Effects Of Allopregnanolone Research Articles

Neurosteroids are endogenous neuroprotectants in an ex vivo glaucoma model.

Allopregnanolone is a neurosteroid and powerful modulator of neuronal excitability. The neuroprotective effects of allopregnanolone involve potentiation of γ-aminobutyric acid (GABA) inhibitory responses. Although glutamate excitotoxicity contributes to ganglion cell death in glaucoma, the role of GABA in glaucoma remains uncertain. The aim of this study was to determine whether allopregnanolone synthesis is induced by high pressure in the retina and whether allopregnanolone modulates pressure-mediated toxicity. Ex vivo rat retinas were exposed to hydrostatic pressure (10, 35, and 75 mm Hg) for 24 hours. Endogenous allopregnanolone production was determined by liquid chromatography and tandem mass spectrometry (LC-MS/MS) and immunochemistry. We also examined the effects of allopregnanolone, finasteride, and dutasteride (inhibitors of 5α-reductase), picrotoxin (a GABA(A) receptor antagonist), and D-2-amino-5-phosphonovalerate (APV, a broad-spectrum N-methyl-D-aspartate receptor [NMDAR] antagonist). Pressure loading at 75 mm Hg significantly increased allopregnanolone levels as measured by LC-MS/MS. Elevated hydrostatic pressure also increased neurosteroid immunofluorescence, especially in the ganglion cell layer and inner nuclear layers. Staining was negligible at lower pressures. Enhanced allopregnanolone levels and immunostaining were substantially blocked by finasteride, but more effectively inhibited by dutasteride and APV. Administration of exogenous allopregnanolone suppressed pressure-induced axonal swelling in a concentration-dependent manner, while picrotoxin overcame these neuroprotective effects. These results indicate that the synthesis of allopregnanolone is enhanced mainly via NMDARs in the pressure-loaded retina, and that allopregnanolone diminishes pressure-mediated retinal degeneration via GABAA receptors. Allopregnanolone and other related neurosteroids may serve as potential novel therapeutic targets for the prevention of pressure-induced retinal damage in glaucoma.

Allopregnanolone and its precursor progesterone do not reduce injury after experimental stroke in hypertensive rats - role of postoperative temperature regulation?

Allopregnanolone is a neurosteroid synthesized from progesterone in brain. It increases inhibition through modulation of the gamma-aminobutyric acid type A (GABA-A) receptor. Both agents are putative neuroprotectants after ischemic stroke. We sought to confirm their effectiveness in a hypertensive rat stroke model, with intra- and post-operative temperature regulation. The primary study compared allopregnanolone, progesterone or vehicle control treatments, administered 105 minutes after induction of temporary middle cerebral artery occlusion in spontaneously hypertensive rats. Temperature was controlled intraoperatively and a heat mat used in the 6 hours postoperatively to permit animal temperature self-regulation. The primary outcome was infarct volume and secondary outcomes were tests of sensory and motor function. There was no significant effect of treatment on any outcome measure. Given prior reports of GABA-A receptor agonists causing hypothermia, follow-up experiments were conducted to examine postoperative temperature regulation. These did not reveal a difference in postoperative temperature in neurosteroid-treated animals compared to control. However, in all rats maintained postoperatively in ambient temperature, moderate hypothermia was observed. This was in contrast to rats maintained over a heat mat. The lowest mean postoperative temperature was between 34.4–34.9°C in all 3 groups. These data do not support a neuroprotective effect of allopregnanolone or progesterone in ischemic stroke in hypertensives in the setting of normothermia. Given previous evidence of synergy between neuroprotective agents and hypothermia, demonstration of neuroprotective effect of these agents in the absence of postoperative hypothermia would be prudent before consideration of these agents for further clinical investigation.

Neurosteroids allopregnanolone sulfate and pregnanolone sulfate have diverse effect on the α subunit of the neuronal voltage-gated sodium channels Nav1.2, Nav1.6, Nav1.7, and Nav1.8 expressed in xenopus oocytes.

The neurosteroids allopregnanolone and pregnanolone are potent positive modulators of γ-aminobutyric acid type A receptors. Antinociceptive effects of allopregnanolone have attracted much attention because recent reports have indicated the potential of allopregnanolone as a therapeutic agent for refractory pain. However, the analgesic mechanisms of allopregnanolone are still unclear. Voltage-gated sodium channels (Nav) are thought to play important roles in inflammatory and neuropathic pain, but there have been few investigations on the effects of allopregnanolone on sodium channels. Using voltage-clamp techniques, the effects of allopregnanolone sulfate (APAS) and pregnanolone sulfate (PAS) on sodium current were examined in Xenopus oocytes expressing Nav1.2, Nav1.6, Nav1.7, and Nav1.8 α subunits. APAS suppressed sodium currents of Nav1.2, Nav1.6, and Nav1.7 at a holding potential causing half-maximal current in a concentration-dependent manner, whereas it markedly enhanced sodium current of Nav1.8 at a holding potential causing maximal current. Half-maximal inhibitory concentration values for Nav1.2, Nav1.6, and Nav1.7 were 12 ± 4 (n = 6), 41 ± 2 (n = 7), and 131 ± 15 (n = 5) μmol/l (mean ± SEM), respectively. The effects of PAS were lower than those of APAS. From gating analysis, two compounds increased inactivation of all α subunits, while they showed different actions on activation of each α subunit. Moreover, two compounds showed a use-dependent block on Nav1.2, Nav1.6, and Nav1.7. APAS and PAS have diverse effects on sodium currents in oocytes expressing four α subunits. APAS inhibited the sodium currents of Nav1.2 most strongly.

Effects of the neuroactive steroid allopregnanolone on intracranial self-stimulation in C57BL/6J mice.

The neuroactive steroid (3α,5α)-3-hydroxy-pregnan-20-one (3α,5α-THP, allopregnanolone) has effects on reward-related behaviors in mice and rats that suggest that it may activate brain reward circuits. Intracranial self-stimulation (ICSS) is an operant behavioral technique that detects changes in the sensitivity of brain reward circuitry following drug administration. To examine the effects of the neuroactive steroid allopregnanolone on ICSS and to compare these effects to those of cocaine. Male C57BL/6J mice implanted with stimulating electrodes implanted into the medial forebrain bundle responded for reinforcement by electrical stimulation (brain stimulation reward (BSR)). Mice received cocaine (n = 11, 3.0-30.0 mg/kg, intraperitoneal (i.p.)) or the neuroactive steroid allopregnanolone (n = 11, 3.0-17.0 mg/kg, i.p.). BSR thresholds (θ 0) and maximum (MAX) operant response rates after drug treatments were compared to those after vehicle injections. Cocaine and allopregnanolone dose dependently lowered BSR thresholds relative to vehicle injections. Cocaine was maximally effective (80 % reduction) in the second 15 min following the 30 mg/kg dose, while allopregnanolone was maximally effective (30 % reduction) 15-45 min after the 17 mg/kg dose. Neither drug had significant effects on MAX response rates. The effects of allopregnanolone on BSR thresholds are consistent with the previously reported effects of benzodiazepines and alcohol, suggesting that positive modulation of GABAA receptors can facilitate reward-related behaviors in C57BL/6J mice.

Allopregnanolone suppresses diabetes-induced neuropathic pain and motor deficit through inhibition of GABAA receptor down-regulation in the spinal cord of diabetic rats.

Painful diabetic neuropathy is associated with hyperexcitability and hyperactivity of spinal cord neurons. However, its underlying pathophysiological mechanisms have not been fully clarified. Induction of excitatory/inhibitory neurotransmission imbalance at the spinal cord seems to account for the abnormal neuronal activity in diabetes. Protective properties of neurosteroids have been demonstrated in numerous cellular and animal models of neurodegeneration. Here, the protective effects of allopregnanolone, a neurosteroid were investigated in an in vivo model of diabetic neuropathy. The tail-flick test was used to assess the nociceptive threshold. Diabetes was induced by injection of 50 mg/kg (IP) streptozotocin. Seven weeks after the induction of diabetes, the dorsal half of the lumbar spinal cord was assayed for the expression of γ2 subunit of GABAA receptor using semiquantitative RT-PCR. The data shows that allopregnanolone (5 and 20 mg/kg) markedly ameliorated diabetes-induced thermal hyperalgesia and motor deficit. The weights of diabetic rats that received 5 and 20 mg/kg allopregnanolone did not significantly reduce during the time course of study. Furthermore, this neurosteroid could inhibit GABAA receptor down-regulation induced by diabetes in the rat spinal cord. The data revealed that allopregnanolone has preventive effects against hyperglycemic-induced neuropathic pain and motor deficit which are related to the inhibition of GABAA receptor down-regulation.

Selective effect of allopregnanolone on corticoliberin-induced anxiety.

We studied the effect of allopregnanolone on stress-induced anxiety caused by intranasal administration of corticoliberin to male Wistar rats. Active and passive rats were selected by their T-maze behavior and then were tested in elevated plus maze for initial anxiety level. Basing on test results, the animals were divided into following groups: active high-anxiety rats, active low-anxiety rats, and passive animals. Rats of the experimental subgroups received subcutaneous injection of allopregnanolone (0.2 mg/kg body weight), control animals received saline. In 30 min, all animals intranasally received corticoliberin (0.5 μg into each nostril) and then were tested in elevated plus maze. Passive animals demonstrated increased anxiety after corticoliberin administration against the background of allopregnanolone. Under the same conditions, low-anxiety rats demonstrated reduced anxiety, while in active high-anxiety animals, only an increase of motor activity was observed. Hence, the effect of allopregnanolone on anxiety level under stress conditions depends on individual typologic features of animal behavior.

Effects of allopregnanolone, a positive allosteric modulation of GABAA receptors upon stress and hypertension in Schlager genetic hypertensive mice (697.3)

Background: Hypertensive Schlager mice (BPH/2J) have neurogenic hypertension associated with differences in GABAA receptors compared to their normotensive counterparts (BPN/3J). Allopregnanolone is an endogenous neurosteroid reduced by chronic stress and when administered, decreases anxiety by positive allosteric modulation of GABAA receptors.Aim: To determine if allopregnanolone reduces the pressor effects of stress and basal MAP in BPH/2J mice.Methods: Male BPN/3J (n=7) and BPH/2J (n=5) mice received vehicle or allopregnanolone (5µg/hour) via subcutaneous minipumps for 2 weeks. Implanted telemetric probes enabled recording of mean arterial pressure (MAP), heart rate (HR) and activity as well as the response to aversive and non‐aversive stress tests and ganglionic blockade with pentolinium before and 7 and 14 days of treatment. Mice were perfused following stress and brains were removed for immunohistochemistry.Results: Allopregnanolone reduced systolic arterial pressure (‐8.8mmHg, P=0.01) and attenuated the depressor response to pentolinium in BPH/2J mice but had no effect upon in BPN/3J. Allopregnanolone produced marked reductions in the pressor response to both cage switch and feeding stress (‐20%, P<0.01) in BPH/2J whilst increasing the pressor response to aversive stress in BPN/3J (P<0.001). Stress induced Fos counts within the medial amygdala and paraventricular nucleus were higher in untreated BPH/2J compared to BPN/3J. Allopregnanolone reduced Fos expression and treatment abolished the difference between strains.Conclusions: The selective antihypertensive and stress inhibitory effects of allopregnanolone in BPH/2J hypertensive mice suggests that allosteric modulation of GABAA receptors at the level of the hypothalamus and amygdala may be a major cause of hypertension in this model and may offer a possible new area for the development of therapy.

Allopregnanolone's attenuation of the lordosis-inhibiting effects of restraint is blocked by the antiprogestin, CDB-4124

A brief restraint experience reduces lordosis behavior in ovariectomized females that have been hormonally primed with estradiol benzoate. The addition of progesterone to the priming prevents the lordosis inhibition. Based on prior studies with an inhibitor of progesterone metabolism, we have implicated the intracellular progesterone receptor, rather than progesterone metabolites, as responsible for this protection. However, the progesterone metabolite, allopregnanolone (3α-hydroxy-5α-pregnan-20-one), also prevents lordosis inhibition after restraint. In a prior study, we reported that the progestin receptor antagonist, RU486 (11β-(4-dimethylamino)phenyl-17β-hydroxy-17-(1-propynyl)estra-4,9-dien-3-one), attenuated the effect of allopregnanolone. Because RU486 can also block the glucocorticoid receptor, in the current studies, we evaluated the effect of the progestin receptor antagonist, CDB-4124 (17α-acetoxy-21-methoxy-11β-[4-N,N-dimethyaminopheny]-19-norpregna-4,9-dione-3,20-dione), which is relatively devoid of antiglucocorticoid activity. Ovariectomized, Fischer rats were injected with 10μg estradiol benzoate. Two days later, rats received either 60mg/kg CDB-4124 or 20% DMSO/propylene glycol vehicle 1h before injection with 4mg/kg allopregnanolone. After a pretest to confirm sexual receptivity, rats were restrained for 5min and immediately tested for sexual behavior. Lordosis behavior was reduced by the restraint and attenuated by allopregnanolone. Pretreatment with CDB-4124 reduced allopregnanolone's effect. These findings support prior suggestions that allopreganolone reduces the response to restraint by mechanisms that require activation of the intracellular progesterone receptor.

Neurosteroid allopregnanolone attenuates high glucose-induced apoptosis and prevents experimental diabetic neuropathic pain: In vitro and in vivo studies

Hyperglycemia plays a critical role in the development of diabetic neuropathy. Hyperglycemia induces oxidative stress in neurons, resulting in neuronal cell apoptosis and dysfunction. Anti-apoptotic properties of neurosteroids have been demonstrated in numerous cellular models of neurodegenerative studies. Here, the protective effects of neurosteroid allopregnanolone were investigated in in vitro and in vivo models of diabetic neuropathy.The data show that glucose decreased the viability of PC12 cells in a concentration-dependent manner. Allopregnanolone at concentrations of 2.5, 5 and 10μM markedly prevented high glucose-induced toxicity in naïve and NGF-treated (neuron-like) PC12 cells. Furthermore, treatment of diabetic rats with allopregnanolone (5 and 20mg/kg) significantly ameliorated diabetic-induced thermal hyperalgesia. Moreover, this neurosteroid inhibited caspase 3 and decreased Bax/Bcl2 ratio in high glucose-treated cells and spinal cord of diabetic rats.In conclusion, the data revealed that allopregnanolone has protective effects against hyperglycemic-induced cellular damage and prevention of cell apoptosis is involved in its mechanisms. Our findings suggest that allopregnanolone has protective effect against pro-apoptotic challenges such as diabetes and hyperglycemia and propose therapeutic potential of neurosteroids in attenuation of diabetic side effects such as neuropathy.

Involvement of cocaine- and amphetamine-regulated transcript peptide in the hyperphagic and body weight promoting effects of allopregnanolone in rats

Allopregnanolone (ALLO), a gamma-aminobutyric acid (GABA) type A receptor active neurosteroid, elicits hyperphagic response in rodents. Since GABA-A receptors are present on the peptidergic neurons in the hypothalamus, we were interested in finding out if ALLO and neuropeptide cocaine- and amphetamine-regulated transcript (CART) interact and influence feeding behavior. While subcutaneous ALLO treatment, for a period of 7 days, produced a significant increase in food intake and body weight, pretreatment with subthreshold dose of CART (intracerebroventricular) attenuated both the effects. On the other hand, subcutaneous administration of dehydroepiandrosterone sulfate (DHEAS; GABA-A inhibitor neurosteroid) for a period of 7 days resulted in a significant reduction in food intake and body weight. These effects of DHEAS were potentiated by intracerebroventricular pretreatment with subeffective dose of CART. The brains of ALLO-treated rats were processed for the immunohistochemical analysis of CART immunoreactive elements. ALLO treatment resulted in a significant reduction in CART immunoreactivity in the hypothalamic arcuate, paraventricular and lateral nuclei, and nucleus accumbens shell. The results of the present study suggest that ALLO and CART might interact in the brain, and influence food intake and body weight. However, further investigations are needed to clarify the precise mechanisms by which ALLO modulate feeding behavior.

Intrathecal neurosteroids and a neurosteroid antagonist: Effects on inflammation-evoked thermal hyperalgesia and tactile allodynia

Neurosteroids regulate neuronal excitability though binding sites associated with the ionotropic γ-aminobutyric acid (GABAA) receptor. We sought to characterize the spinal analgesic actions in rats of two 5α-reduced neurosteroids, allopregnanolone and alphaxalone, on nociceptive processing and to determine whether a putative neurosteroid antagonist attenuates this effect: (3α,5α)-17-phenylandrost-16-en-3-ol (17PA). Intrathecal (IT) injection of allopregnanolone (1–30μg/10μL in 20% cyclodextrin) delivered through lumbar catheters produced a dose-dependent analgesia in rats as measured by thermal thresholds in the ipsilateral (inflamed by intraplantar carrageenan) and in the contralateral (un-inflamed paws). Similar observations were made with alphaxalone (30–60μg in 20% cyclodextrin). Effective doses were not associated with suppressive effects on pinnae, blink or placing and stepping reflex. Effects of allopregnanolone (30μg) on the normal and hyperalgesic paw were completely prevented by IT 17PA (30μg). Reversal by IT 17PA of an equi-analgesic dose of alphaxalone occurred only at higher antagonist dosing. These results suggest that a spinal neurosteroid-binding site with which 17PA interacts may regulate spinal nociceptive processing in normal and inflamed tissue.

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

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. Chow intake was measured after acute administration of allopregnanolone, and feeding behaviour was analysed to detect meal patterns. 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. 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.

Neuroprotection by steroids after neurotrauma in organotypic spinal cord cultures: A key role for progesterone receptors and steroidal modulators of GABAA receptors

Progesterone is neuroprotective after spinal cord injury, however its mechanism of action remains unexplored. Here we used organotypic spinal cord slice cultures from 3 weeks-old mice to evaluate the mechanisms of neuroprotection by progesterone and its 5α-reduced metabolites. In vitro spinal cord injury, using a weight drop model, induced a decrease in the number of motoneurons. This was correlated with an increase in the number of dying cells (PI+ cells) and in LDH release. Addition of 10 μM of progesterone, 5α-dihydroprogesterone (5α-DHP) or allopregnanolone (3α, 5α-tetrahydroprogesterone) to the medium at the time of injury rescued the spinal cord slices from the effects of damage. Progesterone prevented membrane cell damage, motoneuron loss and cell death. These effects were not due to its bioconversion to 5α-DHP nor to allopregnanolone, as supported by the finasteride, an inhibitor of 5α-reductase enzymes, and by the absence of 5α-reduced progesterone metabolites in the slices analyzed by gas chromatography–mass spectrometry. The neuroprotective effects of progesterone required PR as they could not be observed in slices from homozygous knockout PR−/− mice. Allopregnanolone treatment was also neuroprotective. Its effects were not due to its bioconversion back to 5α-DHP, which can activate gene transcription via PR, because they were still observed in slices from knockout PR−/− mice. Allopregnanolone effects involved GABAA receptors, as they were inhibited by the selective GABAA receptor antagonist Gabazine, in both PR+/+ and PR−/− mice. Altogether, these findings identify both PR and GABAA receptors as important targets for neuroprotection by progestagens after spinal cord injury.

Allopregnanolone Reinstates Tyrosine Hydroxylase Immunoreactive Neurons and Motor Performance in an MPTP-Lesioned Mouse Model of Parkinson's Disease

Restorative/protective therapies to restore dopamine neurons in the substantia nigra pars compacta (SNpc) are greatly needed to effectively change the debilitating course of Parkinson's disease. In this study, we tested the therapeutic potential of a neurogenic neurosteroid, allopregnanolone, in the restoration of the components of the nigrostriatal pathway in MPTP-lesioned mice by measuring striatal dopamine levels, total and tyrosine hydroxylase immunoreactive neuron numbers and BrdU-positive cells in the SNpc. An acute treatment (once/week for two weeks) with allopregnanolone restored the number of tyrosine hydroxylase-positive and total cell numbers in the SNpc of MPTP-lesioned mice, even though this did not increase striatal dopamine. It was also noted that MPTP treated mice to which allopregnanolone was administered had an increase in BrdU-positive cells in the SNpc. The effects of allopregnanolone in MPTP-lesioned mice were more apparent in mice that underwent behavioral tests. Interestingly, mice treated with allopregnanolone after MPTP lesion were able to perform at levels similar to that of non-lesioned control mice in a rotarod test. These data demonstrate that allopregnanolone promotes the restoration of tyrosine hydroxylase immunoreactive neurons and total cells in the nigrostriatal tract, improves the motor performance in MPTP-treated mice, and may serve as a therapeutic strategy for Parkinson's disease.

Increased neurosteroid sensitivity – An explanation to symptoms associated with chronic work related stress in women?

Work related psychosocial stress can be accompanied by so called burnout syndrome with symptoms of mental exhaustion, physical fatigue, and cognitive dysfunction. Underlying mechanisms for acquiring burnout syndrome are not clear. Animal studies show that chronic stress is associated with altered release of GABA-A receptor modulating steroids (GAMS), altered composition of the GABA-A receptor and altered sensitivity to GAMS. In the present study we investigated if such changes occur in women with burnout syndrome. We further asked whether flumazenil (a benzodiazepine antagonist, but with positive modulating effects on GABA-A receptors with altered subunit composition) can block the effect of the GAMS allopregnanolone. Ten women with occupational psychosocial stress and burnout syndrome were compared with twelve healthy controls in an experimental setting. Saccadic eye velocity (SEV) was measured after an injection of allopregnanolone, followed by an injection of flumazenil and a second injection of allopregnanolone. The sensitivity to allopregnanolone was significantly higher in the patients compared to controls after the first injection (p=0.04) and the difference increased when the response per allopregnanolone concentration unit was compared (p=0.006). Following the flumazenil injection the burnout patients (p=0.016), but not controls, showed a decrease in SEV and flumazenil acted like a positive modulator that is agonistic. There was no significant difference between the groups after second allopregnanolone injection. In conclusion, patients with work related psychosocial stress and burnout syndrome show a different response to GABA-A receptor modulators than controls suggesting a changed GABA-A receptor function in these patients. More precisely we hypothesize that the α4 and delta subunits are up-regulated elevating the responsiveness to allopregnanolone and change the effect of flumazenil, which provides a potential explanation to the burnout syndrome. Flumazenil does not block the effect of allopregnanolone.

RU486 blocks effects of allopregnanolone on the response to restraint stress

These experiments were designed to provide information about the potential involvement of progesterone receptors in the ability of allopregnanolone (3α-hydroxy-5α-pregnan-20-one) to reduce the lordosis-inhibiting effects of restraint stress. Ovariectomized Fischer rats were hormonally primed with 10μg estradiol benzoate and 4mg/kg allopregnanolone or vehicle. One hour before allopregnanolone, rats were injected with the progesterone receptor antagonist, RU486 (11β-(4-dimethylamino)phenyl-17β-hydroxy-17-(1-propynyl)estra-4,9-dien-3-one), or vehicle. Four hours after allopregnanolone or vehicle, sexual behavior was examined before and after a 5-min restraint stress. Lordosis behavior of rats primed only with estradiol benzoate declined after the 5min of restraint while allopregnanolone prevented this decline. RU486 attenuated the ability of allopregnanolone to prevent the restraint-induced decline in lordosis behavior. These findings are consistent with earlier suggestions that progesterone receptors are involved in allopregnanolone's ability to reduce the effects of restraint stress.

The effect of intra-nucleus accumbens administration of allopregnanolone on δ and γ2 GABAA receptor subunit mRNA expression in the hippocampus and on depressive-like and grooming behaviors in rats

Alterations in GABAA receptor expression have been associated with the allopregnanolone (3α-hydroxy-5α-pregnan-20-one; 3α,5α-THP) antidepressant-like effect in rats. The present study aimed to verify the effect of bilateral, intra-nucleus accumbens core (intra-AcbC) administration of the neurosteroid allopregnanolone on behaviors in the forced swim and grooming microstructure tests and in the δ and γ2 GABAA receptor subunit mRNA expression in right and left hippocampus of rats. The results of this study showed that bilateral, intra-AcbC allopregnanolone administration (5μg/rat) presented antidepressant‐like activity in the forced swim test concomitant with an increase in climbing. Allopregnanolone at doses of 1.25 and 5μg/rat also decreased the percentage of correct transitions in the grooming microstructure test. Both δ and γ2 GABAA subunit expressions increased in the rat hippocampus after allopregnanolone intra-AcbC treatment. Our findings point to asymmetrical GABAA receptor expression changes in the hippocampus of animals treated with allopregnanolone. Further investigation should evaluate the antidepressant-like effect of allopregnanolone not only in other directly infused regions but also with respect to changes in other brain areas of the limbic system to understand allopregnanolone's mechanism of action.

Parallel Session Abstracts

Allopregnanolone effects in women : clinical studies in relation to the menstrual cycle, premenstrual dysphoric disorder and oral contraceptive use

Allopregnanolone Prevents Dieldrin-Induced NMDA Receptor Internalization and Neurotoxicity by Preserving GABAA Receptor Function

Dieldrin is an endocrine disruptor that accumulates in mammalian adipose tissue and brain. It induces convulsions due to its antagonism of the γ-aminobutyric acid A receptor (GABA(A)R). We have previously reported that long-term exposure to dieldrin causes the internalization of the N-methyl-D-aspartate receptor (NMDAR) as a result of persistent GABA(A)R inhibition. Because the neurosteroids 17β-estradiol (E2) and allopregnanolone are known to modulate the function and trafficking of GABA(A)R and NMDAR, we examined the effects of E2 and allopregnanolone on dieldrin-induced GABA(A)R inhibition, NMDAR internalization, and neuronal death in cortical neurons. We found that 1 nM E2 increased the membrane expression of NR1/NR2B receptors and postsynaptic density 95 but did not induce their physical association. In contrast, 10 nM E2 had no effect on these proteins but reduced NR2A membrane expression. We also found that exposure to 60 nM dieldrin for 6 d in vitro caused the internalization of NR1 and NR2B but not NR2A. Treatment with either 1 nM E2 or 10 μM allopregnanolone prevented the dieldrin-induced reduction in membrane levels of the NR1/NR2B receptors. Furthermore, prolonged exposure to 200 nM dieldrin down-regulated the expression of NR2A; this was inhibited only by allopregnanolone. Although both hormones restored NMDAR function, as measured by the NMDA-induced rise in intracellular calcium, allopregnanolone (but not E2) reversed the inhibition of GABA(A)R and neuronal death caused by prolonged exposure to dieldrin. Our results indicate that allopregnanolone protects cortical neurons against the neurotoxicity caused by long-term exposure to dieldrin by maintaining GABA(A)R and NMDAR functionality.

Effect of maternal administration of allopregnanolone before birth asphyxia on neonatal hippocampal function in the spiny mouse

Clinically, treatment options where fetal distress is anticipated or identified are limited. Allopregnanolone is an endogenous steroid, that positively modulates the GABA A receptor, and that has anti-apoptotic and anti-excitotoxic actions, reducing brain damage in adult animal models of brain injury. We sought to determine if prophylactic treatment of the pregnant female with a single dose of this steroid could reduce birth asphyxia-induced losses in hippocampal function at 5 days of age (P5) in spiny mouse neonates ( Acomys cahirinus). At 37 days gestation (term = 39 days) and 1 h before inducing birth asphyxia, spiny mice dams were injected subcutaneously (0.2 ml) with either 3 mg/kg allopregnanolone or 20% w/v β-cyclodextrin vehicle. One hour later, fetuses were either delivered immediately by caesarean section (control group) or exposed to 7.5 min of in utero asphyxia, causing acidosis and hypoxia. At P5, ex vivo hippocampal plasticity was assessed, or brains collected to determine cell proliferation (proliferating cell nuclear antigen; PCNA) or calcium channel expression (inositol trisphosphate receptor type 1; IP 3R1) using immunohistochemistry. Allopregnanolone partially prevented the decrease in long term potentiation at P5, and the asphyxia-induced increase in IP 3R1 expression in CA1 pyramidal neurons. There was no effect of allopregnanolone on the asphyxia induced impairment of the input/output (I/O) curve and paired-pulse facilitation (PPF). In control birth pups, maternal allopregnanolone treatment caused significant changes in short term post-synaptic plasticity and also reduced hippocampal proliferation at P5. These findings show that allopregnanolone can modulate hippocampal development and synaptic function in a normoxic or hypoxic environment, possibly by modifying calcium metabolism. Best practice for treatment dose and timing of treatment will need to be carefully considered.