Effect of hysterectomy on pain in women with endometriosis: a population-based registry study.

OBJECTIVE: To assess pain symptoms before and after hysterectomy in women with endometriosis. DESIGN: A population-based registry study. SETTING: Sweden. POPULATION: Women aged 18-45 years who underwent hysterectomy for endometriosis between 2010 and 2015. METHODS: Pain symptoms before hysterectomy and 12 months after surgery were collected from the Swedish National Quality Register for Gynaecological Surgery (GynOp). Pain symptoms were also assessed by follow-up surveys after a median follow-up period of 63 months. MAIN OUTCOME MEASURES: Pelvic or lower abdominal pain after hysterectomy. RESULTS: The study included 137 women. The proportion of women experiencing pain of any severity decreased by 28% after hysterectomy (P < 0.001). The proportion of women with severe pain symptoms decreased by 76% after hysterectomy (P < 0.001). The majority of women (84%) were satisfied with the surgical result. Presence of severe pain symptoms after the hysterectomy was associated with less satisfaction (P < 0.001). Pain symptoms after surgery, patient satisfaction and the patient's perceived improvement were not significantly different between women whose ovarian tissue was preserved and women who underwent bilateral oophorectomy. CONCLUSIONS: We observed a significant, long-lasting reduction in pain symptoms after hysterectomy among women with endometriosis. Hysterectomy, with the possibility of ovarian preservation, may be a valuable option for women with endometriosis who suffer from severe pain symptoms. TWEETABLE ABSTRACT: Hysterectomy is a valuable option for women with endometriosis and severe pain symptoms.

GABA-A receptor modulating steroids in acute and chronic stress; relevance for cognition and dementia?

Cognitive dysfunction, dementia and Alzheimer's disease (AD) are increasing as the population worldwide ages. Therapeutics for these conditions is an unmet need. This review focuses on the role of the positive GABA-A receptor modulating steroid allopregnanolone (APα), it's role in underlying mechanisms for impaired cognition and of AD, and to determine options for therapy of AD. On one hand, APα given intermittently promotes neurogenesis, decreases AD-related pathology and improves cognition. On the other, continuous exposure of APα impairs cognition and deteriorates AD pathology. The disparity between these two outcomes led our groups to analyze the mechanisms underlying the difference. We conclude that the effects of APα depend on administration pattern and that chronic slightly increased APα exposure is harmful to cognitive function and worsens AD pathology whereas single administrations with longer intervals improve cognition and decrease AD pathology. These collaborative assessments provide insights for the therapeutic development of APα and APα antagonists for AD and provide a model for cross laboratory collaborations aimed at generating translatable data for human clinical trials.

Allopregnanolone involvement in feeding regulation, overeating and obesity.

Obesity is strongly associated with ill health, primarily caused by consumption of excessive calories, and promoted (inter alia) by gamma-amino-butyric-acid (GABA) stimulating food intake by activating GABAA receptors (primarily with α3 and α2 subunits) in the hypothalamic arcuate nucleus and paraventricular nucleus. Allopregnanolone is a potent positive GABAA receptor modulating steroid (GAMS). As reviewed here, elevated allopregnanolone levels are associated with increases in food intake, preferences for energy-rich food, and obesity in humans and other mammals. In women with polycystic ovarian disease, high serum allopregnanolone concentrations are linked to uncontrolled eating, and perturbed sensitivity to allopregnanolone. Increases in weight during pregnancy also correlate with increases in allopregnanolone levels. Moreover, Prader-Willis syndrome is associated with massive overeating, absence of a GABAA receptor (with compensatory >12-, >5- and >1.5-fold increases in α4, γ2, and α1, α3 subunits), and increases in the α4, ßx, δ receptor subtype, which is highly sensitive to allopregnanolone. GABA and positive GABA-A receptor modulating steroids like allopregnanolone stimulates food intake and weight gain.

Effects of GABA active steroids in the female brain with a focus on the premenstrual dysphoric disorder.

Premenstrual dysphoric disorder (PMDD) afflicts 3%-5% of women of childbearing age, and is characterised by recurrent negative mood symptoms (eg, irritability, depression, anxiety and emotional lability) during the luteal phase of the menstrual cycle. The aetiology of PMDD is unknown, although a temporal association with circulating ovarian steroids, in particular progesterone and its metabolite allopregnanolone, has been established during the luteal phase. Allopregnanolone is a positive modulator of the GABAA receptor: it is sedative in high concentrations but may precipitate paradoxical adverse effects on mood at levels corresponding to luteal phase concentrations in susceptible women. Saccadic eye velocity (SEV) is a measure of GABAA receptor sensitivity; in experimental studies of healthy women, i.v. allopregnanolone decreases SEV. Women with PMDD display an altered sensitivity to an i.v. injection of allopregnanolone compared to healthy controls in this model. In functional magnetic resonance imaging (fMRI) studies, women with PMDD react differently to emotional stimuli in contrast to controls. A consistent finding in PMDD patients is increased amygdala reactivity during the luteal phase. Post-mortem studies in humans have revealed that allopregnanolone concentrations vary across different brain regions, although mean levels in the brain also reflect variations in peripheral serum concentrations. The amygdala processes emotions such as anxiety and aggression. This is interesting because allopregnanolone is detected at high concentrations within the region into which marked increases in blood flow are measured with fMRI following progesterone/allopregnanolone administration. Allopregnanolone effects are antagonised by its isomer isoallopregnanolone (UC1010), which significantly reduces negative mood symptoms in women with PMDD when administered s.c. in the premenstrual phase. This was shown in a randomised, placebo-controlled clinical trial in which the primary outcome was change in symptom scoring on the Daily Rating of Severity of Problems (DRSP): the treatment reduced negative mood scores (P < .005), as well as total DRSP scores (P < .01), compared to placebo in women with PMDD. In conclusion, the underlying studies of this review provide evidence that allopregnanolone is the provoking factor behind the negative mood symptoms in PMDD and that isoallopregnanolone could ameliorate the symptoms as a result of its ability to antagonise the allopregnanolone effect on the GABAA receptor.

Acute intermittent porphyria symptoms during the menstrual cycle.

BACKGROUND: Acute intermittent porphyria (AIP), a life-threatening form of the disease, is accompanied by several pain, mental and physical symptoms. AIMS: In this study, we evaluated the cyclicity of AIP and premenstrual syndrome (PMS) symptoms in 32 women with DNA-diagnosed AIP during their menstrual cycles, in northern Sweden. METHODS: The cyclicity of AIP symptoms and differences in them between the follicular and luteal phases, and the cyclicity of each symptom in each individual woman in different phases of her menstrual cycle were analysed with a prospective daily rating questionnaire. PMS symptoms were also evaluated in the patients on a daily rating scale. RESULTS: Of the 32 women, 30 showed significant cyclicity in at least one AIP or PMS symptom (P < 0.05-0.001). Back pain (10/32) was the most frequent AIP pain symptom and sweet craving (10/15) was the most frequent PMS symptom. Pelvic pain (F = 4.823, P = 0.036), irritability (F = 7.399, P = 0.011), cheerfulness (F = 5.563, P = 0.025), sexual desire (F = 8.298, P = 0.007), friendliness (F = 6.157, P = 0.019), breast tenderness (F = 21.888, P = 0.000) and abdominal swelling (F = 16.982, P = 0.000) showed significant cyclicity. Pelvic pain and abdominal swelling (rs = 0.337, P < 0.001) showed the strongest correlation. The age of women with latent AIP was strongly correlated with abdominal swelling during the luteal phase (rs = 0.493, P < 0.01). CONCLUSION: Our results suggest that the symptoms of AIP patients change during their menstrual cycles.

Behavioural responses in a net restraint test predict interrenal reactivity in Arctic charr Salvelinus alpinus.

In this study, a 1 min net restraint test was evaluated as a method to predict stress-coping style in Arctic charr Salvelinus alpinus, by investigating the relationship between behaviour during the test and levels of plasma cortisol sampled after 30 min confinement. In two separate groups of S. alpinus, general linearized model revealed significant correlations between cortisol levels and principal component scores extracted from principal component analysis, combining measures of activity in the tests. With the use of glmulti, the model selection ruled out any effects of size, sex and order of capture on interrenal reactivity. In general, S. alpinus that were more active in the net restraint test also had low levels of circulating cortisol, suggesting a proactive coping style. The results from two repeated runs were not correlated, but both runs, performed eight days apart, show a negative correlation between post-stress cortisol level and activity in the net. The lack of consistency could be explained by different treatments before each run and individual differences in behavioural plasticity. The net restraint test is thus predictive of stress-coping style in S. alpinus, and has the benefit of being less time-consuming than the commonly used confinement stress test.

Repeated allopregnanolone exposure induces weight gain in schedule fed rats on high fat diet.

Ingestion of energy rich high fat diets is one of the determining factors associated with the obesity epidemic. Therefore, much can be learned from studies of obesity-related substances given to animals fed a high fat diet. The progesterone metabolite allopregnanolone is a potent positive modulator of the gamma-aminobutyric acid (GABA)A-receptor, and both allopregnanolone and GABA have been implicated in evoking hyperphagia. In this study, food intake and body weight gain were investigated during repeated allopregnanolone exposure. Male Wistar rats were studied when fed chow ad libitum, with chow access for 4h per day or with 45% high fat pellets for 4h per day. Rats on the high fat diet were separated into obesity prone and obesity resistant individuals. Subcutaneous injections of allopregnanolone were given once daily over five consecutive days. Repeated exposure to allopregnanolone lead to increased weight gain, significantly so in schedule fed rats on a high fat diet. The increased weight gain was correlated to an increased energy intake. Both obesity resistant and obesity prone rats responded to allopregnanolone with increased weight gain. Obesity resistant rats treated with allopregnanolone increased their energy intake and ate as much as vehicle treated obesity prone rats. Their weight gain was also increased to the level of obesity prone rats injected with just the vehicle carrier oil. Thus, it appears that allopregnanolone may be one of the endogenous factors involved in weight gain, especially when the diet is rich in fat.

Behaviour, physiology and carotenoid pigmentation in Arctic charr Salvelinus alpinus.

The behaviour during an exploration task and the response to a confinement stress of Arctic charr Salvelinus alpinus were evaluated. Behaviour of individuals during 90 min of exploration was classified into high and low activity. High-activity individuals had higher plasma cortisol levels following stress compared to low-activity individuals. This indicates that high- and low-activity individuals correspond to reactive and proactive stress-coping styles. Further, a pigmentation analysis showed that high-activity individuals had a higher number of carotenoid spots cm⁻² than low-activity individuals. Thus, carotenoid pigmentation, as melanin pigmentation in other salmonids, could be linked to stress-coping style in S. alpinus.

Allopregnanolone and mood disorders.

UNLABELLED: Certain women experience negative mood symptoms during the menstrual cycle and progesterone addition in estrogen treatments. In women with PMDD increased negative mood symptoms related to allopregnanolone increase during the luteal phase of ovulatory menstrual cycles. In anovulatory cycles no symptom or sex steroid increase occurs. This is unexpected as positive modulators of the GABA-A receptor are generally increasing mood. This paradoxical effect has brought forward a hypothesis that the symptoms are provoked by allopregnanolone the GABA-A receptor system. GABA-A is the major inhibitory system in the brain. Positive modulators of the GABA-A receptor include the progesterone metabolites allopregnanolone and pregnanolone, benzodiazepines, barbiturates, and alcohol. GABA-A receptor modulators are known, in low concentrations to induce adverse, anxiogenic effects whereas in higher concentrations show beneficial, calming properties. Positive GABA-A receptor modulators induce strong paradoxical effects e.g. negative mood in 3-8% of those exposed, while up to 25% have moderate symptoms thus similar as the prevalence of PMDD, 3-8% among women in fertile ages, and up to 25% have moderate symptoms of premenstrual syndrome (PMS). The mechanism behind paradoxical reaction might be similar among them who react on positive GABA-A receptor modulators and in women with PMDD. In women the severity of these mood symptoms are related to the allopregnanolone serum concentrations in an inverted U-shaped curve. Negative mood symptoms occur when the serum concentration of allopregnanolone is similar to endogenous luteal phase levels, while low and high concentrations have less effect on mood. Low to moderate progesterone/allopregnanolone concentrations in women increases the activity in the amygdala (measured with fMRI) similar to the changes seen during anxiety reactions. Higher concentrations give decreased amygdala activity similar as seen during benzodiazepine treatment with calming anxiolytic effects. Patients with PMDD show decreased sensitivity in GABA-A receptor sensitivity to diazepam and pregnanolone while increased sensitivity to allopregnanolone. This agrees with findings in animals showing a relation between changes in alpha4 and delta subunits of the GABA-A receptor and anxiogenic effects of allopregnanolone. CONCLUSION: These findings suggest that negative mood symptoms in women with PMDD are caused by the paradoxical effect of allopregnanolone mediated via the GABA-A receptor.

Allopregnanolone induces a diurnally dependent hyperphagic effect and alters feeding latency and duration in male Wistar rats.

AIM: Gamma-aminobutyric acid (GABA)-ergic transmission from the hypothalamus is essential for normal feeding regulation, and hyperphagia can be induced by local application of GABAA -receptor agonists to different feeding-associated brain areas. The food intake in rats varies diurnally and that may influence the effect of GABAA -receptor active compounds. The progesterone metabolite allopregnanolone is a highly potent endogenous positive modulator of the GABAA receptor. Therefore, it is easy to envisage that allopregnanolone would have a hyperphagic effect, but earlier reports in rat have given ambiguous results. However, a contributing factor for the discrepancy may be the time point of the diurnal cycle in which the experiments were performed. The aim of this study was to investigate the effect of allopregnanolone on intake of standard chow in male Wistar rats at different time points of the day. METHODS: Chow intake was measured after acute administration of allopregnanolone, and feeding behaviour was analysed to detect meal patterns. RESULTS: We found that allopregnanolone increased chow intake by up to four times in the dark part of the 24-h cycle. The rats ate significantly more, and the effect of allopregnanolone was more prominent in the active (dark) compared with the inactive (light) period. Allopregnanolone also reduced feeding latency and prolonged the meal duration compared with vehicle. CONCLUSION: Allopregnanolone seems to act at several levels of feeding regulation, that is, to initiate feeding and to prolong the duration of a meal, thereby increasing the meal size, especially in the dark period of the 24-h cycle.

Frequent administration of levodopa/carbidopa microtablets vs levodopa/carbidopa/entacapone in healthy volunteers.

OBJECTIVES: An oral dispersible microtablet formulation of levodopa/carbidopa 5/1.25 mg (LC-5) was developed for individualized repeated dosing. The aim was to compare pharmacokinetic profiles of LC-5 and levodopa/carbidopa/entacapone (LCE). MATERIALS AND METHODS: A randomized, crossover study was carried out in 11 healthy subjects. Plasma concentrations of levodopa, carbidopa and 3-O-methyldopa were determined after intake of 300 mg levodopa during the day, either as three intakes of 100/25/200 mg LCE or as a morning dose of 75/18.25 mg followed by five repeated doses of 45/11.25 mg LC-5. RESULTS: Repeated dosing (2.4-hourly) with LC-5 microtablets compared to LCE (6-hourly) avoided long periods with low plasma levodopa levels. Time to maximum plasma concentrations was significantly shorter for LC-5. LC-5 showed lower fluctuation index (FI) in plasma compared to LCE (ANOVA P = 0.0028). FI for dose 2-5 was on average 1.26 for levodopa in LC-5, and 2.23 for dose 1-2 of LCE. The ratio between the two mean FI:s is 0.565; that is, LC-5 gave nearly half the FI as compared to LCE. CONCLUSIONS: Fractionation of levodopa with LC-5 into small, frequent administrations as compared to standard administrations of LCE decreased the FI in plasma for both levodopa and carbidopa by nearly half.

Gonadal hormone regulation of the emotion circuitry in humans.

Gonadal hormones are known to influence the regulation of emotional responses and affective states. Whereas fluctuations in progesterone and estradiol are associated with increased vulnerability for mood disorders, testosterone is mainly associated with social dominance, aggressive, and antisocial behavior. Here, we review recent functional neuroimaging studies that have started to elucidate how these hormones modulate the neural circuitry that is important for emotion regulation, which includes the amygdala and the medial prefrontal (mPFC) and orbitofrontal cortex (OFC). The amygdala is thought to generate emotional responses, and the prefrontal brain regions to regulate those responses. Overall, studies that have investigated women during different phases of the menstrual cycle suggest that progesterone and estradiol may have opposing actions on the amygdala and prefrontal cortex. In addition, the influence of exogenous progesterone appears to be dose-dependent. Endogenous testosterone concentrations are generally positively correlated to amygdala and OFC responses, and exogenous testosterone increases amygdala reactivity. Whereas the administration of progesterone increases amygdala reactivity and its connectivity with the mPFC, testosterone administration increases amygdala reactivity but decreases its connectivity with the OFC. We propose that this opposing influence on amygdala-prefrontal coupling may contribute to the divergent effects of progesterone and testosterone on emotion regulation and behavioral inhibition, respectively, which may promote the differential vulnerability to various psychiatric disorders between women and men. This article is part of a Special Issue entitled: Neuroactive Steroids: Focus on Human Brain.

Paradoxical effects of GABA-A modulators may explain sex steroid induced negative mood symptoms in some persons.

Some women have negative mood symptoms, caused by progestagens in hormonal contraceptives or sequential hormone therapy or by progesterone in the luteal phase of the menstrual cycle, which may be attributed to metabolites acting on the GABA-A receptor. The GABA system is the major inhibitory system in the adult CNS and most positive modulators of the GABA-A receptor (benzodiazepines, barbiturates, alcohol, GABA steroids), induce inhibitory (e.g. anesthetic, sedative, anticonvulsant, anxiolytic) effects. However, some individuals have adverse effects (seizures, increased pain, anxiety, irritability, aggression) upon exposure. Positive GABA-A receptor modulators induce strong paradoxical effects including negative mood in 3%-8% of those exposed, while up to 25% have moderate symptoms. The effect is biphasic: low concentrations induce an adverse anxiogenic effect while higher concentrations decrease this effect and show inhibitory, calming properties. The prevalence of premenstrual dysphoric disorder (PMDD) is also 3%-8% among women in fertile ages, and up to 25% have more moderate symptoms of premenstrual syndrome (PMS). Patients with PMDD have severe luteal phase-related symptoms and show changes in GABA-A receptor sensitivity and GABA concentrations. Findings suggest that negative mood symptoms in women with PMDD are caused by the paradoxical effect of allopregnanolone mediated via the GABA-A receptor, which may be explained by one or more of three hypotheses regarding the paradoxical effect of GABA steroids on behavior: (1) under certain conditions, such as puberty, the relative fraction of certain GABA-A receptor subtypes may be altered, and at those subtypes the GABA steroids may act as negative modulators in contrast to their usual role as positive modulators; (2) in certain brain areas of vulnerable women the transmembrane Cl(-) gradient may be altered by factors such as estrogens that favor excitability; (3) inhibition of inhibitory neurons may promote disinhibition, and hence excitability. This article is part of a Special Issue entitled: Neuroactive Steroids: Focus on Human Brain.

Cytochrome P450 induction by rifampicin in healthy subjects: determination using the Karolinska cocktail and the endogenous CYP3A4 marker 4beta-hydroxycholesterol.

The Karolinska cocktail, comprising caffeine, losartan, omeprazole, and quinine, was given before and after administration of rifampicin (20, 100, or 500 mg daily) to measure induction of cytochrome P450 (P450) enzymes. Rifampicin was given for 14 days to eight healthy subjects (all of whom possessed at least one wild-type CYP2C9 and one wild-type CYP2C19 gene) in each dose group. 4beta-hydroxycholesterol was assessed as an endogenous marker of CYP3A4 induction. A fourfold induction of CYP3A4 was seen at the highest dose by both quinine:3'-hydroxyquinine and 4beta-hydroxycholesterol measurements (P < 0.001). CYP3A4 was also induced at the two lower doses of rifampicin when measured by these two markers (P < 0.01 or P < 0.001). CYP1A2, CYP2C9, and CYP2C19 were induced after 500 mg rifampicin daily (1.2-fold, P < 0.05; 1.4-fold, P < 0.05; and 4.2-fold, P < 0.01, respectively). In conclusion, we have shown that the Karolinska cocktail and 4beta-hydroxycholesterol can be used for an initial screening of the induction properties of a drug candidate.

Progesterone selectively increases amygdala reactivity in women.

The acute neural effects of progesterone are mediated by its neuroactive metabolites allopregnanolone and pregnanolone. These neurosteroids potentiate the inhibitory actions of gamma-aminobutyric acid (GABA). Progesterone is known to produce anxiolytic effects in animals, but recent animal studies suggest that pregnanolone increases anxiety after a period of low allopregnanolone concentration. This effect is potentially mediated by the amygdala and related to the negative mood symptoms in humans that are observed during increased allopregnanolone levels. Therefore, we investigated with functional magnetic resonance imaging (MRI) whether a single progesterone administration to healthy young women in their follicular phase modulates the amygdala response to salient, biologically relevant stimuli. The progesterone administration increased the plasma concentrations of progesterone and allopregnanolone to levels that are reached during the luteal phase and early pregnancy. The imaging results show that progesterone selectively increased amygdala reactivity. Furthermore, functional connectivity analyses indicate that progesterone modulated functional coupling of the amygdala with distant brain regions. These results reveal a neural mechanism by which progesterone may mediate adverse effects on anxiety and mood.

Effects of allopregnanolone on sedation in men, and in women on oral contraceptives.

Allopregnanolone is a known GABA(A) receptor agonist not previously given to men, or to women using oral contraceptives (OC). The effects of metabolites of sex hormones on the GABA(A) receptor are different between men and women. OC are known to change GABA(A) receptor subunit composition. These factors might play a role in the differential effect of allopregnanolone in men and women, and in women with or without OC. To study the sedative effect of and sensitivity to allopregnanolone in men and in women with OC, nine healthy men (mean age 24.6 years) and nine healthy women on OC (mean age 21.8 years) were given three, increasing, intravenous dosages (0.015, 0.03, and 0.045 mg/kg) of allopregnanolone. Saccadic eye velocity (SEV), subjective ratings, and electroencephalography (EEG) were used to evaluate the response to allopregnanolone. Repeated blood samples for analyses of serum allopregnanolone levels were drawn throughout the study day. Allopregnanolone decreased SEV more in women than in men, and increased subjective ratings of 'sedation'. The results in women on OC are similar to earlier results in women without OC. Subjective ratings of 'contentedness' decreased in men but increased in women. Serum levels of allopregnanolone were more highly increased in men compared to women. Other pharmacokinetic parameters were not different between sexes. On the EEG, beta power increased in men. In conclusion, men and women on OC reacted differently to allopregnanolone.

Oral progesterone decreases saccadic eye velocity and increases sedation in women.

The aim of this study was to investigate the neurophysiological and behavioural effects of a single dose of progesterone in women. Allopregnanolone is a metabolite of progesterone and a potent positive modulator of the GABA(A) receptor and produces sedative and anxiolytic effects. This study was designed to examine the effect of oral progesterone and the metabolite allopregnanolone in women. Women (n=15) in their follicular phase received oral progesterone (400mg) or placebo. Dependent measures included plasma levels of progesterone and allopregnanolone, saccadic eye velocity (SEV), subjective ratings (visual analogue scales), and reaction time. Administration of progesterone decreased SEV and increased sedation. This effect is probably due to enhanced GABA activity.

Pregnenolone sulphate and Zn2+ inhibit recombinant rat GABA(A) receptor through different channel property.

AIMS: We compared the antagonistic effects of state-dependent gamma-aminobutyric acid A (GABA(A)) receptor blockers picrotoxin, Zn(2+) and pregnenolone sulphate (PS) on GABA- and pentobarbital-activated currents in recombinant rat GABA(A) receptors in Xenopus oocytes. METHODS: Experiments were performed with wild type rat alpha1 beta2 gamma2L and alpha1beta2 receptors, mutants alpha1V256S beta2 gamma2L and alpha1beta2A252Sgamma2L receptors by the two-electrode voltage-clamp technique. RESULTS: In contrast to respective 3840- and 56-fold increases in Zn(2+) potencies to inhibit GABA- and pentobarbital-activated currents in the alpha1beta2 receptor, the corresponding potencies of PS remained unchanged in comparison with the alpha1 beta2 gamma2L receptor. A homologous mutation of the residue at 2' position closest to the cytoplasmic end of the M(2) helix to serine on both alpha1 and beta2 subunit, alpha1V256S and beta2A252S, abolished the inhibition of GABA(A) receptor by PS. In comparison with the wild type alpha1beta2gamma2L receptor, mutants alpha1V256S beta2 gamma2L and alpha1beta2 A252S gamma2L receptors did not affect the Zn(2+) inhibition. Furthermore, a significant increase in GABA potency was observed in the mutant alpha1V256S beta2 gamma2L receptor (P < 0.05), but not the mutant alpha1beta2 A252S gamma2L receptor compared with the wild type receptor. CONCLUSIONS: Pregnenolone sulphate was a gamma2-subunit independent inhibitor in the GABA(A) receptor, whereas the Zn(2+) antagonism was profoundly influenced by the gamma2-subunit. It is likely that the 2' residue closest to the N-terminus of the protein at M(2) helix on both alpha1 and beta2 subunit are critical to the inhibitory actions of PS and the function of Cl(-) channels. These results are consistent with the hypothesis that PS behaves as a Cl(-) channel blocker that does not share with Zn(2+), the coincident channel property in the GABA(A) receptors.

Neurosteroid modulation of allopregnanolone and GABA effect on the GABA-A receptor.

The neurosteroid allopregnanolone (ALLO) or 3alpha-OH-5alpha-pregnane-20-one interacts with the GABA type A receptor chloride ion channel complex and enhances the effect of GABA. Animal and human studies suggest that ALLO plays an important role in several disorders including premenstrual syndrome, anxiety, and memory impairment. In contrast to ALLO, steroids with a hydroxy group in the 3beta position usually exert a reducing effect and have recently attracted interest due to their suggested role in counteracting the negative action of ALLO. In this study, five different 3beta-steroids were tested for their ability to modulate GABA-mediated chloride ion uptake in the absence and presence of ALLO in rat brain microsacs preparations. In addition, the effects of the 3beta-steroids and their interaction with ALLO were investigated by patch-clamp recordings of spontaneous inhibitory postsynaptic currents (sIPSCs) in rat hypothalamic neurons from the medial preoptic nucleus (MPN). All tested 3beta-steroids reduced the ALLO-enhanced GABA response in cerebral cortex, in hippocampus and in MPN. In cerebellum, only one had this effect. However, in the absence of ALLO, two of the 3beta-steroids potentiated GABA-evoked chloride ion uptake and prolonged the sIPSCs decay time, whereas the others had little or no effect. Therefore, it is possible that at least some 3beta-steroids can act as positive GABA(A) receptor modulators as well as negative modulators depending on whether or not ALLO is present. Finally, these results suggest that the 3beta-steroids could be of interest as pharmacological agents that could counteract the negative effects of ALLO.

GABA-site antagonism and pentobarbital actions do not depend on the alpha-subunit type in the recombinant rat GABA receptor.

AIM: The roles of alpha-subunits on the gamma-aminobutyric acid (GABA)-site antagonism and pentobarbital actions were examined in rat recombinant GABA(A) receptors in Xenopus oocytes. METHODS: Experiments were performed with binary and ternary GABA(A) receptors containing alpha1-, alpha4- or alpha5-subunit by the two-electrode voltage-clamp technique. RESULTS: The potency of GABA was significantly higher in the alpha1beta2, alpha4beta2 and alpha5beta2 receptors compared with the alpha1beta2gamma2L, alpha4beta2gamma2L and alpha5beta2gamma2L receptors. However, the alpha5beta2 receptor possessed significantly lower GABA efficacy compared with the alpha5beta2gamma2L receptor. While the gamma2-subunit was essential to the potency of GABA, its influence on the apparent GABA-site antagonism was less profound. The antagonist affinity constants (K(B)) of bicuculline inhibition and slopes of Schild plots were similar between all types of ternary and binary receptors except alpha5beta2 receptor which was not tested. The pK(B)s and IC(50)s of the GABA-site antagonism were not significantly different between the alpha1beta2gamma2L, alpha4beta2gamma2L and alpha5beta2gamma2L receptors. Bicuculline blocked pentobarbital-activated currents in a reversible and non-competitive manner with the alpha1beta2gamma2L, alpha4beta2gamma2L, and alpha5beta2gamma2L receptors, indicating an allosteric inhibition of the GABA-site. No significant difference of bicuculline potencies in inhibiting GABA- and pentobarbital-activated currents was found between the alpha1beta2gamma2L, alpha4beta2gamma2L and alpha5beta2gamma2L receptors. CONCLUSION: The GABA-site antagonism does not depend on the subtype of alpha-subunits. Similarly, pentobarbital activates ternary receptors composed of different alpha-subunits in a bicuculline-sensitive manner. The potencies of bicuculline to inhibit pentobarbital-activated currents are identical with receptors containing alpha1, alpha4 or alpha5-subunit. The alpha1beta2 and alpha4beta2 receptors possess higher GABA potencies compared with the alpha1beta2gamma2L and alpha4beta2gamma2L receptors.