Estrogen reduces acute striatal dopamine responses in vivo to the neurotoxin MPP+ in female, but not male rats

The effects of in vivo estrogen treatment upon MPP⁺-induced dopamine (DA) release were determined using in vivo microdialysis in female and male rats. Ovariectomized female rats were implanted or not with an estrogen pellet (0.1 mg, 17β estradiol) and subjected to microdialysis 6 days later. After baseline DA release was determined, 5 mM MPP⁺ was infused through the microdialysis probe for one 20-min interval. Perfusion resumed with normal medium for the duration of the experiment. A significant attenuation of MPP⁺-induced DA release was obtained in estrogen-treated females. One week later, striatal DA and dihydroxyphenylacetic acid (DOPAC) concentrations were determined for the lesioned and non-lesioned striata of each animal. MPP⁺ infusion significantly decreased striatal DA concentrations, however, there was no effect of estrogen treatment on striatal DA depletion. This experiment was repeated using orchidectomized male rats treated with 0, 0.1, or 5 mg estradiol. In contrast to the females, no differences in MPP⁺-induced DA release were seen among these males, and there was no significant effect of the varying estrogen treatments on striatal DA or DOPAC concentrations. These results demonstrate that in vivo estrogen treatment attenuates MPP⁺-induced striatal DA release in gonadectomized female, but not male, rats.     

Estrogen decreases corpus striatal neurotoxicity in response to 6-hydroxydopamine

Ovariectomized rats treated or not with an estradiol pellet were subjected to an unilateral intrastriatal infusion of 6-hydroxydopamine (6-OHDA). Various parameters of nigrostriatal dopaminergic function as derived from measurements of dopamine and dihydroxyphenylacetic acid (DOPAC) concentrations were determined from the 6-OHDA lesioned and non-lesioned sides of the corpus striatum in these animals. Dopamine concentrations within the 6-OHDA lesioned striatum of estrogen-treated rats were significantly greater than non-estrogen-treated rats. There were no differences in striatal dopamine concentrations between estrogen- versus non-estrogen-treated rats on their non-lesioned side. In contrast to that of dopamine, no differences in DOPAC concentrations between estrogen and non-estrogen-treated rats were obtained within the 6-OHDA-lesioned side. The DOPAC concentrations on the non-lesioned side of the striatum were significantly greater in the non-estrogen-treated rats. These results demonstrate that estrogen significantly diminishes the depletion of striatal dopamine resulting from the neurotoxin 6-OHDA. The data obtained from the DOPAC determinations imply that this capacity of estrogen may be exerted through actions upon uptake processes of striatal dopaminergic neurons. Such findings suggest that estrogen may function as an important modulatory factor capable of attenuating degeneration within the corpus striatum, and in this way serve as a neuroprotectant of the nigrostriatal dopaminergic system.     

Lowering ambient or core body temperature elevates striatal MPP+ levels and enhances toxicity to dopamine neurons in MPTP-treated mice

The neuroprotective effects of lowering body temperature have been well documented in various models of neuronal injury. The present study investigated the effects a lower ambient or core body temperature would have on damage to striatal dopamine (DA) neurons produced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Mice received systemic MPTP treatment at two different temperatures, 4°C and 22°C. MPTP-treated mice maintained at 4°C demonstrated (1) a greater hypothermic response, (2) a significant reduction in striatal DA content and tyrosine hydroxylase (TH) activity, and (3) significantly greater striatal 1-methyl-4-phenylpyridinium (MPP+) levels, as compared to mice dosed with MPTP at room temperature. Parallel studies with methamphetamine (METH) were conducted since temperature appears to play a pivotal role in the mediation of damage to DA neurons by this CNS stimulant in rodents. As previously reported, METH-induced hyperthermia and the subsequent loss of striatal DA content were attenuated in animals dosed at 4°C. We also evaluated the effects a hypothermic state induced by pharmacological agents would have on striatal neurochemistry and MPP+ levels following MPTP treatment. Concurrent administration of MK-801 or 8-OHDPAT increased the striatal MPP+ levels following MPTP treatment. However, only 8-OHDPAT potentiated the MPTP-induced decrements of striatal DA content and TH activity; MK-801 did not affect MPTP decreases in these striatal markers of dopaminergic damage. Altogether, these findings indicate that temperature has a profound effect on striatal MPP+ levels and MPTP-induced damage to DA neurons in mice.     

Use of in vitro superfusion to assess the dynamics of striatal dopamine clearance: influence of estrogen

To determine the feasibility of assessing dopamine uptake using in vitro superfusion, striatal tissue from ovariectomized female rats was infused with dopamine (1 microM), nomifensine (1 mM), or a combination of dopamine and nomifensine. Treatment with nomifensine or dopamine/nomifensine increased the recovery of dopamine in the effluent samples as compared to treatment with dopamine alone. In Experiment 2, the striatal tissue was treated with varying concentrations (0, 3, 30 or 300 nM) estradiol throughout the superfusion and subsequently given a dopamine (1 microM) challenge. The recovery of dopamine was enhanced in the presence of 3 and 30 nM estradiol. These results show that (1) in vitro superfusion can be used to dynamically evaluate dopamine recovery, and (2) estradiol, like nomifensine, increases the recovery of exogenously applied dopamine from the striata of ovariectomized female rats. Such increases in dopamine recovery with estrogen and similarities to that obtained with nomifensine suggest that estrogen may be inhibiting dopamine uptake from these striatal tissue fragments. Moreover, the doses at which estrogen can exert these effects insinuates a physiological role for this process. Our data provide a clear functional demonstration for one of the mechanisms by which estradiol can modulate striatal dopamine neurons, that of an uptake inhibitor. Such a mechanism has important implications with regard to estradiol's capacity to function as a neuroprotectant of the nigrostriatal dopaminergic system through inhibition of uptake of neurotoxins which can produce neurodegeneration of striatal dopamine neurons.     

Differential neurotoxicity induced by -DOPA and dopamine in cultured striatal neurons

The neurotoxicity of -DOPA and dopamine (DA) on striatal neurons was examined by using primary cultures of rat striatum. Exposure to -DOPA and DA at concentrations of 30–300 μM induced dose-dependent cell death in both younger cultures (3 days in culture, 3 DIC) and elder cultures (10 days in culture, 10 DIC). The cytotoxicity of -DOPA and DA was also dependent on the exposure time (6–24 h). Ascorbic acid (200 μM) inhibited both -DOPA- and DA-induced cytotoxicity in 3 DIC cultures, whereas it provided significant protection against DA- but not -DOPA-induced cytotoxicity in 10 DIC cultures. The -DOPA cytotoxicity in 10 DIC cultures was prevented by a non-NMDA receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), and by an NMDA receptor antagonist, MK-801. Neither antagonist prevented DA cytotoxicity. -DOPA did not affect the viability of 10 DIC cultures, though it elicited marked toxicity in 3 DIC cultures. These results suggest that there are two components in the mechanisms that mediate the -DOPA neurotoxicity on striatal neurons: one is autoxidation-relevant and the other is autoxidation-irrelevant. With respect to the latter, glutamate receptor stimulation may be involved. In contrast, autoxidation plays an important role in DA neurotoxicity.     

Receptor subtype-specific modulation by dopamine of glutamatergic responses in striatal medium spiny neurons

The output of GABAergic medium-sized spiny neurons in the dorsal striatum is controlled in part by glutamatergic input from the neocortex and the thalamus, and dopaminergic input from ventral midbrain. We acutely isolated these neurons from juvenile (P14-24) rats to study the consequences of the interaction between glutamate and dopamine for neuronal excitability. Single-cell RT-PCR analysis was used to identify the expression patterns of dopamine receptors. D1 and D2 dopamine receptor mRNA was detected in 11/22 and 3/22 of isolated neurons, respectively. Receptor mRNA co-expression was detected in 1/22 cells tested. Whole-cell voltage clamp recording (V(h)=-70 mV) was combined with local or bath application of dopaminergic and glutamatergic agonists to explore dopamine receptor modulation of glutamatergic excitation. Glutamate-evoked inward currents (5 microM, Mg(2+)-free, 1 microM glycine) were attenuated by dopamine (5 microM) to 83.2+/-3.6% (n=31). NMDA-evoked (20 microM), APV-sensitive currents were attenuated by dopamine to 80.9+/-4.5% (n=24). NMDA-induced responses were also attenuated by the D1 receptor agonist SKF 38393 (1 microM; n=28), while the D2/3 receptor agonist quinpirole (10 microM) had no effect. The currents evoked by application of AMPA (5 microM) displayed a steady rundown. Application of dopamine abolished or significantly reduced the rundown in the cells tested (n=17). A similar effect was observed after the application of SKF 38393 (1 microM), while quinpirole (10 microM) had no significant effect. Our results provide direct evidence for modulation by dopamine of glutamatergic responses of striatal medium spiny neurons, and demonstrate that the effects of this neuromodulator are receptor subtype specific. Disruption of this modulatory effect is likely to contribute to movement disorders associated with Parkinson's disease.     

Antioxidant compounds protect dopamine neurons from death due to oxidative stress in vitro

Using tissue culture models of oxidative stress caused by serum deprivation or MPTP/MPP+ toxicity, the present study establishes that the antioxidants epigallocatechin gallate, lazaroids U74389G and U83836E, reservatrol, MnTBAP, MCI 186, trolox, and melatonin protect 68-100% of dopamine (DA) neurons from cell death. In contrast, the nitric oxide inhibitor LY83583, the caspase inhibitors Z-VAD-FMK, Ac-DQMD-CHO and Z-DEVD-FMK, and the CDK-5 inhibitor, roscovotine were not neuroprotective, although death was often delayed by 1 day in vitro. We conclude that antioxidants are more effective at preventing cell death in vitro than are inhibitors at later stages in the death cascade.     

MPP(+) and glutamate in the degeneration of nigral dopaminergic neurons

Glutamate neurotoxicity can be an experimental oxidative stress, and we investigated glutamate toxicity against cultured rat mesencephalic neurons. Although glutamate showed similar toxicity against dopaminergic and nondopaminergic neurons, nitric oxide (NO) showed neurotoxicity restricted exclusively in nondopaminergic neurons. An inhibitor of NO synthase had no significant effect on the glutamate toxicity against dopaminergic neurons, however, it had a significant antagonistic effect on that against nondopaminergic neurons. These findings indicate the presence of two mechanisms of glutamate neurotoxicity, one being not mediated by NO, found in dopaminergic neurons, and the other being mediated via NO, found in nondopaminergic neurons. In contrast to NO, peroxynitrite (ONOO⁻), an active metabolite of NO, caused significant cytotoxicity against dopaminergic and nondopaminergic neurons, suggesting that conversion of NO to ONOO⁻ is suppressed in dopaminergic neurons. After pretreatment with small doses of methyl-4-phenylpyridium ion (MPP⁺), NO caused significant cytotoxicity against dopaminergic neurons, and glutamate toxicity was enhanced only against dopaminergic neurons. Therefore, sublethal dose of MPP⁺ enhances glutamate toxicity against dopaminergic neurons, probably by the facilitation of suppressed NO conversion to ONOO⁻ in dopaminergic neurons. Finally, to provide basic data for neuroprotective therapy in Parkinson's disease, we investigated neuroprotection against glutamate toxicity by dopamine agonists. Preincubation with the D2 type dopamine agonists provides neuroprotection against glutamate neurotoxicity and the protective effects blocked by a D2 antagonist, indicating that D2 agonists provide protection mediated not only by the inhibition of dopamine turnover, but also via D2 type dopamine receptor.     

Mesolimbic dopamine projection modulates amygdala-evoked EPSP in nucleus accumbens neurons: an in vivo study

In urethane anesthetized rats, excitatory postsynaptic potential (EPSP) recorded intracellularly from nucleus accumbens neurons following stimulation of the amygdala was attenuated by repetitive stimulation of the ventral tegmental area (VTA). VTA stimulation also depolarized the resting membrane potential of accumbens neurons. Attenuation of the EPSP and membrane depolarization were frequently dissociated but both were blocked by haloperidol, a dopamine antagonist.     

Partial characterization of kainic acid-induced striatal dopamine release using in vivo microdialysis

The aim of this study was to characterize interactions between striatal kainate (KA) receptors and dopamine (DA) release using in vivo microdialysis. After insertion of a microdialysis probe and establishment of baseline DA release, each preparation was standardized with a pulse of an iso-osmotic solution of 100 mM KCl in Ringer's solution. DA release following pharmacological manipulation was compared to potassium-induced release and expressed as a percent value. In one group of animals, KA (12.5 mM in Ringer's solution) was administered via the microdialysis probe in 2, 3, 5 or 10 min pulses 30 min following standardization with potassium resulting in release of DA which was15.7 ± 3.9, 30.3 ± 11.3, 67.5 ± 15.0and92.9 ± 19.8% of potassium-induced DA release, respectively. Perfusion of CdCl₂ (0.6 mM in Ringer's solution) 30–45 min prior to a 10 min KA pulse significantly reduced KA-induced DA release compared to control values. Intrastriatal administration of kynurenate (Kyn) attenuated KA-induced DA release in a dose-dependent manner. Levels of DA metabolites in striatal perfusates were significantly reduced following KA administration. This effect was partially reversed by cadmium pretreatment but not affected by Kyn pretreatment. Findings of this study indicate that KA induces striatal DA release in a dose-dependent manner, and this effect is at least partially dependent upon activation of calcium channels. Results also indicate dose-dependent inhibition of KA-induced striatal DA release by the excitatory amino acid receptor antagonist, Kyn, suggesting that this compound interacts with striatal KA receptors and that these receptors are involved with modulating striatal DA release in vivo.     

Neuroprotective effects of dopamine D2 receptor agonist on neuroinflammatory injury in olfactory bulb neurons in vitro and in vivo in a mouse model of allergic rhinitis

Available evidence indicates that dopamine D2 receptor modulates the neurotoxic effects induced by glutamate. However, neurotoxicity mediated by AMPA-subtype glutamate receptor has rarely been studied in the olfactory bulb. This study mainly explores the neuroprotective effects of dopamine D2 receptor agonist on AMPA receptor-mediated neurotoxicity in the olfactory bulb in a mouse model of allergic rhinitis (AR) with olfactory dysfunction (OD). In our study, we found that AR with OD was closely associated with increased surface expression of the AMPA receptor GluR1, reduced surface expression of GluR2, and apoptosis damage in the olfactory bulb in vivo. Quinpirole (a dopamine D2 receptor agonist) improved olfactory function in mice, ameliorated apoptosis injury in the olfactory bulb but not in the olfactory mucosa, and inhibited the internalization of GluR2-containing AMPA receptor in vitro and in vivo. In addition, phosphorylation plays a crucial role in the regulation of AMPA receptor trafficking. Our results showed that quinpirole reduced the phosphorylation of GluR1 S845 and GluR2 S880 in olfactory bulb neurons in vitro, but it had no obvious effect on GluR1 S831. Therefore, dopamine D2 receptor agonist may inhibit the phosphorylation of GluR1 S845 and GluR2 S880, thereby reducing AMPA receptor-mediated neurotoxicity and alleviating neurotoxic injury to the olfactory bulb caused by AR.     

左旋多巴对体外培养大鼠纹状体的神经毒性及其保护方法的研究

目的 研究左旋多巴（Ｌ－ｄｏｐａ）的神经毒性及其神经保护方法。方法 用胎盼兰染色法检测Ｌ－ｄｏｐａ和抗氧化剂还原型谷胱甘肽（ＧＳＨ）、ＮＭＤＡ受体拮抗剂ＭＫ８０１对体外培养大鼠纹状体的神经元活力的影响。结果 Ｌ－ｄｏｐａ使大鼠纹状体神经元活力明显下降（Ｐ〈０．０１）,ＧＳＨ和ＭＫ８０１可明显减轻Ｌ－ｄｏｐａ引起的纹状体神经元活力下降（Ｐ〈０．０１）。结论 Ｌ－ｄｏｐａ可能通过氧化应激和兴奋ＮＭＤＡ受体两个途径发挥其神经毒性,抗氧化剂或ＮＭＤＡ受体拮抗剂与Ｌ－ｄｏｐａ合用可能减轻Ｌ－ｄｏｐａ的神经毒性。     

Thermal responses of ventromedial hypothalamic neurons in vivo and in vitro

The ventromedial hypothalamic neurons were recorded in anesthetized rats and in slice preparations. Whereas almost all thermosensitive neurons (27 out of 28) in the slice preparations showed warm sensitivity, cold-responsive neurons were recorded more frequently than warm-responsive ones during preoptic thermal stimulation in anesthetized rats. These results suggest that most cold-responsive neurons excited by preoptic cooling in the anesthetized rats were influenced by synaptic inputs.     

In vivo and in vitro responses of neurons in the ventrolateral medulla to hypoxia

Neurons in the ventrolateral medulla (VLM) are known to be involved in several cardiorespiratory reflexes and to provide tonic drive to sympathetic preganglionic neurons. Recent studies have suggested that VLM neurons modulate the respiratory responses to hypoxia and to hypercapnia. The purpose of the present study was to determine with electrophysiological techniques if the discharge of these neurons is altered by hypoxia and/or by hypercapnia both in vivo and in vitro. Extracellular single-unit activity of VLM neurons (n = 39) was recorded during inhalation of a hypoxic gas (10% O₂) and during inhalation of a hypercapnic gas (5% CO₂) in anesthetized, spontaneously breathing rats (n = 16). Hypoxia elicited an increase in the discharge frequency in 64% of the VLM neurons studied; hypercapnia stimulated 42% of the neurons. Fifty-two percent of the neurons were stimulated by both hypoxia and hypercapnia. Signal averaging revealed that 76% of the hypoxia-stimulated neurons had a resting discharge related to the cardiac and/or respiratory cycle. Similar percentages of VLM neurons (35/54) were stimulated by hypoxia in a second group of animals (n = 14) that were studied after sinoaortic denervation. A rat brain slice preparation was then used to determine if hypoxia exerts a direct effect upon neurons in the VLM. Perfusing a hypoxic gas over the surface of medullary slices evoked an increase in the discharge frequency in the majority (39/49) of VLM neurons studied; responses were graded in relation to the magnitude of the hypoxic stimulus. Similar responses to hypoxia were observed in VLM neurons studied during perfusion with a synaptic blockade medium. Retrograde labeling of VLM neurons with rhodamine tagged microspheres injected into the thoracic intermediolateral cell column demonstrated that the hypoxia sensitive neurons were located in a region of the VLM that projects to the thoracic spinal cord. These results demonstrate that neurons in the ventrolateral medulla are excited by a direct effect of hypoxia; these neurons may play a critical role in the cardiorespiratory responses to hypoxia.     

Electrophysiological and pharmacological characterization of identified nigrostriatal and mesoaccumbens dopamine neurons in the rat

Extracellular single-unit recording techniques were used to compare the basal activity and pharmacological responsiveness of identified nigrostriatal and mesoaccumbens dopamine (DA)-containing neurons. The projection area of each DA cell was determined by antidromic activation techniques. The forebrain stimulation used for the cell identification procedure did not alter the pharmacological responsiveness of DA neurons; the inhibitory effect of apomorphine (and d-amphetamine) was identical when stimulation was applied either prior to or following drug administration. Analysis of the spike discharge pattern revealed that a higher proportion of mesoaccumbens DA cells exhibited burst-firing activity. Although the firing pattern of the two populations of burst-firing DA cells was similar in many regards, mesoaccumbens DA cells exhibited a longer postburst inhibition than did nigrostriatal DA cells. Each of the DA agonists, apomorphine, pergolide, B-HT 920, and d-amphetamine, inhibited nigrostriatal and mesoaccumbens DA neuronal activity in a similar fashion. However, there was a marked population difference in the recovery of cell firing in the 10 minutes following apomorphine-induced inhibition; the recovery of mesoaccumbens spike discharges was considerably slower. Although this population difference was apparent to some extent following administration of pergolide or B-HT 920 (but not d-amphetamine), it was considerably less marked. The present findings are discussed with respect to the known regulatory control of midbrain DA neurons.     

Endogenous kynurenate controls the vulnerability of striatal neurons to quinolinate: Implications for Huntington's disease

Excessive activation of NMDA receptors results in excitotoxic nerve cell loss, which is believed to play a critical role in the pathophysiology of Huntington's disease (HD) and several other catastrophic neurodegenerative diseases. Kynurenic acid (KYNA), a neuroinhibitory tryptophan metabolite, has neuroprotective properties and may serve as an endogenous anti-excitotoxic agent. This hypothesis was tested in the striatum, using mice with a targeted deletion of kynurenine aminotransferase II (KAT II), a major biosynthetic enzyme of KYNA in the mammalian brain. On post-natal day (PND) 14, the striatum of mkat-2-/- mice showed a reduction in KYNA levels but contained normal concentrations of the metabolically related neurotoxins 3-hydroxykynurenine and quinolinic acid (QUIN). Intrastriatal injections of QUIN, a NMDA receptor agonist, caused significantly larger lesions in these immature mutant mice than in age-matched wild-type animals. This lesion enlargement was not observed when mkat-2-/- mice were acutely pre-treated with the kynurenine 3-hydroxylase inhibitor UPF 648, which counteracted the striatal KYNA deficit. Moreover, no increased vulnerability to QUIN was observed in 2-month-old mkat-2-/- mice, which present with normal brain KYNA levels. Intrastriatal injections of the non-NMDA receptor agonist kainate caused similar lesion sizes in both genotypes regardless of age. These results indicate that endogenous KYNA preferentially controls the vulnerability of striatal neurons to QUIN. Our data suggest that timely pharmacological interventions resulting in an up-regulation of brain KYNA levels may benefit patients suffering from HD or other neurodegenerative diseases.     

Electrophysiological characterization of rat striatal neurons in vitro following a unilateral lesion of dopamine cells

The effects of a unilateral 6 to 19-week lesion of dopamine cells on the excitability of rat striatal neurons were investigated in vitro using the intracellularly recorded membrane properties of neurons obtained ipsilateral and contralateral to 6-hydroxydopamine (6-OHDA) injection sites. Neurons ipsilateral to the lesion site and in striatal tissue depleted of dopamine exhibited resting membrane potentials and membrane resistances similar to those recorded in contralateral striatal neurons. Denervation appeared to have no appreciable effect on the proportion of neurons exhibiting various patterns of neuronal spiking (repetitive, bursting, or single spike) evoked by depolarizing current pulses. Current-voltage determinations revealed nominal rectification in the majority of neurons and marked nonlinearty consistent with inward rectification at potentials hyperpolarized and depolarized to rest in a large proportion of the remaining neurons. Neurons ipsilateral to 6-OHDA lesion sites exhibited these relationships in the same proportion as contralateral control cells. However, ipsilateral neurons with nominal rectification exhibited an average rate constant for the early onset of small hyperpolarizing membrane transients which was significantly smaller than that of controls. This finding suggests that intrinsic membrane parameters regulating the excitability of certain striatal neurons may be under the influence of dopamine or other factors closely associated with nigrostriatal nerve terminals. Published 1993 Wiley-Liss, Inc.     

Intracellularly recorded response of rat striatal neurons in vitro to fenoldopam and SKF 38393 following lesions of midbrain dopamine cells

The effect of long-term (6–19 weeks) 6-hydroxydopamine-induced (6-OHDA) lesions of midbrain dopamine cells on dopamine D₁-like agonist-induced changes in the excitability of rat striatal neurons was investigated in vitro using tissue slices and intracellular recording techniques. Fenoldopam and (±)-SKF 38393 predominantly decreased excitability in control preparations including striatal neurons located contralateral to 6-OHDA injection sites and neurons obtained from rats receiving sham injections or no treatment. Fenoldopam also inhibited neurons ipsilateral to lesions of midbrain dopamine cells. (±)-SKF 33393, unlike fenoldopam, produced predominantly increases in the excitability of ipsilateral striatal neurons. Superfusion of the D₁ receptor antagonist, SCH 23390, blocked fenoldopam-induced decreases in excitability but not the (±)-SKF 38393-induced excitation of neurons ipsilateral to the lesion. Sequential application of fenoldopam and quinpirole, a D₂/D₃ receptor agonist, produced responses to both drugs in a majority of neurons. The results demonstrate that inhibitory responses to fenoldopam are mediated by D₁ receptors, while excitatory effects of (±)-SKF 38393 in the striatum ipsilateral to the lesion are apparently not dependent on D₁ receptor activation. These findings also suggest that dopamine D₁ and D2/D3 receptors are able to concurrently influence the excitability of striatal neurons in the dopamine deafferentated striatum. Similar regulation of striatal neurons in vivo may contribute to dopaminergic regulation of basal ganglia output and the ability of dopaminomimetic agents to ameliorate symptoms of dopaminergic deficiency in Parkinson's disease. © 1994 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America.

     

Pregnanolone, a metabolite of progesterone, stimulates LH-RH release: in vitro and in vivo studies

The effect of 5 beta- and 5 alpha-reduced progestins on luteinizing hormone-releasing hormone (LH-RH) release was examined using either an in vitro superfusion or an in vivo push-pull perfusion (PPP) technique. Ovariectomized rats (at least 6 days post operation: OVX) were implanted subcutaneously in the neck region with a silastic capsule containing 17 beta-estradiol (235 micrograms/ml: E2). Two days afterwards, these OVX + E2 rats were subjected to experimental conditions. Pregnanolone(5 beta-pregnane-3 beta-ol-20-one) at a low concentration of 0.01 ng/ml and an apparent latency of 1 h stimulated in vitro LH-RH release from superfused hypothalamic fragments containing the preoptic-anterior hypothalamic-mediobasal hypothalamic area (POA-AHA-MBH). The stimulatory effect of pregnanolone required estrogen-priming and a pulsatile mode of administration. In addition, the effect appeared quite specific since other closely related steroids such as epipregnanolone, R-5020 and 5 alpha-DHP were ineffective. In OVX + E2 rats bearing a push-pull cannula (PPC) in the hypothalamic area, in vivo infusion of repetitive pulses of pregnanolone through the PPC was also effective, confirming in vivo the stimulatory effect of pregnanolone on LH-RH release obtained from in vitro preparations. In these experiments, the same OVX + E2 rats served both as an experimental animal receiving pulses of pregnanolone and as a control subject when perfused only with medium after a week interval. The basal release rate of LH-RH and the response to pulsatile pregnanolone were variable among animals apparently due to different locations of PPC in the hypothalamus.(ABSTRACT TRUNCATED AT 250 WORDS)