Disruptions in the hypothalamic-pituitary-gonadal axis in rat offspring following prenatal maternal exposure to lipopolysaccharide.

Postnatal treatment with bacterial endotoxin lipopolysaccharide (LPS) changes the activity of the hypothalamic-pituitary-gonadal (HPG) axis and the gonadotropin-releasing hormone (GnRH) surge in rats. Exposure to an immune challenge in the critical periods of development has profound and long-lasting effects on the stress response, immune, metabolic, and reproductive functions. Prenatal LPS treatment delays the migration of GnRH neurons associated with increased cytokine release in maternal and fetal compartments. We investigated the effects of a single maternal exposure to LPS (18 µg/kg, i.p.) on day 12 (embryonic day (E)12) of pregnancy on reproductive parameters in rat offspring. Hypothalamic GnRH content, plasma luteinizing hormone (LH), testosterone, and estradiol concentrations were measured in both male and female offsprings at different stages of postnatal development by RIA and ELISA (n = 10 each per group). Body weight and in females day of vaginal opening (VO) were recorded. In offspring exposed to LPS prenatally, compared with controls, body weight was decreased in both sexes at P5 and P30; in females, VO was delayed; hypothalamic GnRH content was decreased at postnatal days 30-60 (P30-P60) in both sexes; plasma LH concentration was decreased at P14-P60 in females; plasma concentrations of testosterone/estradiol were increased at P14 in females, and plasma estradiol was increased at P14 in males. Hence activation of the maternal immune system by LPS treatment at a prenatal critical period leads to decreased GnRH and LH levels in pre- and postpubertal life and sex steroid imbalance in the prepubertal period, and delayed sexual maturation of female offspring.

Gonadotropin-releasing hormone stimulates the biosynthesis of pregnenolone sulfate and dehydroepiandrosterone sulfate in the hypothalamus.

The sulfated neurosteroids pregnenolone sulfate (Δ(5)PS) and dehydroepiandrosterone sulfate (DHEAS) are known to play a role in the control of reproductive behavior. In the frog Pelophylax ridibundus, the enzyme hydroxysteroid sulfotransferase (HST), responsible for the biosynthesis of Δ(5)PS and DHEAS, is expressed in the magnocellular nucleus and the anterior preoptic area, two hypothalamic regions that are richly innervated by GnRH1-containing fibers. This observation suggests that GnRH1 may regulate the formation of sulfated neurosteroids to control sexual activity. Double labeling of frog brain slices with HST and GnRH1 antibodies revealed that GnRH1-immunoreactive fibers are located in close vicinity of HST-positive neurons. The cDNAs encoding 3 GnRH receptors (designated riGnRHR-1, -2, and -3) were cloned from the frog brain. RT-PCR analyses revealed that riGnRHR-1 is strongly expressed in the hypothalamus and the pituitary whereas riGnRHR-2 and -3 are primarily expressed in the brain. In situ hybridization histochemistry indicated that GnRHR-1 and GnRHR-3 mRNAs are particularly abundant in preoptic area and magnocellular nucleus whereas the concentration of GnRHR-2 mRNA in these 2 nuclei is much lower. Pulse-chase experiments using tritiated Δ(5)P and DHEA as steroid precursors, and 3'-phosphoadenosine 5'-phosphosulfate as a sulfonate moiety donor, showed that GnRH1 stimulates, in a dose-dependent manner, the biosynthesis of Δ(5)PS and DHEAS in frog diencephalic explants. Because Δ(5)PS and DHEAS, like GnRH, stimulate sexual activity, our data strongly suggest that some of the behavioral effects of GnRH could be mediated via the modulation of sulfated neurosteroid production.

Behavioural and neurobiological effects of colostrum ingestion in the newborn lamb associated with filial bonding.

In sheep, the onset of filial bonding relies on early intake of colostrum. The aim of our work was to describe in the newborn lamb housed with its mother the immediate post-ingestive effects of colostrum intake, in terms of behaviour and brain activity. In Experiment 1, lambs received five nasogastric infusions of colostrum, or saline, or sham intubations during the first 6 h after birth. Mother-young interactions were recorded before and after the first, third and fifth infusions. The activity of the dam and of the young, which diminished over time in all groups, was temporarily increased in both partners just after each intubation procedure. The number of high-pitched bleats was significantly lower in lambs that received colostrum than in the sham group, suggesting soothing or satiating properties of colostrum. In Experiment 2, newborn lambs received a single nasogastric infusion of colostrum or saline 4.5 h after birth, or were sham intubated. Neuronal activation was investigated 1.5 h later for maximum c-Fos activity. Infusion of colostrum and saline induced different patterns of c-Fos-like immunoreactivity in the paraventricular and supraoptic nuclei of the hypothalamus as compared with the sham group. A specific oxytocinergic/vasopressinergic (OT/VSP) cell population in the paraventricular nucleus was activated following colostrum and saline infusion, but not sham intubation. Only colostrum induced the activation of the cortical amygdala and insular cortex, two structures involved in learning, associative processes, reward and emotion. We hypothesize that filial bonding may be triggered through colostrum-rewarded learning/calming processes and that the OT/VSP system may play a role.

[Hypothalamic neuropeptides and control of GnRH neurones. Neuroanatomical study in the ewe]. / Les neuropeptides hypothalamiques dans le contrôle des neurones à GnRH. Etude neuroanatomique chez la brebis.

Reproduction in mammals is directly controlled by GnRH neurons. These neurons are regulated by many external and internal factors, among which sexual steroids, in particular oestradiol, play an important part. However the mechanisms through which these steroids regulate GnRH secretion are largely unappreciated, and the neurochemical identity of central neurons liable to transmit the steroidal information to GnRH neurons is not completely clarified. Many functional neuroanatomy studies have been carried out on the ovine model, which is particularly favorable to understand the neuroendocrine mechanisms controlling reproduction. These studies have brought about the identification of some of the potential actors in this regulation. The present review reports the major results concerning two recently discovered neuropeptides, galanin and kisspeptin, which appear to be major actors in integration of signals regulating reproduction, among which steroids. These results have revealed the major interaction sites between neurons expressing these neuropeptides and GnRH neurons.

Prepubertal changes in the synthesis, storage and release of the somatostatin in the hypothalamus of female lambs: a morphofunctional study.

It is assumed that hypothalamic somatostatin plays a role in the preovulatory phase of the oestrous cycle in sheep. The aim of the study was to investigate the processes of synthesis, storage and release of somatostatin in hypothalamic neurons, in immature female lambs, in the period approaching to puberty. Experiments were carried out on 10 prepubertal (17 weeks old) and 10 peripubertal (32 weeks old) ovary-intact lambs. Morphofunctional changes in the somatostatin neurons were assayed with immunohistochemistry and hybridisation in situ. Computer image analysis was used to determine the density of both reactions and the percentage of the area exhibiting immunohistochemical staining. These parameters express the content of immunoreactive (ir) somatostatin and expression of mRNA for pre-pro-somatostatin (PPS). Two populations of ir somatostatin perikarya were localized in the hypothalamus: a very large number of perikarya in the periventricular (PEV) nucleus, and single cell bodies in the arcuate (ARC) nucleus. Only ir somatostatin fibres, but no perikarya were seen in the ventromedial (VM) nucleus and preoptic area. The analysis of mRNA PPS showed perikarya filled with silver grains localized in the PEV, ARC and VM. There were differences in the content of ir somatostatin and the intensity of the PPS mRNA signal between the two periods investigated. In the median eminence, the content of ir somatostatin in the terminals decreased in the peripubertal compared to the prepubertal group (P<0.001). In the PEV, the content of ir somatostatin in the perikarya and the expression of PPS mRNA decreased in the peripubertal compared to the prepubertal group (P<0.001). In the ARC, the content of ir somatostatin in the perikarya increased (P<0.001), but expression of PPS mRNA decreased (P<0.001) in the peripubertal compared to the prepubertal group. There were no differences in the expression of PPS mRNA in the VM. We concluded, that the different secretory activity of the two hypothalamic populations of somatostatin neurons can be related to their different physiological functions in the prepubertal period of female lambs.

Efficacy and safety of three ophthalmic inserts for topical anaesthesia of the cornea. An exploratory comparative dose-ranging, double-blind, randomized trial in healthy volunteers.

BACKGROUND: Cataract surgery requires prolonged anaesthesia, concomitant with permanent hydration and lubrication of the cornea, in order to provide a clear view of the operation area. AIMS: The primary objective of the study was to assess several formulae of a soluble ophthalmic insert: TOPICSERT [bupivacaine (Bupi) + hyaluronic acid (HA) or sodium hyaluronate] in terms of complete and long-lasting anaesthesia of the cornea. The hydration properties of HA were not assessed in this study. METHODS: In a prospective double-blind, cross-over, randomized study, with latin-square allocation of treatments, 16 healthy volunteers received a single dose of each formula (A, 1 mg Bupi and 0.1 mg HA; B, 0.5 mg Bupi and 0.1 mg HA; C, 1 mg Bupi and 0 mg HA, and D acting as a placebo) via the ocular route with 1 week of wash-out between each period. Corneal anaesthesia was measured using a Cochet-Bonnet esthesiometer. RESULTS: There was a statistically significant difference between treatments with regard to the main criterion (complete anaesthesia lasting at least 20 min) when general association statistics were used (Mantel-Haenzel test, P < 0.0001): 68.75% (n = 11) of subjects receiving treatment A, 37.5% (n = 6) receiving treatment B, and 87.5% (n = 14) on treatment C reached complete and satisfactory anaesthesia, while this was not achieved in any of the subjects receiving placebo. Ninety-five percent confidence intervals of the difference between treatments were as follows: treatment A vs. B (-0.03, 0.66), treatment A vs. C (-0.47, 0.10), treatment B vs. C (-0.84, - 0.16). Only the difference between B and C was statistically significant (adjusted probability by the method of Bonferroni, P < 0.001). When complete anaesthesia was reached, mean (+/-SD) duration of anaesthesia was as follows: 20.7 (+/-6.5), 15.3 (+/-11.4) and 24.7 (+/-7.6) min for treatments A, B, C, respectively. CONCLUSIONS: Bupivacaine 1 mg seems to be the efficient and safe dose. The value of hyaluronic acid as a corneal hydration agent and used in association with bupivacaine will be the subject of further studies.

Differential c-Fos expression in the newborn lamb nucleus tractus solitarius and area postrema following ingestion of colostrum or saline.

Visceral stimuli and the gut-brain axis play a crucial role in the control of ingestion even in the neonate. The aim of this study was to assess the neuronal activation in the nucleus tractus solitarius (NTS) and the area postrema (AP) following nutritional and non-nutritional stimulations. Lambs received a single gastric infusion of colostrum or saline at 5% birth weight or were sham infused. Infusion of either liquid led to c-Fos-like immunoreactivity (c-FLI) in the NTS and AP. Differences were observed along the sections of the NTS rostro-caudal axis according to the nature of the stimulation, suggesting a specificity of certain afferents and/or NTS areas for nutritional or non-nutritional signals. In the AP, the neuronal activation induced by colostrum was much higher than that induced by saline. A higher number of TH-immunoreactive cells were activated following colostrum infusion, suggesting a specific involvement of the catecholaminergic pathway in the treatment of meal-related stimuli. In spite of functional convergence, the two medullary structures observed responded differently according to the stimulation, indicating a complementary role in the integration of visceral signals.