Central injection of ketone body suppresses luteinizing hormone release via the catecholaminergic pathway in female rats

Ketosis is found in various pathophysiological conditions, including diabetes and starvation, that are accompanied by suppression of gonadal activity. The aim of the present study was to determine the role of ketone body in the brain in regulating pulsatile luteinizing hormone (LH) secretion in female rats. Injection of 3-hydroxybutyrate (3HB), a ketone body, into the fourth cerebroventricle (4V) induced suppression of pulsatile LH secretion in a dose-dependent manner in ovariectomized (OVX) rats with an estradiol (E2) implant producing diestrus plasma E2 levels. Plasma glucose and corticosterone levels increased immediately after the 4V 3HB injection, suggesting that the treatment caused a hunger response. The 3HB-induced suppression of LH pulses might be mediated by noradrenergic inputs to the hypothalamic paraventricular nucleus (PVN) because a local injection of α-methyl- p-tyrosine, a catecholamine synthesis inhibitor, into the PVN blocked 3HB-induced suppression of LH pulses and PVN noradrenaline release was increased by 4V 3HB injection in E2-primed OVX rats. These results suggest that ketone body sensed by a central energy sensor in the hindbrain may suppress gonadotropin release via noradrenergic inputs to the PVN under ketosis.     

Role of noradrenergic receptors in the bed nucleus of the stria terminalis in regulating pulsatile luteinizing hormone secretion in female rats

The bed nucleus of the stria terminalis (BNST) is one of the brain areas densely innervated by noradrenergic neurons originating in the brain stem. The present study aims to determine the role of noradrenergic receptors in the BNST in regulating pulsatile luteinizing hormone (LH) secretion in female rats. Ovariectomized (OVX) or estrogen-primed OVX (OVX+E2) rats received three 1-h-interval injections of 0.05 micromol of noradrenaline (NA), phenylephrine (alpha1-adrenergic receptor agonist), clonidine (alpha2-agonist), or isoproterenol (beta-agonist) into the BNST. Injection of NA or alpha1-adrenergic agonist into the BNST strongly suppressed pulsatile LH secretion in OVX+E2 rats with a significant (P < 0.05) decrease in the mean LH level for 3 h and LH pulse frequency, but alpha2-and beta-agonists did not affect any of the LH pulse parameters. In OVX animals, alpha1- and alpha2-adrenergic agonists caused a significant change in LH pulse frequency and amplitude, respectively, though the effect was not as apparent as the NA- or alpha1-agonist-induced changes in OVX+E2 animals. These results indicate that NA inputs to the BNST suppress pulsatile LH secretion via alpha1-adrenergic receptors and that estrogen enhances this suppression.     

Acute lipoprivation suppresses pulsatile luteinizing hormone secretion without affecting food intake in female rats

The present study examined the effect of acute lipoprivation on pulsatile luteinizing hormone (LH) secretion in both normal-fat diet, ad libitum-fed and fasted female rats. To produce an acute lipoprivic condition, mercaptoacetate (MA), an inhibitor of fatty acid oxidation, was administered intraperitoneally to ad libitum-fed or 24-h fasted ovariectomized (OVX) rats with or without an estradiol (E2) implant, that produces a negative feedback effect on LH pulses. The steroid treatment was performed to determine the effect of estrogen on lipoprivic changes in LH release, because estrogen enhances fasting- or glucoprivation-induced suppression of LH pulses. Pulsatile LH secretion was suppressed by MA administration in a dose-dependent manner in the ad libitum-fed OVX and OVX+E2 rats. LH pulses were more severely suppressed in the 24-h-fasted OVX and OVX+E2 rats compared to the ad libitum-fed rats. Estrogen slightly enhanced lipoprivic suppression but the effect was not significant. In the present study, increased plasma glucose and free-fatty acid concentrations may indicate a blockade of fatty acid metabolism by the MA treatment, but food intake was not affected by any of the MA doses. Acute vagotomy did not block lipoprivic suppression of LH pulses. Thus, the present study indicates that lipid metabolism is important for maintenance of normal reproductive function even in rats fed a normal-fat diet and lipoprivation may be more critical in fasted animals that probably rely more heavily on fatty acid oxidation to maintain appropriate metabolic fuel levels. In addition, failure of blockade of lipoprivic LH inhibition by vagotomy suggests that lipoprivic information resulting in LH suppression is not transmitted to the brain via the vagus nerve.     

Paraventricular alpha1- and alpha2-adrenergic receptors mediate hindbrain lipoprivation-induced suppression of luteinizing hormone pulses in female rats

Acute central lipoprivation suppresses pulsatile luteinizing hormone (LH) release and increases blood glucose levels through noradrenergic input to the hypothalamic paraventricular nucleus (PVN) in female rats. The present study was conducted to identify adrenergic receptor subtypes involved in central lipoprivation-induced suppression of pulsatile LH secretion and increases in plasma glucose levels in female rats. Acute hindbrain lipoprivation was produced by injection into the fourth cerebroventricle (4V) of 2-mercaptoacetate (MA), an inhibitor of fatty acid oxidation, in estradiol-implanted ovariectomized rats. Two min before MA injection, alpha1-, alpha2- or beta-adrenergic receptor antagonist was injected into the PVN. Injection of MA into the 4V suppresses pulsatile LH release in PVN vehicle-treated rats, whereas pretreatment of animals with injection of alpha1- or alpha2-adrenergic antagonist into the PVN blocked the effect of the 4V MA injection on LH pulses. beta-Adrenergic antagonist did not affect MA-induced suppression of LH pulses. The counter-regulatory increase in plasma glucose levels after 4V MA injection was also partially blocked by pretreatment with alpha1- and alpha2-adrenergic receptor antagonists. These results suggest that alpha1- and alpha2-adrenergic receptors in the PVN mediate hindbrain lipoprivation-induced suppression of LH release and counter-regulatory increases in plasma glucose levels in female rats.     

Central estrogen action sites involved in prepubertal restraint of pulsatile luteinizing hormone release in female rats

The present study aimed to determine estrogen feedback action sites to mediate prepubertal restraint of gonadotropin-releasing hormone (GnRH)/luteinizing hormone (LH) release in female rats. Wistar-Imamichi strain rats were ovariectomized (OVX) and received a local estradiol-17β (estradiol) or cholesterol microimplant in several brain areas, such as the medial preoptic area (mPOA), paraventricular nucleus, ventromedial nucleus and arcuate nucleus (ARC), at 20 or 35 days of age. Six days after receiving the estradiol microimplant, animals were bled to detect LH pulses at 26 or 41 days of age, representing the pre- or postpubertal period, respectively. Estradiol microimplants in the mPOA or ARC, but not in other brain regions, suppressed LH pulses in prepubertal OVX rats. Apparent LH pulses were found in the postpubertal period in all animals bearing estradiol or cholesterol implants. It is unlikely that pubertal changes in responsiveness to estrogen are due to a change in estrogen receptor (ER) expression, because the number of ERα-immunoreactive cells and mRNA levels of Esr1, Esr2 and Gpr30 in the mPOA and ARC were comparable between the pre- and postpubertal periods. In addition, kisspeptin or GnRH injection overrode estradiol-dependent prepubertal LH suppression, suggesting that estrogen inhibits the kisspeptin-GnRH cascade during the prepubertal period. Thus, estrogen-responsive neurons located in the mPOA and ARC may play key roles in estrogen-dependent prepubertal restraint of GnRH/LH secretion in female rats.     

Negative feedback control of luteinizing hormone secretion in prepubertal beef heifers at 60 and 200 days of age

Prepubertal beef heifers at 60 and 200 d of age, born in the fall or spring, were assigned randomly to one of three treatment groups: (1) intact = 1; (2) bilateral ovariectomy (OVX); or (3) OVX plus estradiol-17 beta(E2) administered in silastic implants (OVX + E2). Luteinizing hormone (LH) was measured in serum samples collected at 20-min intervals for 4 h from heifers on -1, +7, +21, +35 and +49 d after OVX. Luteinizing hormone concentrations increased in the serum by 7 d after OVX in heifers at both 60 and 200 d of age (P less than .001; time X treatment). Prior to OVX, the LH patterns were characterized by low levels and infrequent episodic pulses. By 49 d after OVX, the mean LH concentrations increased and the pattern changed to one of rhythmic LH pulses with a periodicity of 1 h (P less than .001; time X treatment). Estradiol-treated OVX heifers did not exhibit a postovariectomy rise in serum LH concentrations. Serum E2 concentration 49 d after OVX in OVX heifers was threefold greater than in 1 or OVX heifers, thus demonstrating that E2 exerted negative feedback on pituitary LH secretion in prepubertal heifers. There was no measurable difference in serum E2 concentrations between I and OVX heifers; however, the contrast in the concentration and pattern of serum LH between the two groups was dramatic and suggested gonadal factors in addition to E2 are involved in controlling LH secretion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dynorphin-Kappa Opioid Receptor Signaling Partly Mediates Estrogen Negative Feedback Effect on LH Pulses in Female Rats

Accumulating evidence suggests that the arcuate nucleus (ARC) kisspeptin/neurokinin B (NKB)/dynorphin (KNDy) neurons play a role in estrogen negative feedback action on pulsatile gonadotropin-releasing hormone (GnRH)/luteinizing hormone (LH) release. The present study aimed to determine if dynorphin (Dyn) is involved in estrogen negative feedback on pulsatile GnRH/LH release. The effect of the injection of nor-binaltorphimine (nor-BNI), a kappa-opioid receptor (KOR) antagonist, into the third cerebroventricle (3V) on LH pulses was determined in ovariectomized (OVX) adult female rats with/without replacement of negative feedback levels of estradiol (low E₂). The mean LH concentrations and baseline levels of LH secretion in nor-BNI-injected, low E₂-treated rats were significantly higher compared with vehicle-treated controls. On the other hand, the nor-BNI treatment failed to affect any LH pulse parameters in OVX rats without low E₂ treatment. These results suggest that Dyn is involved in the estrogen negative feedback regulation of pulsatile GnRH/LH release. The low E₂ treatment had no significant effect on the numbers of ARC Pdyn (Dyn gene)-,Kiss1- and Tac2 (NKB gene)-expressing cells. The treatment also did not affect mRNA levels of Pdyn and Oprk1 (KOR gene) in the ARC-median eminence region, but significantly increased the ARC kisspeptin immunoreactivity. These findings suggest that the negative feedback level of estrogen suppresses kisspeptin release from the ARC KNDy neurons through an unknown mechanism without affecting the Dyn and KOR expressions in the ARC. Taken together, the present result suggests that Dyn-KOR signaling is a part of estrogen negative feedback action on GnRH/LH pulses by reducing the kisspeptin release in female rats.     

Endocrine changes during restoration of estrous cycles following induction of anestrus by restricted nutrient intake in beef heifers

The working hypotheses in this experiment were: that ovarian estradiol would inhibit luteinizing hormone (LH) secretion in heifers that were anestrus as a result of restricted dietary energy intake and the responsiveness of LH secretion to estradiol negative feedback would decrease during the period when restoration of estrous cycles occurred following feeding of diets adequate in energy. Fifteen heifers weighing 341 +/- 12 (mean +/- SE) kg were fed a diet containing 50% of the energy required for maintenance until 40 to 50 d following cessation of estrous cycles. Heifers were assigned to intact control (C, n = 5), ovariectomized (OVX, n = 5) or ovariectomized-estradiol-17 beta-implanted (OVX + E2, n = 5) treatments. Heifers were subsequently provided a high-energy (HE) diet until termination of the study. Progesterone concentrations indicating cessation of corpus luteum function were detected after heifers had lost 71 +/- 8 kg body weight over 186 +/- 28 d. Control heifers re-initiated estrous cycles as indicated by increased progesterone concentrations in serum at 49 +/- 9 d after initiation of feeding the HE diet (360 +/- 18 kg body weight). Initiation of pulsatile LH secretion was observed in heifers by d 12 following OVX. Estradiol suppressed LH secretion in OVX + E2 heifers during the period of nutritional anestrus in C heifers. Suppressive effects of E2 on LH secretion continued in OVX heifers after C heifers had initiated corpus luteum function. Therefore, the working hypothesis that LH secretion is inhibited by E2 in the nutritionally anestrous heifer is accepted but responsiveness to estradiol does not subside with re-initiation of estrous cycles, thus this working hypothesis is rejected.     

Luteinizing hormone secretion in ovariectomized gilts: effects of age, reproductive state and estrogen replacement

Crossbred gilts were ovariectomized (OVX) at 120, 150, 180 and 210 d of age to determine whether various characteristics of luteinizing hormone (LH) concentrations are influenced by age and reproductive state (prepuberal vs postpuberal). All 120-d-old gilts were prepuberal and all 210-d-old animals were postpuberal, whereas gilts 150 and 180 d old included both prepuberal and postpuberal animals. Blood was collected at 15-min intervals for 2 h, 2 d before OVX (d -2), and 2 (d +2), 8 (d +8) and 14 (d +14) d after OVX. Mean LH concentrations for prepuberal gilts were similar among age groups (P greater than .05) on d -2 and +2; however, LH increased (P greater than .05) from d -2 to +2. No change in LH secretion was found in postpuberal gilts during these two periods. After OVX, LH increased from d +2 to +14 in both prepuberal and postpuberal gilts in all age groups. In postpuberal gilts, LH increased linearly (P less than .05) between d +2 and +14; rate of increase accelerated with advancing age (P less than .01). In prepuberal gilts, LH increased in a nonlinear manner, but it did not increase between d +2 and +8. The increase observed in prepuberal and postpuberal gilts after OVX resulted primarily from an increase in magnitude of peak concentrations of LH. Implants of estradiol-17 beta (E2) were used to determine whether the postovariectomy increase in LH is affected differently by E2 in prepuberal and postpuberal gilts during advancing ages.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of Estradiol and Progesterone on the Numbers and Distribution of Mastocytes in the Uterus of Ovariectomized Rats

The purpose of this study was to investigate the effects of estradiol(E)and progesterone(P)on mastocyte distribution in the uterus of ovariectomized rats.Thirty-five adult female rats were divided randomly into seven groups:one sham operated control group(SHAM);one ovariectomized group(OVX);three ovariectomized plus E treatment groups(OVX+E 20,100,or 500 μg/kg body weight·d);and two ovariectomized plus P groups(OVX+P 2 or 10 mg/kg body weight·d).Seven days after treatment,the contents of estradiol and progesterone in serum were detected by radioimmunoassay,and mastocytes in the uterus were stained by toluidine blue staining.Results were as following:① Compared to ovariectomized rat,the concent ration of estradiol in serum increased by 97.13 % in OVX+E 20(P0.05),204.84 % in OVX+E 100(P0.05),and 936.45 % in OVX + E 500 group(P0.05);the progesterone concent ration increased by 77.25 % in OVX+P 2(P0.05)and 235.25 %in OVX+P 10 group(P0.05).② Compared to ovariectomized rat,the number of mast cells in uteri decreased by 32.65% in OVX+E 20,64.50 % in OVX+E 100(P0.05),74.49 % in OVX+E 500(P0.05)and 70.67 % in OVX+P 10 groups(P0.05).However,the number of mast cells increased by 66.73% in OVX+P 2 group(P0.05)compared with OVX.The trend of mast cells number in the rat uterus was decreased gradually with the increase of estrogen or progesterone concent ration.The number of mast cells in ovariectomized rat uterus was affected by estrogen or progesterone.These results demonstrated that estrogen or progesterone directly affected the number of mast cells in the uterus of rat.     

Involvement of anteroventral periventricular metastin/kisspeptin neurons in estrogen positive feedback action on luteinizing hormone release in female rats

Metastin/kisspeptin, the KiSS-1 gene product, has been identified as an endogenous ligand of GPR54 that reportedly regulates GnRH/LH surges and estrous cyclicity in female rats. The aim of the present study was to determine if metastin/kisspeptin neurons are a target of estrogen positive feedback to induce GnRH/LH surges. We demonstrated that preoptic area (POA) infusion of the anti-rat metastin/kisspeptin monoclonal antibody blocked the estrogen-induced LH surge, indicating that endogenous metastin/kisspeptin released around the POA mediates the estrogen positive feedback effect on GnRH/LH release. Metastin/kisspeptin neurons in the anteroventral periventricular nucleus (AVPV) may be responsible for mediating the feedback effect because the percentage of c-Fos-expressing KiSS-1 mRNA-positive cells to total KiSS-1 mRNA-positive cells was significantly higher in the afternoon than in the morning in the anteroventral periventricular nucleus (AVPV) of high estradiol (E(2))-treated females. The percentage of c-Fos-expressing metastin/kisspeptin neurons was not different between the afternoon and morning in the arcuate nucleus (ARC). Most of the KiSS-1 mRNA expressing cells contain ERalpha immunoreactivity in the AVPV and ARC. In addition, AVPV KiSS-1 mRNA expressions were highest in the proestrous afternoon and lowest in the diestrus 1 in females and were increased by estrogen treatment in ovariectomized animals. On the other hand, the ARC KiSS-1 mRNA expressions were highest at diestrus 2 and lowest at proestrous afternoon and were increased by ovariectomy and decreased by high estrogen treatment. Males lacking the surge mode of GnRH/LH release showed no obvious cluster of metastin/kisspeptin-immunoreactive neurons in the AVPV when compared with high E(2)-treated females, which showed a much greater density of these neurons. Taken together, the present study demonstrates that the AVPV metastin/kisspeptin neurons are a target of estrogen positive feedback to induce GnRH/LH surges in female rats.     

The role of noradrenaline in the generation of the preovulatory LH surge in the ewe

Increasing plasma estrogen (E) levels during the follicular phase of the estrous cycle trigger the pre-ovulatory surge of gonadotropin-releasing hormone (GnRH)/LH. Noradrenaline (NA)-producing cells of the brain stem are involved in regulating GnRH cells and project to the preoptic area (POA) and bed nucleus of stria terminalis (BnST). Input to GnRH cells may be direct or indirect, via relay neurons in the POA/BnST. To investigate this, we ascertained whether an ₁-adrenergic antagonist would block/delay the LH surge in ovariectomised (OVX), E-treated ewes. E benzoate (EB) (50 μg) was injected (i.m.) and Doxazosin (100 nmol/h) or vehicle was infused into the third ventricle 2–26 h after EB injection. Doxazosin reduced the magnitude of the LH surge, but did not affect timing. To determine if NA is released in the POA/BnST of cyclic ewes, we immunostained dopamine-β-hydroxylase (DBH) in terminal fields. Reduced numbers of varicosities staining for DBH indicates release of NA. The number of varicosities immunostained for DBH was reduced in the dorsal and lateral BnST during the follicular phase and during the preovulatory LH surge compared to the luteal phase. These data suggest that noradrenergic mechanisms are involved in generation of the GnRH/LH surge via projections to the BnST and relay to GnRH cells. Since Doxasozin reduced the magnitude of the LH surge in the E-treated OVX ewe, and release of NA in cyclic ewes occurred during the follicular phase of the estrous cycle, we speculate that NA is a permissive factor in surge generation. Thus, increased noradrenergic activity is not a trigger mechanism for initiation of the surge.     

Immunohistochemical distribution of inwardly rectifying K+ channels in the medulla oblongata of the rat

The inwardly rectifying K+ channels, Kir1.1, Kir2.3 and Kir4.1-Kir5.1, are the candidate chemosensory molecules for CO2/H+. We determined the mRNA expression and immunohistochemical localization of these channels in the medulla oblongata of the rat. RT-PCR analysis revealed mRNAs of Kir1.1, Kir2.3, Kir4.1 and Kir5.1 were detected in the medulla. The immunoreactivities for Kir1.1, Kir2.3, Kir4.1, and Kir5.1 were observed in the medulla, and immunolabeling pattern was varied by the subunit. Immunoreactivities for Kir1.1 and Kir2.3 were observed in the nerve cell bodies and glial cells both in the chemosensory areas [nucleus tractus solitarius (NTS), nucleus raphe obscurus (RO), pre-B?tzinger complex (PreB?tC)] and non-chemosensory area [hypoglossal nucleus (XII), inferior olive nucleus (IO)]. Kir4.1 immunoreactivity was observed in the glial cells and neuropil, especially in XII and IO. Kir5.1 immunoreactivity was observed in the nerve cell bodies in the XII, RO, and PreB?tC, but not in the NTS or IO. In the NTS, a dense network of varicose nerve fibers showed immunoreactivity for Kir5.1. Our findings suggest that Kir channels may not act specific to the central chemoreception, but regulate the ionic properties of cellular membranes in various neurons and glial cells.     

Reproductive seasonality in female goats adapted to a subtropical environment persists independently of food availability

The aim of this study was to determine whether the seasonal change in food availability provides a predictive cue that synchronizes the breeding season in goats adapted to the subtropical conditions of Northern Mexico. Groups of multiparous intact (n = 7) and ovariectomized does bearing a subcutaneous implant constantly releasing estradiol-l7β (OVX+E; n = 12) were allocated in two pens and received a constant amount of feed. Another group of OVX+E does (n = 10) was incorporated into a flock raised under natural grazing conditions with seasonal fluctuations in food availability. Blood samples were taken twice weekly. Ovulation of intact goats was inferred from plasma progesterone levels. In OVX+E does plasma levels of LH were measured. Intact does displayed seasonal variations in ovulation and the breeding season lasted from September to February. OVX+E goats displayed large seasonal variations in LH secretion regardless of whether they were fed constantly indoors or kept under natural grazing conditions (P < 0.0001). The mean date of increase in LH secretion varied between years (P < 0.0001) but did not differ between groups (naturally grazed: August 13 ± 10 days; constantly fed: August 26 ± 10 days). In contrast, the seasonal decline in LH secretion differed between groups: it was consistently delayed in constantly fed animals compared to those feeding naturally (naturally grazed: January 28 ± 4 days; constantly fed: February 24 ± 6 days, group effect, P < 0.01). These results show that local female goats from subtropical Mexico display large reproductive seasonality independently of food availability.     

Central somatostatin-somatostatin receptor 2 signaling mediates lactational suppression of luteinizing hormone release via the inhibition of glutamatergic interneurons during late lactation in rats

Reproductive function is suppressed during lactation owing to the suckling-induced suppression of the kisspeptin gene (Kiss1) expression in the arcuate nucleus (ARC) and subsequent suppression of luteinizing hormone (LH) release. Our previous study revealed that somatostatin (SST) neurons mediate suckling-induced suppression of LH release via SST receptor 2 (SSTR2) in ovariectomized lactating rats during early lactation. This study examined whether central SST-SSTR2 signaling mediates the inhibition of ARC Kiss1 expression and LH release in lactating rats during late lactation and whether the inhibition of glutamatergic neurons, stimulators of LH release, is involved in the suppression of LH release mediated by central SST-SSTR2 signaling in lactating rats. A central injection of the SSTR2 antagonist CYN154806 (CYN) significantly increased ARC Kiss1 expression in lactating rats on day 16 of lactation. Dual in situ hybridization revealed that few ARC Kiss1-positive cells co-expressed Sstr2, and some of the ARC Slc17a6 (a glutamatergic neuronal marker)-positive cells co-expressed Sstr2. Furthermore, almost all ARC Kiss1-positive cells co-expressed Grin1, a subunit of N-methyl-D-aspartate (NMDA) receptors. The numbers of Slc17a6/Sstr2 double-labeled and Slc17a6 single-labeled cells were significantly lower in lactating dams than in non-lactating rats whose pups had been removed after parturition. A central injection of an NMDA antagonist reversed the CYN-induced increase in LH release in lactating rats. Overall, these results suggest that central SST-SSTR2 signaling, at least partly, mediates the suppression of ARC Kiss1 expression and LH release by inhibiting ARC glutamatergic interneurons in lactating rats.     

Estrogen Increases c-Fos expression in the paraventricular nucleus along with its anorexic effect in developing rats

Estrogen inhibits food intake in cycling females in a variety of species. To determine how the development of the anorexic system by estrogen is regulated, rat pups at four developmental stages, postnatal day 11 (P11)-13, P20-22, P25-27 and P29-31, and adult ovariectomized (OVX) rats received a daily subcutaneous injection of 20 μg/kg of estradiol benzoate (EB) or vehicle for three days. Food intake, body weight gain and immunohistochemical c-Fos expression in the brain were measured after each injection. EB treatment decreased both food intake and body weight gain from P27 onwards and significantly increased c-Fos expression in the parvocellular division of the paraventricular nucleus of the hypothalamus (pPVN), which is coincident with its anorexic effect in developing rats. The pattern of EB-induced c-Fos activation in other feeding-related nuclei did not coincide with its anorexic effect in developing pups. However, in adult OVX rats, EB treatment increased c-Fos expression in the nucleus tractus solitarius (NTS), the central nucleus of the amygdala (CeA), and, to a lesser degree, the ventromedial nucleus of the hypothalamus (VMH). These results suggested that the pPVN is an essential site in the brain for controlling the anorexic effect of estrogen and that the feeding system of rat begins to respond to estrogen before the onset of puberty (P25-28).     

Fourth ventricular alloxan injection suppresses pulsatile luteinizing hormone release in female rats

Previous studies have suggested the presence of a glucose-sensing mechanism in the hindbrain that appears to regulate reproductive function as well as feeding behavior. The ependymocytes lining the ventricular wall of the hindbrain showed immunoreactivities to pancreatic glucokinase (GK), a key enzyme for glucose sensing in pancreatic B cells. Our goal in the present study was to test whether the GK-immunopositive ependymocytes in the wall of the fourth cerebroventricle (4V) play a role in regulating gonadal activity. Our approach was to determine the effect of injecting alloxan, a GK inhibitor, into the 4V on pulsatile luteinizing hormone (LH) secretion. Estrogen-primed ovariectomized rats received an injection of alloxan (10 or 20 microg/animal) into the 4V and blood samples were collected every 6 min for 3 h for measurement of blood LH, corticosterone and glucose levels. Pulsatile LH secretion was suppressed after alloxan injection and all pulse parameters were significantly (P<0.05) inhibited by 20 microg alloxan. Plasma corticosterone levels were increased significantly (P<0.05) by 20 microg alloxan, suggesting that LH pulse suppression by alloxan may be at least partly mediated by activation of the hypothalamo-pituitary-adrenal axis. The present results suggest that acute suppression of GK activity in the hindbrain inhibits pulsatile LH secretion in female rats, and supports the idea that GK-immunopositive ependymocytes may sense glucose levels in the cerebrospinal fluid and play a role in regulation of LH secretion.     

The effects of GnRH and adrenergic agents on PRL and beta-endorphin secretion by porcine pituitary cells in vitro

The direct effects of alpha- and beta-adrenergic agents on PRL and beta-endorphin (beta-END) secretion in vitro by porcine pituitary cells have been investigated. Pituitary glands were obtained from mature gilts, which were ovariectomised (OVX) one month before slaughter. Ovariectomised gilts, assigned to four groups, were primed with: (1) vehicle (OVX); (2) and (3) oestradiol benzoate (EB; 2.5 mg/100 kg b.w.) at 30-36 h (OVX+EB I) and 60-66 h (OVX+EB II) before slaughter, respectively; and (4) progesterone (P4; 120 mg/100 kg b.w.) for 5 consecutive days before slaughter (OVX+P4). Isolated anterior pituitary cells were submitted to 3.5 h incubation in the presence of GnRH, alpha- and beta-adrenergic agonists [phenylephrine (PHEN) and isoproterenol (ISOP), respectively], or alpha- and beta-adrenergic blockers [phentolamine (PHENT) and propranolol (PROP), respectively]. The culture media were assayed for PRL (exp. I) and beta-endorphin-like immunoreactivity (beta-END-LI) (experiment II). In experiment I, GnRH did not influence PRL release by pituitary cells in all experimental groups. Some of tested doses of adrenergic agonists, PHEN and ISOP, increased PRL release from pituitary cells of OVX gilts, but not from those of OVX+EB I animals. In the OVX+EB II group, PHEN alone, but ISOP with PROP, potentiated PRL secretion by the cells. In OVX+P4 animals, PHEN alone or in combination with PHENT and also ISOP alone or with PROP enhanced PRL output from the cells. In experiment II, addition of GnRH increased beta-END-LI release from pituitary cells only in the OVX+EB II group. PHEN and PHENT potentiated beta-END-LI secretion by pituitary cells in OVX+EB II and OVX+P4 groups, while ISOP and PROP increased beta-END-LI secretion by the cells of OVX and OVX+EB II animals. In turn, in the OVX+EB I group, effect of PHENT and PROP on PRL secretion by pituitary cells was inhibitory. In conclusion, our results suggest that adrenergic agents can modulate PRL and beta-END secretion by porcine pituitary cells in a manner dependent on the hormonal status of gilts.     

Changes in luteinizing hormone secretion after estradiol treatment in prepubertal Nelore heifers

Changes in luteinizing hormone (LH) secretion after 17β-estradiol (E(2)) injection were evaluated during sexual maturation in 10 prepubertal Nelore heifers. Heifers were divided into 2 groups: intact (I) and ovariectomized (OVX). 17β-estradiol (2 μg/kg) was administered to both groups at 10, 13, and 17 mo of age. Only at 10 mo of age was there a greater mean LH concentration in OVX heifers (1.33 ± 0.29 ng/mL) compared with the I group (0.57 ± 0.15 ng/mL). At 13 and 17 mo of age there was no significant difference between the 2 groups in any of the evaluated variables (number of peaks, total peak area, greatest peak area, and time to greatest peak occurrence). This suggests a decrease in negative E(2) feedback associated with an increase in positive feedback to LH secretion during sexual maturation, and these were likely the key factors that determined the time of first ovulation in Nelore heifers.     

滴度及注射方式对2型腺相关病毒载体表达效果的影响

由于腺相关病毒载体(adeno-associated viral vector,AAV)本身的侵染不具有神经元特异性,其在神经系统相关研究中会存在侵染范围不符合试验要求的情况,因此本试验拟研究不同滴度和注射方式对病毒侵染范围的影响.结果显示,在注射较高滴度的AAV2-CMV-EGFP(1.3×101 mg/L)3周后,...