The Effects of Neuroactive Steroids on Myelin in Health and Disease.

Myelin plays a pivotal role in the efficient transmission of nerve impulses. Disruptions in myelin integrity are associated with numerous neurological disorders, including multiple sclerosis. In the central nervous system (CNS), myelin is formed by oligodendrocytes. Remyelination refers to the re-formation of the damaged myelin sheath by newly formed oligodendrocytes. Steroids have gained attention for their potential modulatory effects on myelin in both health and disease. Steroids are traditionally associated with endocrine functions, but their local synthesis within the nervous system has generated significant interest. The term "neuroactive steroids" refers to steroids that can act on cells of the nervous system. In the healthy state, neuroactive steroids promote myelin formation, maintenance, and repair by enhancing oligodendrocyte differentiation and maturation. In pathological conditions, such as demyelination injury, multiple neuroactive steroids have shown promise in promoting remyelination. Understanding the effects of neuroactive steroids on myelin could lead to novel therapeutic approaches for demyelinating diseases and neurodegenerative disorders. This review highlights the potential therapeutic significance of neuroactive steroids in myelin-related health and diseases. We review the synthesis of steroids by neurons and glial cells and discuss the roles of neuroactive steroids on myelin structure and function in health and disease. We emphasize the potential promyelinating effects of the varying levels of neuroactive steroids during different female physiological states such as the menstrual cycle, pregnancy, lactation, and postmenopause.

Role of progesterone on dexamethasone-induced alterations in placental vascularization and progesterone receptors in rats†.

BACKGROUND: Intrauterine growth restriction (IUGR) is manifested by lower maternal progesterone levels, smaller placental size, and decreased placental vascularity indicated by lower expression of vascular endothelial growth factor (VEGF). Studies showed that progesterone increases angiogenesis and induces VEGF expression in different tissues. Therefore, the aim of the present study is to evaluate the effect of progesterone on placental vascular bed and VEGF expression and the modulation of nuclear and membranous progesterone receptors (PR) in dexamethasone-induced rat IUGR model. METHODS: Pregnant Sprague-Dawley rats were allocated into four groups and given intraperitoneal injections of either saline, dexamethasone, dexamethasone, and progesterone or progesterone. Injections started on gestation day (DG) 15 and lasted until the days of euthanization (19 and 21 DG). Enzyme-linked immunosorbent assay was used to evaluate plasma progesterone levels. Real-time PCR and western blotting were used to evaluate gene and protein expressions of VEGF, and PR in labyrinth and basal placental zones. Immunohistochemistry was used to locate VEGF and different PRs in placental cells. Immunofluorescence was used to monitor the expression of blood vessel marker (αSMA). RESULTS: Dexamethasone decreased the vascular bed fraction and the expression of VEGF in both placental zones. Progesterone co-treatment with dexamethasone prevented this reduction. Nuclear and membrane PRs showed tissue-specific expression in different placental zones and responded differently to both dexamethasone and progesterone. CONCLUSIONS: Progesterone treatment improves the outcomes in IUGR pregnancy. Progesterone alleviated DEX-induced IUGR probably by promoting placental VEGF and angiogenesis.

Progesterone partially recovers placental glucose transporters in dexamethasone-induced intrauterine growth restriction.

RESEARCH QUESTION: How does progesterone improve fetal outcome and change the expression of placental glucose transporters (GLUT) in dexamethasone-induced intrauterine growth restriction (IUGR)? DESIGN: A total of 64 rats were divided randomly into four different treatment groups based on daily i.p. injections of either saline or dexamethasone in the presence or absence of progesterone. Injections started on the 15th day of gestation (15dg) and lasted until the day of sacrifice at 19dg or 21dg. Maternal plasma progesterone concentrations were measured by enzyme-linked immunosorbent assay. The gene and protein expression of placental GLUT1 and GLUT3 were evaluated in the placental labyrinth and basal zones by real-time polymerase chain reaction and Western blotting, respectively. The localization of GLUT1 and GLUT3 was evaluated by immunohistochemistry. RESULTS: Dexamethasone induced significant decreases in maternal serum progesterone concentrations (P = 0.029) and placental (P < 0.001) and fetal body (P = 0.009) weights. Dexamethasone also reduced the expression of GLUT1 in the labyrinth zone (P = 0.028) and GLUT3 in both the labyrinth (P = 0.002) and basal zones (P = 0.026). Coadministration of dexamethasone and progesterone prevented the reduction in fetal body weight, placental weight and placental GLUT expression compared with that seen in dexamethasone-treated groups. CONCLUSION: These results suggest that progesterone prevents the significant reduction in fetal and placental weights in dexamethasone-induced IUGR, possibly through improving the expression of placental GLUT.

Maternal Interleukin-6 Hampers Hippocampal Neurogenesis in Adult Rat Offspring in a Sex-Dependent Manner.

Maternal immune activation (MIA) during pregnancy leads to long-lasting effects on brain development and function. Several lines of evidence suggest that the maternal inflammatory cytokine interleukin (IL)-6 plays a crucial role in the long-lasting effects of MIA on adult offspring. IL-6 is naturally produced during pregnancy in the absence of any underlying immune activation. The objective of this study was to assess whether this naturally occurring IL-6 has long-lasting effects on brain plasticity and function. Therefore, pregnant rats were given either an IL-6-neutralizing antibody (IL-6Ab) or vehicle during the third week of pregnancy. Newly born (doublecortin) and mature neurons (NeuN) were monitored in the hippocampus of adult male and female offspring. Prenatal IL-6Ab led to an enhanced number of newly born and mature neurons in the dentate gyrus of the hippocampus of male but not female adult offspring. This enhanced neurogenesis was associated with an increased propensity in memory acquisition in male offspring. Blunting the naturally occurring IL-6 during pregnancy did not have a significant long-lasting impact on astrocyte cell density (GFAP), or on anxiety-like behavior as assessed with elevated plus maze and open field tests. Taken together, these data suggest that maternal IL-6 contributes, at least in part, to the programming of the brain's development in a sex-dependent manner.

Prenatal Inflammation Dampens Neurogenesis and Enhances Serotonin Transporter Expression in the Hippocampus of Adult Female Rats.

BACKGROUND/AIMS: Prenatal exposure to lipopolysaccharide (LPS) dampens hippocampal neurogenesis. This effect is associated with increased anxiety-like behavior in adult offspring. Furthermore, blocking serotonin transporters (SERT) promotes adult neurogenesis. Previous studies were performed largely in males. Therefore, we explored the impact of prenatal LPS on neurogenesis, SERT expression in the hippocampus, and anxiety-like behavior in female rats during prepubertal and adulthood stages. MATERIALS AND METHODS: Timed pregnant rats were injected with either saline or LPS (100 µg/kg, i.p.) on gestational days 15, 17, and 19. Newly born neurons were monitored by immunohistochemistry, and anxiety-like behavior was monitored using the elevated plus maze and open-field test. SERT expression in the hippocampus was assessed by Western blot and immunofluorescence. RESULTS: Prenatal LPS led to reduced hippocampal neurogenesis in adult but not in prepubertal female offspring. This reduced neurogenesis was associated with enhanced hippocampal expression of SERT protein. However, there was no significant impact of prenatal LPS on anxiety-like behavior. CONCLUSIONS: Prenatal LPS-induced reduction in neurogenesis was dissociated from anxiety-like behavior in adult female rats. Furthermore, the long-lasting impact of prenatal LPS on neurogenesis in female offspring was age-dependent.

Toll-like receptor 4-mediated immune stress in pregnant rats activates STAT3 in the fetal brain: role of interleukin-6.

BACKGROUND: Prenatal exposure to pathogens induces long lasting effect on brain function and plasticity. It is unclear how maternal immune stress impacts fetal brain development. Immune challenged pregnant rats induce the production of inflammatory cytokines including tumor necrosis factor (TNF)α, interleukin (IL)1ß, and IL-6. IL-6 crosses the placenta but its mechanism of action on fetal brain is unclear. METHODS: Gestation day 15 (GD15) rats were given a single injection of lipopolysaccharide (LPS) (100 µg/kg) in the presence or the absence of an IL-6 neutralizing antibody (IL-6Ab, 10 µg/kg). The activation of the intracellular signal of IL-6; signal transducer and activator of transcription (STAT3) and levels of glucocorticoids (GCs) were monitored in fetal brains. RESULTS: LPS administration to GD15 rats significantly increased the phosphorylation levels of STAT3 in fetal brains. Such activation was blunted by IL-6Ab. LPS induced a significant rise in GCs in the plasma of dams but not in fetal brains. IL-6Ab significantly reduced LPS-induced GCs in maternal plasma. CONCLUSION: Toll-like receptor 4 (TLR4)-induced activation of the maternal innate immune system affects fetal brains likely via the mobilization of IL-6/STAT3 pathway. In contrast, TLR4-stimulated maternal GCs release is less likely to play a significant role in fetal brain development.

The promyelinating properties of androstenediol in gliotoxin-induced demyelination in rat corpus callosum.

AIMS: Experimental evidence has shown that the adrenal steroid hormone, androstenediol, dampens the symptoms of demyelination. However, the cellular and molecular effects of androstenediol are not yet known. In the present study, we investigated the cellular and subcellular effects of this hormone in a gliotoxin-induced demyelination model. METHODS: Male Sprague Dawley rats received 2 µl of either saline or the gliotoxin ethidium bromide (EB, 0.04%) into the corpus callosum. These rats received daily subcutaneous injections of either oil or androstenediol (5 mg/kg). Their brains were collected at 2, 7, 14 and 28 days post-EB injection. Demyelinated lesions were assessed using Luxol fast blue staining. Immunofluorescent staining was used to investigate the number of oligodendrocyte progenitor cells, their maturation and microglial activation at the lesion site. Remyelination was further explored using transmission electron microscopy. The expression levels of total and phosphorylated MBP isoforms and CNPase were explored using western blot. RESULTS: Androstenediol decreased the size of demyelinated lesions in the corpus callosum at 7 and 14 days post-EB injection. It enhanced the number of oligodendrocyte precursor cells, promoted an increase in the number of mature oligodendrocytes and reduced microglial activation. Androstenediol also stimulated the phosphorylation of MBP at the site of the lesion and promoted remyelination of the affected axons. CONCLUSIONS: These data strongly suggest that androstenediol is endowed with promyelinating properties in a model of focal gliotoxin-induced demyelination. It induces its promyelinating effects by enhancing the number of oligodendrocyte precursor cells and their maturation at the lesion site.

Glucocorticoid-induced fetal brain growth restriction is associated with p73 gene activation.

Fetal exposure to excessive amounts of glucocorticoids (GCs) hampers proper brain development. The molecular mechanism(s) underlying these GCs effects are not well understood. We explored the impact of fetal exposure to maternal GCs on fetal brain expression of p63 and p73 transactivation (TA) and dominant negative (ΔN) gene variants that promote neural cell death (TA) and cell survival programs (ΔN). The fetoplacental enzyme 11ß-hydroxysteroid dehydrogenase 2, which shields fetuses from maternal glucocorticoids, was inhibited throughout pregnancy by daily injection of carbenoxolone to pregnant dams. The expression of p63 and p73 gene variants and proteins was monitored by real-time rtPCR and Western blot in the brains of male and female fetuses. Carbenoxolone administration led to an overall enhanced level of corticosterone in the amniotic fluid of both male and female fetuses at late pregnancy. These enhanced corticosterone levels were associated with a significant reduction in fetal brain weights and a significant increase in TAp73 mRNA and p73 protein levels. However, the expression levels of TAp63 mRNA and p63 proteins were either suppressed or unaffected. The pro-neural survival gene variant ΔNp73 was significantly reduced in female and enhanced in male fetal brains, whereas ΔNp63 was significantly reduced in the brains of both genders. These data suggest that the GCs-induced negative impact on fetal brain development likely is due, at least in part, to their action of the pro-neural cell death gene variant TAp73 and to the modulation of the pro-survival ΔNp63 and ΔNp73 gene variants in a gender-dependent fashion.

Long-lasting impact of early life immune stress on neuroimmune functions.

Fever is one major cardinal sign of disease. It results from an intricate interplay between the immune system and the central nervous system. Bacterial or viral infections activate peripheral immune competent organs which send inflammatory signals to the brain and lead to an increase in body temperature. The increased body temperature creates a conducive environment to optimize the body's fight against the infection. A large body of experimental evidence suggests that early life bacterial or viral infections can lead to a long-lasting impact on this natural febrile response. The early life pathogenic encounter heightens the hypothalamic-pituitary-adrenal axis response, dampens the innate immune system, and consequently reduces the febrile response to a subsequent immune challenge during adulthood. This 'programming' effect operates only when such early life immune challenges occur during a critical window of either prenatal or postnatal development. In this review, the mechanisms underlying the long-lasting impact of perinatal immune challenge on adult fever are addressed.

Breastfeeding promotes oligodendrocyte precursor cells division and myelination in the demyelinated corpus callosum.

Demyelination alters the conduction of neuronal signals and hampers sensory-motor functions. Experimental and clinical evidence suggest that breastfeeding exerts a promyelinating impact on the maternal brain. The mechanism underlying this neuroprotective effect is not well-understood. In the present paper, we assessed the impact of rat lactation on lysolecithin-induced demyelination injury within the corpus callosum of lactating and non-lactating postpartum rats. We show that lactation enhanced the cell density of oligodendrocyte precursor cells (OPCs), but not that of activated microglia and astrocytes, within the demyelination lesion. Lactation also increased the expression of myelin markers involved in the initial stage of myelin recovery (Myelin-associated glycoprotein and 2',3'-cyclic nucleotide 3'-phosphodiesterase) and reduced the demyelination injury. Altogether, these data suggest that lactation creates a conducive promyelinating environment through increased OPCs cell division, enhanced expression of select myelin proteins, and reduced number of non-myelinated axons.

Prenatal immune stress in rats dampens fever during adulthood.

Fever is a major component of the host's defense against infection. Inadequate febrile response can predispose an individual to the deleterious effects of infection. Neonatal exposure to infectious agents such as bacterial lipopolysaccharide (LPS) permanently dampens the adult febrile response. Whether prenatal immune challenge alters febrile response during adulthood is still not known. In the present study, LPS (100 µg/kg, i.p.) or pyrogen-free saline was administered to pregnant rats on either gestation day (GD) 12, 15 or 19 and the febrile response of their respective adult offspring was monitored. During adulthood (>70 days old), the rats born to LPS-injected dams on GD15 displayed a significantly attenuated febrile response to LPS (50 µg/kg, i.p.) compared to their control counterparts born to dams given saline on GD15. Immune challenge during either early (GD12) or late (GD19) pregnancy did not have a significant impact on fever in the adult offspring. Immune challenge on GD15, but not on GD12 or 19, heightened the plasma corticosterone response to a subsequent LPS injection to the adult offspring but did not have a significant effect on their basal plasma corticosterone levels. Finally, LPS-induced COX-2 in the fever-controlling regions of the hypothalamus was significantly reduced in the adult rats born to dams given LPS on GD15 compared to their counterparts born to dams given saline on GD15. Such COX-2 reduction was not observed in the adult offspring born to dams given LPS on either GD12 or 19. Taken together, these data suggest that a single immune challenge during a critical window of pregnancy alters the neuroimmune response in adult offspring.

Muscle Hypertrophy in a Newly Developed Resistance Exercise Model for Rats.

Clinical evidence suggests that resistance exercise exerts health benefit. The mechanisms underlying such health benefits is largely explored in experimental animals. Available experimental models have several shortcomings such as the need for noxious stimuli that could affect the physiological readouts. In this study, we describe a simple-to-use experimental model of resistance exercise. In this resistance exercise, rats pull pre-determined weights using a tunnel and pulley system. We show that resistance-exercised rats developed a larger pulling strength when compared to those seen in either control rats or in rats subjected to traditional treadmill exercise. Histological examination revealed that resistance exercise led to a larger fiber cross-sectional area in the plantaris muscle, but not in the gastrocnemius or the soleus muscles. Similarly, the percentage of type-II muscle fibers in the plantaris was increased in resistance exercised rats when compared to those seen in plantaris muscles of either control or treadmill-exercised rat groups. Furthermore, this resistance exercise led to a significant increase in the expression levels of the phosphorylated protein kinase B; a marker of muscle hypertrophy in the plantaris muscle. Such effects were not seen in treadmill-trained rats. In conclusion, we developed an experimental model that can be amenable for experimental exploration of the mechanisms underlying the beneficial effects of resistance exercise. We further provide evidence that this resistance exercise model enhanced muscle strength and muscle hypertrophy.

Prenatal Activation of Glucocorticoid Receptors Induces Memory Impairment in a Sex-Dependent Manner: Role of Cyclooxygenase-2.

Prenatal exposure to dexamethasone (DEX) results in long-lasting effects on cognitive functions such as learning and memory impairment. However, the mechanisms underlying these DEX-induced deleterious effects are not well known. Here, we assessed whether cyclooxygenase-2 (COX-2) is involved in the impact of prenatal exposure to DEX on learning and memory during adulthood. Pregnant Sprague-Dawley rats received daily injections of either DEX (0.2 mg/kg; i.p.) or saline from gestation day (GD) 14 until GD21. Gene and protein expression of COX-2, as well as presynaptic (synaptophysin) and postsynaptic (postsynaptic density protein-95) proteins, were monitored in the dorsal and ventral hippocampi of adult male and female offspring. A different cohort of adult male and female rat offspring was given daily injections of either vehicle or a specific COX-2 inhibitor (celecoxib 10 mg/kg, i.p.) for 5 consecutive days and was subsequently subjected to Morris water maze memory test. Prenatal DEX enhanced the expression of COX-2 protein and cox-2 mRNA in the dorsal hippocampus of adult female but not male rats. This enhanced COX-2 expression was associated with reduced expression in pre- and postsynaptic proteins and altered memory acquisition and retention. Administration of COX-2-specific inhibitor alleviated prenatal DEX-induced memory impairment in adult female rats. This study suggests that prenatal activation of glucocorticoid receptors stimulates COX-2 gene and protein expression and impairs hippocampal-dependent spatial memory in female but not male rat offspring. Furthermore, COX-2 selective inhibitors can be used to alleviate the long-lasting deleterious effects of corticosteroid medication during pregnancy.

Astrocytes and Microglia in Stress-Induced Neuroinflammation: The African Perspective.

Background: Africa is laden with a youthful population, vast mineral resources and rich fauna. However, decades of unfortunate historical, sociocultural and leadership challenges make the continent a hotspot for poverty, indoor and outdoor pollutants with attendant stress factors such as violence, malnutrition, infectious outbreaks and psychological perturbations. The burden of these stressors initiate neuroinflammatory responses but the pattern and mechanisms of glial activation in these scenarios are yet to be properly elucidated. Africa is therefore most vulnerable to neurological stressors when placed against a backdrop of demographics that favor explosive childbearing, a vast population of unemployed youths making up a projected 42% of global youth population by 2030, repressive sociocultural policies towards women, poor access to healthcare, malnutrition, rapid urbanization, climate change and pollution. Early life stress, whether physical or psychological, induces neuroinflammatory response in developing nervous system and consequently leads to the emergence of mental health problems during adulthood. Brain inflammatory response is driven largely by inflammatory mediators released by glial cells; namely astrocytes and microglia. These inflammatory mediators alter the developmental trajectory of fetal and neonatal brain and results in long-lasting maladaptive behaviors and cognitive deficits. This review seeks to highlight the patterns and mechanisms of stressors such as poverty, developmental stress, environmental pollutions as well as malnutrition stress on astrocytes and microglia in neuroinflammation within the African context.

Early life activation of toll-like receptor 4 reprograms neural anti-inflammatory pathways.

A single postnatal exposure to the bacterial endotoxin, lipopolysaccharide (LPS), reduces the neuroimmune response to a subsequent LPS exposure in the adult rat. The attenuated fever and proinflammatory response is caused by a paradoxical, amplified, early corticosterone response to LPS. Here we identify the mechanisms underlying the heightened corticosterone response to LPS in adults after early life exposure to LPS. In postnatal LPS-treated rats, hypothalamic corticotrophin-releasing hormone mRNA, pituitary proopiomelanocortin mRNA, and circulating adrenocorticotrophic hormone were all increased after adult exposure to LPS without significant modification to hippocampal or hypothalamic glucocorticoid receptor mRNA or protein or vagally mediated afferent signaling to the brain. Postnatal LPS administration did cause a persistent upregulation of the LPS Toll-like receptor-4 (TLR4) mRNA in liver and spleen, but not in brain, pituitary, or adrenal gland. In addition, cyclooxygenase-2 (COX-2), which is a prostaglandin biosynthetic enzyme and is normally undetectable in most peripheral tissue, was constitutively expressed in the liver. Adult immune activation of the upregulated TLR4 and COX-2 caused a rapid, amplified rise in circulating, but not brain, prostaglandin E(2) that induced an early, enhanced activation of the hypothalamic-pituitary-adrenal (HPA) axis. Thus, postnatal LPS reprograms the neuroimmune axis by priming peripheral tissues to create a novel, prostaglandin-mediated activation of the HPA axis brought about by increased constitutive expression of TLR4 and COX-2.

Effects of the noradrenergic system in rat white matter exposed to oxygen-glucose deprivation in vitro.

Norepinephrine (NE) is released in excess into the extracellular space during oxygen-glucose deprivation (OGD) in brain, increasing neuronal metabolism and aggravating glutamate excitoxicity. We used isolated rat optic nerve and spinal cord dorsal columns to determine whether the noradrenergic system influences axonal damage in white matter. Tissue was studied electrophysiologically by recording the compound action potential (CAP) before and after exposure to 60 min of OGD at 36 degrees C. Depleting catecholamine stores with reserpine was protective and improved CAP recovery after 1 h of reperfusion from 17% (control) to 35%. Adding NE during OGD decreased CAP recovery to 8%, and adding NE to reserpine during OGD eliminated the protective effect of the latter. Selective inhibitors of Na(+)-dependent norepinephrine transport desipramine and nisoxetine improved recovery to 58% and 44%, respectively. alpha2 adrenergic receptor agonists UK14,304 and medetomidine improved CAP recovery to 41% and 46% after 1 h of OGD. Curiously, alpha2 antagonists alone were also highly protective (e.g., atipamezole: 86% CAP recovery), at concentrations that did not affect baseline excitability. The protective effect of alpha2 receptor modulation was corroborated by imaging fluorescent Ca(2+) and Na(+) indicators within axons during OGD. Both agonists and antagonists significantly reduced axonal Ca(2+) and Na(+) accumulation in injured axons. These data suggest that the noradrenergic system plays an active role in the pathophysiology of axonal ischemia and that alpha2 receptor modulation may be useful against white matter injury.

Ganaxolone enhances microglial clearance activity and promotes remyelination in focal demyelination in the corpus callosum of ovariectomized rats.

AIM: Experimental studies have shown that the progesterone metabolite, allopregnanolone, is endowed with promyelinating effects. The mechanisms underlying these promyelinating effects are not well understood. Therefore, we explored the impact of allopregnanolone's synthetic analogue, ganaxolone, on remyelination and microglial activation following focal demyelination in the corpus callosum of ovariectomized rats. METHODS: Ovariectomized adult Sprague Dawley rats received a stereotaxic injection of 2 µL of 1% lysolecithin solution in the corpus callosum followed by daily injections of either ganaxolone (intraperitoneal injection [i.p.], 2.5 mg/kg) or vehicle. The demyelination lesion was assessed 3 and 7 days postdemyelination insult using Luxol fast blue staining and transmission electron microscopy. The expression levels of myelin proteins (MBP, MAG, MOG, CNPase) were explored using Western blot. The inflammatory response and clearance of damaged myelin were evaluated using immunofluorescent staining (Iba1, dMBP, GFAP) and multiplex enzyme-linked immunosorbent assay (IL-1ß, TNF-α, IL-4, IL-10, IL-6). RESULTS: Systemic administration of ganaxolone promoted remyelination of lysolecithin-induced demyelination, upregulated the expression of major myelin proteins, and enhanced microglial clearance of damaged myelin. Astrocytosis, as well as locally produced pro- and antiinflammatory cytokines, was not affected by ganaxolone treatment. CONCLUSION: Ganaxolone promotes remyelination in response to focal demyelination of the corpus callosum of ovariectomized rats. This effect is, at least in part, mediated by enhancing microglial clearance of myelin debris, which creates a conducive environment for a successful remyelination process.

Postnatal inflammation increases seizure susceptibility in adult rats.

There are critical postnatal periods during which even subtle interventions can have long-lasting effects on adult physiology. We asked whether an immune challenge during early postnatal development can alter neuronal excitability and seizure susceptibility in adults. Postnatal day 14 (P14) male Sprague Dawley rats were injected with the bacterial endotoxin lipopolysaccharide (LPS), and control animals received sterile saline. Three weeks later, extracellular recordings from hippocampal slices revealed enhanced field EPSP slopes after Schaffer collateral stimulation and increased epileptiform burst-firing activity in CA1 after 4-aminopyridine application. Six to 8 weeks after postnatal LPS injection, seizure susceptibility was assessed in response to lithium-pilocarpine, kainic acid, and pentylenetetrazol. Rats treated with LPS showed significantly greater adult seizure susceptibility to all convulsants, as well as increased cytokine release and enhanced neuronal degeneration within the hippocampus after limbic seizures. These persistent increases in seizure susceptibility occurred only when LPS was given during a critical postnatal period (P7 and P14) and not before (P1) or after (P20). This early effect of LPS on adult seizures was blocked by concurrent intracerebroventricular administration of a tumor necrosis factor alpha (TNFalpha) antibody and mimicked by intracerebroventricular injection of rat recombinant TNFalpha. Postnatal LPS injection did not result in permanent changes in microglial (Iba1) activity or hippocampal cytokine [IL-1beta (interleukin-1beta) and TNFalpha] levels, but caused a slight increase in astrocyte (GFAP) numbers. These novel results indicate that a single LPS injection during a critical postnatal period causes a long-lasting increase in seizure susceptibility that is strongly dependent on TNFalpha.

Enhanced remyelination during late pregnancy: involvement of the GABAergic system.

Pregnant women with MS experience fewer relapses, especially during the third trimester. In this study, we explore the cellular and molecular events that bring about the protective effect of late pregnancy on the course of de/remyelination in rats. Using cellular, molecular, and ultrastructural methods, we explored remyelination in response to a focal demyelination in the corpus callosum of late pregnant, virgin, and postpartum rats. We further explored the role of GABAA receptor (GABAAR) in the promyelinating effect observed during late pregnancy. Remyelination in response to a gliotoxin-induced demyelination in the corpus callosum was enhanced in late pregnant rats when compared to that seen in virgin and postpartum rats. This pregnancy-associated promyelinating effect was lost when either the GABAAR was blocked or when 5α-reductase, the rate limiting enzyme for the endogenous GABAAR activator allopregnanolone, was inhibited. Taken together, these data suggest that the pregnancy-associated pro-myelination operates, at least in part, through a GABAergic activated system.

Differential Release of Exocytosis Marker Dyes Indicates Stimulation-Dependent Regulation of Synaptic Activity.

There is a general consensus that synaptic vesicular release by a full collapse process is the primary machinery of synaptic transmission. However, competing view suggests that synaptic vesicular release operates via a kiss-and-run mechanism. By monitoring the release dynamics of a synaptic vesicular marker, FM1-43 from individual synapses in hippocampal neurons, we found evidence that the release of synaptic vesicle was delayed by several seconds after the start of field stimulation. This phenomenon was associated with modified opening kinetics of fusion pores. Detailed analysis revealed that some synapses were completely inactive for a few seconds after stimulation, despite immediate calcium influx. This delay in vesicular release was modulated by various stimulation protocols and different frequencies, indicating an activity-dependent regulation mechanism for neurotransmitter exocytosis. Staurosporine, a drug known to induce "kiss-and-run" exocytosis, increased the proportion of delayed synapses as well as the delay duration, while fluoxetine acted contrarily. Besides being a serotonin reuptake inhibitor, it directly enhanced vesicle mobilization and reduced synaptic fatigue. Exocytosis was never delayed, when it was monitored with pH-sensitive probes, synaptopHlourin and αSyt-CypHerE5 antibody, indicating an instantaneous formation of a fusion pore that allowed rapid equilibration of vesicular lumenal pH but prevented FM1-43 release because of its slow dissociation from the inner vesicular membrane. Our observations suggest that synapses operate via a sequential "kiss-and-run" and "full-collapse" exocytosis mechanism. The initially narrow vesicular pore allows the equilibration of intravesicular pH which then progresses toward full fusion, causing FM1-43 release.