Differential effects of glutamate agonists and D-aspartate on oxytocin release from hypothalamus and posterior pituitary of male rats.

In order to determine whether ionotropic (iGluRs) and metabotropic (mGluRs) glutamate receptor activation modulates oxytocin release in male rats, we investigated the effect of agonists of both types of glutamate receptors on oxytocin release from hypothalamus and posterior pituitary. Kainate and quisqualate (1 mM) increased hypothalamic oxytocin release. Their effects were prevented by selective AMPA/kainate receptor antagonists. NMDA (0.01-1 mM) did not modify hypothalamic oxytocin release. Group I mGluR agonists, such as quisqualate and 3-HPG, significantly increased hypothalamic oxytocin release. These effects were blocked by AIDA (a selective antagonist of group I mGluRs). In the posterior pituitary, oxytocin release was not modified by kainate, quisqualate, trans-ACPD (a broad-spectrum mGluR agonist) and L-SOP (a group III mGluR agonist). However, NMDA (0.1 mM) significantly decreased oxytocin release from posterior pituitary. D-Aspartate significantly increased oxytocin release from the hypothalamus, while it decreased oxytocin release from posterior pituitary. AP-5 (a specific NMDA receptor antagonist) reduced the D-Aspartate effect in the hypothalamus, but not in the posterior pituitary. Our data indicate that the activation of non-NMDA receptors and group I mGluRs stimulates oxytocin release from hypothalamic nuclei, whereas NMDA inhibits oxytocinergic terminals in the posterior pituitary. D-Aspartate also has a dual effect on oxytocin release: stimulatory at the hypothalamus and inhibitory at the posterior pituitary. These results suggest that excitatory amino acids differentially modulate the secretion of oxytocin at the hypothalamic and posterior pituitary levels.

Role of phosphodiesterase and protein kinase G on nitric oxide-induced inhibition of prolactin release from the rat anterior pituitary.

OBJECTIVE: In order to determine the mechanism by which nitric oxide (NO) inhibits prolactin release, we investigated the participation of cGMP-dependent cAMP-phosphodiesterases (PDEs) and protein kinase G (PKG) in this effect of NO. METHODS: Anterior pituitary glands of male rats were incubated with inhibitors of PDE and PKG with or without sodium nitroprusside (NP). Prolactin release, and cAMP and cGMP concentrations were determined by RIA. RESULTS AND CONCLUSIONS: The inhibitory effect of NP (0.5 mmol/l) on prolactin release and cAMP concentration was blocked by EHNA (10(-4)mol/l) and HL-725 (10(-4)mol/l), inhibitors of cGMP-stimulated cAMP-PDE (PDE2). 8-Br-cGMP (10(-4) and 10(-3)mol/l), which mimics cGMP as a mediator of NP effects on prolactin release, also decreased cAMP concentration. Zaprinast (10(-4)mol/l), a selective inhibitor of specific cGMP-PDE (PDE5), potentiated the NP effect on cAMP concentration. Rp-8-[(4-chlorophenyl)thio]-cGMP triethylamine (Rp-8-cGMP, 10(-7)-10(-6)mol/l), an inhibitor of PKG, reversed the effect of NP on prolactin release. The present study suggests that several mechanisms are involved in the inhibitory effect of NO on prolactin release. The activation of PDE2 by cGMP may mediate the inhibitory effect of NO on cAMP concentration and therefore on prolactin release. NO-activated PKG may also be participating in the inhibitory effect of NO on prolactin release.

The effect of excitatory aminoacids on GABA release from mediobasal hypothalamus of female rats.

The purpose of the present study was to examine the in vitro effect of L-glutamate and its agonists on basal and potassium-evoked GABA release from incubated mediobasal hypothalamus (MBH) of intact, ovariectomized (OVX) and OVX-estrogenized female rats. L-glutamate (100 microM) decreased evoked GABA release from MBH of intact female rats in diestrus. NMDA and quisqualate (10 and 100 microM) modified neither basal nor evoked hypothalamic GABA release of intact rats. However, kainate (10 and 100 microM) decreased hypothalamic basal and evoked GABA release of intact rats. Kainate induced no changes in basal or in evoked GABA release from hypothalami of OVX rats, but decreased GABA release in chronically estrogenized rats. DNQX (6,7-dinitroquinoxaline-2,3-dione), a non-NMDA receptor antagonist, failed to affect GABA release but blocked the inhibitory effect of kainate. The kainate effect was not Mg2+-sensitive and was not inhibited by D-AP5 (D(-)-2-amino-5-phosphonopentanoic acid), an NMDA-specific receptor antagonist. Kainate induced no changes in nitric oxide synthase activity in MBH of either intact or estrogenized rats. These data indicate that kainate decreases GABA release from MBH of female rats through a non-NMDA receptor subtype, and provide evidence to support the view that kainate-mediated decrease of the hypothalamic GABAergic tone is affected by estrogens.

Nitric oxide inhibits the release of norepinephrine and dopamine from the medial basal hypothalamus of the rat.

Previous research indicates that norepinephrine and dopamine stimulate release of luteinizing hormone (LH)-releasing hormone (LHRH), which then reaches the adenohypophysis via the hypophyseal portal vessels to release LH. Norepinephrine exerts its effect via alpha 1-adrenergic receptors, which stimulate the release of nitric oxide (NO) from nitricoxidergic (NOergic) neurons in the medial basal hypothalamus (MBH). The NO activates guanylate cyclase and cyclooxygenase, thereby inducing release of LHRH into the hypophyseal portal vessels. We tested the hypothesis that these two catecholamines modulate NO release by local feedback. MBH explants were incubated in the presence of sodium nitroprusside (NP), a releaser of NO, and the effect on release of catecholamines was determined. NP inhibited release of norepinephrine. Basal release was increased by incubation of the tissue with the NO scavenger hemoglobin (20 micrograms/ml). Hemoglobin also blocked the inhibitory effect of NP. In the presence of high-potassium (40 mM) medium to depolarize cell membranes, norepinephrine release was increased by a factor of 3, and this was significantly inhibited by NP. Hemoglobin again produced a further increase in norepinephrine release and also blocked the action of NP. When constitutive NO synthase was inhibited by the competitive inhibitor NG-monomethyl-L-arginine (NMMA) at 300 microM, basal release of norepinephrine was increased, as was potassium-evoked release, and this was associated in the latter instance with a decrease in tissue concentration, presumably because synthesis did not keep up with the increased release in the presence of NMMA. The results were very similar with dopamine, except that reduction of potassium-evoked dopamine release by NP was not significant. However, the increase following incubation with hemoglobin was significant, and hemoglobin, when incubated with NP, caused a significant elevation in dopamine release above that with NP alone. In this case, NP increased tissue concentration of dopamine along with inhibiting release, suggesting that synthesis continued, thereby raising the tissue concentration in the face of diminished release. When the tissue was incubated with NP plus hemoglobin, which caused an increase in release above that obtained with NP alone, the tissue concentration decreased significantly compared with that in the absence of hemoglobin, indicating that, with increased release, release exceeded synthesis, causing a fall in tissue concentration. When NO synthase was blocked by NMMA, the release of dopamine, under either basal or potassium-evoked conditions, was increased. Again, in the latter instance the tissue concentration declined significantly, presumably because synthesis did not match release. Therefore, the results were very similar with both catecholamines and indicate that NO acts to suppress release of both amines. Since both catecholamines activate the release of LHRH, the inhibition of their release by NO serves as an ultra-short-loop negative feedback by which NO inhibits the release of the catecholamines, thereby reducing the activation of the NOergic neurons and decreasing the release of LHRH. This may be an important means for terminating the pulses of release of LHRH, which generate the pulsatile release of LH that stimulates gonadal function in both male and female mammals.

Involvement of hypothalamic substance P in the effect of prolactin on dopamine release.

In order to examine the role of hypothalamic SP in the feedback regulation of prolactin, we studied the effect of prolactin and dopamine on SP concentration and release, and the effect of SP on dopamine release. Hypothalamic fragments from male Wistar rats were incubated in the presence of prolactin, dopamine or SP under basal and K(+)-stimulated conditions. SP (10(-7) M) stimulated dopamine release, while dopamine (10(-7) M) decreased SP content and release. Prolactin (100 ng ml-1) increased SP content and release. An increase in hypothalamic SP content was also found during suckling. In addition, a specific antagonist for SP, Win 62,577, blocked the effect of prolactin and dopamine release. These results show an interaction between SP and dopamine at the hypothalamic level and suggest that SP could mediate the feedback action of prolactin on dopamine release.

Possible mechanisms of action of ethanol-induced release of prolactin from rat anterior pituitary.

We have demonstrated previously that ethanol increases the in vitro synthesis and release of prolactin by the anterior pituitary gland. In the present study we have examined the possible role of calcium and calmodulin in the ethanol-stimulation of in vitro prolactin release from hemipituitary glands. We also investigated the effect of inhibitors of arachidonic acid metabolism on the release of prolactin induced by ethanol. In addition we studied the effect of ethanol on the release of prostaglandins and the conversion of arachidonic acid to hydroperoxides. Basal and ethanol-stimulated prolactin release was almost blocked completely by omitting calcium from the incubation medium. In vitro addition of calmodulin-blocking agents such as pimozide and trifluoperazine reduced basal and ethanol-induced prolactin release. Acetylsalicylic acid, an inhibitor of the cycloxygenase pathway, had no effect on prolactin release. Nordihydroguaiaretic acid, an inhibitor of the lipoxygenase pathway, blocked completely the release of prolactin induced by ethanol. Ethanol (5 and 10 mM) significantly increased the release of prostaglandin E2 and prostaglandin F2 alpha and the conversion of [14C]arachidonic acid to 5-hydroxyeicosatetraenoic acid. The results of this study show that ethanol increases prolactin release by a calcium-dependent mechanism. Besides, our data suggest that arachidonic acid metabolism is involved in ethanol-stimulated prolactin secretion and that lipoxygenase pathway metabolites are at least partially responsible for this effect.

Mechanisms of endogenous GABA release from hypothalamic fragments. Effect of prolactin.

The efflux of endogenous gamma-aminobutyric acid (GABA) has been studied using small hypothalamic fragments containing arcuate-paraventricular nuclei and median eminence from the rat brain. The amount of GABA present in the medium and the tissue GABA content were quantified by radioreceptor assay. The endogenous GABA efflux was found to be dependent upon the Ca2+ and K+ concentrations in the incubation medium, and it required synthesis of GABA, indicating neuronal origin of the released neurotransmitter. Nipecotic acid, an inhibitor of neuronal and glial uptake, prevented reuptake of released GABA. Prolactin in concentrations of 250 and 1,000 ng/ml augmented the K+-evoked efflux of GABA. The effect of prolactin was dependent on the presence of Ca2+ and on the synthesis of GABA. In addition, prolactin seems to alter the reuptake of endogenous GABA and the uptake of [3H]-GABA. In conclusion, these results suggest that prolactin may influence its own secretion by stimulating the release of hypothalamic GABA, both through an increase of its synthesis and a modification of its reuptake.

Pituitary response to luteinizing hormone-releasing hormone during haloperidol-induced hyperprolactinemia.

The effects of a 6-hour infusion with haloperidol on serum prolactin and luteinizing hormone (LH) levels was studied in a group of male subjects. Five hours after starting the infusions, a study of the pituitary responses to LH-releasing hormone (LH-RH) was carried out. Control patients received infusions of 0.9% NaCl solution. During the course of haloperidol infusions, significant hyperprolactinemia was found, together with an abolished pituitary response to LH-RH, as compared with responses of control subjects.