The study was designed to determine the in vitro effect of bacterial endotoxin – lipopolysaccharide (LPS) – on luteinizing hormone (LH) secretion from anterior pituitary (AP) explants collected from saline-treated ‘healthy’ and LPS-treated ewes in the follicular phase. In the AP explants, the expression of LH-β, gonadotropin-releasing hormone receptor (GnRH-R) and Toll-like receptor 4 (TLR4) genes was also assayed. It was found that explants incubated alone with LPS, as well as together with LPS-binding protein (LBP), abolished the stimulatory effect of GnRH on LH release. LPS added without LBP did not suppress LH-β gene expression in ‘healthy’ APs, however. Moreover, LBP intensified the inhibitory effect of LPS on gene expression in ‘healthy’ APs. These results show that LPS is a potent negative modulator of LH secretion and suggest that its direct action on the pituitary gland could be one of the mechanisms via which an immune/inflammatory challenge inhibits the reproductive process.
Bacterial endotoxin, LPS, is recognized by Toll-like receptor-4 (TLR4) and induces a signaling cascade leading to synthesis of proinflammatory cytokines and induction of sickness behavior in animals. Transduction of the TLR4 signal is controlled by a potent negative regulator—Toll-interacting protein. The presented study concerns the effect of intravenously injected LPS on the level of expression of TLR4 and Tollip genes in the hypothalamus of ewes. Endotoxin increased (P < 0.01) cortisol release and expression of TLR4 and Tollip genes in the preoptic area (1.87 ± 0.42 and 1.31 ± 0.15), anterior hypothalamus (1.77 ± 0.22 and 1.27 ± 0.13), medial basal hypothalamus (2.53 0.65 and 1.43 ± 0.15), and median eminence (2.93 ± 0.46 and 1.73 ± 0.10), respectively, in comparison with non-treated animals. Our results show that immune stress increases TLR4 gene expression in the hypothalamus. Increased transcription of Tollip may be an attempt to reduce the effect of TLR4 stimulation.
One of the most important inhibitory neurotransmitter in the control of gonadoliberin (GnRH) secretion is gamma-aminobutyric acid (GABA). To examine the role of GABAA receptor mediating systems in the control of GnRH/LH release from the preoptic area and ventromedial hypothalamus of ewes in different reproductive stages (seasonal anestrus, follicular and luteal phase of the estrous cycle) the extracellular concentration of GnRH, ß-endorphin, noradrenaline, dopamine and metabolites of catecholamines: MHPG, DOPAC were quantified during local stimulation or blockade of GABAA receptors with muscimol or bicuculline, respectively. Stimulation of GABAA receptors in the preoptic area and ventromedial hypothalamus markedly attenuatted GnRH/LH release in the anestrous ewes and in sheep during follicular phase of the estrous cycle but did not affect these hormones secretion in animals during the luteal phase of the estrous cycle. The activation of GABAA receptors in these structures in different way affected ß-endorphinergic and catecholaininergic system activity in anestrous ewes arid animals during the follicular phase; lack of changes in the release of ß-endorphin, noradrenaline and dopamine was noted in ewes during the luteal phase. With the exception of preoptic area in anestrous ewes the blockade of GABAA receptors in the preoptic and ventromedial hypothalamus affected in specifical way the activity of ß-endorphinergic and catecholaminergic systems in these structure in all animals. In conclusion: presented results indicate that activation of GABAA receptors in the preoptic area and ventromedial hypothalamus of anestrous ewes and sheep during follicular phase of the estrous cycle inhibits GnRH/LH secretion. Presented results indicate that different neural mechanisms may be involved in the suppression of GnRH/LH by GABA directly through the GABAA receptor mechanism on the perikaria or on the axon terminals of GnRH cells, and/or indirectly through GABAA receptor processes on ß-endorphinergic and catecholaminergic systems. Lack of changes in GnRH/LH release during GABAA receptor activation in ewes during luteal phase of the estrous cycle indicates that the influence of GABA on GnRH-ergic activity depends on physiological state of animals. Blockade of GABAA receptors in the anestrous and cycling ewes does not change GnRH/LH release; it is suggested that blockade of GABAA recreceptors may change GnRH neurons response to ß-endorphin and dopamine.
To examine the role of GABAA receptor mediating systems in the control of gonadotropin-releasing hormone (GnRH) release from the ventromedial- infundibular region (VEN/NI) in ewes during luteal phase, the extracellular concentrations of GnRH, ß-endorphin, noradrenaline (NE), dopamine (DA), and their metabolites: MHPG and DOPAC were quantified by local stimulation or blockade of GABAA receptors with muscimol or bicuculline, respectively. Stimulation of GABAA receptors in the VEN/NI did not affect GnRH, ß-endorphin release or catecholaminergic system activity. Blockade of GABAA receptors decreased ß-endorphinergic and dopaminergic activity, and lowered the extracellular concentration of MHPG. It did not affect GnRH release or luteinizing hormone (LH) secretion. It is suggested that progesterone-induced GABAergic activity during the luteal phase may desensitize GABAA receptors to muscimol. Lack of changes in GnRH/LH secretion with concomitant depressed ß-endorphinergic activity corroborated the conclusion that ß-endorphin does not inhibit GnRH release from the VEN/NI during the luteal phase. The physiological significance of changes in the catecholaminergic system activity under GABAA receptor blockade in the control of GnRH secretion awaits to be established.
Salsolinol with its derivatives has been considered as a potential neurotoxin for the dopaminergic system in the human and rat brain. Investigating a sheep model for studies on the action of salsolinol within the central nervous system we examined whether this compound is able to affect the hypothalamic neuroendocrine dopaminergic (NEDA) system during its high seasonal activity, when sheep entered to anestrus under the long day conditions. Therefore, salsolinol was infused into the third ventricle of the brain in combination with the in vivo push-pull perfusion of the mediobasal hypothalamus/median eminence (MBH/ME). The effects of this drug on either perfusate noradrenaline (NA) or plasma prolactin concentration were also studied. The infusion of salsolinol resulted in rapid and permanent diminution in dopamine (DA) release into the extracellular spaces of the MBH/ME up to an undetectable level and in the 57% decrease in DA metabolite 3,4- dihydroxyphenylacetic acid concentration, compared to the control. This effect of salsolinol was accompanied by the significant enhancement of the pituitary prolactin release into circulation. The concentration of other DA metabolite, homovanillic acid, as well as NA in the MBH/ME was not affected. Thus, our results in the anestrous sheep underline the role played by salsolinol as a neuromodulator for the hypothalamic NEDA system and as a signal transmitter for the pituitary prolactin release. We suggest that the hypothalamic NEDA system of anestrous sheep during its high secretory activity may be set as a model for studies on the salsolinol-dopamine relationship.
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