular nucleus (MVN) the effect of exogenous administration of estrogenic (17β-estradiol, E2) and androgenic (5α-dihydrotestosterone, DHT) steroids on the synaptic response to vestibular afferent stimulation. By whole cell patch clamp recordings we showed that E2 induced synaptic long-term potentiation (LTP) that was cancelled by the subsequent administration of DHT. Conversely, DHT induced synaptic long-term depression (LTD) that was partially reversed by E2. The electrophysiological findings were supported by immunohistochemical analysis showing the presence of estrogen (ER: α and β) and androgen receptors (AR) in the MVN neurons. We found that a large number of neurons were immunoreactive for ERα, ERβ, and AR and most of them co-localized ERβ and AR. We also showed the presence of P450-aromatase (ARO) in the MVN neurons, clearly proving that E2 can be locally synthesized in the MVN. On the whole, these results demonstrate a role of estrogenic and androgenic signals in modulating vestibular synaptic plasticity and suggest that the enhancement or depression of vestibular synaptic response may depend on the local conversion of T into E2 or DHT., (Copyright © 2013 Elsevier Inc. All rights reserved.)

A noninvasive method for quantifying adrenal gland vascular patterns could be helpful for improving detection of adrenal gland disease in dogs. The purpose of this retrospective study was to compare the contrast-enhanced ultrasound (CEUS) characteristics of adrenal glands in 18 dogs with pituitary-dependent hyperadrenocorticism (PDH) vs. four clinically healthy dogs. Each dog received a bolus of the contrast agent (SonoVue®, 0.03 ml/kg of body weight) into the cephalic vein, immediately followed by a 5 ml saline flush. Dynamic contrast enhancement was analyzed using time-intensity curves in two regions of interest drawn manually in the caudal part of the adrenal cortex and medulla, respectively. In healthy dogs, contrast enhancement distribution was homogeneous and exhibited increased intensity from the medulla to the cortex. In the washout phase, there was a gradual and homogeneous decrease of enhancement of the adrenal gland. For all dogs with PDH, there was rapid, chaotic, and simultaneous contrast enhancement in both the medulla and cortex. Three distinct perfusion patterns were observed. Peak perfusion intensity was approximately twice as high (P < 0.05) in dogs with PDH compared with that of healthy dogs (28.90 ± 10.36 vs. 48.47 ± 15.28, respectively). In dogs with PDH, adrenal blood flow and blood volume values were approximately two- to fourfold (P < 0.05) greater than those of controls. Findings from the present study support the use of CEUS as a clinical tool for characterizing canine adrenal gland disease based on changes in vascular patterns., (© 2013 Veterinary Radiology & Ultrasound.)

The study was designed to examine the aglepristone (RU534) mechanisms affecting the corpora lutea (CL) lifespan in pseudopregnant rabbits. Aglepristone (10 mg/kg b.w.) was injected subcutaneously twice at either early- or mid-luteal phase (Days 3 and 4, or Days 8 and 9, respectively) after induction of ovulation with GnRH (Day 0). Corpora lutea and uteri, explanted at days 6 and 11, were evaluated for immunohistochemistry and Western blotting of progesterone (PR) and estrogen (ER) receptors, cyclooxygenase 1 (COX1), COX2, and PGE2-9-ketoreductase (PGE2-9-K) enzymatic activities, and progesterone, PGF2α, and PGE2 in vitro synthesis. Independent of luteal stage, aglepristone prolonged the functional luteal phase by 3 Days over that of controls as assessed by blood progesterone profiles. Aglepristone decreased protein for ER during both luteal-stages in CL and uteri. Progesterone receptor protein was decreased by RU354 at Days 6 in the uterus and at Days 11 in CL, whereas RU534 increased PR at Days 11 in uteri. In the CL, RU534 enhanced progesterone production at Days 6 and 11, whereas it decreased PGF2α and increased PGE2 at Day 11. In the uteri, RU534 decreased PGF2α and increased PGE2 synthesis at both days. COX2 and PGE2-9K activities were decreased by RU534 in the CL at Day 11, whereas in the uteri COX2 increased and PGE2-9-K decreased at Days 6 and 11. In conclusion, these data on aglepristone effects suggest that progesterone has a regulatory role on luteal function through direct and uterine-mediated mechanisms in pseudopregnant rabbits., (Copyright © 2013 Elsevier B.V. All rights reserved.)

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 3beta-hydroxysteroid 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 15d-PGJ2 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.

The aim of the study was to further characterize the relationship between hemodynamic changes in the ovary and luteal function in pregnant and nonpregnant bitches. Fourteen German Shepherd bitches were monitored three times a week from the first day of cytological diestrus (D1) until parturition or the end of diestrus (progesterone <2 ng/mL) by color Doppler, pulsed wave spectral Doppler, and power Doppler (PD) ultrasonography. By means of PD the total number of color pixels were calculated. The Doppler parameters evaluated were: peak systolic velocity (PSV), end diastolic velocity (EDV), and both resistive and pulsatility indices. Blood samples were collected three times a week throughout the experiment to determine progesterone (P4) concentrations. The length of diestrus in pregnant versus nonpregnant group was significantly shorter (P < 0.01; 57 ± 1 vs. 63 ± 1, respectively). By means of pulsed wave spectral Doppler the waveform showed a typical pattern of a low-resistive vessel characterized by a rapid systolic peak followed by a slow telediastolic decrease with a relatively high end-diastolic velocity. Blood flow parameters did not differ between left and right ovary. In both groups PSV and EDV showed a gradual decrease with the progress of diestrus; however, the values of PSV and EDV were significantly higher (P < 0.05) in the pregnant group versus nonpregnant group from D31 to D61 and from D49 to D58 respectively. Moreover, a significantly decrease (P < 0.05) of PSV and EDV in the pregnant group was observed from D46 to D58 and from D49 to D55, respectively. The resistive and pulsatility indices showed an increase during diestrus and the values were significantly lower (P < 0.05) in the pregnant group from D49 to D61. By means of PD, the pixel number was significantly higher (P < 0.05) in the pregnant versus nonpregnant group from D40 to D61. In particular, a significant decrease (P < 0.05) in the pixel number in the pregnant group was observed from D46 to D61. The comparison of the P4 values with the ovarian pixel number in the pregnant and nonpregnant group showed a direct correlation (r = 0.792, N = 59 and r = 0.774, N = 59, respectively). In particular, the P4 values were higher (P < 0.05) in the pregnant than in the nonpregnant group from D37 to D52. In conclusion, significant physiological differences between pregnant and nonpregnant bitches in terms of P4 and ovarian blood supply are reported. In addition it was possible to define that blood flow pattern during diestrus in pregnant bitches is not always closely related with P4 production., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The aim of the present study was to investigate the presence and the distribution of leptin (Ob) and its receptor (ObR) in the feline placenta at term by means of immunohistochemical techniques. A few Ob-positive cells were observed scattered in the lamellae of the labyrinthine placenta. These cells had the morphological characteristics typical of the very abundant cells in the placenta of cats that can be considered as being decidual and, in some cases, syncytiotrophoblastic cells. A few ObR-positive cells were observed in the same placental portion and were mainly localized in the lamellae, showing morphological features typical of decidual and syncytiotrophoblastic cells. No other structure of the placenta or the uterine wall showed positive reaction to the antibodies used. Our results confirm what has already been demonstrated in humans and laboratory animals, but not in domestic animals. Together with other emerging data on the secretory activities of the feline placenta, our study underlines its relevance in the production of molecules long known to be involved in appetite control and, probably, with potential effects on the developing fetus., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

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.

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 (+0.41 kg and +0.6 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 (+0.3 mm; P < 0.05). In comparison with FCF, CAL females had smaller and thinner live born litters (-2.5 kits and -139 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 (-1.7 kits; P < 0.05) and the greater mortality of their litters at birth (+13.9%; 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 +0.15 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 aim of the present study was to investigate the presence and distribution of cells containing orexin A (OXA), and orexin type 1 and 2 receptors (OX1R and OX2R, respectively) in the feline placenta by means of immunohistochemical technique. OXA was identified in several decidual and syncytiotrophoblastic cells present in the lamellar portion of the placenta. In the same placental structures, few decidual and syncytiotrophoblastic cells showed the presence of OX1R-like immunoreactivity. Characteristically, immunopositivity for OX2R, but not for OX1R, was evidenced in the cells of the glandular layer. The orexinic system was not expressed in the uterine structures that were not engaged by the chorion. Our results provide the first evidence of the presence of a placental orexinic system in a mammalian species. Orexin A and both OX1R and OX2R are unequally distributed within the cat placenta. Local OXA production and the presence of specific receptors, differentially expressed in the placental structures of the cat, suggest that the orexinic system may participate in placental growth and development as well as in the regulation of its steroidogenic capacity via endocrine, paracrine and/or autocrine mechanisms., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The aim of the present work was to study, by means of immunohistochemical and RT-PCR techniques, the presence and distribution of immunopositivity for orexin A and B (OXA and OXB) and orexin type 1 and 2 receptors (OX(1)R and OX(2)R) in the lacrimal gland of sheep as well as the gene expressions for prepro-orexin (PPOX) and cognate receptors. In serial sections, positive staining for OXA and OXB were localized in the same nervous fibers within the connective tissue septa. Positive staining for OX(1)R was evidenced in the wall of small arteries while that for OX(2)R was observed in the secretory portion of the acinar gland cells with a characteristic localization in the apical cytoplasm. RT-PCR analysis showed the presence of transcripts for PPOX, OX(1)R and OX(2)R in the sheep lacrimal gland; the gene expression of OX(1)R was two-fold greater (p<0.01) than that of OX(2)R. Taken together the present findings raise intriguing questions on the potential role of the orexinergic system in the regulation of lacrimal gland functions that require further investigations., (Copyright © 2012 Elsevier Inc. All rights reserved.)

We investigated the possible influence of sex and estrous cycle on the synaptic responses of neurons in the medial vestibular nucleus (MVN) and their long-term modifications. In brain stem slices of male and female rats during proestrus (PE) and diestrus (DE), we evaluated the field potential evoked in the MVN by vestibular afferent stimulation. Here we find that in PE females the field potential had a lower threshold and higher amplitude than in DE females and in males and also that the stimulus-response curve was shifted to the left. Such difference is related to the level and cyclic fluctuation of circulating 17β-estradiol (E(2)). This is supported by the exogenous administration of E(2) in DE females and males, with low levels of circulating E(2) that enhanced the field potential amplitude to values close to those of PE females. Sex and estrous cycle also influence the MVN synaptic plasticity. This has been shown by investigating the effect of testosterone (T) on the induction of long-term effects, since T is the precursor for the neural synthesis of E(2) (estrogenic pathway), which is involved in the induction of fast long-term potentiation (LTP), or of 5α-dihydrotestosterone (DHT, androgenic pathway) which mediates slow LTP and long-term depression (LTD). We found that T mostly induced LTD in PE females and no effect in DE females, while it only provoked fast LTP in males. We suggest that high level of circulating E(2) may interfere with the conversion of T, by inhibiting the neural estrogenic pathway and facilitating the androgenic one. On the whole these results demonstrate an influence of circulating E(2) on vestibular synaptic transmission and plasticity that in some cases may contribute to the sex and menstrual cycle dependence of symptoms in human vestibular pathology., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The aim of the present study was to investigate by immunohistochemistry the presence and distribution of the orexin system in the stomach and gut of fallow deer. Abundant orexin A-positive cells were localized in the middle and basal portions of the mucosal glands of the cardial and fundic regions of the stomach. In the same gastric areas, orexin B-positive cells were also found, mainly localized in the basal portion of glands. In the intestinal tract, orexin-containing cells were occasionally found in the duodenal epithelium and in the rectal intestinal glands. Immunoreactivity for orexin receptors, type 1 and 2 (OX1R and OX2R), was not detected in the same stomach regions. OX1R-immunopositivity was observed in the enteric neuron ganglia localized in the submucosal and muscular intestinal layers, while OX2R-immunopositivity was found close in contact with the cytoplasmic membrane of epithelial cells in the small intestine., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

In veterinary medicine, there is an increasing interest in the study of the endo-cannabinoid system and the possible use of the cannabinoids for the treatment of several diseases. Cannabinoid receptors (CB) are widely distributed in human and laboratory animal tissues, justifying the involvement of the endo-cannabinoid system in a great number of metabolic ways. Since there are no data regarding cannabinoid receptors in hair follicles of domestic animals, we investigated the presence and localization of CB1 receptor in dog hair follicles. By using a goat anti-CB1 polyclonal antibody, we observed CB1 receptor in the proximal part of both primary and secondary hair follicles. Staining was localized in the inner root sheath cells. We suppose that the endo-cannabinoid system is involved in the molecular mechanisms regulating hair follicle activity in dog. The identification of CB1 receptor at the level of the inner root sheath may help in the understanding of hair follicle biology and the possibility that cannabinoid molecules could be considered as suitable therapeutic tools in dog., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

The aim of the present work was to characterize the expression and cell type distribution of the orexinic system in the major swine salivary glands by RT-PCR and immunohistochemistry techniques. Tissue samples of the mandibular (MSG), sub-lingual, and parotid glands were obtained from six pigs. Marked differences in the expression of the orexinergic system were observed among the salivary glands. Prepro-orexin (PPOX) mRNA abundance was approximately 15-fold higher (P<0.05) in MSG than in the others salivary glands. The expression level of OX1R mRNA did not differ among the salivary glands, while that for OX2R transcript was undetectable. The hypothalamus, used as positive control tissues, showed the highest expression level for each component of the orexinic system. Immunoreactivity (IR) for both orexins A and B (OXA and OXB), and cognate receptors (OX1R and OX2R) were identified only in the excretory striated ducts of the MSG while acinar cells were not immunoreactive. Both sub-lingual and parotid glands completely lacked IR for any component of the orexinergic system. In the MSG, parasympathetic neurons and axons of local salivary gland ganglia were IR to both OXA and OXB and also to their receptors., (Copyright © 2011 Elsevier B.V. All rights reserved.)