Virgin Rats Research Articles

CNS Relaxin activates spinally projecting and vasopressin containing cells in the hypothalamic paraventricular nucleus

Relaxin (RLX), an ovarian hormone which is secreted in pregnancy, activates the subfornical organ (SFO) and hypothalamic regions associated with control of blood volume and sympathetic nerve activity (SNA). The current experiments phenotyped cells in the paraventricular nucleus (PVN) which are activated by RLX. Carotid and femoral artery and vein catheters were implanted in female virgin rats. Spinally projecting cells were retrogradely labeled by microinjection of fluorogold (FG) or cholera toxin‐b (CTb) (90nl) into the intermediolateral cell column. After 5 days human RLX 2 (1μg/hr) or saline (Sal, 1ml/hr) was infused (1.5 hr) into the forebrain circulation (intra‐carotid artery, ica) of conscious rats. RLX (n=3) increased mean arterial pressure (MAP, 9±2 mmHg) while Sal (n=5) had no effect (+1 ± 1mmHg). Rats were euthanized, perfused with 4% paraformaldehyde, brains sectioned (30 μm), and Fos‐, CTb‐, and vasopressin (VP‐) immunoreactivity (IR) were evaluated. Following RLX, cells in the lateral margins of the SFO expressed Fos‐IR, consistent with activation of PVN projecting neurons. Forty ±11% of VP‐IR cells (bregma −1.8mm) and 36±7% of spinally projecting cells (bregma −2.12mm) were activated (Fos‐IR) by ica RLX. In contrast, Fos‐IR was minimal in Sal treated rats. These data provide an anatomical substrate for a role of RLX in the adaptations in regulation of VP and SNA in pregnancy. HL091164(CMH)

Cellular localization and protein concentration of the SNAT2 transporter in rat mammary gland during pregnancy and lactation

The sodium coupled amino acid transporter SNAT2 is expressed in mammary gland (MG). There is a special amino acid demand for cellular differentiation during pregnancy and for protein milk secretion during lactation in the MG. The purpose of the present work was to assess the amount and localization of SNAT along pregnancy and lactation in rat MG. Western blot analysis in MG of virgin rats showed the presence of a single band with a MW of 60 kD corresponding to SNAT2 protein. The intensity of this band was maintained during pregnancy and decreased dramatically one day before delivery. During lactation, SNAT2 increased rapidly after 5 days and remains constant along lactation and decreased after post-weaning. Immunohistochemical analysis revealed that MG SNAT2 of virgin rats was exclusively located in adipose cells. During gestation, there was an increase in the staining of SNAT2 in epithelial cells that will synthesize milk and decrease before delivery. During lactation there was a significant increase in the amount of SNAT2 in epithelial cells. Electron immunomicroscopy analysis of SNAT2 in MG during peak of lactation indicated that the SNAT2 protein was retained in vesicles originated from the endoplasmic reticulum and then translocated to the basolateral membrane of the epithelial cells. Our results suggested that presence of SNAT2 in the different cells and stages depends on the needs of the MG. CONACYT grant No 46134

Environmental enrichment delays pup‐induced maternal behavior in rats

Adult, virgin rats do not spontaneously display maternal behavior when exposed to foster pups. However, continuous daily exposure of the female to foster pups for about 5-7 days can induce a set of maternal behaviors similar to those shown by postpartum dams. Induction latencies depend upon a number of factors, including the stress and anxiety levels of the female. The goal of this study was to attempt to mitigate the likely stressfulness of being singly housed during testing by enriching the rat's home cage environment and to determine if the concomitant environmental change would alter the latency to express maternal behavior. In addition, the effect of varying the number of test pups used for testing was examined. Two groups of virgin Sprague-Dawley rats were first tested on the elevated plus maze after 1 week of exposure to either control (standard housing) or enriched conditions. One week later, maternal behavior testing began using one or three pups. Upon completion of maternal behavior testing, plasma corticosterone concentrations were determined following a mild stressor. The data indicate that enrichment tends to increase anxiety-like behaviors in the elevated plus maze. In addition, enrichment delayed the onset of maternal behavior irrespective of the number of test pups. There were no effects of environmental enrichment on plasma corticosterone levels following exposure to a stressor. These results indicate that what is considered a modestly enriched environment delays the expression of pup-oriented responses and does not apparently reduce stress or improve performance on all behavioral tasks.

Open field behavior and intra-nucleus accumbens dopamine release in vivo in virgin and lactating rats.

behavioral, endocrine, and neurochemical implications. This experience causes behavioral and neurochemical changes that involve several brain areas important for the expression of maternal behavior. The present study showed that lactating rats exhibited reduced general locomotor activity in the open field test compared with virgin animals. Our hypothesis was that nucleus accumbens dopamine, which regulates maternal behavior in lactating rats, is also involved in the low expression of maternal locomotion in the open field test observed during the early stages of lactation and reflects decreased motivation. Initially we compared open field behavior in virgin and lactating rats to confirm our previous data. Thus, the in vivo release of dopamine in the nucleus accumbens in virgin and lactating female rats was measured. Perfusate concentrations of extracellular dopamine and its metabolites showed no differences between virgin and lactating rats. Thus, the reduced general activity observed in lactating rats might not be related to intra-nucleus accumbens dopamine control. Keywords: maternal behavior, virgin rats, lactating rats, nucleus accumbens, dopamine.

Leptin mRNA expression in the rat mammary gland at different activation stages

Leptin is expressed in various tissues, suggesting that this protein is effective not only at the central nervous system level, but also peripherically. Recent studies have shown leptin production by other tissues, including the placenta, stomach, and mammary tissues, but there is no information available concerning expression levels of leptin in the rat mammary gland at different activation stages. We used semi-quantitative RT-PCR to investigate leptin mRNA expression levels in the rat mammary gland at different activity stages. Rat mammary gland samples were collected from virgin females and on days 6, 12, 18 of pregnancy and of lactation (six rats per group). The expression levels of leptin mRNA were measured by semi-quantitative RT-PCR, with β-actin as an internal control. Leptin mRNA was highly expressed in virgin rat mammary glands (leptin(IOD)/β-actin(IOD) = 1.60). It decreased gradually during pregnancy, being lowest at 18 days of pregnancy, when the levels were significantly lower than in virgin mammary tissue. Leptin mRNA increased slightly during lactation, but the difference was not significant. By day 18 of lactation, expression levels of leptin mRNA reached the same values as in virgin mammary tissue (leptin(IOD)/β-actin(IOD) = 1.65). Based on these results, we suggest that leptin has an important regulation role in rat mammary gland activation.

Impaired Baroreflex Gain During Pregnancy in Conscious Rats

Pregnancy impairs baroreflex gain, but the mechanism is incompletely understood. To test the hypothesis that reductions in brain insulin contribute, we determined whether pregnant rats exhibit lower cerebrospinal fluid (CSF) insulin concentrations and whether intracerebroventricular infusion of insulin normalizes gain of baroreflex control of heart rate in conscious pregnant rats. CSF insulin was lower in pregnant (68 ± 21 pg/mL) compared to virgin (169 ± 25 pg/mL) rats (P < 0.05). Pregnancy reduced baroreflex gain (pregnant 2.4 ± 0.2 bpm/mm Hg, virgin 4.6 ± 0.3 bpm/mm Hg; P < 0.0001) and the maximum heart rate elicited by hypotension (pregnant 455 ± 15 bpm, virgin 507 ± 12 bpm; P = 0.01). Infusion of insulin (100 μU/min) intracerebroventricularly increased baroreflex gain in pregnant (2.4 ± 0.4 to 3.9 ± 0.5 bpm/mm Hg; P < 0.01) but not virgin (4.6 ± 0.4 to 4.2 ± 0.4 bpm/mm Hg; NS) rats. Maximum heart rate was not altered by intracerebroventricular insulin in either group. Interestingly, while in pregnant rats the baroreflex was unchanged by intracerebroventricular infusion of the artificial CSF vehicle, in virgin rats, vehicle infusion lowered baroreflex gain (4.7 ± 0.3 to 3.9 ± 0.3 bpm/mm Hg; P < 0.05) and the maximum baroreflex heart rate (495 ± 19 to 444 ± 21 bpm; P < 0.05). These data support the hypothesis that brain insulin is required to support optimal baroreflex function and that a decrease in brain insulin contributes to the fall in baroreflex gain during pregnancy.

Inhibitory effect of curcumin on angiogenesis in ectopic endometrium of rats with experimental endometriosis

The aim of this study was to observe the inhibitory effect of curcumin on endometriosis (EMS) and to determine its influence on vascular endothelial growth factor (VEGF) and microvessel density (MVD) in eutopic and ectopic endometrium of experimental rats, thus exploring the pathogenesis of EMS offering more experimental evidence for the clinical use of curcumin. Forty-eight female virgin rats were subjected to autotransplantation of endometrium during the estrus stage. After four weeks, 8 rats were randomly sacrificed to confirm that the rat model was successful. The remaining rats were randomly divided into four groups. Three groups were intragastrically administered curcumin (50, 100 and 150 mg/kg), and the model group was intragastrically administered vehicle alone. All rats were treated daily for four continuous weeks and examined by histology and immunohistochemical staining for MVD of eutopic and ectopic endometrium. Our results revealed that the cubic capacity of focal tissue in gross appearance was high in the model group and dose-dependently diminished after treatment with curcumin (P<0.05). There was an increase in MVD and VEGF in the ectopic endometrium, which was decreased significantly after treatment with curcumin (P<0.05); the effects being dose-dependent. The correlation between MVD and VEGF was positive. In conclusion, heterogeneity was found to exist between eutopic and ectopic endometrium due to differences noted in MVD and the expression of VEGF between the eutopic and ectopic endometrium in the model group. Curcumin decreased the quantity of microvessels and VEGF protein expression in the heterotopic endometrium of rats with EMS.

Mammary gland proliferation in female rats: Effects of the estrous cycle, pseudo-pregnancy and age

Assessment of mammary gland proliferation in rats is an important endpoint in preclinical safety studies of pharmaceutical compounds. However, existing data on mammary gland proliferation in rats during the estrous cycle is conflicting, and it is unknown whether mammary gland proliferation differs between young and mature female virgin rats. Additionally, it is unclear which of the commonly applied markers of proliferating cells that is optimal for assessment of rat mammary gland proliferation. In this study the caudal thoracic, the abdominal and the cranial inguinal (i.e., the 3rd the 4th and the 5th) mammary gland were collected from 29 young and 26 mature non-treated, virgin female Sprague Dawley rats. Estrous cycle stage was determined from repeated vaginal smears and histological examination of the reproductive organs. Proliferation of mammary epithelium was assessed by immunohistochemistry using three markers: PCNA, Ki67, and BrdU. Proliferation of the mammary epithelium occurred mainly in the terminal end buds in the young animals. Epithelial proliferation was significantly increased during metestrus compared to the other phases. Mammary gland proliferation in pseudo-pregnant females was increased compared to proestrus, estrus and diestrus, but not metestrus. Except during estrus no difference in mammary gland proliferation was observed between young and mature female rats, and no significant differences was observed between different mammary glands. The percentages of PCNA-, Ki67- and BrdU-positive epithelial cells were significantly correlated. In conclusion, the variation in normal proliferation between estrous cycle stages and animals with an irregular estrous cycle should be considered in toxico-pathological studies of mammary gland proliferation.

Increased renal α-epithelial sodium channel (ENAC) protein and increased ENAC activity in normal pregnancy

Pregnancy-mediated sodium (Na) retention is required to provide an increase in plasma volume for the growing fetus. The mechanisms responsible for this Na retention are not clear. We first used a targeted proteomics approach and found that there were no changes in the protein abundance compared with virgin rats of the β or γ ENaC, type 3 Na(+)/H(+) exchanger (NHE3), bumetanide-sensitive cotransporter (NKCC2), or NaCl cotransporter (NCC) in mid- or late pregnancy. In contrast, we observed marked increases in the abundance of the α-ENaC subunit. The plasma volume increased progressively during pregnancy with the greatest plasma volume being evident in late pregnancy. ENaC inhibition abolished the difference in plasma volume status between virgin and pregnant rats. To determine the in vivo activity of ENaC, we conducted in vivo studies of rats in late pregnancy (days 18-20) and virgin rats to measure the natriuretic response to ENaC blockade (with benzamil). The in vivo activity of ENaC (U(Na)V postbenzamil-U(Na)V postvehicle) was markedly increased in late pregnancy, and this difference was abolished by pretreatment with the mineralocorticoid receptor antagonist, eplerenone. These findings demonstrate that the increased α-ENaC subunit of pregnancy is associated with an mineralocorticoid-dependent increase in ENaC activity. Further, we show that ENaC activity is a major contributor of plasma volume status in late pregnancy. These changes are likely to contribute to the renal sodium retention and plasma volume expansion required for an optimal pregnancy.

Dopamine receptor alterations in female rats with diet-induced decreased brain docosahexaenoic acid (DHA): interactions with reproductive status

Decreased tissue levels of n-3 (omega-3) fatty acids, particularly docosahexaenoic acid (DHA), are implicated in the etiologies of non-puerperal and postpartum depression. This study examined the effects of a diet-induced loss of brain DHA content and concurrent reproductive status on dopaminergic parameters in adult female Long–Evans rats. An α-linolenic acid-deficient diet and breeding protocols were used to produce virgin and parous female rats with cortical phospholipid DHA levels 20–22% lower than those fed a control diet containing adequate α-linolenic acid. Decreased brain DHA produced a significant main effect of decreased density of ventral striatal D2-like receptors. Virgin females with decreased DHA also exhibited higher density of D1-like receptors in the caudate nucleus than virgin females with normal DHA. These receptor alterations are similar to those found in several rodent models of depression, and are consistent with the proposed hypodopaminergic basis for anhedonia and motivational deficits in depression.

Increased renal phosphodiesterase-5 activity mediates the blunted natriuretic response to a nitric oxide donor in the pregnant rat

Pregnancy is characterized by plasma volume expansion and renal sodium retention with loss of natriuretic response to atrial natriuretic peptide due to increased medullary phosphodiesterase-5 (PDE5). Here, we determined whether natriuretic responses to nitric oxide (NO) are also blunted in pregnancy due to increased PDE5. Anesthetized 16-day pregnant and virgin rats were studied at baseline and during intrarenal infusion of the NO donor spermine NONOate (2.5 nmol/min), the PDE5 inhibitor sildenafil (SILD; 0.5 μg/min), or a combination. The right (noninfused) kidney served as a control. Intrarenal NONOate had no effect on mean arterial pressure (MAP); however, SILD reduced MAP in virgin rats, and the combination of NONOate+SILD reduced MAP in both virgin and pregnant rats. Neither NONOate nor SILD altered glomerular filtration rate. NONOate and SILD each stimulated sodium excretion (U(Na)V) and fractional excretion of sodium (FE(Na)) in virgin rats, but the combination did not result in an additional natriuretic response. However, NONOate infusion did not increase U(Na)V or FE(Na) in pregnant rats, but the natriuretic response to NONOate was restored with SILD, and SILD alone produced a natriuresis during pregnancy. Sodium nitroprusside (10(-4) mol/l)-stimulated cGMP accumulation from inner medullary collecting duct cells was blunted in cells from pregnant vs. virgin or postpartum rats and was restored by treatment with the PDE5 inhibitor DMPPO (10(-7) mol/l). Therefore, increased intrarenal PDE5 mediates the blunted natriuretic response to NO, and loss of responsiveness to the cGMP-dependent, natriuretic agents may contribute to volume expansion during pregnancy.

Renal nitric oxide production in rat pregnancy: role of constitutive nitric oxide synthases.

Functional studies show that increased renal nitric oxide (NO) mediates the renal vasodilation and increased glomerular filtration rate that occur during normal pregnancy. We investigated whether changes in the constitutive NO synthases (NOS), endothelial (eNOS) and neuronal (nNOS), were associated with the increased renal NO production in normal midterm pregnancy in the rat. In kidneys from midterm pregnant (MP: 11-13 days gestation), late-term pregnant (LP: 18-20 days gestation), and similarly aged virgin (V) rats, transcript and protein abundance for eNOS and the nNOSα and nNOSβ splice variants, as well as the rate of L-arginine-to-L-citrulline conversion, were determined as a measure of NOS activity. At MP, renal cortical abundance of the total eNOS protein and phosphorylated (Ser(1177)) eNOS was reduced, and L-arginine-to-L-citrulline conversion in the cortical membrane fraction was decreased; these declines were also seen in LP. There were no changes in the eNOS transcript. In contrast, L-arginine-to-L-citrulline conversion in the soluble fraction of renal cortex increased at MP and then declined at LP. This MP increase was ablated by S-methylthiocitrulline, a nNOS inhibitor. Using Western blotting, we did not detect a change in the protein abundance or transcript of the 160-kDa nNOSα, but protein abundance and transcript of the nNOSβ were increased at MP in cortex. Collectively, these studies suggest that the soluble nNOSβ is responsible for the increased renal cortical NO production during pregnancy.

Pregnancy- and delivery-induced biomechanical changes in rat vagina persist postpartum

We sought to define changes in vaginal distensibility (VD) induced by pregnancy and vaginal delivery using a novel in vivo biomechanical testing protocol. Under sedation, a balloon was inserted into the vagina of 27 virgin, pregnant and 4-week postpartum Long-Evans rats and incrementally distended. Pressure-volume curves were generated with slopes characterizing VD (higher slope = less distensible). One-way ANOVA with a Bonferroni post-hoc test were used for statistical analyses. Mean pressures at an infusion volume of 1 cc were lower in pregnant and postpartum rats than in virgins (P < 0.001). VD was increased in pregnant vs. virgin rats (P < 0.001) and did not recover to virgin levels post partum (P < 0.001). We have developed a test that measures VD in vivo under clinically relevant loading conditions. The increased VD in the late postpartum period defines a persistent change in biomechanical behavior of the vagina related to pregnancy and vaginal delivery.

Increased Sensitivity of Monoamine Release in the Supraoptic Nucleus in Late Pregnancy: Region‐ and Stimulus‐Dependent Responses

Oxytocin neurone activation at birth depends upon noradrenaline-mediated signals from the uterus via a brainstem pathway, as well as on factors within the supraoptic nucleus (SON), including oxytocin itself, and the system adapts during pregnancy to optimise the delivery process. We determined whether noradrenaline release in the SON in response to stimuli activating brainstem inputs or antidromically activating magnocellular neurones is enhanced at term pregnancy. Noradrenaline, serotonin and dopamine concentrations were measured in microdialysis samples collected from the dorsal and ventral SON before, during and after either i.v. cholecystokinin (CCK) or neural stalk stimulation in virgin and late pregnant rats. Each stimulus transiently increased noradrenaline and serotonin but not dopamine concentration in the dorsal SON, and responses were increased on days 21 and 22 of pregnancy compared to day 20 pregnant and virgin rats. Neural stalk stimulation induced sensitisation to subsequent stalk stimulation and so the responses in the dorsal SON were doubled; on day 22 of pregnancy, the area under the curve of monoamine concentration was 3.4-fold greater than in virgins, suggesting that adaptations perinatally enhance responsiveness. In conclusion, there are enhanced responses of noradrenaline and serotonin release in the SON that can generate very high, transient extracellular concentrations at term. This may be a consequence of neuroendocrine adaptations in late pregnancy and probably contributes to optimal oxytocin neurone activation during parturition.

Decreased Venous Contraction and Enhanced Relaxation and Nitric Oxide Production in the Inferior Vena Cava of Pregnant Rat

Normal pregnancy (Norm‐Preg) is associated with increased cardiac output and plasma volume, partly due to increased nitric oxide (NO) production by endothelial cells in various arterial beds; however, the role of venous endothelium is unclear. We tested whether Norm‐Preg is associated with increases in venous relaxation and NO production. Segments of inferior vena cava (IVC) from Norm‐Preg (day 19) and virgin Sprague‐Dawley rats were used to measure isometric contraction/relaxation. The IVC was loaded with the NO‐sensitive DAF‐FM diacetate (DAF) for simultaneous measurement of NO production. The α‐adrenergic agonist phenylephrine (PHE, 10−5 M) and membrane depolarization by 96mM KCl caused less contraction in IVC of Norm‐Preg (20.64±8.75 and 230.07±31.40) than virgin rats (70.90±15.60 and 308.32±95.65 mg/mg tissue). Acetylcholine (Ach, 10−5 M) caused greater relaxation of KCl‐induced contraction in IVC of Norm‐Preg (46.85±12.95) than virgin rats (36.05±8.07%). Basal NO was slightly greater in IVC of Norm‐Preg (0.12±0.02) than virgin rats (0.10±0.02). In IVC precontracted with PHE, Ach (10−5 M) caused greater NO production in Norm‐Preg (0.37±0.08) than virgin rats (0.22±0.03). Thus Norm‐Preg is associated with decreased venous contraction and enhanced Ach‐induced relaxation and NO production, which could lead to more distensible veins and venous insufficiency during pregnancy.

It's not my nerves! Augmented pressor response to stress in multiparous rats is not associated with increased sympathetic outflow.

There is epidemiological evidence that multiparity (multiple pregnancies) increases the long‐term risk for cardiovascular disease. We have shown that multiparous rats have an augmented pressor response to stress. This is due, at least in part, to an oxidative stress induced increase in vascular reactivity. We proposed that the augmented pressor response in the multiparous rats was also the result of a greater increase in sympathetic outflow. Age‐matched multiparous and virgin rats were implanted with telemetric devices to monitor renal sympathetic nerve activity (Telemetry Research) and blood pressure (DSI). One week later, they were exposed to 10 sec air jet. The stress‐induced increase in integrated mean amplitude of nerve activity was significantly higher in the virgin animals (Virgin: 3.2±1.6 uV to 36.5±12.2 uV; Multiparous: 0.8±0.6 uV to 8.9±6.2 uV; P&lt;0.05). There was no difference between the groups with respect to the increase in spike frequency (Virgin: 28.2±7.8 to 36.6±10.0 spikes/sec; Multiparous: 16.6±2.8 to 28.6±1.4 spikes/sec; P&gt;0.05). We conclude that, contrary to our hypothesis, the augmented pressor response of multiparous rats is due solely to increased vascular reactivity, not to a greater increase in sympathetic outflow. (HSFC)

Parity-Induced Decrease in Systemic Growth Hormone Alters Mammary Gland Signaling: A Potential Role in Pregnancy Protection from Breast Cancer

Early full-term pregnancy is an effective natural protection against breast cancer in both humans and experimental rodents. The protective effect of an early pregnancy is, in part, linked to changes in circulating hormones that are involved in both normal breast development and breast cancer. For example, a reduction in circulating growth hormone (GH) has been shown to protect rats from carcinogen-induced mammary tumors. We examined the ability of a full-term pregnancy to alter the endocrine GH/insulin-like growth factor-I (IGF-I) axis and how this change affected normal mammary gland function in two commonly used rat models (Sprague-Dawley and Wistar Furth). Circulating GH and IGF-I were measured in blood drawn every 30 minutes from parous and age-matched virgin female rats. Mean serum GH levels were significantly decreased (P < 0.01) in parous compared with age-matched virgin rats for both strains. Changes in GH levels were independent of estrous cycle, indicated by a significant (P < 0.05) reduction in circulating levels of GH during estrus and diestrus in both parous strains. Despite the decrease in circulating GH, pituitary GH mRNA levels were unaltered in parous rats. Circulating IGF-I and hepatic IGF-I mRNA were also unaltered by parity in either rat strain. Immunoblot analysis of mammary glands showed decreases in phosphorylation of signal transducer and activator of transcription 5A and Janus-activated kinase 2, suggesting reduced action of GH in the mammary gland. Therefore, although the parity reduction in circulating GH does not affect circulating IGF-I levels, it is possible that reduced GH acts directly at the mammary gland and may play a role in pregnancy protection from breast cancer.

Clitoral stimulation modulates appetitive sexual behavior and facilitates reproduction in rats

In rats, sexual reward, appetitive sexual behaviors and reproduction are modulated by the amount and rate of vaginocervical stimulation. Here the effect of clitoral stimulation (CLS) on proceptivity was assessed. In Exp 1, ovariectomized, hormone-primed Wistar females formed three groups: G1 (1 CLS every second), G2 (1 CLS every 5 s) and G3 (no CLS). Precopulatory CLS consisted of 5 cycles of 1 min of stimulation with the tip of a cotton swab connected to a vibrator device, followed by 1–2 min of rest. CLS increased proceptive behavior in G1 compared to G2, but not compared to G3. In Exp 2, gonadally-intact rats in late proestrous received CLS prior to copulation. No differences in sexual behavior were detected between the groups, but CLS enhanced reproduction in females that received > 9 intromissions. 28, 66 and 10% of females became pregnant in G1, G2, and G3, respectively. These data indicate that precopulatory CLS affects proceptive behaviors depending on the pattern and rhythm of stimulation in hormone-primed females. In virgin rats that have received sufficient vagino cervical stimulation CLS also increases fertility.

Short term hypothyroidism affects ovarian function in the cycling rat

BackgroundRats made hypothyroid with propilthyouracil start showing abnormal cycling on the second cycle after the start of the treatment, with a high proportion of spontaneous pseudopregnancies and reduced fertility.MethodsTo investigate some of the mechanisms involved in these reproductive abnormalities, hypothyroidism was induced in virgin rats by propilthyouracil (0.1 g/L in the drinking water) and we determined circulating hormones by radioimmunoassay and whole ovary expression of ovarian hormone receptors, growth factors and steroidogenic enzymes using semi-quantitative RT-PCR.The study was performed on days 6 to 9 of treatment, corresponding to diestrus I (at 20.00-22.00 h), diestrus II (at 20.00-22.00 h), proestrus and estrus (both at 8.00-10.00 h and 20.00-22.00 h) of the second estrous cycle after beginning propilthyouracil treatment. Another group of rats was mated on day 8 and the treatment continued through the entire pregnancy to evaluate reproductive performance.ResultsHypothyroidism increased circulating prolactin and estradiol on estrus 5 to 7-fold and 1.2 to 1.4-fold respectively. Growth hormone and insulin-like growth factor 1 diminished 60 and 20% respectively on proestrus morning. Hypothyroidism doubled the ovarian mRNA contents of estrogen receptor-beta on proestrus and estrus evenings, cyp19A1 aromatase mRNA on estrus evening and of growth hormone receptor on proestrus evening. Hypothyroidism did not influence ovulation rate or the number of corpora lutea at term, but a diminished number of implantation sites and pups per litter were observed (Hypothyroid: 11.7 +/- 0.8 vs. Control: 13.9 +/- 0.7).ConclusionsShort term hypothyroidism alters normal hormone profile in the cycling rat increasing the expression of estrogen receptor-beta and cyp19A1 aromatase on estrus, which in turn may stimulate estradiol and prolactin secretion, favouring corpus luteum survival and the subsequent instauration of pseudopregnancy.

Glia-Dependent Switch of Kainate Receptor Presynaptic Action

Presynaptic kainate receptors (KARs) exert a modulatory action on transmitter release. This effect can be switched from facilitation to inhibition by an increased concentration of KAR agonists. We here report that activation of presynaptic GluK1-containing KARs facilitates GABA release on oxytocin and vasopressin neurons in the supraoptic nucleus of the hypothalamus. Increase in ambient levels of glutamate associated with the physiological reduction of astrocytic coverage of oxytocin neurons in lactating rats switches this KAR-mediated facilitation to inhibition of GABAergic transmission. This effect was reproduced in both oxytocin and vasopressin neurons of virgin rats when glutamate transporters were blocked pharmacologically, thereby establishing that enhanced levels of extracellular glutamate induce the switch in KAR-mediated action. The facilitation of GABA release was inhibited with philanthotoxin, a Ca(2+)-permeable KAR antagonist, suggesting that this effect was associated with an ionotropic mode of action. Conversely, KAR-mediated inhibition was compromised in the presence of U73122, a phospholipase C inhibitor, in agreement with the involvement of a metabotropic pathway. We thus reveal that physiological astrocytic plasticity modifies the mode of action of presynaptic KARs, thereby inversing their coupling with GABA release.