RESUMO
BACKGROUND: Sympathetic stress stimulates norepinephrine (NE) release from sympathetic nerves. During pregnancy, it modifies the fetal environment, increases NE to the fetus through the placental NE transporter, and affects adult physiological functions. Gestating rats were exposed to stress, and then the heart function and sensitivity to in vivo adrenergic stimulation were studied in male progeny. METHODS: Pregnant Sprague-Dawley rats were exposed to cold stress (4 °C/3 h/day); rats' male progeny were euthanized at 20 and 60 days old, and their hearts were used to determine the ß-adrenergic receptor (ßAR) (radioligand binding) and NE concentration. The in vivo arterial pressure response to isoproterenol (ISO, 1 mg/kg weight/day/10 days) was monitored in real time (microchip in the descending aorta). RESULTS: Stressed male progeny presented no differences in ventricular weight, the cardiac NE was lower, and high corticosterone plasma levels were recorded at 20 and 60 days old. The relative abundance of ß1 adrenergic receptors decreased by 36% and 45%, respectively (p < 0.01), determined by Western blot analysis without changes in ß2 adrenergic receptors. A decrease in the ratio between ß1/ß2 receptors was found. Displacement of 3H-dihydroalprenolol (DHA) from a membrane fraction with propranolol (ß antagonist), atenolol (ß1 antagonist), or zinterol (ß2 agonist) shows decreased affinity but no changes in the ß-adrenergic receptor number. In vivo exposure to ISO to induce a ß-adrenergic overload provoked death in 50% of stressed males by day 3 of ISO treatment. CONCLUSION: These data suggest permanent changes to the heart's adrenergic response after rat progeny were stressed in the uterus.
Assuntos
Mães , Placenta , Ratos , Feminino , Masculino , Gravidez , Animais , Humanos , Ratos Sprague-Dawley , Placenta/metabolismo , Norepinefrina , Receptores Adrenérgicos beta/metabolismo , AdrenérgicosRESUMO
Acetylcholine (ACh) may be involved in the regulation of ovarian functions. A previous systemic study in rats showed that a 4-week, intrabursal local delivery of the ACh-esterase blocker Huperzine-A increased intraovarian ACh levels and changed ovarian follicular development, as evidenced by increased healthy antral follicle numbers and corpora lutea, as well as enhanced fertility. To further characterize the ovarian cholinergic system in the rat, we studied whether innervation may contribute to intraovarian ACh. We explored the cellular distribution of three muscarinic receptors (MRs; M1, M3, and M5), analyzed the involvement of MRs in ovarian steroidogenesis, and examined their roles in ovarian follicular development in normal conditions and in animals exposed to stressful conditions by employing the muscarinic antagonist, atropine. Denervation studies decreased ovarian norepinephrine, but ovarian ACh was not affected, evidencing a local, nonneuronal source of ACh. M1 was located on granulosa cells (GCs), especially in large antral follicles. M5 was associated with the ovarian vascular system and only traces of M3 were found. Ex vivo ovary organo-typic incubations showed that the MR agonist Carbachol did not modify steroid production or expression of steroid biosynthetic enzymes. Intrabursal, in vivo application of atropine (an MR antagonist) for 4 weeks, however, increased atresia of the secondary follicles. The results support the existence of an intraovarian cholinergic system in the rat ovary, located mainly in follicular GCs, which is not involved in steroid production but rather via MRs exerts trophic functions and regulates follicular atresia.
Assuntos
Atresia Folicular , Ovário , Animais , Feminino , Ratos , Ovário/metabolismo , Receptores Muscarínicos/metabolismo , Acetilcolina/fisiologia , Atropina/farmacologia , Antagonistas Muscarínicos/farmacologia , Esteroides/metabolismoRESUMO
The exposure to sympathetic stress during the entire period of gestation (4 °C/3 h/day) strongly affects the postnatal reproductive performance of the first generation of female offspring and their fertility capacity. The aim of this work was to determine whether this exposure to sympathetic stress affects the reproductive capacity of the next three generations of female offspring as adults. Adult female Sprague-Dawley rats were mated with males of proven fertility. We studied the reproductive capacity of the second, third, and fourth generations of female offspring (the percentage of pregnancy and the number and weight of female offspring). The estrus cycle activity of the progenies was studied, and a morphological analysis of the ovaries was carried out to study the follicular population. The second generation had a lower number of pups per litter and a 20% decrease in fertile capacity. The estrus cycle activity of the third generation decreased even more, and they had a 50% decrease in their fertile capacity, and their ovaries presented polycystic morphology. The fourth generation however, recovered their reproductive capacity but not the amount of newborns pups. Most probably, the chronic intrauterine exposure to the sympathetic stress programs the female gonads to be stressed in a stressful environment; since the fourth generation was the first born with no direct exposure to stress during development, it opens studies on intrauterine factors affecting early follicular development.
Assuntos
Fertilidade , Efeitos Tardios da Exposição Pré-Natal , Animais , Estro , Feminino , Masculino , Ovário , Gravidez , Ratos , Ratos Sprague-Dawley , ReproduçãoRESUMO
The functioning of the ovary is influenced by the autonomic system (sympathetic and cholinergic intraovarian system) which contributes to the regulation of steroid secretion, follicular development, and ovulation. There is no information on the primary signal that activates both systems. The nerve growth factor (NGF) was the first neurotrophic factor found to regulate ovarian noradrenergic neurons and the cholinergic neurons in the central nervous system. The aim of this study was to determine whether NGF is one of the participating neurotrophic factors in the activation of the sympathetic and cholinergic system of the ovary in vivo and its role in follicular development during normal or pathological states. The administration of estradiol valerate (a polycystic ovary [PCO] phenotype model) increased norepinephrine (NE) (through an NGF-dependent mechanism) and acetylcholine (ACh) levels. Intraovarian exposure of rats for 28 days to NGF (by means of an osmotic minipump) increased the expression of tyrosine hydroxylase and acetylcholinesterase (AChE, the enzyme that degrades ACh) without affecting enzyme activity but reduced ovarian ACh levels. In vitro exposure of the ovary to NGF (100 ng/ml for 3 h) increased both choline acetyl transferase and vesicular ACh transporter expression in the ovary, with no effect in ACh level. In vivo NGF led to an anovulatory condition with the appearance of follicular cysts and decreased number of corpora lutea (corresponding to noradrenergic activation). To determine whether the predominance of a NE-induced polycystic condition after NGF is responsible for the PCO phenotype, rats were exposed to an intraovarian administration of carbachol (100 µM), a muscarinic cholinergic agonist not degraded by AChE. Decreased the number of follicular cysts and increased the number of corpora lutea, reinforcing that cholinergic activity of the ovary participates in controlling its functions. Although NGF increased the biosynthetic capacity for ACh, it was not available to act in the ovary. Hence, NGF also regulates the ovarian cholinergic system, implying that NGF is the main regulator of the dual autonomic control. These findings highlight the need for research in the treatment of PCO syndrome by modification of locally produced ACh as an in vivo regulator of follicular development.
Assuntos
Fator de Crescimento Neural/metabolismo , Ovário/metabolismo , Receptores Adrenérgicos/metabolismo , Receptores Colinérgicos/metabolismo , Acetilcolina/metabolismo , Acetilcolinesterase/metabolismo , Animais , Sistema Nervoso Autônomo , Carbacol/metabolismo , Colina O-Acetiltransferase/metabolismo , Estradiol/sangue , Estradiol/farmacologia , Estro , Feminino , Norepinefrina/metabolismo , Osmose , Ovulação/metabolismo , Fenótipo , Síndrome do Ovário Policístico/tratamento farmacológico , Isoformas de Proteínas , Ratos , Ratos Sprague-Dawley , Esteroides/metabolismo , Sistema Nervoso SimpáticoRESUMO
Chronic cold stress affects ovarian morphology and impairs fertility in rats. It causes an ovarian polycystic ovary (PCOS)-like phenotype, which resembles PCOS in women. The mechanism of cold stress action involves increased ovarian noradrenaline (NA) levels, which remain elevated after cessation of cold stress. By contrast, ovarian acetylcholine (ACh) levels are only transiently elevated and returned to control levels after a 28-day post stress period. Because ACh can exert trophic actions in the ovary, we hypothesised that a sustained elevation of ovarian ACh levels by intraovarian exposure to the ACh-esterase blocker huperzine-A (Hup-A) may interfere with cold stress-induced ovarian changes. This possibility was examined in female Sprague-Dawley rats exposed to cold stress (4°C for 3 h day-1 for 28 days), followed by a 28-day period without stress. To elevate ACh, in a second group Hup-A was delivered into the ovary of cold stress-exposed rats. A third group was not exposed to cold stress. As expected, cold stress elevated ovarian NA, reduced the number of corpora lutea and increased the number of follicular cysts. It increased plasma testosterone and oestradiol but decreased plasma levels of progesterone. In the Hup-A group, ovarian levels of both, NA and ACh, were elevated, there were fewer cysts and normal testosterone and oestradiol plasma levels were found. However, progesterone levels remained low. Most likely, low progesterone was associated with impaired mating behaviour and low pregnancy rate. We propose that elevated intraovarian levels of ACh are involved in the rescue of ovarian function, opening a target to control ovarian diseases affecting follicular development.
Assuntos
Alcaloides/farmacologia , Inibidores da Colinesterase/farmacologia , Norepinefrina/metabolismo , Ovário/efeitos dos fármacos , Sesquiterpenos/farmacologia , Estresse Fisiológico/fisiologia , Sistema Nervoso Simpático/fisiopatologia , Acetilcolina/metabolismo , Animais , Temperatura Baixa , Modelos Animais de Doenças , Estradiol/sangue , Feminino , Ovário/metabolismo , Ovário/fisiopatologia , Síndrome do Ovário Policístico/metabolismo , Síndrome do Ovário Policístico/fisiopatologia , Progesterona/sangue , Ratos , Ratos Sprague-Dawley , Sistema Nervoso Simpático/metabolismo , Testosterona/sangueRESUMO
An increase in the sympathetic tone in the rat ovary induces a polycystic ovary (PCOS-like) phenotype. No information exists about its impact on fertility. In contrast, increased follicular development and improved fertility in rats were found after pharmacological inhibition of acetylcholinesterase, which increased intraovarian acetylcholine (ACh). Now, we studied the impact of sympathetic stress, followed by a recovery period without stress, on the cholinergic and noradrenergic systems of the rat ovary and on fertility. To activate ovarian sympathetic nerves, female Sprague-Dawley rats were exposed to cold stress (4°C/3 h day for 28 days; first period), followed by a 28-day period without cold stress (second period). No changes in estrous cyclicity during the first period was found. At the end of this period, ovarian levels of NA and ACh were increased. Morphometric analysis showed lower numbers of secondary and antral follicles, enhanced follicular atresia and fewer corpora lutea. Plasma progesterone was lower and testosterone was higher than that in controls. At end of the second period, ovarian ACh levels had returned to control levels, but NA levels remained elevated. The second period was also characterized by the presence of cystic follicles in the ovary, by elevated plasma testosterone and estradiol levels, while progesterone levels were decreased. Estrous cyclicity and ovulation during that period were irregular and fertility decreased. Thus, cold stress initially activated both ovarian noradrenergic and cholinergic system. After stress, the ovary did not fully recover and activation of the noradrenergic system persisted and correlated with cystic ovarian morphology and decreased fertility.
Assuntos
Acetilcolina/metabolismo , Resposta ao Choque Frio/fisiologia , Ovário/inervação , Ovário/metabolismo , Sistema Nervoso Simpático/fisiologia , Animais , Ciclo Estral/fisiologia , Feminino , Fertilidade/fisiologia , Humanos , Norepinefrina/metabolismo , Ratos Sprague-DawleyRESUMO
Sympathetic innervation of the ovary in rodents occurs via two routes: the superior ovarian nerve (SON), which runs along the ovarian ligament, and the plexus nerve (PN), which is mainly associated with the vasculature. SON and ovarian norepinephrine (NE) levels play a major role in regulating ovarian cystic health. Although it was previously described that the polycystic ovarian phenotype (PCO) is causally related to hyperstimulation of the sympathetic nerves of the ovary, much less is known, however, regarding the role of PN in ovarian physiology. We studied the role of SON and PN in relation to the maintenance of the PCO phenotype induced in the rat by exposure to estradiol valerate (EV). EV exposure at 24 days old (juvenile exposure) increases NE in the ovary for up to 90 days after EV injection. SON or PN denervation (SONX and PNX) decreased NE. SONXreversed the acyclic condition from 30 days after surgery (p < 0.05), but PNXdid not. SONX was more effective than PNX to downregulate the increased number of cysts induced by EV, with the presence of the corpora lutea (CL, signifying ovulation) in the SONX group. Seventy percent of SONX rats presented with pregnancy at 60 days post-EV (6 of the 7 sperm-positive rats were pregnant); however, SONX rats had a reduced number (half) of pups compared with vehicle-treated rats and 60% more pups than EV rats. These data suggest that the SON plays a predominant role in follicular development, ovulation and pregnancy during ovarian diseases.
RESUMO
Previous work has demonstrated that the increase in the activity of sympathetic nerves, which occurs during the subfertility period in female rats, causes an increase in follicular cyst development and impairs follicular development. In addition, the increase in ovarian sympathetic activity of aged rats correlates with an increased expression of kisspeptin (KISS1) in the ovary. This increase in KISS1 could participate in the decrease in follicular development that occurs during the subfertility period. We aimed to determine whether the blockade of ovarian sympathetic tone prevents the increase in KISS1 expression during reproductive aging and improves follicular development. We performed 2 experiments in rats: (1) an in vivo blockade of beta-adrenergic receptor with propranolol (5.0 mg/kg) and (2) an ovarian surgical denervation to modulate the sympathetic system at these ages. We measured Kisspeptin and follicle-stimulating hormone receptor (FSHR) mRNA and protein levels by qRT-PCR and western blot and counted primordial, primary and secondary follicles at 8, 10 and 12 months of age. The results showed that ovarian KISS1 decreased but FSHR increased after both propranolol administration and the surgical denervation in rats of 8, 10 and 12 months of age. An increase in FSHR was related to an increase in the number of smaller secondary follicles and a decreased number of primordial follicles at 8, 10 and 12 months of age. These results suggest that intraovarian KISS1 is regulated by sympathetic nerves via a beta-adrenergic receptor and participates locally in ovarian follicular development in reproductive aging.
Assuntos
Envelhecimento/metabolismo , Regulação da Expressão Gênica , Kisspeptinas/metabolismo , Ovário/metabolismo , Sistema Nervoso Simpático/metabolismo , Antagonistas Adrenérgicos beta/farmacologia , Animais , Feminino , Kisspeptinas/genética , Ovário/efeitos dos fármacos , Propranolol/farmacologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores do FSH/genética , Receptores do FSH/metabolismo , Sistema Nervoso Simpático/efeitos dos fármacosRESUMO
Chronic cold stress produces adrenergic overload that can affect fetal development. The placental norepinephrine transporter (NET) clears norepinephrine (NE) from both maternal circulation and the fetus during gestation. If this system fails, NE clearance can be reduced, leading to high fetal exposure to NE. The main aim of this study was to determine the changes in NET expression during gestation and their relationship with the functional capacity of NET to transport NE under stressful conditions. Additionally, this study correlated these findings with the reproductive capacity of 2nd-generation progeny. Pregnant rats were subjected to chronic cold stress at 4°C for 3 h each day throughout their pregnancies. We found that exposure of pregnant rats to sympathetic stress caused the following effects: increased NE and corticosterone levels throughout pregnancy, decreased capacity of the placenta to clear NE from the fetus to the mother's circulation, altered NET protein levels depending on the sex of the fetus and increased placental and body weights of pups. For the first time, we also described the disrupted fertility of progeny as adults. Increased NE plasma levels during pregnancy under sympathetic stress conditions correlated with decreased NET functionality that provoked changes in the development of progeny and their fertility in adulthood.
Assuntos
Fertilidade/fisiologia , Proteínas da Membrana Plasmática de Transporte de Norepinefrina/fisiologia , Placenta/química , Complicações na Gravidez/fisiopatologia , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Estresse Fisiológico/fisiologia , Animais , Temperatura Baixa , Corticosterona/sangue , Feminino , Masculino , Troca Materno-Fetal , Norepinefrina/sangue , Norepinefrina/metabolismo , Gravidez , Ratos , Ratos Sprague-Dawley , Fatores SexuaisRESUMO
Growth and differentiation of ovarian follicles are regulated by systemic and local factors, which may include acetylcholine (ACh). Granulosa cells (GCs) of growing follicles and luteal cells produce ACh and in cultured GCs it exerts trophic actions via muscarinic receptors. However, such actions were not studied in vivo. After having established that rat ovarian GCs and luteal cells express the ACh-metabolizing enzyme ACh esterase (AChE), we examined the consequences of local application of an AChE inhibitor, huperzine A (HupA), by osmotic minipump delivery into the ovarian bursa of hemiovariectomized rats. Saline was used in the control group. Local delivery of HupA for 4 weeks increased ovarian ACh content. Estrus cyclicity was not changed indicating a locally restricted range of HupA action. The number of primordial and primary follicles was unaffected, but small secondary follicles significantly increased in the HupA group. Furthermore, a significant increase in the number of corpora lutea suggested increased ovulatory events. In support, as shown upon mating, HupA-treated females had significantly increased implantation sites and more pups. Thus the data are in support of a trophic role of ACh in follicular development and ovulation and point to an important role of ACh in female fertility.