Martha F. Gimeno

Interleukin 1 alpha inhibits prostaglandin E2 release to suppress pulsatile release of luteinizing hormone but not follicle-stimulating hormone.

Interleukin 1 alpha (IL-1 alpha), a powerful endogenous pyrogen released from monocytes and macrophages by bacterial endotoxin, stimulates corticotropin, prolactin, and somatotropin release and inhibits thyrotropin release by hypothalamic action. We injected recombinant human IL-1 alpha into the third cerebral ventricle, to study its effect on the pulsatile release of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in conscious, freely moving, ovariectomized rats. Intraventricular injection of 0.25 pmol of IL-1 alpha caused an almost immediate reduction of plasma LH concentration; this decrease was statistically significant 20 min after injection and occurred through a highly significant reduction in the number of LH pulses, with no effect on pulse amplitude. In contrast, there was no change in pulse frequency but a small significant elevation in amplitude of FSH pulses. Intraventricular injection of the diluent had no effect on gonadotropin release. The results provide further evidence for separate hypothalamic control mechanisms for FSH and LH release. To determine the mechanism of the suppression of LH release, mediobasal hypothalamic fragments were incubated in vitro with IL-1 alpha (10 pM) and the release of LH-releasing hormone (LHRH) and prostaglandin E2 into the medium was measured by RIA in the presence or absence of norepinephrine (50 microM). IL-1 alpha reduced basal LHRH release and blocked LHRH release induced by norepinephrine. It had no effect on the basal release of prostaglandin E2; however, it completely inhibited the release of PGE2 evoked by norepinephrine. To evaluate the possibility that IL-1 alpha might also interfere with the epoxygenase pathway of arachidonic acid metabolism, epoxyeicosatrienoic acids were also measured. IL-1 alpha had no effect on the content of epoxyeicosatrienoic acids in the hypothalamic fragments as measured by gas chromatography and mass spectrometry. In conclusion, IL-1 alpha suppresses LH but not FSH release by an almost complete cessation of pulsatile release of LH in the castrated rat. The mechanism of this effect appears to be by inhibition of prostaglandin E2-mediated release of LHRH.

Melatonin, 5-methoxytryptamine and some of their analogs as cyclo-oxygenase inhibitors in rat medial basal hypothalamus

Melatonin, and its analogs 6-chloro- and 6-fluoromelatonin inhibited in a dose-dependent way (10(-8)-10(-5) M) labeled prostaglandin (PG) E2, PGF2 alpha, thromboxane (Tx) B2 and 6-keto-PGF1 alpha production from [14C]arachidonate by rat medial basal hypothalamus (MBH). 5-Methoxytryptamine also depressed arachidonate metabolism; at 10(-8) M concentrations the effect of 5-methoxytryptamine on PGE2, PGF2 alpha and TxB2 synthesis (93-96% inhibition), and 6-keto-PGF1 alpha (75% inhibition) was greater than that observed for melatonin (51-56% and 44% inhibition, respectively). Neither 6-hydroxymelatonin nor serotonin affected MBH cyclo-oxygenase pathway in vitro.

Role of Nitric Oxide in Control of Growth Hormone Release in the Rat

Previous experiments in this and other laboratories have revealed that nitric oxids (NO) plays a role in controlling the release of corticotropin-releasing hormone (CRH) and luteinizing-hormone-releasing hormone (LHRH). Therefore, we have investigated its role in control of growth hormone (GH) release in conscious rats by microinjecting NG-monomethyl-L-arginine (NMMA), an inhibitor of NO synthase (NOS), into the third ventricle (3V) of conscious, freely moving castrate male rats. An initial blood sample (0.3 ml) was drawn from an indwelling intra-atrial catheter just prior to injection of NMMA [1 mg in 5 microliters of 0.9% NaCl (saline)] into the 3V. To maintain the inhibitory action on NOS, a second injection of NMMA was administered into the 3V 60 min after the first. Additional blood samples (0.3 ml) were removed at 10 min intervals for 120 min. Other animals received injections of the diluent at the same times and volumes as NMMA. Interleukin (IL)-1 alpha (0.06 pmol in 2 microliters saline) was injected into the 3V immediately after the first injection of NMMA, whereas other animals received the NMMA diluent followed by IL-1 alpha. The effects of IL-1 alpha were almost identical to those of NMMA in that there was a dramatic lowering of plasma GH achieved primarily by a reduction in height of the GH pulses without a significant reduction in their number.(ABSTRACT TRUNCATED AT 250 WORDS)

Blockade by lnterleukin-1-Alpha of Nitricoxidergic Control of Luteinizing Hormone-Releasing Hormone Release in vivo and in vitro

Nitric oxide (NO) synthase (NOS), the enzyme that converts arginine into citrulline plus NO, the latter a highly active free radical, occurs in a large number of neurons in the brain, including certain neurons in the hypothalamus. Our previous experiments have shown that norepinephrine (NE)-induced prostaglandin E2 (PGE2) release from medial basal hypothalamic explants (MBH) is mediated by NO. Because release of luteinizing hormone (LH)-releasing hormone (LHRH) is also driven by NE and PGE2, we hypothesized that NO controls pulsatile release of LHRH in vivo, which in turn induces pulsatile LH release. Indeed, in vivo and in vitro experiments using an inhibitor of NOS (NG-monomethyl-L-arginine; NMMA) demonstrated that pulsatile LH release is mediated by NO; LHRH release in vitro is also mediated by this free radical. Cytokines that are released from cells of the immune system during infection also inhibit LHRH release. We compared the action of one such cytokine, interleukin-1 alpha (IL-1 alpha), on LHRH release with that of substances which inhibit or induce NO release. Microinjection of IL-1 alpha (0.06 pmol in 2 microliters) into the third cerebral ventricle (3V) of conscious, castrated male rats had an action similar to that of 3V microinjection of NMMA (1 mg in 5 microliters): it blocked pulsatile LH, but not follicle-stimulating hormone (FSH) release. The only difference between the responses to NMMA and IL-1 alpha was that the latency to onset was greater with IL-1 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of prostaglandin F2α (PGF2α) on oviductal nitric oxide synthase (NOS) activity: possible role of endogenous NO on PGF2α-induced contractions in rat oviduct

Abstract In this paper, we evaluated the hypothesis that prostaglandin F 2α (PGF 2α ) regulates NOS activity and we also investigated, by means of nitric oxide inhibitors (N-monomethyl- l -arginine monoacetate, l -NMMA) the role of endogenous NO on PGF 2α -induced contractions in rat oviduct. NOS activity was determined by measuring the conversion of 14 [C]- l -arginine to 14 [C]- l -citrulline on oviductal homogenates from estrogenized rats (1 μg/rat). The presence of PGF 2α (10 −8 M) in the incubation medium produced an increase in NOS activity ( p ≤ 0.05). The effect of the prostanoid was blocked completely by the presence of two NOS inhibitors: N-nitro- l -arginine methyl ester ( l -NAME, 0.6 mM) and aminoguanidine (Ag, 0.5 mM). These results suggested that PGF 2α could be modulating the Ca 2+ -independent NOS activity. We determined NOS activity using 1 mM EGTA, a chelator of Ca 2+ , in a free Ca 2+ medium. These results indicated that PGF 2α produces an increase in Ca 2+ -independent NOS activity ( p ≤ 0.05). PGF 2α induces contraction of the oviductal smooth muscle in a concentration dependent manner. l -NMMA enhanced PGF 2α induced contraction of the oviduct, providing indirect evidence that there is a basal release of NO in the oviduct, which may reduce and/or modulate the contractile effects of PGF 2α .

Physiologic and Pharmacologic Studies on the Motility of Isolated Guinea Pig Ovaries

The spontaneous motility of ovaries isolated from guinea pigs in early and late proestrus and estrus was explored. The influences of oxytocin, prostaglandin F2alpha (PGF2alpha), and adrenergic agonists and antagonists were also studied. Spontaneous ovarian isometric developed tension (IDT) and rate of contractions (RC) were greater in late proestrus than in early proestrus or estrus. Furthermore, in late proestrus, oxytocin and PGF2alpha induced ovarian motilities of comparable magnitude and significantly greater than those elicited in other stages of the cycle. Norepinephrine and phenylephrine either failed to alter or inhibited IDT and RC of ovaries in estrus, but stimulated motility in the presence of propranolol. The depressive influence of norephinephrine upon the IDT of ovaries in early proestrus was not seen in late proestrus, when the neurotransmitter was clearly stimulating. The hormonal status of guinea pigs seems to influence spontaneous ovarian motility as well as pharmacologic reactivity to several agents, including those presumably acting upon alpha- and beta-adrenergic receptors.

A Positive Influence of Frequency on the Inotropic Response of Isolated Rat Atria to Certain Adrenergic Amines. Effects of Cocaine and U-0521 (3′, 4′-Dihydroxy-2-methyl Propiophenone)

Summary The effects of several inotropic interventions on the isometric developed tension (IDT) of isolated left rat atrium, electrically driven at several frequencies were investigated. The experimental stimulating procedure allowed the attainment at all the rates tested (25, 50, 100, and 200 per min), similar magnitudes of IDT prior to drug addition. Two types of contractile responses to the inotropic agents at different beating frequencies were found: (1) Norepinephrine and isoproterenol enhanced the IDT less at the slower rates (25 and 50 per min) than at the highest rate (200 per min); (2) phenylephrine and CaCl2 similarly augmented atrial IDT at all driving rates. The presence of cocaine potentiated the effect of norepinephrine at lower frequencies but not at 200 beats per min; in addition, it did not modify the inotropic influence of isoproterenol at any of the rates explored. U-0521 enhanced the positive inotropism of both norepinephrine and isoproterenol only at low stimulating frequencies. The results suggest that: (a) The dissimilar effects of norepinephrine and isoproterenol at low and high rates could be associated with a different importance of the inactivating processes of these amines at the various frequencies, namely, presynaptic neuronal uptake and metabolization via COMT. Both mechanisms appear to be more important at lower rates and this could be the reason underlying a diminished influence of norepinephrine or isoproterenol over slowly beating atria. (b) The positive influence of frequency over the inotropic effects of norepinephrine and isoproterenol is not due to the magnitude of atrial contractions existing before their addition.

Role of Nitric Oxide in the Actions of Cytokines on Hypothalamic Releasing Hormone Secretion

Interleukins act within the hypothalamus to cause the release of corticotropin-releasing hormone (CRH) and inhibit release of luteinizing hormone-releasing hormone (LHRH). Nitric oxide synthase (NOS), the enzyme that converts arginine into citrulline plus nitric oxide (NO), the latter a highly active free radical, occurs in a large number of neurons in the brain, including certain neurons in the hypothalamus. Our in vivo experiments in rats employing NG-monomethyl-L-arginine (NMMA), an inhibitor of NOS and nitroprusside, a releaser of NO, have determined that interleukin-2 (IL-2) activates CRH release by acting on its receptors on cholinergic neurons which stimulate NOergic neurons by muscarinic receptors. The NO diffuses to the CRH neurons and activates them as revealed by in vitro experiments with hypothalamic explants. On the other hand, norepinephrine or glutamic acid activate LHRH release by stimulating NOergic neurons that in turn induce LHRH release by activation of guanylate cyclase and cyclooxygenase. IL-lα acts directly on the LHRH neuron to block its response to NO on the basis of in vitro experiments. In vivo, NO stimulates LHRH release which induces pulsatile LH but not FSH release on the one hand and induces mating behavior on the other. The IL-1 -induced blockade of LHRH release by NO may account for the reduction in libido in infection. In vivo experiments employing injection of NMMA into the 3rd ventricle have revealed that NO mediates the prolactin-releasing action of interleukins. It also mediates pulsatile growth hormone (GH) release via stimulation of growth hormone-releasing hormone (GRH) release. IL-1 inhibits pulsatile GH release by blocking the response of the GRH neurons to NO. IL-1 also stimulates somatostatin release.

Spontaneous and Prostaglandin- or Oxytocin-Induced Motility of Rat Ovaries Isolated During Different Stages of the Estrous Cycle: Effects of Norepinephrine**Supported in part by Grants 6154 and 6638 from the Consejo Nacional de Investigaciones Científicas y Técnicas de la República Argentina.

Ovaries isolated from rats in different stages of the sex cycle were explored for spontaneous or drug-induced contractile activity. The number of spontaneously active ovaries as well as the magnitude of the isometrically developed tension and frequency of contractions were greater during the periovulatory interval (late proestrus and estrus) than during early proestrus or metestrus. Furthermore, during estrus or late proestrus the left ovaries exhibited a mechanical activity significantly greater than that of the right ovaries. The oxytocin-triggered motility was clearly more marked immediately prior to ovulation (late proestrus) and greater in left ovaries than in right ovaries. In contrast, the contractions induced by prostaglandin F2alpha were similar during early proestrus and late proestrus. Ovarian contractile reactivity to norepinephrine indicated the presence in the tissue of alpha- and beta-adrenergic receptors. During early proestrus this agent stimulated the motility of left and right ovaries, whereas close to the ovulatory interval (late proestrus) it depressed the contractions of left ovaries. This last influence was blocked by propranolol. The existence of a close relationship between ovarian contractile activity and ovulation is reinforced by the present results in the rat. A tentative participation of oxytocin is also suggested. In addition, the influences of other possible regulatory agents of ovarian contraction, such as catecholamines and prostaglandins, are presented and discussed.

Physiologic and Pharmacologic Studies on the Motility of Isolated Guinea Pig Cauda Epididymidis**Supported in part by Grants 6154 and 6638 from the Consejo Nacional de Investigaciones Cientifícas y Técnicas de la República Argentina.

The effects of norepinephrine, phentalamine, oxytocin, vasopressin, several prostaglandins, and indomethacin on the spontaneous motility of isolated guinea pig cauda epididymidis were explored. Phentolamine and indomethacin reduced the isometric peak tension of spontaneous epididymal contractions. Phentolamine also depressed the frequency. Both findings suggest that catecholamines and endogenous prostaglandins are in some way regulators of the spontaneous motility of the cauda epididymidis. Norepinephrine resulted in the development of a distinct, sustained, tonic contraction without phasic activity, whereas prostaglandins E1, E2, and F2 alpha elicited a tonic increase accompanied by frequent, superimposed, phasic contractions. Both oxytocin and vasopressin comparably enhanced epididymal motility, producing contractile responses similar to those observed with prostaglandins. Since the epididymal contractions can influence the time spent by spermatozoa in passing through the ductus epididymidis, the above-mentioned compounds could play an important role in spermatozoal transport via modulation of epididymal contractile activity. In addition, such naturally occurring substances might regulate the release of sperm from the last portion of the epididymis into the ductus deferens.