Estradiol rapidly modulates spinogenesis in hippocampal dentate gyrus: Involvement of kinase networks.

This article is part of a Special Issue "Estradiol and cognition". Estradiol (E2) is locally synthesized within the hippocampus and the gonads. Rapid modulation of hippocampal synaptic plasticity by E2 is essential for synaptic regulation. The molecular mechanisms of modulation through the synaptic estrogen receptor (ER) and its downstream signaling, however, are largely unknown in the dentate gyrus (DG). We investigated the E2-induced modulation of dendritic spines in male adult rat hippocampal slices by imaging Lucifer Yellow-injected DG granule cells. Treatments with 1 nM E2 increased the density of spines by approximately 1.4-fold within 2h. Spine head diameter analysis showed that the density of middle-head spines (0.4-0.5 µm) was significantly increased. The E2-induced spine density increase was suppressed by blocking Erk MAPK, PKA, PKC and LIMK. These suppressive effects by kinase inhibitors are not non-specific ones because the GSK-3ß antagonist did not inhibit E2-induced spine increase. The ER antagonist ICI 182,780 also blocked the E2-induced spine increase. Taken together, these results suggest that E2 rapidly increases the density of spines through kinase networks that are driven by synaptic ER.

Estradiol rapidly rescues synaptic transmission from corticosterone-induced suppression via synaptic/extranuclear steroid receptors in the hippocampus.

We investigated rapid protection effect by estradiol on corticosterone (CORT)-induced suppression of synaptic transmission. Rapid suppression by 1 µM CORT of long-term potentiation (LTP) at CA3-CA1 synapses was abolished via coperfusion of 1 nM estradiol. N-methyl-D-aspartate (NMDA) receptor-derived field excitatory postsynaptic potential (NMDA-R-fEPSP) was used to analyze the mechanisms of these events. Estradiol abolished CORT-induced suppression of NMDA-R-fEPSP slope. This CORT-induced suppression was abolished by calcineurin inhibitor, and the rescue effect by estradiol on the CORT-induced suppression was inhibited by mitogen-activated protein (MAP) kinase inhibitor. The CORT-induced suppressions of LTP and NMDA-R-fEPSP slope were abolished by glucocorticoid receptor (GR) antagonist, and the restorative effects by estradiol on these processes were mimicked by estrogen receptor α (ERα) and ERß agonists. Taken together, estradiol rapidly rescued LTP and NMDA-R-fEPSP slope from CORT-induced suppressions. A GR→calcineurin pathway is involved in these suppressive effects. The rescue effects by estradiol are driven via ERα or ERß→MAP kinase pathway. Synaptic/extranuclear GR, ERα, and ERß probably participate in these rapid events. Mass-spectrometric analysis determined that acute hippocampal slices used for electrophysiological measurements contained 0.48 nM estradiol less than exogenously applied 1 nM. In vivo physiological level of 8 nM estradiol could protect the intact hippocampus against acute stress-induced neural suppression.

Bisphenol A significantly modulates long-term depression in the hippocampus as observed by multi-electrode system.

OBJECTIVE: Low dose exposure to endocrine disrupters (environmental chemicals) may induce hormone-like effects on wildlife and humans. bisphenol A (BPA) might disturb the neuronal signaling regulated by endogenous estrogens. We investigated the rapid modulation effects of 10nM BPA, a typical endocrine disruptor, on long-term depression (LTD) of adult rat hippocampal slices. METHOD: LTD was induced by a transient perfusion of 30 µM NMDA for 3 min. And measured with multielectrode probes. RESULTS: A 30 min perfusion of 10 nM BPA rapidly enhanced LTD in CA1, however, BPA suppressed LTD in dentate gyrus (DG). An ERRγ antagonist, 4-OH-tamoxifen, suppressed LTD in CA1 and DG. Inhibitor of estrogen receptor ICI 182,780 did not disturb BPA effects. On the other hand, tributyltin (TBT), another endocrine disruptor, did not have any effect on LTD in CA1 and DG. CONCLUSION: ERRγ, but not estrogen receptors, is a high affinity BPA receptor in LTD processes, since the effect of BPA on LTD was suppressed by an ERRγ antagonist. A possible mechanisms of BPA-induced enhancement of LTD could be described with ERRγ, MAPK activation and phosphorylation of MMDA receptors.

Automated analysis of spines from confocal laser microscopy images: application to the discrimination of androgen and estrogen effects on spinogenesis.

Accurate 3D determination of postsynaptic structures is essential to our understanding memory-related function and pathology in neurons. However, current methods of spine analysis require time-consuming and labor-intensive manual spine identification in large image data sets. Therefore, a realistic implementation of algorithm is necessary to replace manual identification. Here, we describe a new method for the automated detection of spines and dendrites based on analysis of geometrical features. Our "Spiso-3D" software carries out automated dendrite reconstruction and spine detection using both eigenvalue images and information of brightness, avoiding detection of pseudo-spines. To demonstrate the potential application of Spiso-3D automated analysis, we distinguished the rapid effects of androgen and estrogen on rapid modulation of spine head diameter in the hippocampus. These findings advance our understanding of neurotrophic function of brain sex steroids. Our method is expected to be valuable to analyze vast amounts of dendritic spines in neurons in the mammalian cerebral cortex.

Modulation of synaptic plasticity by brain estrogen in the hippocampus.

The hippocampus is a center for learning and memory as well as a target of Alzheimer's disease in aged humans. Synaptic modulation by estrogen is essential to understand the molecular mechanisms of estrogen replacement therapy. Because the local synthesis of estrogen occurs in the hippocampus of both sexes, in addition to the estrogen supply from the gonads, its functions are attracting much attention. Hippocampal estrogen modulates memory-related synaptic plasticity not only slowly but also rapidly. Slow actions of 17ß-estradiol (17ß-E2) occur via classical nuclear receptors (ERα or ERß), while rapid E2 actions occur via synapse-localized ERα or ERß. Elevation or decrease of the E2 concentration changes rapidly the density and morphology of spines in CA1-CA3 neurons. ERα, but not ERß, drives this enhancement/suppression of spinogenesis. Kinase networks are involved downstream of ERα. The long-term depression but not the long-term potentiation is modulated rapidly by changes of E2 level. Determination of the E2 concentration in the hippocampus is enabled by mass-spectrometry in combination with derivatization methods. The E2 level in the hippocampus is as high as approx. 8 nM for the male and 0.5-2 nM for the female, which is much higher than that in circulation. Therefore, hippocampus-derived E2 plays a major role in modulation of synaptic plasticity. Many hippocampal slice experiments measure the restorative effects of E2 by supplementation of E2 to E2-depleted slices. Accordingly, isolated slice experiments can be used as in vitro models of in vivo estrogen replacement therapy for ovariectomized female animals with depleted circulating estrogen.

Comparison of sex-steroid synthesis between neonatal and adult rat hippocampus.

Sex-steroid synthesis in the hippocampus had been thought to be much more active at the neonatal stage than at the adult stage. However, the detailed comparison between these two stages had not been demonstrated yet. Here we performed the comparison about the mRNA level of steroidogenic enzymes and the rate of steroid metabolism between these two stages of the hippocampus. The relative expression level of P450(17alpha), 17beta- or 3beta-hydroxysteroid dehydrogenase, or P450arom was approximately 1.3-1.5-fold higher at the neonatal than at the adult stage. The rate of sex-steroid metabolism (from dehydroepiandrosterone to estradiol) was 2-7-fold (depending on different steps) more rapid at the neonatal than at the adult stage. Taken together, neonatal steroidogenesis is moderately more active than adult steroidogenesis.

Rapid modulation of synaptic plasticity by estrogens as well as endocrine disrupters in hippocampal neurons.

Estrogen modulates memory-related synaptic plasticity not only slowly but also rapidly in the hippocampus. However, molecular mechanisms of the rapid action are yet largely unknown. We here describe rapid modulation of representative synaptic plasticity, i.e., long-term depression (LTD), long-term potentiation (LTP) and spinogenesis, by 17beta-estradiol, selective estrogen agonists as well as endocrine disrupters. The authors demonstrated that 1-10 nM estradiol induced rapid enhancement of LTD within 1 h in not only CA1 but also CA3 and dentate gyrus (DG). On the other hand, the modulation of LTP by estradiol was not statistically significant. The total density of spines was increased in CA1 pyramidal neurons, within 2 h after application of estradiol. The total density of thorns (postsynaptic spine-like structure) was, however, decreased by estradiol in CA3 pyramidal neurons. Both the increase of spines in CA1 and the decrease of thorns in CA3 were completely suppressed by Erk MAP kinase inhibitor. Only ERalpha agonist PPT induced the same enhancement/suppression effect as estradiol on both LTD and spinogenesis in CA1 and CA3. ERbeta agonist DPN induced completely different results. ERalpha localized in spines and presynapses of principal glutamatergic neurons in CA1, CA3 and DG. The same ERalpha was also located in nuclei and cytoplasm. Identification of ERalpha was successfully performed using purified RC-19 antibody. Non-purified ERalpha antisera, however, reacted significantly with unknown proteins, resulting in wrong immunostaining different from real ERalpha distribution. An issue of 'endocrine disrupters' (1-100 nM low dose of environmental chemicals), which are artificial xenoestrogenic or anti-xenoestrogenic substances, has emerged as a social and environmental problem. Endocrine disrupters were found to significantly modulate LTD and spinogenesis. Bisphenol A (BPA) and diethylstilbestrol (DES) enhanced LTD in CA1 and CA3. The total spine density was significantly increased by BPA and DES in CA1. Most probable receptors for BPA and DES may be Ralpha; however, other receptors might also be involved.

Estrogen synthesis in the brain--role in synaptic plasticity and memory.

Estrogen and androgen are synthesized from cholesterol locally in hippocampal neurons of adult animals. These neurosteroids are synthesized by cytochrome P450s and hydroxysteroid dehydrogenases (HSDs) and 5alpha-reductase. The expression levels of enzymes are as low as 1/200-1/50,000 of those in endocrine organs, however these numbers are high enough for local synthesis. Localization of P450(17alpha), P450arom, 17beta-HSD and 5alpha-reductase is observed in principal glutamatergic neurons in CA1, CA3 and the dendate gyrus. Several nanomolar levels of estrogen and androgen are observed in the hippocampus. Estrogen modulates memory-related synaptic plasticity not only slowly but also rapidly in the hippocampus. Rapid action of 17beta-estradiol via membrane receptors is demonstrated for spinogenesis and long-term depression (LTD). The enhancement of LTD by 1-10nM estradiol occurs within 1 h. The density of spine is increased in CA1 pyramidal neurons within 2h after application of estradiol. The density of spine-like structure is, however, decreased by estradiol in CA3 pyramidal neurons. ERalpha, but not ERbeta, induces the same enhancement/suppression effects on both spinogenesis and LTD.

Local production of sex hormones and their modulation of hippocampal synaptic plasticity.

It is believed that sex hormones are synthesized in the gonads and reach the brain via the blood circulation. In contrast with this view, the authors have demonstrated that sex hormones are also synthesized locally in the hippocampus and that these steroids act rapidly to modulate neuronal synaptic plasticity. The authors demonstrated that estrogens are locally synthesized from cholesterol through dehydroepiandrosterone and testosterone in adult hippocampal neurons. Significant expression of mRNA for P450(17alpha), P450arom, and other steroidogenic enzymes was demonstrated. Localization of P450(17alpha) and P450arom was observed in synapses of principal neurons. In contrast to the slow action of gonadal estradiol, hippocampal neuron-derived estradiol may act locally and rapidly within the neurons. For example, 1 to 10 nM estradiol rapidly enhances long-term depression (LTD). The density of thin spines is selectively increased within two hours upon application of estradiol in pyramidal neurons. Estrogen receptor ERalpha agonist has the same enhancing effect as estradiol on both LTD and spinogenesis. Localization of ERalpha in spines in addition to nuclei of principal neurons implies that synaptic ERalpha is responsible for rapid modulation of synaptic plasticity by endogenous estradiol. Activin A, a peptide sex hormone, may also play a role as a local endogenous modulator of synaptic plasticity.

Rapid Ca(2+) signaling induced by Bisphenol A in cultured rat hippocampal neurons.

OBJECTIVES: Bisphenol A (BPA) is a typical endocrine disrupter. We investigated the mechanisms of rapid Ca(2+) signaling induced by a low dose BPA application in cultured hippocampal neurons. MATERIALS AND METHODS: The primary culture of hippocampal neurons were prepared from postnatal 3 to 5-day-old rats. Cells were loaded with Calcium Green-1 fluorophore. Ca(2+) imaging and analysis were performed by Argus system. RESULTS: The application of BPA at 10-100 nM induced a transient increase in the intracellular Ca(2+) of N-methyl-D-aspartate (NMDA)-responsive neurons. The Ca(2+) transient occurred within 30 sec after the BPA application. The proportion of BPA-responsive neurons was 9.6 % and 8.5 % of the total NMDA-responsive neurons, respectively, upon 10 nM and 100 nM BPA application. The pre-treatment of neurons with Ca(2+) channel blockers, thapsigargin and nifedipine, considerably decreased the proportion of BPA-responsive neurons to 0.7 % and 3.7%, respectively. The treatment of neurons with an antagonist of estrogen receptor, ICI 182,780, also significantly decreased the proportion of BPA-responsive neurons down to 1.1 %. CONCLUSION: These results suggest that a low dose BPA application rapidly drives the Ca(2+) signaling system via activation of non-genomic pathway including estrogen receptors.

Hippocampal cytochrome P450s synthesize brain neurosteroids which are paracrine neuromodulators of synaptic signal transduction.

Hippocampal pyramidal neurons and granule neurons of adult male rats are equipped with a complete machinery for the synthesis of pregnenolone, dehydroepiandrosterone, 17beta-estradiol and testosterone as well as their sulfate esters. These brain neurosteroids are synthesized by cytochrome P450s (P450scc, P45017alpha and P450arom) from endogenous cholesterol. Synthesis is acutely dependent on the Ca(2+) influx attendant upon neuron-neuron communication via N-methyl-D-aspartate (NMDA) receptors. Pregnenolone sulfate, estradiol and corticosterone rapidly modulate neuronal signal transduction and the induction of long-term potentiation via NMDA receptors and putative membrane steroid receptors. Brain neurosteroids are therefore promising neuromodulators that may either activate or inactivate neuron-neuron communication, thereby mediating learning and memory in the hippocampus.

Rapid increase of spines by dihydrotestosterone and testosterone in hippocampal neurons: Dependence on synaptic androgen receptor and kinase networks.

Rapid modulation of hippocampal synaptic plasticity by locally synthesized androgen is important in addition to circulating androgen. Here, we investigated the rapid changes of dendritic spines in response to the elevation of dihydrotestosterone (DHT) and testosterone (T), by using hippocampal slices from adult male rats, in order to clarify whether these signaling processes include synaptic/extranuclear androgen receptor (AR) and activation of kinases. We found that the application of 10nM DHT and 10nM T increased the total density of spines by approximately 1.3-fold within 2h, by imaging Lucifer Yellow-injected CA1 pyramidal neurons. Interestingly, DHT and T increased different head-sized spines. While DHT increased middle- and large-head spines, T increased small-head spines. Androgen-induced spinogenesis was suppressed by individually blocking Erk MAPK, PKA, PKC, p38 MAPK, LIMK or calcineurin. On the other hand, blocking CaMKII did not inhibit spinogenesis. Blocking PI3K altered the spine head diameter distribution, but did not change the total spine density. Blocking mRNA and protein synthesis did not suppress the enhancing effects induced by DHT or T. The enhanced spinogenesis by androgens was blocked by AR antagonist, which AR was localized postsynaptically. Taken together, these results imply that enhanced spinogenesis by DHT and T is mediated by synaptic/extranuclear AR which rapidly drives the kinase networks. This article is part of a Special Issue entitled SI: Brain and Memory.

Estrogen receptor KO mice study on rapid modulation of spines and long-term depression in the hippocampus.

Rapid modulation of hippocampal synaptic plasticity through synaptic estrogen receptors is an essential topic. We analyzed estradiol-induced modulation of CA1 dendritic spines using adult male ERαKO and ERßKO mice. A 2h treatment of estradiol particularly increased the density of middle-head spines (diameter 0.3-0.4 µm) in wild type mouse hippocampal slices. The enhancement of spinogenesis was completely suppressed by MAP kinase inhibitor. Estradiol-induced increase in middle-head spines was observed in ERßKO mice (which express ERα), but not in ERαKO, indicating that ERα is necessary for the spinogenesis. Direct observation of the dynamic estradiol-induced enhancing effect on rapid spinogenesis was performed using time-lapse imaging of spines in hippocampal live slices from yellow fluorescent protein expressed mice. Both appearance and disappearance of spines occurred, and the number of newly appeared spines was significantly greater than that of disappeared spines, resulting in the net increase of the spine density within 2h. As another type of synaptic modulation, we observed that estradiol rapidly enhanced N-methyl-D-aspartate (NMDA)-induced long-term depression (LTD) in CA1 of the wild type mouse hippocampus. In contrast, estradiol did not enhance NMDA-LTD in ERαKO mice, indicating the involvement of ERα in the estrogen signaling. This article is part of a Special Issue entitled SI: Brain and Memory.

Aging-induced changes in sex-steroidogenic enzymes and sex-steroid receptors in the cortex, hypothalamus and cerebellum.

We investigated age-induced changes in mRNA expression profiles of sex-steroidogenic enzymes and sex-steroid receptors in 3-, 12-, and 24-month-old male rat brain subregions [cerebral cortex (CC), hypothalamus (Hy) and cerebellum (CL)]. In many cases, the expression levels of mRNA decreased with age for androgen synthesis enzyme systems, including Cyp17a1, Hsd17b and Srd5a in the CC and CL, but not in the Hy. Estradiol synthase Cyp19a1 did not show age-induced decline in the Hy, and nearly no expression of Cyp19a1 was observed in the CC and CL over 3-24 m. Androgen receptor Ar increased in the Hy but decreased in the CC with age. Estrogen receptor Esr1 increased in the CC and Hy, and did not change in the CL with age. Esr2 did not change in the CC and Hy, but decreased in the CL with age. As a comparison, age-induced changes of brain-derived neurotrophic factor mRNA were also investigated.

Corticosterone rapidly increases thorns of CA3 neurons via synaptic/extranuclear glucocorticoid receptor in rat hippocampus.

Modulation of synapses under acute stress is attracting much attention. Exposure to acute stress induces corticosterone (CORT) secretion from the adrenal cortex, resulting in rapid increase of CORT levels in plasma and the hippocampus. We tried to test whether rapid CORT effects involve activation of essential kinases as non-genomic processes. We demonstrated rapid effects (~1 h) of CORT on the density of thorns, by imaging Lucifer Yellow-injected neurons in adult male rat hippocampal slices. Thorns of thorny excrescences of CA3 hippocampal neurons are post-synaptic regions whose presynaptic partners are mossy fiber terminals. The application of CORT at 100, 500, and 1000 nM induced a rapid increase in the density of thorns in the stratum lucidum of CA3 pyramidal neurons. Co-administration of RU486, an antagonist of glucocorticoid receptor (GR), abolished the effect of CORT. Blocking a single kinase, including MAPK, PKA, or PKC, suppressed CORT-induced enhancement of thorn-genesis. On the other hand, GSK-3ß was not involved in the signaling of thorn-genesis. Blocking AMPA receptors suppressed the CORT effect. Expression of CA3 synaptic/extranuclear GR was demonstrated by immunogold electron microscopic analysis. From these results, stress levels of CORT (100-1000 nM) might drive the rapid thorn-genesis via synaptic/extranuclear GR and multiple kinase pathways, although a role of nuclear GRs cannot be completely excluded.

Modulation of synaptic plasticity in the hippocampus by hippocampus-derived estrogen and androgen.

The hippocampus synthesizes estrogen and androgen in addition to the circulating sex steroids. Synaptic modulation by hippocampus-derived estrogen or androgen is essential to maintain healthy memory processes. Rapid actions (1-2h) of 17ß-estradiol (17ß-E2) occur via synapse-localized receptors (ERα or ERß), while slow genomic E2 actions (6-48h) occur via classical nuclear receptors (ERα or ERß). The long-term potentiation (LTP), induced by strong tetanus or theta-burst stimulation, is not further enhanced by E2 perfusion in adult rats. Interestingly, E2 perfusion can rescue corticosterone (stress hormone)-induced suppression of LTP. The long-term depression is modulated rapidly by E2 perfusion. Elevation of the E2 concentration changes rapidly the density and head structure of spines in neurons. ERα, but not ERß, drives this enhancement of spinogenesis. Kinase networks are involved downstream of ERα. Testosterone (T) or dihydrotestosterone (DHT) also rapidly modulates spinogenesis. Newly developed Spiso-3D mathematical analysis is used to distinguish these complex effects by sex steroids and kinases. It has been doubted that the level of hippocampus-derived estrogen and androgen may not be high enough to modulate synaptic plasticity. Determination of the accurate concentration of E2, T or DHT in the hippocampus is enabled by mass-spectrometric analysis in combination with new steroid-derivatization methods. The E2 level in the hippocampus is approximately 8nM for the male and 0.5-2nM for the female, which is much higher than that in circulation. The level of T and DHT is also higher than that in circulation. Taken together, hippocampus-derived E2, T, and DHT play a major role in modulation of synaptic plasticity.

Nanomolar dose of bisphenol A rapidly modulates spinogenesis in adult hippocampal neurons.

We demonstrated the rapid effects of 10nM bisphenol A (BPA) on the spinogenesis of adult rat hippocampal slices. The density of spines was analyzed by imaging Lucifer Yellow-injected CA1 neurons in slices. Not only the total spine density but also the head diameter distribution of spine was quantitatively analyzed. Spinogenesis was significantly enhanced by BPA within 2h. In particular, the density of middle-head spine (with head diameter of 0.4-0.5µm) was significantly increased. Hydroxytamoxifen, an antagonist of both estrogen-related receptor gamma (ERRγ) and estrogen receptors (ERα/ERß), blocked the BPA-induced enhancement of the spine density. However, ICI 182,780, an antagonist of ERα/ERß, did not suppress the BPA effects. Therefore, ERRγ is deduced to be a high affinity receptor of BPA, responsible for modulation of spinogenesis. The BPA-induced enhancement of spinogenesis was also suppressed by MAP kinase inhibitor, PD98059, and the blocker of NMDA receptors, MK-801. Washout of BPA for additional 2h after 2h BPA treatment abolished the BPA-induced enhancement of spinogenesis, suggesting that the BPA effect was reversible. ERRγ was localized at synapses as well as cell bodies of principal neurons. ERRγ at synapses may contribute to the observed rapid effect. The level of BPA in the hippocampal slices was determined by mass-spectrometric analysis.

Corticosterone induces rapid spinogenesis via synaptic glucocorticoid receptors and kinase networks in hippocampus.

BACKGROUND: Modulation of dendritic spines under acute stress is attracting much attention. Exposure to acute stress induces corticosterone (CORT) secretion from the adrenal cortex, resulting in rapid increase of CORT levels in plasma and the hippocampus. METHODOLOGY/PRINCIPAL FINDINGS: Here we demonstrated the mechanisms of rapid effect (∼1 h) of CORT on the density and morphology of spines by imaging neurons in adult male rat hippocampal slices. The application of CORT at 100-1000 nM induced a rapid increase in the density of spines of CA1 pyramidal neurons. The density of small-head spines (0.2-0.4 µm) was increased even at low CORT levels (100-200 nM). The density of middle-head spines (0.4-0.5 µm) was increased at high CORT levels between 400-1000 nM. The density of large-head spines (0.5-1.0 µm) was increased only at 1000 nM CORT. Co-administration of RU486, an antagonist of glucocorticoid receptor (GR), abolished the effect of CORT. Blocking a single kinase, such as MAPK, PKA, PKC or PI3K, suppressed CORT-induced enhancement of spinogenesis. Blocking NMDA receptors suppressed the CORT effect. CONCLUSIONS/SIGNIFICANCE: These results imply that stress levels of CORT (100-1000 nM) drive the spinogenesis via synaptic GR and multiple kinase pathways.

Endogenous synthesis of corticosteroids in the hippocampus.

BACKGROUND: Brain synthesis of steroids including sex-steroids is attracting much attention. The endogenous synthesis of corticosteroids in the hippocampus, however, has been doubted because of the inability to detect deoxycorticosterone (DOC) synthase, cytochrome P450(c21). METHODOLOGY/PRINCIPAL FINDINGS: The expression of P450(c21) was demonstrated using mRNA analysis and immmunogold electron microscopic analysis in the adult male rat hippocampus. DOC production from progesterone (PROG) was demonstrated by metabolism analysis of (3)H-steroids. All the enzymes required for corticosteroid synthesis including P450(c21), P450(2D4), P450(11ß1) and 3ß-hydroxysteroid dehydrogenase (3ß-HSD) were localized in the hippocampal principal neurons as shown via in situ hybridization and immunoelectron microscopic analysis. Accurate corticosteroid concentrations in rat hippocampus were determined by liquid chromatography-tandem mass spectrometry. In adrenalectomized rats, net hippocampus-synthesized corticosterone (CORT) and DOC were determined to 6.9 and 5.8 nM, respectively. Enhanced spinogenesis was observed in the hippocampus following application of low nanomolar (10 nM) doses of CORT for 1 h. CONCLUSIONS/SIGNIFICANCE: These results imply the complete pathway of corticosteroid synthesis of 'pregnenolone →PROG→DOC→CORT' in the hippocampal neurons. Both P450(c21) and P450(2D4) can catalyze conversion of PROG to DOC. The low nanomolar level of CORT synthesized in hippocampal neurons may play a role in modulation of synaptic plasticity, in contrast to the stress effects by micromolar CORT from adrenal glands.

Regulation of synaptic plasticity by hippocampus synthesized estradiol.

Estradiol is synthesized from cholesterol in hippocampal neurons of adult rats by cytochrome P450 and hydroxysteroid dehydrogenase enzymes. These enzymes are expressed in the glutamatergic neurons of the hippocampus. Surprisingly, the concentration of estradiol and androgen in the hippocampus is significantly higher than that in circulation. Locally synthesized estradiol rapidly and potently modulates synaptic plasticity within the hippocampus. E2 rapidly potentiates long-term depression and induces spinogenesis through synaptic estrogen receptors and kinases. The rapid effects of estradiol are followed by slow genomic effects mediated by both estrogen receptors located at the synapse and nucleus, modulating long-term potentiation and promoting the formation of new functional synaptic contacts. Age-related changes in hippocampally derived estradiol synthesis and distribution of estrogen receptors may alter synaptic plasticity, and could potentially contribute to age-related cognitive decline. Understanding factors which regulate hippocampal estradiol synthesis could lead to the identification of alternatives to conventional hormone therapy to protect against age-related cognitive decline.