Administration Of Allopregnanolone Research Articles

Synergistic Modulation of γ-Aminobutyric Acid Type A Receptor-Mediated Synaptic Inhibition in Cortical Networks by Allopregnanolone and Propofol

The neuroactive steroid allopregnanolone (ALLO) is an endogenous allosteric modulator of γ-aminobutyric acid type A (GABAA) receptors. There is evidence that ALLO, at physiologically relevant concentrations, modulates GABAA receptor function in the cerebral cortex. The widely used anesthetic agent propofol and ALLO share a similar mode of molecular action. Here, we ask how GABAA receptor-mediated synaptic inhibition and action potential firing of neurons in cultured cortical slices are altered by either ALLO or propofol or by coapplying both agents. We explored the effects of ALLO and propofol on spontaneous action potential activity of neocortical neurons in organotypic slices cultured from C57BL6 mice by performing extracellular multiunit recordings. Furthermore, we carried out whole-cell voltage-clamp experiments to quantify the drug effects on GABAA receptor-mediated tonic and phasic currents. We found that ALLO (100 nM) decreased multiunit action potential firing of neocortical neurons by approximately 21%. Moreover, the duration of GABAA receptor-mediated inhibitory postsynaptic currents (IPSCs) was prolonged (mean Δdecay time prolongation: 12.9 ± 2.2 milliseconds; n = 23), and a bicuculline-sensitive tonic current was induced (mean Δbaseline shift: -24.6 ± 13.6 pA; P = .002; n = 6). A subanesthetic concentration of propofol (250 nM) decreased the discharge rates of cortical neurons to a similar degree as ALLO (100 nM). ALLO and propofol administered in combination acted in an additive manner to reduce action potential firing. However, during ALLO administration, propofol was significantly more effective in enhancing GABAergic synaptic transmission. Propofol (250 nM) prolonged the inhibitory postsynaptic currents decay times by 10.4 ± 6.1 milliseconds (n = 9) with ALLO added to the bathing solution; in the absence of ALLO, however, propofol prolonged the decay time by only 3.8 ± 2 milliseconds (n = 13). In cortical neurons, GABAA receptor-mediated synaptic transmission is potentiated by ALLO and propofol in a synergistic manner, whereas the effects on spontaneous action potential activity appear additive. A coapplication of neurosteroids and propofol in general anesthesia and intensive care medicine may open new ways to reduce anesthetic dose requirements and, thus, avoid undesired anesthetic-induced side effects.

P.4.c.001 - G-protein coupled receptor 55 in the central amygdala modulates anxiety-like behaviors in rats

Since allopregnanolone (ALLO) elicits anxiolytic-like action and increases neuropeptide Y Y1 (NPY Y1) receptors gene expression in the amygdala, we were interested in studying the involvement of NPY Y1 receptors in the anxiolytic-like actions of ALLO. The anxiety-like behavior was evaluated in mice using Vogel's conflict test (VCT), in which number of shocks were measured. ALLO and NPYergic agents, alone or in combinations, were administered bilaterally into the central nucleus of amygdala (CeA). The intra-CeA administration of ALLO, NPY or NPY Y1/Y5 receptors agonist [Leu31, Pro34]-NPY resulted in dose-dependent increase in the number of shocks in VCT, indicating anxiolytic-like effect. However, opposite effect was observed following administration of selective NPY Y1 receptors antagonist BIBP3226. While prior administration with NPY or [Leu31, Pro34]-NPY, at the subeffective dose, potentiated the ALLO-induced anxiolytic-like effect, the same was antagonized by BIBP3226. Further, the effect of acute ALLO (30 mg/kg, intraperitoneal) on the endogenous NPY system in the CeA, ventral part of lateral division of bed nucleus of the stria terminalis (BSTLV), nucleus accumbens core (AcbC) and arcuate nucleus (ARC) was studied with immunocytochemistry. Acute ALLO treatment significantly decreased the population of NPY-immunoreactive cells in the CeA and also in the ARC. While NPY-immunoreactive fibers were slightly increased in the AcbC and BSTLV, the cells in AcbC and fibers in ARC did not respond. We suggest that NPY may mediate ALLO induced anxiolytic-like behavior in the neuroanatomical framework of the CeA, possibly via NPY Y1 receptors.

Why may allopregnanolone help alleviate loneliness?

Impaired biosynthesis of Allopregnanolone (ALLO), a brain endogenous neurosteroid, has been associated with numerous behavioral dysfunctions, which range from anxiety- and depressive-like behaviors to aggressive behavior and changes in responses to contextual fear conditioning in rodent models of emotional dysfunction. Recent animal research also demonstrates a critical role of ALLO in social isolation. Although there are likely aspects of perceived social isolation that are uniquely human, there is also continuity across species. Both human and animal research show that perceived social isolation (which can be defined behaviorally in animals and humans) has detrimental effects on physical health, such as increased hypothalamic pituitary adrenal (HPA) activity, decreased brain-derived neurotrophic factor (BDNF) expression, and increased depressive behavior. The similarities between animal and human research suggest that perceived social isolation (loneliness) may also be associated with a reduction in the synthesis of ALLO, potentially by reducing BDNF regulation and increasing HPA activity through the hippocampus, amygdala, and bed nucleus of the stria terminalis (BNST), especially during social threat processing. Accordingly, exogenous administration of ALLO (or ALLO precursor, such as pregnenolone), in humans may help alleviate loneliness. Congruent with our hypothesis, exogenous administration of ALLO (or ALLO precursors) in humans has been shown to improve various stress-related disorders that show similarities between animals and humans i.e., post-traumatic stress disorders, traumatic brain injuries. Because a growing body of evidence demonstrates the benefits of ALLO in socially isolated animals, we believe our ALLO hypothesis can be applied to loneliness in humans, as well.

Postnatal Administration of Allopregnanolone Modifies Glutamate Release but Not BDNF Content in Striatum Samples of Rats Prenatally Exposed to Ethanol

Ethanol consumption during pregnancy may induce profound changes in fetal CNS development. We postulate that some of the effects of ethanol on striatal glutamatergic transmission and neurotrophin expression could be modulated by allopregnanolone, a neurosteroid modulator of GABAA receptor activity. We describe the acute pharmacological effect of allopregnanolone (65 μg/kg, s.c.) administered to juvenile male rats (day 21 of age) on the corticostriatal glutamatergic pathway, in both control and prenatally ethanol-exposed rats (two ip injections of 2.9 g/kg in 24% v/v saline solution on gestational day 8). Prenatal ethanol administration decreased the K+-induced release of glutamate regarding the control group. Interestingly, this effect was reverted by allopregnanolone. Regarding BDNF, allopregnanolone decreases the content of this neurotrophic factor in the striatum of control groups. However, both ethanol alone and ethanol plus allopregnanolone treated animals did not show any change regarding control values. We suggest that prenatal ethanol exposure may produce an alteration of GABAA receptors which blocks the GABA agonist-like effect of allopregnanolone on rapid glutamate release, thus disturbing normal neural transmission. Furthermore, the reciprocal interactions found between GABAergic neurosteroids and BDNF could underlie mechanisms operating during the neuronal plasticity of fetal development.

Progesterone Exerts a Neuromodulatory Effect on Turning Behavior of Hemiparkinsonian Male Rats: Expression of 3 α -Hydroxysteroid Oxidoreductase and Allopregnanolone as Suggestive of GABAA Receptors Involvement.

There is a growing amount of evidence for a neuroprotective role of progesterone and its neuroactive metabolite, allopregnanolone, in animal models of neurodegenerative diseases. By using a model of hemiparkinsonism in male rats, injection of the neurotoxic 6-OHDA in left striatum, we studied progesterone's effects on rotational behavior induced by amphetamine or apomorphine. Also, in order to find potential explanatory mechanisms, we studied expression and activity of nigrostriatal 3α-hydroxysteroid oxidoreductase, the enzyme that catalyzes progesterone to its active metabolite allopregnanolone. Coherently, we tested allopregnanolone for a possible neuromodulatory effect on rotational behavior. Also, since allopregnanolone is known as a GABAA modulator, we finally examined the action of GABAA antagonist bicuculline. We found that progesterone, in addition to an apparent neuroprotective effect, also increased ipsilateral expression and activity of 3α-hydroxysteroid oxidoreductase. It was interesting to note that ipsilateral administration of allopregnanolone reversed a clear sign of motor neurodegeneration, that is, contralateral rotational behavior. A possible GABAA involvement modulated by allopregnanolone was shown by the blocking effect of bicuculline. Our results suggest that early administration of progesterone possibly activates genomic mechanisms that promote neuroprotection subchronically. This, in turn, could be partially mediated by fast, nongenomic, actions of allopregnanolone acting as an acute modulator of GABAergic transmission.

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.

Isoallopregnanolone antagonize allopregnanolone-induced effects on saccadic eye velocity and self-reported sedation in humans

Allopregnanolone (AP) is an endogenous neurosteroid. It modulates the effect of γ-amino-butyric acid (GABA) on the GABA type A (GABAA) receptor, which leads to increased receptor activity. Since the GABA-system is mainly inhibitory, increased AP activity leads to modulation of neuronal activity. In vitro studies of GABAA receptor activity and in vivo animal studies of sedation have shown that AP-induced effects can be inhibited by another endogenous steroid, namely isoallopregnanolone (ISO). In this study we investigated if ISO can antagonize AP-induced effects in healthy female volunteers, via measurements of saccadic eye velocity (SEV) and self-rated sedation. With a single-blind cross-over design, 12 women were studied on three separate occasions; given AP alone or AP in combination with one of two ISO doses. Congruent with previous reports, AP administration decreased SEV and induced sedation and these effects were diminished by simultaneous ISO administration. Also, the ISO effect modulation was seemingly stronger for SEV than for sedation. These effects were observed already at an ISO dose exposure that was approximately half of that of AP. In conclusion, ISO antagonized AP-induced decrease in SEV and self-reported sedation, probably in a non-competitive manner.

Allopregnanolone and neuroinflammation: a focus on multiple sclerosis.

The progesterone derivative allopregnanolone (ALLO) is one of the most widely studied compounds among neurosteroids. Through interactions with GABA-A receptors expressed by neurons and glial cells, ALLO has been shown to affect diverse aspects of neural cell physiology, including cell proliferation and survival, migration, and gene expression. Recent data point to important roles for ALLO in different neurodegenerative disorders, including Alzheimer's disease, Parkinson's disease, and multiple sclerosis (MS). Dysregulation in ALLO biosynthesis pathways has been reported in brain tissue from MS patients as well as in the central nervous system (CNS) tissue derived from MS animal models. Administration of ALLO has been shown to ameliorate neurobehavioral deficits together with neuropathology and inflammation in the CNS of animals with autoimmune demyelination. These findings are in line with previous reports indicating growth- and differentiation-promoting actions of ALLO on neurons and glial cells as well as its neuroprotective effects in the context of other CNS diseases. Nonetheless, these findings have also raised the possibility that ALLO might influence leukocyte biology and associated neuroinflammatory mechanisms independent of its neuroregenerative properties. Herein, we review the current knowledge regarding the role of ALLO in the pathogenesis of MS, and discuss the potential cellular and molecular pathways that might be influenced by ALLO in the context of disease.

Biosynthesis and biological action of pineal allopregnanolone.

The pineal gland transduces photoperiodic changes to the neuroendocrine system by rhythmic secretion of melatonin. We recently provided new evidence that the pineal gland is a major neurosteroidogenic organ and actively produces a variety of neurosteroids de novo from cholesterol in birds. Notably, allopregnanolone is a major pineal neurosteroid that is far more actively produced in the pineal gland than the brain and secreted by the pineal gland in juvenile birds. Subsequently, we have demonstrated the biological action of pineal allopregnanolone on Purkinje cells in the cerebellum during development in juvenile birds. Pinealectomy (Px) induces apoptosis of Purkinje cells, whereas allopregnanolone administration to Px chicks prevents cell death. Furthermore, Px increases the number of Purkinje cells that express active caspase-3, a crucial mediator of apoptosis, and allopregnanolone administration to Px chicks decreases the number of Purkinje cells expressing active caspase-3. It thus appears that pineal allopregnanolone prevents cell death of Purkinje cells by suppressing the activity of caspase-3 during development. This paper highlights new aspects of the biosynthesis and biological action of pineal allopregnanolone.

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.

Interaction between neonatal allopregnanolone administration and early maternal separation: Effects on adolescent and adult behaviors in male rat

Endogenous neurosteroid level fluctuations are related to several emotional and behavioral alterations. Neurosteroids also have important roles during neurodevelopment, with there being a relationship between modification of their levels in neurodevelopmental periods and behavioral alterations in adolescence and adulthood. Early maternal separation (EMS) is a stressful event that also alters neurodevelopment and adolescent and adult behaviors. The aim of the present study is to analyze the interaction between the effects of the neonatal alteration of allopregnanolone (AlloP), neurosteroid that increase its levels after acute stress presentation, and EMS on adolescent exploration and adult anxiety and sensorimotor gating in male rats. AlloP (10mg/kgs.c.) was administrated between postnatal day 5 (PN5) and PN9, and a single 24-hour period of EMS was carried out on PN9. Exploration was analyzed at PN40 and PN60. At adult age (PN85), anxiety was tested by means of the elevated plus-maze test (EPM), and sensorimotor gating by means of prepulse inhibition test (PPI). PPI deterioration has been considered as a reliable indicator of diseases such as schizophrenia. Results showed that the previous neonatal AlloP administration neutralized the effects of EMS in the adolescent exploration (increase of traveled distance and decrease of head-dips). In adult age, an anxiolytic-like profile was observed as a consequence of EMS. Finally, EMS and neonatal AlloP disrupted PPI. Taken together, these data show the important role that physiological neonatal AlloP levels and stressful events play in neural development, adult behavior and vulnerability to neurodevelopmental disorders such as schizophrenia.

Allopregnanolone Elevations Following Pregnenolone Administration Are Associated with Enhanced Activation of Emotion Regulation Neurocircuits

The neurosteroid allopregnanolone is a potent allosteric modulator of the gamma-aminobutyric acid type A receptor with anxiolytic properties. Exogenous administration of allopregnanolone reduces anxiety, and allopregnanolone blockade impairs social and affective functioning. However, the neural mechanism whereby allopregnanolone improves mood and reduces anxiety is unknown. In particular, brain imaging has not been used to link neurosteroid effects to emotion regulation neurocircuitry. To investigate the brain basis of allopregnanolone's impact on emotion regulation, participants were administered 400 mg of pregnenolone (n=16) or placebo (n=15) and underwent 3T functional magnetic resonance imaging while performing the shifted-attention emotion appraisal task, which probes emotional processing and regulation. Compared with placebo, allopregnanolone was associated with reduced activity in the amygdala and insula across all conditions. During the appraisal condition, allopregnanolone increased activity in the dorsal medial prefrontal cortex and enhanced connectivity between the amygdala and dorsal medial prefrontal cortex, an effect that was associated with reduced self-reported anxiety. These results demonstrate that in response to emotional stimuli, allopregnanolone reduces activity in regions associated with generation of negative emotion. Furthermore, allopregnanolone may enhance activity in regions linked to regulatory processes. Aberrant activity in these regions has been linked to anxiety psychopathology. These results thus provide initial neuroimaging evidence that allopregnanolone may be a target for pharmacologic intervention in the treatment of anxiety disorders and suggest potential future directions for research into neurosteroid effects on emotion regulation neurocircuitry.

Possible role of pineal allopregnanolone in Purkinje cell survival

It is believed that neurosteroids are produced in the brain and other nervous systems. Here, we show that allopregnanolone (ALLO), a neurosteroid, is exceedingly produced in the pineal gland compared with the brain and that pineal ALLO acts on the Purkinje cell, a principal cerebellar neuron, to prevent apoptosis in the juvenile quail. We first demonstrated that the pineal gland is a major organ of neurosteroidogenesis. A series of experiments using molecular and biochemical techniques has further demonstrated that the pineal gland produces a variety of neurosteroids de novo from cholesterol in the juvenile quail. Importantly, ALLO was far more actively produced in the pineal gland than in the brain. Pinealectomy (Px) decreased ALLO concentration in the cerebellum and induced apoptosis of Purkinje cells, whereas administration of ALLO to Px quail chicks prevented apoptosis of Purkinje cells. We further found that Px significantly increased the number of Purkinje cells that expressed active caspase-3, a key protease in apoptotic pathway, and daily injection of ALLO to Px quail chicks decreased the number of Purkinje cells expressing active caspase-3. These results indicate that the neuroprotective effect of pineal ALLO is associated with the decrease in caspase-3 activity during the early stage of neuronal development. We thus provide evidence that the pineal gland is an important neurosteroidogenic organ and that pineal ALLO may be involved in Purkinje cell survival during development. This is an important function of the pineal gland in the formation of neuronal circuits in the developing cerebellum.

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.

Allopregnanolone Alters the Luteinizing Hormone, Prolactin, and Progesterone Serum Levels Interfering with the Regression and Apoptosis in Rat Corpus Luteum

Steroids synthesized in the central nervous system are termed "neurosteroids". They are synthesized and metabolized in several brain areas. The objective of this work was to determine if 1 intracerebroventricular allopregnanolone injection in rats can interfere in luteal regression in a close relationship with modifications in LH, progesterone, and prolactin serum concentrations. Allopregnanolone was injected during proestrus morning and the animals were sacrificed on oestrous morning. Ovulation test and histological analysis were performed in the oestrus morning with light and electron microscopy. Serum prolactin, LH, and progesterone levels were measured by radioimmunoassay. The allopregnanolone injection significantly decreased luteinizing hormone serum level and the number of oocytes on oestrus. Progesterone and prolactin serum levels were increased after this injection. The inhibition of apoptotic figures due to allopregnanolone administration was detected in the already formed corpora lutea belonging to the previous ovary cycle and it was significantly lower than in vehicle group (control). When the GABA(A) antagonist (bicuculline) was administered alone or previously to allopregnanolone, no effect on the ovulation rate was observed. No changes in the apoptotic cell numbers were observed with respect to those of vehicle group. These results show that the effect of centrally injected allopreganolone over reproductive function could be due to a centrally originated LH mediated effect over ovarian function that affects luteal regression, through the inhibition of apoptosis and stimulation of progesterone and prolactin release.

Allopregnanolone prevents memory impairment: Effect on mRNA expression and enzymatic activity of hippocampal 3-α hydroxysteroid oxide-reductase

In this work we investigated how the neurosteroid allopregnanolone can modulate learning and memory processes. For this purpose, we used ovariectomized (OVX) rats subcutaneously injected with oestradiol benzoate (E) alone or E and progesterone (P). Then, rats were injected in dorsal hippocampus with allopregnanolone or vehicle. Animals were tested in inhibitory avoidance task (IA task). After behavioural test hippocampal mRNA expression and enzymatic activity of 3α-HOR, the enzyme responsible of allopregnanolone synthesis, were analysed. In IA task OVX-EP rats spent less time on platform, compared to those OVX or OVX-E. Regression analyses revealed that there was a significant negative relationship between E-P infusion and performance in this task. Pre-training allopregnanolone administration to OVX-EP rats increased the time spent on the platform. Interestingly, when enzymatic activity of 3α-HOR was tested, OVX-EP rats showed a significant decrease in the enzymatic activity, compared with OVX and OVX-E rats. In addition, OVX-EP group showed a significant increase in the enzymatic activity after intrahippocampal infusion of allopregnanolone. On the other hand, when mRNA expression of 3α-HOR was analysed no differences were observed when the hippocampal allopregnanolone injection was done. These results suggest that E and P have amnesic effects on female rats, being reversed by allopregnanolone through its modulation on hippocampal 3α-HOR activity.

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.

Effects of Early Postnatal Allopregnanolone Administration on Elevated Plus Maze Anxiety Scores in Adult Male Wistar Rats

Background/Aims: Recent findings suggest that neurosteroids are involved in brain development. The present study focused on the long-term effects of developmentally altered allopregnanolone (AlloP) levels on anxiety-like behavior in adulthood. Method: We administered AlloP (10 mg/kg) to rat pups once a day from the 5th to the 9th day after birth. A dose-response study on midazolam in the elevated plus maze test was carried out in adulthood (experiment 1) in order to screen GABAA-benzodiazepine function alterations. Given that the anxiety-like responses were not affected by AlloP, we doubled the initial AlloP dose (experiment 2). One group of pups was left undisturbed with their dams in order to control the effects of daily handling. Only males were behaviorally tested. Results: Neonatal AlloP administration (10 mg/kg) did not alter the behavioral response to midazolam in adulthood at the doses tested. Neonatal AlloP administration at the higher dose (20 mg/kg) induced an anxiolytic-like profile in adulthood (increased entries into and time spent in the open arms), without affecting motor activity. The behavioral effects of neonatal AlloP administration were both selective and independent of daily handling. Conclusion: Alterations in AlloP levels during maturation could partly explain the interindividual differences shown by adult subjects in response to environmental stress.

Allopregnanolone Potentiates the Glutamate-Mediated Seizures Induced by 4-Aminopyridine in Rat Hippocampus in vivo

Excitatory and inhibitory neurotransmission in the central nervous system can be modulated by neurosteroids. We previously found that in rat hippocampal slices allopregnanolone (3α-hydroxy-5α-pregnan-20-one), a positive GABA(A) receptor modulator, suppresses the epileptic discharges induced by 4-aminopyridine (4-AP), a convulsant K(+) channel blocker that stimulates glutamate release. Here, we tested the action of allopregnanolone on the epileptogenic and excitotoxic effects of the intrahippocampal administration of 4-AP in vivo. Drugs were perfused by a microdialysis cannula-electrode in the dorsal hippocampus and the EEG was recorded. Extracellular levels of aspartate, glutamate and GABA were analyzed by HPLC in the microdialysis fractions, and 24h after the experiment the hippocampus was studied histologically. 4-AP induced intense epileptic discharges, increased the extracellular levels of aspartate, glutamate, and GABA by 383, 420, and 245%, respectively, and produced a notable neurodegeneration in CA1 and CA3 areas. Allopregnanolone administration alone did not affect the electrical activity, amino acids levels or cellular morphology, but when co-infused with 4-AP incremented 55-77% the duration of the epileptic discharges, and potentiated 32-49% the release of glutamate in comparison with 4-AP alone. The 4-AP-induced neurodegeneration was not modified by allopregnanolone. The NMDA receptor antagonist MK-801 protected against the epilepsy and neurodegeneration produced by 4-AP, and allopregnanolone did not affect this protection. We conclude that, differently from the observations in vitro, allopregnanolone potentiated the stimulatory effect of 4-AP on glutamate release and that this may explain the potentiation of the epileptogenic effect of 4-AP in vivo.

Role of the neurosteroid allopregnanolone in the hyperalgesic behavior induced by painful nerve injury in rats

The neurosteroid allopregnanolone (AP) influences the excitability of the central nervous system by acting as a positive allosteric modulator of γ-aminobutyric acid type A (GABA(A)) receptors. Here, we investigated the role of AP and its therapeutic potential in rats that showed hyperalgesic behavior after undergoing spinal nerve ligation (SNL). AP levels measured in the spinal cord and brain of rats that underwent SNL were greater than the corresponding levels in control animals. More importantly, spinal AP levels in hyperalgesic rats were lower than those in the rats that did not develop hyperalgesia following SNL; in contrast, brain AP levels were comparable among these groups. No differences in serum AP levels were observed among the groups. In addition, intrathecal exogenous administration of AP showed the antihyperalgesic effects in hyperalgesic rats after SNL. These findings suggest that changes in spinal AP biosynthesis are involved in the pathogenesis of neuropathic pain following peripheral nerve injury, and pharmacological manipulation of this phenomenon may provide a potential therapeutic target for neuropathic pain.