Neurosteroid modulation of embryonic neuronal survival in vitro following anoxia

Neurosteroids are synthesized de novo in the brain from cholesterol or peripheral steroid precursors and modulate inhibitory γ-aminobutyric acid (GABA_A) and excitatory N-methyl-d-aspartate (NMDA) receptors. Evidence indicates that neurosteroids are neuroprotective and important during neurodevelopment. We tested the hypothesis that neurosteroids increase embryonic neuronal survival following anoxia in rat embryonic day 18 cerebral cortical cultures to examine potential neurosteroid modulation of this insult during early development. Twenty-four hours after plating in serum-free medium, cultures were exposed to DHEA, DHEAS, or allopregnanolone (10⁻¹⁰, 10⁻⁸, or 10⁻⁶ M), or vehicle, for 24 h (n=9 per treatment condition). Cultures were then subjected to anoxia for 2 h and subsequently reincubated for 24 h prior to neuron immunostaining with microtubule-associated protein 2 (MAP-2) antibody. Supernatant from DHEA and DHEAS-exposed cultures was tested for 17β-estradiol metabolite formation by radioimmunoassay. DHEA 10⁻⁶ and 10⁻⁸ M significantly increased neuron survival by 85–87% following anoxia. DHEAS 10⁻⁶ M significantly increased neuron survival by 74% following anoxia, but DHEAS 10⁻¹⁰ M decreased neuron survival after this insult. Allopregnanolone had modest effects on neuron survival that did not attain statistical significance. 17β-Estradiol concentrations were below the limit of detection in all specimens tested (sensitivity 4.7 nM). Our data indicate that pretreatment with DHEA and DHEAS at physiologically relevant concentrations promotes neuronal survival following anoxia in embryonic rat cerebral cortical cultures, and that these effects are not secondary to 17β-estradiol metabolite formation. DHEA and DHEAS modulation of anoxia in embryonic neurons may be relevant to disorders of neurodevelopment involving this insult.     

Noradrenergic innervation of human pineal gland: abnormalities in aging and Alzheimer''s disease

Using previously characterized polyclonal antibody directed against dopamine beta-hydroxylase (DBH), immunoreactive fibers were demonstrable in pineals of 6 controls of various ages and of 3 individuals with Alzheimer's disease (AD). Abnormal, swollen axons were present in pineals from aged individuals and from individuals with AD. The pathology of noradrenergic axons in a structure innervated by the superior cervical ganglion suggests that peripheral noradrenergic systems may be affected in aging and in AD.     

Interactions of methylestrogens with cytoplasmic and nuclear estrogen receptors in rat pituitary gland, hypothalamus and uterus

The in vitro affinities to cytoplasmic estrogen receptors of the methylestrogens 2-methylestradiol-17 beta, 4-methylestradiol-17 beta and 4-hydroxy-2-methylestradiol-17 beta, which are useful probes to test the biological importance of 2- or 4-hydroxylation of estradiol-17 beta (catecholestrogen formation), have been determined in hypothalamic, pituitary and uterine tissue of the ovariectomized rat. Moreover, the in vivo capacity of these compounds to translocate estrogen receptors into the cell nucleus of pituitary and uterine tissue has been studied. Methylestrogens exhibited estrogen receptor affinities which were not significantly different from the binding affinity of estradiol-17 beta. When given at a high dose (100 micrograms/animal) their nuclear translocation capacity was equal (4-methylestradiol-17 beta) or even higher (2-methylestradiol-17 beta) than that of estradiol-17 beta. However, at a low dose (5 micrograms/animal) 4-methylestradiol was completely ineffective in both the pituitary gland and the uterus, and 2-methylestradiol-17 beta was less potent than estradiol-17 beta in the pituitary gland.     

Differentiation-dependent progesterone synthesis and metabolism in NT2-N human neurons

Human embryonic teratocarcinoma-derived Ntera2/cl.D1 (NT2) cells recapitulate many features of embryonic neuronal progenitor cells. Upon retinoic acid (RA) treatment they terminally differentiate into post-mitotic neuron-like cells (NT2-N), akin to human fetal neurons, thus representing an in vitro model of human neuron terminal differentiation. Experimental evidence also indicate NT2-N cultures as a potential source for cell transplantation therapy. The neurosteroids progesterone and its metabolite 3α-hydroxy-5α-pregnan-20-one (3α,5α-THP) promote neurogenesis and show anti-neurodegenerative properties. This study's aim was to assess the neurosteroidogenic competence of NT2 cells during RA-induced neuronal differentiation. Radioimmunoassay measurements revealed progesterone only in NT2-N cultures (4 week RA). Accordingly, progesterone synthesis from ³H-pregnenolone was absent in NT2 cells and increased during RA exposure, being highest in NT2-N. [³H]-pregnenolone metabolism, yielding [³H]-progesterone and [³H]-5α-dihydroprogesterone ([³H]-5α-DHP), was time-dependent and inhibited by trilostane, a 3β-hydroxysteroid-dehydrogenase (3β-HSD) inhibitor. Conversely, ³H-progesterone metabolism, which yielded [³H]-5α-DHP > [³H]-3β,5α-THP > [³H]-3α,5α-THP, occurred at all time points examined, though showing a nadir in cultures treated with RA for 1 and 2 weeks. The differentiation-dependent increase of progesterone accumulation matched 3β-HSD type I mRNA expression and 3β-HSD immunoreactivity, that co-localized with Map2a/b- and GAD67 in NT2-N. Hence, in vitro differentiated human neurons, while retaining progesterone metabolic activity, also become competent in progesterone synthesis. These findings suggest an autocrine/paracrine role of neuronal progesterone, either on its own or through its 5α-reduced metabolites, in fetal brain development and allow speculation that NT2-N-produced neurosteroids may contribute to the encouraging results of NT2-N transplants in animal models of neurodegenerative diseases.     

Localization of neurons containing pro-opiomelanocortin-related peptides in the hypothalamus and midbrain of the lizard, Anolis carolinensis evidence for region-specific processing of β-endorphin

Immunohistochemical analyses of the lizard brain, following colchicine pretreatment, revealed two populations of POMC-producing cell bodies located in medial-basal hypothalamus and the mesencephalic tegmentum. Analyses of extracts of lizard brain regions by radioimmunoassay and gel filtration chromatography indicate that β-endorphin-sized and α-MSH-sized peptides are the major POMC-related end products. Evidence is presented for region-specific processing of β-endorphin in the lizard brain.     

Competition by monophenolic estrogens and catecholestrogens for high-affinity uptake of [H](−)-norepinephrine into synaptosomes from rat cerebral cortex and hypothalamus

High affinity uptake of [³H](−)-norepinephrine (NE) was investigated in synaptosomes from rat cerebral cortex (K_m=360±30nM) and hypothalamus (K_m=307±90nM). Estrogens but not androgens, glucocorticoids or progestin interfered competitively with NE uptake. Ethinylestradiol was the most effective competitor tested, its K_i value being 200 nM in the cortex and 144 nM in the hypothalamus. Stereospecificity of the inhibitory effect of estradiol-17β with a preference for the 17β-hydroxy group was indicated by the ineffectiveness of estradiol-17α and estrone as competitors. A-ring substitution of estradiol-17β or ethinylestradiol by hydroxyl groups in positions 2 and 4 (yielding catecholestrogens) or methyl substitution in positions 2 and 4 (yielding methylestrogens) significantly reduced the inhibitory potency of the estrogen. Methoxylation in positions 2,4 or 11β completely abolished the competitive action of estradiol-17β or ethinylestradiol on NE uptake.     

GABAA Receptor subunits have differential distributions in the rat retinae: In situ hybridization and immunohistochemistry

The distributions of nine different subunits of the gamma-aminobutyric acid_A (GABA_A) receptor (α₁, α₂, α₃, ₅; β₁, β₂, β₃; γ₂; δ) were investigated in the rat retina using immunocytochemistry and in situ hybridization. With the exception of the α₅ subunit, all subunits could be localized. Each subunit was expressed in characteristic strata within the inner plexiform layer (IPL). Some subunits (e.g., γ₂) showed a ubiquitous distribution, while others (e.g., δ) were restricted to narrow sublayers. Double labeling experiments using different combinations of the subunit-specific antibodies revealed colocalizations of subunits within individual neurons. Additionally, GABA_A receptor subunits were mapped to distinct populations of retinal neurons by coapplication of defined immunocytochemical markers and subunit-specific antibodies. Cholinergic amacrine cells were found to express the α₂, β1, β_2/3 and δ subunits, while dopaminergic amacrine cells express the α₂, α₃ and γ₂ subunits. Dissociated rod bipolar cells express the γ₂ subunits. In summary, this study provides evidence for the existence of multiple GABA_A receptor subtypes in the retina. The distinct stratification pattern of the subunits in the IPL suggests that different functional circuits involve specific subtypes of GABAA receptors. © 1995 Wiley-Liss, Inc.     

Distribution of glycinergic terminals on lumbar motoneurons of the adult cat: an ultrastructural study

The distribution of glycine-like immunoreactivity on cat lumbar motoneurons was examined in electron microscopy, using pre-embedding immunocytochemistry. In the dorsolateral portion of the ventral horn, numerous labeled axon terminals were presynaptic to somatic and dendritic profiles of alpha-motoneurons. Most of the glycinergic boutons contained pleomorphic vesicles and showed symmetrical contacts. On the somatic and proximal dendritic compartments, glycinergic terminals accounted for, respectively, 24.6 and 26.6% of the total number of terminals. There were very few glycinergic terminals on gamma-motoneurons. Immunoreactive axons, dendrites and cell bodies were also observed near the motoneurons. These results support the view that glycine plays a major role in the inhibition of alpha-motoneurons and suggest that inhibitory mechanisms occur on the soma as well as on dendrites.     

Pro-opiocortin fragments in human and rat brain: β-endorphin and α-MSH are the predominant peptides

The ‘pro-opiocortin’ fragments, β-lipotropin, β-endorphin, ACTH and α-MSH, were estimated in discrete areas of rat and human brain and pituitaries by means of radioimmunoassay in combination with gelfiltration. These peptides exihibited parallel patterns of distribution, but with β-endorphin and α-MSH predominant in the brain of rat and man, and, in contrast, their respective precursors, β-LPH and ACTH predominant in the adenohypophysis of rat and man. These data may be indicative of important differences in post-translational processing of ‘pro-opiocortin’ between these contrasting tissues.     

Metabolites of the β-amyloid precursor protein generated by β-secretase localise to the trans-golgi network and late endosome in 293 cells

Deposition of β-amyloid occurs in the brains of all sufferers of Alzheimer's disease. β-amyloid is proteolytically derived from the β-amyloid precursor protein by as yet unidentified enzymes termed secretases. We have generated and characterised antisera to the carboxy-terminal domain and β-secretase cleavage site of the Alzheimer's amyloid precursor protein. The β-secretase cleavage event occurs at the extreme N-terminus of the β-amyloid peptide. Our antiserum to the N-terminus of the β-amyloid peptide (NTβ4) specifically recognises β-secretase cleaved species as opposed to intact βAPP. NTβ4 specifically immunoprecipitates a 13 kDa fragment of βAPP (p13) which is potentially amyloidogenic. We have used these anti-sera in confocal laser scanning immunofluorescence microscopy to localise the intracellular location of potentially amyloidogenic βAPP processing fragments such as p13. Using a number of marker antisera of known intracellular location, we have defined the major location of βAPP fragments possessing the Asp-1 N-terminus of β-amyloid as the trans-Golgi network or late endosome on the basis of colocalisation with a monoclonal antibody to the cation-independent mannose-6-phosphate receptor. The colocalisation was further investigated using brefeldin A which demonstrated that the p13 fragment and mannose-6-phosphate receptor are trafficked by alternative pathways from the trans-Golgi network. © 1996 Wiley-Liss, Inc.     

Age- and region-specific expressions of the messenger RNAs encoding for steroidogenic enzymes P450scc, P450c17 and 3β-HSD in the postnatal rat brain

Neurosteroids are now known to be synthesized de novo in the nervous system through mechanisms at least partly independent of peripheral steroidogenic glands. In mammals, the presence of the cholesterol side-chain cleavage enzyme (cytochrome P450scc) and the enzyme 3β-hydroxysteroid dehydrogenase/Δ⁵-Δ⁴-isomerase (3β-HSD) has been well established in the brain, whereas limited information has been available on the enzyme 17α-hydroxylase/c17, 20-lyase (cytochrome P450c17), which converts pregnenolone to dehydroepiandrosterone, one of the most abundant neurosteroids. In addition, little is known regarding developmental changes in these steroidogenic enzymes during postnatal life. Thus, the pathway of neurosteroid formation in the brain is still incomplete. Therefore, we examined expressions of the messenger RNAs (mRNAs) encoding for three key enzymes, P450scc, P450c17 and 3β-HSD, in the rat brain at different postnatal ages using RT-PCR analysis. The expression of P450scc mRNA was found throughout the brain at the same level, while the 3β-HSD mRNA expression was higher in the cerebellum and cerebrum than in other brain regions. The P450c17 mRNA was highly expressed in the mesencephalon. On the other hand, higher expressions of the cerebellar and cerebral 3β-HSD mRNAs were observed only in neonatal life. In contrast, the expression of P450scc mRNA was relatively constant during neonatal life and in adulthood. A similar constant expression of the P450c17 mRNA was evident in the mesencephalon. Serial Southern hybridization in this study confirmed the specific mRNA expression corresponding to each enzyme. These results suggest that in the postnatal rat the expression of 3β-HSD or P450c17 mRNA may be age- or region-dependent, unlike the P450scc mRNA expression.     

Regional distribution of β-lipotropin converting enzymes in rat pituitary and brain

Among the brain areas studied only pars distalis and pars intermedia are found to contain β-lipotropin activating enzyme indicating that these may be the exclusive organs for a physiologically significant conversion of β-lipotropin into β-endorphin. β-Endorphin inactivating enzyme is found to be rather uniformly distributed in all the pituitary and brain regions, α- and γ-endorphins are presumably formed by the action of this enzyme on β-endorphin.     

Effects of sex steroids on the growth of neuronal processes in neonatal rat hypothalamus-preoptic area and cerebral cortex in primary culture

Dissociated cells of neonatal rat hypothalamus-preoptic area and cerebral cortex were grown in primary culture, and the effects of sex steroids on neurons were morphometrically studied. In cultures of the hypothalamus-preoptic area estradiol-17β or testosterone propionate significantly increased the total neuronal process length as compared with control cultures given no steroids in the medium. Significant stimulatory effects were detected on 1, 2 and 3 days after plating, while, 5a-dihydrotestosterone failed to show any stimulatory influence. In contrast, none of these steroids was effective in cultures of cerebral cortical cells. Based on the number of processes arising from the soma, neurons were divided into unipolar, bipolar and multipolar neurons. Percentage of multipolar neurons of the hypothalamus-preoptic area was more in cultures treated with estradiol-17β or testosterone propionate than the controls. Significant effect was not noted in cultures with 5α-dihydrotestosterone, while, in cultures of the cerebral cortical cells, effects on sprouting of these steroids were far less marked than in cultures of the hypothalamus-preoptic area. The present findings substantiate the importance of aromatization of androgen to exert its influence on neuronal process growth in cell culture.     

Cognitive impairment and increased brain neurosteroids in adult rats perinatally exposed to low millimolar blood alcohol concentrations

Epidemiological evidence suggests that adolescents and adults perinatally exposed to alcohol, even at low doses, show high prevalence of cognitive impairment and social behavior deficits, which may be in part related to alcohol-induced changes of the gamma-aminobutyric acid (GABA)ergic neurotransmission. The endogenous neurosteroid 3alpha-hydroxy,5alpha-pregnan-20-one (3alpha,5alpha-tetrahydroprogesterone/3alpha,5alpha-THP), a potent positive allosteric modulator of GABA(A) receptor function, is implicated in the physiological tuning of GABA-mediated fast inhibition and in various alcohol's actions in the brain. This study was undertaken to determine whether perinatal exposure to low millimolar blood alcohol concentrations alters cognitive skills (social discrimination and inhibitory avoidance tests), emotional reactivity (elevated plus maze test), and neurosteroid content in brain cortex and hippocampus of adult male offspring. Dams had access to a 3% alcohol solution or to an equicaloric sucrose solution from gestational day 15 to postnatal day 9. Eighty-day old alcohol-exposed male offspring exhibited impaired social recognition memory, but unchanged inhibitory avoidance performance and normal behavior on the elevated-plus maze. The concentrations of 3alpha,5alpha-THP and its precursor progesterone were more than doubled in brain cortex and hippocampus of alcohol-exposed rats, whereas in plasma only progesterone was increased. Thus, exposure to low millimolar blood alcohol concentrations has a long-lasting impact on the developing brain as it causes an impairment of social recognition as well as an increase of brain neurosteroid content in mature animals. The latter may be consequent to altered expression/activity of brain steroidogenic enzymes, as reflected by the enduring increase of the GABA(A) receptor-active neurosteroid 3alpha,5alpha-THP in brain cortex and hippocampus, but not in plasma. It is speculated that, by inducing a greater amplification of GABA(A) receptor function, the elevation of 3alpha,5alpha-THP brain content contributes to the cognitive impairment exhibited by adult alcohol-exposed offspring.     

Cooperative regulation of nerve growth factor synthesis and secretion in fibroblasts and astrocytes by fibroblast growth factor and other cytokines

Acidic fibroblast growth factor (aFGF) enhances nerve growth factor (NGF) synthesis by astrocytes obtained from various brain regions. NGF secretion by fibrous-shaped astrocytes transformed by dibutyryl-cAMP (db-cAMP) pretreatment was less than that by untreated astrocytes. However, aFGF also enhanced NGF secretion by fibrous-shaped astrocytes. The effects of various kinds of intracellular signaling modulators on NGF synthesis were examined. None of the following second messenger effectors had an effect on NGF synthesis: protein kinase C (PKC) agonist (phorbol myristate acetate (PMA)) or antagonist (sphingosine (SP)). LiCl, and ionomycin (Iono). Further, increases of intracellular cAMP by forskolin (FK) or db-cAMP have no significant effect on NGF synthesis in astrocytes under a standard culture condition. However, NGF synthesis by astrocytes in the presence of aFGF was significantly enhanced by db-cAMP, but not by FK or sodium butyrate. These results indicate that an excessive amount of cAMP enhances the effect of aFGF on NGF synthesis in astrocytes. NGF synthesis in astrocytes was not affected by treatment with anti-aFGF or anti-bFGF neutralizing antibodies, indicating that FGFs are not involved in the autocrine regulation of NGF synthesis in astrocytes. Transforming growth factor-beta 1 (TGF-beta 1), which inhibits some effects of FGFs, increased NGF synthesis in concert with aFGF. Furthermore, the highest NGF synthesis was observed when astrocytes were stimulated by all of the following cytokines: aFGF, interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha) and TGF-beta 1. The mechanism regulating NGF synthesis in fibroblasts obtained from prenatal rat skin was also investigated. Acidic FGF, basic FGF (bFGF), epidermal growth factor (EGF), platelet-derived growth factor (PDGF), transforming growth factor-alpha (TGF-alpha), TGF-beta 1, IL-1 beta, and TNF-alpha were found to be regulators of NGF synthesis in skin fibroblasts. Among these cytokines, aFGF is the most potent regulator of NGF synthesis in fibroblasts. NGF synthesis by skin fibroblasts, either in the presence or absence of aFGF, was not modified by any of the following: FK, PMA, SP, LiCl, and Iono. However, db-cAMP significantly enhanced NGF synthesis in both conditions. Sodium butyrate enhanced NGF synthesis in the presence of aFGF, but not in the absence of aFGF. These results suggest that an excessive amount of cAMP and butyrate moiety regulate NGF synthesis in skin fibroblasts in different ways.(ABSTRACT TRUNCATED AT 400 WORDS)     

Alzheimer's β-amyloid peptides induce inflammatory cascade in human vascular cells: the roles of cytokines and CD40

Accumulating evidence suggests that β-amyloid (Aβ)-induced inflammatory reactions may partially drive the pathogenesis of Alzheimer's disease (AD). Recent data also implicate similar inflammatory processes in cerebral amyloid angiopathy (CAA). To evaluate the roles of Aβ in the inflammatory processes in vascular tissues, we have tested the ability of Aβ to trigger inflammatory responses in cultured human vascular cells. We found that stimulation with Aβ dose-dependently increased the expression of CD40, and secretion of interferon-γ (IFN-γ) and interleukin-1β (IL-1β) in endothelial cells. Aβ also induced expression of IFN-γ receptor (IFN-γR) both in endothelial and smooth muscle cells. Characterization of the Aβ-induced inflammatory responses in the vascular cells showed that the ligation of CD40 further increased cytokine production and/or the expression of IFN-γR. Moreover, IL-1β and IFN-γ synergistically increased the Aβ-induced expression of CD40 and IFN-γR. We have recently found that Aβ induces expression of adhesion molecules, and that cytokine production and interaction of CD40–CD40 ligand (CD40L) further increase the Aβ-induced expression of adhesion molecules in these same cells. These results suggest that Aβ can function as an inflammatory stimulator to activate vascular cells and induces an auto-amplified inflammatory molecular cascade, through interactions among adhesion molecules, CD40–CD40L and cytokines. Additionally, Aβ_1–42, the more pathologic form of Aβ, induces much stronger effects in endothelial cells than in smooth muscle cells, while the reverse is true for Aβ_1–40. Collectively, these findings support the hypothesis that the Aβ-induced inflammatory responses in vascular cells may play a significant role in the pathogenesis of CAA and AD.     

Neurosteroidogenesis Today: Novel Targets for Neuroactive Steroid Synthesis and Action and Their Relevance for Translational Research

Neuroactive steroids are endogenous neuromodulators synthesised in the brain that rapidly alter neuronal excitability by binding to membrane receptors, in addition to the regulation of gene expression via intracellular steroid receptors. Neuroactive steroids induce potent anxiolytic, antidepressant, anticonvulsant, sedative, analgesic and amnesic effects, mainly through interaction with the GABA_A receptor. They also exert neuroprotective, neurotrophic and antiapoptotic effects in several animal models of neurodegenerative diseases. Neuroactive steroids regulate many physiological functions, such as the stress response, puberty, the ovarian cycle, pregnancy and reward. Their levels are altered in several neuropsychiatric and neurological diseases and both preclinical and clinical studies emphasise a therapeutic potential of neuroactive steroids for these diseases, whereby symptomatology ameliorates upon restoration of neuroactive steroid concentrations. However, direct administration of neuroactive steroids has several challenges, including pharmacokinetics, low bioavailability, addiction potential, safety and tolerability, which limit its therapeutic use. Therefore, modulation of neurosteroidogenesis to restore the altered endogenous neuroactive steroid tone may represent a better therapeutic approach. This review summarises recent approaches that target the neuroactive steroid biosynthetic pathway at different levels aiming to promote neurosteroidogenesis. These include modulation of neurosteroidogenesis through ligands of the translocator protein 18 kDa and the pregnane xenobiotic receptor, as well as targeting of specific neurosteroidogenic enzymes such as 17β‐hydroxysteroid dehydrogenase type 10 or P450 side chain cleavage. Enhanced neurosteroidogenesis through these targets may be beneficial not only for neurodegenerative diseases, such as Alzheimer's disease and age‐related dementia, but also for neuropsychiatric diseases, including alcohol use disorders.     

Pharmacological evidence that α-ketoisovaleric acid induces convulsions through GABAergic and glutamatergic mechanisms in rats

Neurological dysfunction is common in patients with maple syrup urine disease (MSUD). However, the mechanisms underlying the pathophysiology of this disorder are poorly known. In the present study we investigated the effect of intrastriatal administration of the α-keto acids accumulating in MSUD on the behavior of adult rats. After cannula placing, rats received unilateral intrastriatal injections of α-ketoisocaproic acid (KIC, 8 μmol), α-ketoisovaleric acid (KIV, 8 μmol), α-keto-β-methylvaleric acid (KMV, 6 μmol) or NaCl. KIV elicited clonic convulsions in a dose–response manner, whereas KIC and KMV did not induce seizure-like behavior. Convulsions provoked by KIV were prevented by intrastriatal preadministration of muscimol (46 pmol) and MK-801 (3 nmol), but not by the preadministration of DNQX (8 nmol). These results indicate that among the keto acids that accumulate in MSUD, KIV is the only metabolite capable of causing convulsions in the present animal model and indicates that KIV is an important excitatory metabolite. Moreover, the participation of GABAergic and glutamatergic NMDA mechanisms in the KIV-induced convulsant behavior is suggested, since KIV-induced convulsions are attenuated by muscimol and MK-801. The authors suggest that KIV may play an important role in the convulsions observed in MSUD, and highlight its relevance to the understanding of the pathophysiology of the neurological dysfunction of MSUD patients.     

Ovulation delay induced by blockade of the cholinergic system on dioestrus-1, is related to changes in dopaminergic activity of the preoptic anterior-hypothalamic area of the rat

One hour after the injection of 100 mg/kg of atropine-sulphate at 1300 h of dioestrus-1, there was an abrupt increase of 17beta-oestradiol plasma level and a significant increase in dopaminergic neural activity in preoptic anterior-hypothalamic area, without changes in luteinizing hormone serum level, in comparison with the saline injected group. Animals injected with atropine-sulphate showed a second increase in dopaminergic neural activity in the preoptic anterior-hypothalamic at 1100 of dioestrus-2 (atropine-sulphate 0.471 +/- 0.7 vs. saline 0.241 +/- 0.03, p < 0.01). In this group of animals, the preovulatory surges of 17beta-oestradiol and luteinizing hormone occurred simultaneously at 1700 h of the expected day of oestrus; spontaneous ovulation was delayed until the expected day of dioestrus-1. Present results suggest that during dioestrus-1 there is a functional relationship between the cholinergic and dopaminergic systems in preoptic anterior-hypothalamic area, regulating the release of luteinizing hormone resulting in ovulation.