Relationship between omega-3 fatty acids and plasma neuroactive steroids in alcoholism, depression and controls

Deficiency in the long-chain omega-3 fatty acid, docosahexaenoic acid (DHA) has been associated with increased corticotropin releasing hormone and may contribute to hypothalamic pituitary axis (HPA) hyperactivity. Elevated levels of the neuroactive steroids, allopregnanolone (3alpha,5alpha-THP) and 3alpha,5alpha-tetrahydrodeoxycorticosterone (THDOC) appear to counter-regulate HPA hyperactivity. Plasma essential fatty acids and neurosteroids were assessed among 18 male healthy controls and among 34 male psychiatric patients with DSM-III alcoholism, depression, or both. Among all subjects, lower plasma DHA was correlated with higher plasma THDOC (r = -0.3, P < 0.05) and dihydroprogesterone (DHP) (r = -0.52, P < 0.05). Among psychiatric patients lower DHA was correlated with higher DHP (r = -0.60, P < 0.01), and among healthy controls lower plasma DHA was correlated with higher THDOC (r = -0.83, P < 0.01) and higher isopregnanolone (3beta,5alpha-THP) (r = -0.55, P < 0.05). In this pilot observational study, lower long-chain omega-3 essential fatty acid status was associated with higher neuroactive steroid concentrations, possibly indicating increased feedback inhibition of the HPA axis.     

Neurosteroids: a new function in the brain

"Neurosteroids" accumulate in the central nervous system independently of supply by peripheral endocrine glands. Dehydroepiandrosterone (DHA) and pregnenolone (delta 5P) were first found in the rat brain. Then, a steroid biosynthetic pathway was demonstrated in oligodendrocytes, mostly by enzyme immunocytochemistry and biochemical studies in primary cultures of glial cells, where the formation, from appropriate radioactive precursors, of delta 5P, delta 5-pregn-3 beta, 20 alpha-diol (20 alpha-DH delta 5P), progesterone (P), 5 alpha-pregnane-3,20-dione (5 alpha-DHP) and 3 alpha-hydroxy-5 alpha-pregnane-20-one (3 alpha, 5 alpha-THP), as well as estrogen-induced nuclear progesterone receptor (PR) was observed. Several biological effects of neurosteroids have been observed, such as electrical stimulation of neurones, involvement in behaviorial activities, modulation of GABAA-receptor (GABAA-R) function (potentiated by 3 alpha, 5 alpha-THP and its 21-hydroxyderivative, antagonized by delta 5P- and DHA-sulfates) and growth/differentiation of glial cells in vitro. Preliminary findings suggest that the neurosteroid concept applies to all mammalian species, including man. Further investigations should assess the pathophysiological significance of the synthesis of neurosteroids and decipher their mechanisms of action via nuclear and membrane receptors.     

Quantification of neurosteroids in rat plasma and brain following swim stress and allopregnanolone administration using negative chemical ionization gas chromatography/mass spectrometry

A simplified method for the quantitative analysis of neurosteroids in rat plasma and brain is described. The method uses negative chemical ionization gas chromatography/mass spectrometry and involves the synthesis of pentafluorobenzyloxime/trimethylsilyl ether derivatives with excellent chromatographic and electron-capturing properties. Deuterium-labeled analogs of the steroids of interest were synthesized and used as internal standards. The steroids (allopregnanolone, epiallopregnanolone, pregnenolone, testosterone, and dehydroepiandrosterone) were isolated from the plasma or brain matrix by a rapid and straightforward solid-phase extraction procedure. The mass spectrometer was operated in a selective ion monitoring mode, allowing for picograms of neurosteroids to be quantified from biological extracts. The method was linear (typical R(2) = 0.999) over the concentration range (100 to 8000 pg from 0.3 ml plasma and 250 to 8000 pg from 100 mg brain tissue) with good precision and accuracy. In experimental protocols, the procedure was suitable for measuring concentrations of endogenous neurosteroids in rat plasma and brain. Significant elevations (P < 0.001) were observed in the frontal cortex for allopregnanolone and pregnenolone following a swim stress and for allopregnanolone and epiallopregnanolone following allopregnanolone injection (8 mg/kg, sc). The present method allows accurate determination of neurosteroids and will be helpful in elucidating the role of neurosteroids in health and disease.     

Pregnanolone isomers,pregnenolone and their polar conjugates around parturition

The levels of four pregnanolone isomers and their polar conjugates and pregnenolone sulfate were measured in the plasma of 13 and 7 women at delivery with subarachnoidal and epidural analgesia, respectively, and in corresponding samples of umbilical plasma using a simple quadrupole GC/MS system with electron impact ionization (pregnenolone isomers), RIA following HPLC separation (pregnenolone) and specific RIA (pregnanolone sulfate). The concentration of epipregnanolone (3beta-hydroxy-5beta-pregnan-20-one) in both maternal and umbilical plasma was much lower than that of other pregnanolone isomers. The levels of 3beta-hydroxy-pregnanolone isomers were significantly higher in the umbilical plasma than in the maternal, while the differences in 3alpha-hydroxy-isomers were insignificant. The differences in conjugates were insignificant with the exception of allopregnanolone, the levels of which were lower in umbilical plasma. In all the pregnanolone isomers, a significantly lower conjugated/unconjugated steroid ratio was found in the umbilical plasma than in the maternal plasma. In addition, time profiles of the steroids were measured around parturition and in the postpartum period in the maternal serum. Similarly, the levels of polar conjugates of all pregnanolone isomers were followed during parturition. Changes in concentrations of free steroids exhibited a similar pattern, with a fall primarily within the first hour after delivery. The decrease in conjugated steroids was shifted to the interval within the first hour and first day after delivery, and the changes were more pronounced. The time profiles of the conjugated/free steroid ratio exhibited a significant decrease within the first hour and the first day after delivery in all of the isomers investigated. A decrease was also observed in the ratio of 3alpha/3beta-isomers and 5alpha/5beta-isomers around parturition. The possible physiological consequences of the findings are indicated.     

Urinary analysis of 16(5alpha)-androsten-3alpha-ol by gas chromatography/combustion/isotope ratio mass spectrometry: implications in anti-doping analysis

We present a method for the analysis of urinary 16(5alpha)-androsten-3alpha-ol together with 5beta-pregnane-3alpha,20alpha-diol and four testosterone metabolites: androsterone (Andro), etiocholanolone (Etio), 5alpha-androstane-3alpha,17beta-diol (5alphaA), 5beta-androstane-3alpha,17beta-diol (5betaA) by means of gas chromatography/combustion/isotopic ratio mass spectrometry (GC/C/IRMS). The within-assay and between-assay precision S.D.s of the investigated steroids were lower than 0.3 and 0.6 per thousand, respectively. A comparative study on a population composed of 20 subjects has shown that the differences of the intra-individual delta(13)C-values for 16(5alpha)-androsten-3alpha-ol and 5beta-pregnane-3alpha,20alpha-diol are less than 0.9 per thousand. Thereafter, the method has been applied in the frame of an excretion study following oral ingestion of 50 mg DHEA initially and oral ingestion of 50mg pregnenolone 48 h later. Our findings show that administration of DHEA does not affect the isotopic ratio values of 16(5alpha)-androsten-3alpha-ol and 5beta-pregnane-3alpha,20alpha-diol, whereas the isotopic ratio values of 5beta-pregnane-3alpha,20alpha-diol vary by more 5 per thousand upon ingestion of pregnenolone. We have observed delta(13)C-value changes lower than 1 per thousand for 16(5alpha)-androsten-3alpha-ol, though pregnenolone is a precursor of the 16-ene steroids. In contrast to 5beta-pregnane-3alpha,20alpha-diol, the 16-ene steroid may be used as an endogenous reference compound when pregnenolone is administered.     

Plasma levels of steroid glucuronides in prepubertal, adult and elderly men

The present study has been designed to measure the levels of C-19 steroids and their glucuronide derivatives (G) in the plasma of males at three different ages: (1) 4-7 years old, (2) 20-35 years old and (3) 55-70 years old. In 4-7 years old, the plasma levels of the free and conjugated C-19 steroids were almost undetectable. From the data obtained in both groups of adult men, it appeared that except for androsterone, the levels of all free C-19 steroids were diminished by 10-70% during aging. In adult men, androsterone-G was the predominant C-19 steroid-G in plasma. The levels of androsterone-G, in the group of 20-35 years old, exceed by 3 and 20-fold the levels of androstane-3 alpha, 17 beta-diol-G and androstane-3 beta, 17 beta-diol-G, respectively. During aging, while the levels of androsterone-G and androstane-3 beta, 17 beta-diol-G were decreased or in the case of androstane-3 alpha,17 beta-diol-G remained unchanged, testosterone-G and dihydrotestosterone-G levels were markedly increased, suggesting a modification in the action of glucuronyl transferase during aging.     

5alpha-reduced C21 steroids are substrates for human cytochrome P450c17

The 5alpha-reduction of testosterone in target tissues is a key step in androgen physiology; however, 5alpha-reduced C(19) steroids are sometimes synthesized in testis via a pathway that does not involve testosterone as an intermediate. We studied the metabolism of 5alpha-reduced C(21) steroids by human cytochrome P450c17 (hCYP17), the enzyme responsible for conversion of C(21) steroids to C(19) steroids via its 17alpha-hydroxylase and 17,20-lyase activities. hCYP17 17alpha-hydroxylates 5alpha-pregnan-3,20-dione, but little androstanedione is formed by 17,20-lyase activity. hCYP17 also 17alpha-hydroxylates 5alpha-pregnan-3alpha-ol-20-one and the 5alpha-pregnan-3alpha,17alpha-diol-20-one intermediate is rapidly converted to androsterone by 17,20-lyase activity. Furthermore, 5alpha-pregnan-3alpha,17alpha-diol-20-one is a better substrate for the 17,20-lyase reaction than the preferred substrate 17alpha-hydroxypregnenolone and cytochrome b(5) stimulates androsterone formation only 3-fold. Both 5alpha-pregnan-3alpha-ol-20-one and 5alpha-pregnan-3alpha,17alpha-diol-20-one bind to hCYP17 with higher affinity than does progesterone. We conclude that 5alpha-reduced, 3alpha-hydroxy-C(21) steroids are excellent, high-affinity substrates for hCYP17. The brisk metabolism of 5alpha-pregnan-3alpha,17alpha-diol-20-one to androsterone by CYP17 explains how, when 5alpha-reductases are present, the testis can produce C(19) steroids androsterone and androstanediol from 17alpha-hydroxyprogesterone without the intermediacy of androstenedione and testosterone.     

Progesterone as a neuroactive neurosteroid, with special reference to the effect of progesterone on myelination

Some steroids are synthesized within the central and peripheral nervous system, mostly by glial cells. These are known as neurosteroids. In the brain, certain neurosteroids have been shown to act directly on membrane receptors for neurotransmitters. For example, progesterone inhibits the neuronal nicotinic acetylcholine receptor, whereas its 3alpha,5alpha-reduced metabolite 3alpha, 5alpha-tetrahydroprogesterone (allopregnanolone) activates the type A gamma-aminobutyric acid receptor complex. Besides these effects, neurosteroids also regulate important glial functions, such as the synthesis of myelin proteins. Thus, in cultures of glial cells prepared from neonatal rat brain, progesterone increases the number of oligodendrocytes expressing the myelin basic protein (MBP) and the 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase). An important role for neurosteroids in myelin repair has been demonstrated in the rodent sciatic nerve, where progesterone and its direct precursor pregnenolone are synthesized by Schwann cells. After cryolesion of the male mouse sciatic nerve, blocking the local synthesis or action of progesterone impairs remyelination of the regenerating axons, whereas administration of progesterone to the lesion site promotes the formation of new myelin sheaths.     

Selective release of luteinizing hormone by 3 alpha-hydroxy-5 alpha-pregnan-20-one in immature ovariectomized estrogen-primed rats

This study investigated the role of 3 alpha-hydroxy-5 alpha-pregnan-20-one (3 alpha,5 alpha-THP) in the modulation of gonadotropin secretion using the immature ovariectomized (OVX) rat primed with a low dose of estradiol. A treatment regimen of either 0.2 or 0.4 mg/kg of 3 alpha,5 alpha-THP given in conjunction with estradiol for 4 days significantly increased levels of serum luteinizing hormone (LH) but had no effect on serum follicle-stimulating hormone (FSH). Estrogen-primed rats receiving a single injection of 3 alpha,5 alpha-THP at 0930 h showed an increase in serum and pituitary LH levels at 1200 h and 1500 h. At 1800 h, only pituitary levels of LH remained significantly higher than controls. An injection of 3 alpha,5 alpha-THP at 1230 h in estrogen-primed rats resulted in enhanced levels of pituitary LH at 1500 h and elevated levels of both serum and pituitary LH at 1800 h. When 3 alpha,5 alpha-THP was given at 0930 h and 1230 h, elevated serum levels of LH were maintained for over 6 h. The administration of pentobarbital (Pb) 30 min after an injection of 3 alpha,5 alpha-THP at 0930 h or 1230 h prevented the increases in serum LH at 1200 h, 1500 h or 1800 h. This suggests that LH-releasing hormone (LHRH) is involved in mediating the LH response by 3 alpha,5 alpha-THP. There was no change in the sensitivity of the pituitary to LHRH following 3 alpha,5 alpha-THP treatment, indicating the absence of a pituitary effect of this steroid.     

DHEA,PREG and Their Sulphate Derivatives on Plasma and Brain After CRH and ACTH Administration

The term neurosteroids applies to steroids that are synthesized in the nervous system, either de novo from cholesterol or from steroid hormone precursors. RIA was used to determine plasma and brain levels of the neurosteroids pregnenolone (PREG), ehydroepiandrosterone (DHEA), and their sulfate derivatives (PREG-S and DHEA-S) in male and female rats after administration of two typical stress hormones: corticotropin-releasing hormone (CRH) and adrenocorticotropin hormone (ACTH). In all cases, the parameters measured were detectable in plasma and brain. PREG, PREG-S, and DHEA increased significantly in plasma and brain after CRH and ACTH administration in males and females. Because neurosteroids play an important role in mammalian physiology, including that of humans, stress situations may alter the physiological functions regulated by these neurosteroids.     

Fetal alcohol exposure alters neurosteroid levels in the developing rat brain

Neurosteroids are modulators of neuronal function that may play important roles in brain maturation. We determined whether chronic prenatal ethanol exposure altered neurosteroid levels in the developing brain. Rat dams were exposed to: (i) a 5% ethanol-containing liquid diet that produces peak maternal blood alcohol levels near the legal intoxication limit (approximately 0.08 g/dL); (ii) an isocaloric liquid diet containing maltose-dextrin instead of ethanol with pair-feeding; (iii) rat chow ad libitum. Neurosteroid levels were assessed in offspring brains using radioimmunoassay or gas chromatography-mass spectrometry techniques. A prenatal ethanol exposure-induced increase in pregnenolone sulfate levels, but not dehydroepiandrosterone sulfate levels, was evident at the earliest time point studied (embryonic day 14). This effect lasted until post-natal day 5. Levels of other neurosteroids were assessed at embryonic day 20; pregnenolone levels, but not allopregnanolone levels, were elevated. Pregnenolone sulfate levels were not altered in the maternal brain. Neither pregnenolone nor pregnenolone sulfate levels were significantly altered in the fetal liver, placenta and maternal blood, indicating that the effect of ethanol is not secondary to accumulation of peripherally-produced steroids. Fetal ethanol exposure has been shown to decrease both cellular and behavioral responsiveness to neurosteroids, and our findings provide a plausible explanation for this effect.     

Studies on neurosteroids XVII. Analysis of stress-induced changes in neurosteroid levels in rat brains using liquid chromatography-electron capture atmospheric pressure chemical ionization-mass spectrometry

The analysis of stress-induced changes in the brain neurosteroid levels by liquid chromatography (LC)-electron capture atmospheric pressure chemical ionization-mass spectrometry (ECAPCI-MS) is described. In the present method, neurosteroids were derivatized with a highly electron-affinitive reagent, 2-nitro-4-trifluoromethylphenylhydrazine (NFPH), to convert them to the corresponding hydrazones. The derivatized steroids showed over a 20-fold higher sensitivity in ECAPCI-MS than intact steroids measured by positive atmospheric pressure chemical ionization (APCI)-MS. Application of this method to the analysis of rat brain samples confirmed the significant increase in the levels of pregnenolone (PREG), progesterone (PROG), 5alpha-dihydroprogesterone (DHPROG), allopregnanolone (3alpha-hydroxy-5alpha-pregn-20-one; AP), and epiallopregnanolone (3beta-hydroxy-5alpha-pregn-20-one; EpiAP) in the fixated rats. The din stress, which we examined as a new short-term mental stress model, also elevated the brain neurosteroid levels. It is known that various types of stress lower the gamma-aminobutyric acid type A (GABA(A)) receptor function and induce the neuronal overexcitation. The increase in the brain level of AP, a potent positive modulator of GABA(A) receptors, may be the defensive response against acute stress. The increase in the brain concentration of its precursors, PREG, PROG, and DHPROG, may be associated with the acceleration of the AP synthesis. Thus, the present studies suggest that changes in the brain levels of neurosteroids may play an important role in the homeostatic mechanisms that counteract the inhibitory effect of stress on the GABA(A) receptor function.     

Validation of an analytical procedure to measure trace amounts of neurosteroids in brain tissue by gas chromatography–mass spectrometry

A selective and extremely sensitive procedure has been developed and optimized, using high-performance liquid chromatography (HPLC), specific derivatization and gas chromatography–mass spectrometry (GC–MS), to simultaneously quantify very small amounts of different neurosteroids from rat brain. Unconjugated and sulfated steroids in brain extracts were separated by solid-phase extraction. The unconjugated fraction was further purified by HPLC, the steroids being collected in a single fraction, and the sulfated fraction was solvolyzed. All steroids were derivatized with heptafluorobutyric acid anhydride and analyzed by GC–MS (electron impact ionization) using selected-ion monitoring. High sensitivity and accuracy were obtained for all steroids. The detection limits were 1 pg for pregnenolone (PREG), dehydroepiandrosterone (DHEA) and their sulfate esters PREG-S and DHEA-S, 2 pg for progesterone (PROG) and 5 pg for 3α,5α-tetrahydroprogesterone (3α,5α-THP). In a pilot study on a rat brain, the concentrations of PREG-S and DHEA-S were 8.26±0.80 and 2.47±0.27 ng/g, respectively. Those of PREG, DHEA and PROG were 4.17±0.22, 0.45±0.02 and 1.95±0.10 ng/g, respectively. Good linearity and accuracy were observed for each steroid. The methodology validated here, allows femtomoles of neurosteroids, including the sulfates, found in small brain samples (at least equal to 10 mg) to be quantified simultaneously.     

Neuroactive steroids.

Neuroactive steroids are natural or synthetic steroids that rapidly alter the excitability of neurons by binding to membrane-bound receptors such as those for inhibitory and (or) excitatory neurotransmitters. The best-studied neuroactive steroids are a series of sedative-hypnotic 3 alpha-hydroxy ring A-reduced pregnane steroids that include the major metabolites of progesterone and deoxycorticosterone, 3 alpha-hydroxy-5 alpha-pregnan-20-one (allopregnanolone) and 3 alpha,21-dihydroxy-5 alpha-pregnan-20-one (allotetrahydroDOC), respectively. These 3 alpha-hydroxysteroids do not interact with classical intracellular steroid receptors but bind stereoselectively and with high affinity to receptors for the major inhibitory neurotransmitter in brain, gamma-amino-butyric acid (GABA). Biochemical and electrophysiological studies have shown that these steroids markedly augment GABA-activated chloride ion currents in a manner similar (but not identical) to that of anesthetic barbiturates. Several steroids have also been observed to have convulsant or proconvulsant properties, including the synthetic amidine 3 alpha-hydroxy-16-imino-5 beta-17-azaandrostan-11-one (RU5135) and the natural sulfate esters of pregnenolone and dehydroepiandrosterone. Several of these have been shown to be bicuculline or picrotoxin-like GABAA receptor antagonists. Examples of steroids that alter neuronal excitability rapidly by augmenting or inhibiting excitatory amino acid receptor-mediated responses have also been reported. Recently, allopregnanolone and allotetrahydroDOC have also been measured in brain and plasma where their levels have been shown to fluctuate in response to stress and during the estrous and menstrual cycles of rats and humans, respectively. Although the major fraction of allopregnanolone in tissue, including brain, is of adrenal and/or ovarian origin, appreciable levels of allopregnanolone can still be measured in the brains of adrenalectomized and/or oophorectomized animals. Receptor-active neurosteroids may represent an important class of neuromodulators that can rapidly alter central nervous system excitability via novel nongenomic mechanisms.     

Metabolism of C19-steroids by homogenates of normal rat and mouse adrenal tissue and of the Snell transplantable rat adrenocortical tumour 494

C(19)-steroid metabolism in homogenates of adrenal tissue from rats and mice has been studied. Production of these compounds from [7alpha-(3)H]cholesterol by rat adrenal tissue appeared to follow a route independent of pregnenolone. The major products of [7alpha-(3)H]-dehydroepiandrosterone metabolism by rat adrenal tissue were 5alpha-reduced steroids, principally androsterone, epiandrosterone and 5alpha-androstanedione. No differences in metabolism of [7alpha-(3)H]dehydroepiandrosterone or [4-(14)C]pregnenolone were detected between adrenal tissue from Sprague-Dawley, Wistar and Osborne-Mendel rats, but experiments with the Snell rat adrenocortical tumour 494 showed that this tissue had low 5alpha-reductase activity. In contrast, the major products of [7alpha-(3)H]dehydroepiandrosterone metabolism by mouse adrenal tissue were 5beta-reduced steroids. Differences were observed between LACA and NH strains of mice in that there was a lower metabolism of androstenedione by NH mouse adrenal and a considerable difference in the proportions of aetiocholanolone and epiaetiocholanolone produced.     

Steroid glucuronides: Human circulatory levels and formation by LNCaP cells

We studied the relationship between circulating androsterone glucuronide, androstane-3,17β-diol glucuronide and androstane-3β,17β-diol glucuronide concentrations and adrenal as well as testicular C-19 steroids in men. Among the three 5-reduced steroid glucuronides, androsterone glucuronide is the predominant C-19 steroid measured in plasma and its levels are markedly elevated compared to those of the non-conjugated steroid. The marked rise in testosterone during puberty was strongly correlated with the increase in both androsterone glucuronide and androstane-3,17β-diol glucuronide, thus suggesting that testicular C-19 steroids are the main precursors of the steroid glucuronides. We also found that the presence of testicular androgen in plasma contributes to approx. 70% of plasma androsterone glucuronide and androstane-3,17β-diol glucuronide. Our data suggest that the adrenal C-19 steroids remaining in circulation after castration in men are converted into potent androgen which are then glucuronidated by UDP-glucuronyltransferase. We also demonstrated that the human prostate cell line LNCaP is capable of converting to a large extent androstenedione into androsterone glucuronide. Our data further confirm that glucuronidation is a major pathway of steroid metabolism in steroid target tissues.     

Chronic cigarette smoking alters circulating sex hormones and neuroactive steroids in premenopausal women

Chronic smoking alters the circulating levels of sex hormones and possibly also the neuroactive steroids. However, the data available is limited. Therefore, a broad spectrum of free and conjugated steroids and related substances was quantified by GC-MS and RIA in premenopausal smokers and in age-matched (38.9+/-7.3 years of age) non-smokers in the follicular (FP) and luteal phases (LP) of menstrual cycle (10 non-smokers and 10 smokers, in the FP, and 10 non-smokers and 8 smokers in the LP). Smokers in both phases of the menstrual cycle showed higher levels of conjugated 17-hydroxypregnenolone, 5alpha-dihydroprogesterone, conjugated isopregnanolone, conjugated 5alpha-pregnane-3beta,20alpha-diol, conjugated androstenediol, androstenedione, testosterone, free testosterone, conjugated 5alpha-androstane-3alpha/beta,17beta-diols, and higher free testosterone index. In the FP, the smokers exhibited higher levels of conjugated pregnenolone, progesterone, conjugated pregnanolone, lutropin, and a higher lutropin/follitropin ratio, but lower levels of cortisol, allopregnanolone, and pregnanolone. In the LP, the smokers exhibited higher levels of free and conjugated 20alpha-dihydropregnenolone, free and conjugated dehydroepiandrosterone, free androstenediol, 5alpha-dihydrotestosterone, free and conjugated androsterone, free and conjugated epiandrosterone, free and conjugated etiocholanolone, 7alpha/beta-hydroxy-dehydroepiandrosterone isomers, and follitropin but lower levels of estradiol and sex hormone binding globulin (SHBG) and lower values of the lutropin/follitropin ratio. In conclusion, chronic cigarette smoking augments serum androgens and their 5alpha/beta-reduced metabolites (including GABAergic substances) but suppresses the levels of estradiol in the LP and SHBG and may induce hyperandrogenism in female smokers. The female smokers had pronouncedly increased serum progestogens but paradoxically suppressed levels of their GABA-ergic metabolites. Further investigation is needed concerning these effects.     

Neurosteroid biosynthesis in the quail brain: a review

The brain traditionally has been considered to be a target site of peripheral steroid hormones. In contrast to this classical concept, new findings over the past decade have shown that the brain itself also has the capability of forming steroids de novo, the so-called "neurosteroids". De novo neurosteroidogenesis in the brain from cholesterol is a conserved property of vertebrates. Our studies using the quail, as an excellent animal model, have demonstrated that the avian brain possesses cytochrome P450 side-chain cleavage enzyme (P450scc), 3beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4)-isomerase (3beta-HSD), cytochrome P450 17alpha-hydroxylase/c17,20-lyase (P450(17alpha,lyase)), 17beta-HSD, etc., and produces pregnenolone, progesterone, 3beta, 5beta-tetrahydroprogesterone, androstenedione, testosterone and estradiol from cholesterol. However, the biosynthetic pathway of neurosteroids in the avian brain from cholesterol may be still incomplete, because we recently found that the quail brain actively produces 7alpha-hydroxypregnenolone, a previously undescribed avian neurosteroid. This paper summarize the advances made in our understanding of biosynthesis of neurosteroids in the avian brain.     

Social isolation-induced increase in the sensitivity of rats to the steroidogenic effect of ethanol

Social isolation of rats for 30 days immediately after weaning results in marked decreases in the cerebrocortical and plasma concentrations of pregnenolone, progesterone, 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha,5alpha-TH PROG), and 3alpha,5alpha-tetrahydrodeoxycorticosterone (3alpha,5alpha-TH DOC), as well as a moderate increase in the plasma concentration of corticosterone. This mildly stressful condition has now been shown to increase the sensitivity of rats to the effect of acute ethanol administration on the cerebrocortical and plasma concentrations of neuroactive steroids. The percentage increases in the brain and plasma concentrations of pregnenolone, progesterone, 3alpha,5alpha-TH PROG, and 3alpha,5alpha-TH DOC, apparent 20 min after a single intraperitoneal injection of ethanol (1 g/kg), were thus markedly greater in isolated rats than in group-housed animals. A subcutaneous injection of isoniazid (300 mg/kg) also induced greater percentage increases in the concentrations of these steroids in isolated rats than in group-housed animals. These results suggest that mild chronic stress, such as that induced by social isolation, enhances the steroidogenic effect of ethanol, a drug abused by humans under stress or affected by neuropsychiatric disorders. Social isolation also induced hyper-responsiveness of the hypothalamic-pituitary-adrenal (HPA) axis, as was apparent after reduction of GABA-mediated inhibitory tone by isoniazid administration.     

Neuroactive steroids alter the circadian system of the Syrian hamster in a phase-dependent manner

Several steroid compounds affect neuronal function, primarily by modulating the GABAA receptor complex. A circadian variation in the brain concentration of neurosteroids has been reported in rats and humans. We have previously reported that natural occurring or synthetic neuroactive steroids such as androsterone and alphaxalone also have a rhythmic effect on behavior (anesthetic and anticonvulsant activity) and GABAergic activity. In the present work, we have tested the ability of neuroactive steroids to phase shift circadian rhythms in hamsters. The GABA(A) negative modulator dehydroepiandrosterone sulphate (DHEAS) elicited phase advances when administered at CT 6, while the positive modulator androsterone lacked any effect at this time. A complete phase response curve for DHEAS revealed a nonphotic-like effect. DHEAS also blocked the circadian effects of light, while androsterone induced photic-like responses. There is also evidence that neurosteroids may be present and even synthesized in the SCN. Collectively, the results so far indicate that some neuroactive steroids might modulate the activity of the circadian clock.