Roles of estrogen receptors alpha and beta in sexually dimorphic neuroprotection against glutamate toxicity.

Although most agree that 17ß-estradiol is neuroprotective via a variety of mechanisms, less is known about the role that biological sex plays in receptor-mediated estradiol neuroprotection. To address this issue we isolated primary cortical neurons from rat pups sorted by sex and assessed the ability of estradiol to protect the neurons from death induced by glutamate. Five-minute pretreatment with 10-50 nM 17ß-estradiol protected female but not male neurons from glutamate toxicity 24 h later. Both estrogen receptor alpha (ERα) and estrogen receptor beta (ERß) are expressed in these cultures. Experiments using an ERα selective agonist or antagonist indicate that this receptor is important for neuroprotection in female cortical neurons. The ERß selective agonist conveys a small degree of neuroprotection to both male and female cortical neurons. Interestingly, we found that 17α estradiol and the novel membrane estrogen receptor (mER) agonist STX, but not bovine serum albumin conjugated estradiol or the GPR30 agonist G1 were neuroprotective in both male and female neurons. Taken together these data highlight a role for ERα in sexually dimorphic neuroprotection.

Estrogen induces rapid translocation of estrogen receptor beta, but not estrogen receptor alpha, to the neuronal plasma membrane.

Estrogen receptors can activate transcription in the nucleus, and activate rapid signal transduction cascades in the cytosol. Multiple reports identify estrogen receptors at the plasma membrane, while others document the dynamic responses of estrogen receptor to ligand binding. However, the function and identity of membrane estrogen receptors remain controversial. We have used confocal microscopy and cell fractionation on the murine hippocampus-derived HT22 cell line and rat primary cortical neurons transfected with estrogen receptor-green fluorescent protein constructs to address the membrane localization of these receptors. We observe translocation of estrogen receptor beta (beta) to the plasma membrane 5 min after exposure to 17beta-estradiol, whereas estrogen receptor alpha (alpha) localization remains unchanged. Membrane localization of estrogen receptor beta is transient, selective for 17beta-estradiol, and is not blocked by ICI182,780. Inhibition of the mitogen-activated protein kinase pathway does not block estrogen-mediated estrogen receptor beta membrane translocation, and in fact prolongs membrane localization. These data suggest that while both estrogen receptor alpha and estrogen receptor beta can be present at the neuronal membrane, their presence is differentially regulated.

17-Beta estradiol rapidly enhances extracellular signal-regulated kinase 2 phosphorylation in the rat brain.

Physiological doses of 17-beta Estradiol (E2) rapidly induce mitogen-activated protein kinase (MAPK) phosphorylation in a variety of cell culture and tissue explant preparations. Rapid MAPK phosphorylation has been implicated as a critical step in estrogen's effects on neuronal activity, gene transcription and neuroprotection. The present series of in vivo experiments were designed to determine whether acute administration of estrogen rapidly increased extracellular signal-regulated protein kinase (ERK) 2 phosphorylation. Brains were harvested 20 min after a single i.p. injection of 15 microg/kg of 17-beta or 17-alpha estradiol. Twelve brain structures were micro-dissected, homogenized and processed for Western blotting. E2-treated rats exhibited a statistically significant increase in ERK2 phosphorylation in the diagonal band of Broca, rostral nucleus accumbens, paraventricular nucleus, arcuate nucleus and anteromedial visual cortex. Administration of the same dose of 17-alpha estradiol did not enhance ERK phosphorylation in any of the brain regions examined. The in vivo data presented here extend previously published in vitro data indicating that E2 rapidly activates MAPK in primary neuronal cultures, explants and cell lines. These data also indicate that MAPK activation is a potential mediator of estrogens effects in some but not all estrogen receptor containing regions of the brain.

Retinal innervation of calbindin-D28K cells in the hamster suprachiasmatic nucleus: ultrastructural characterization.

The authors have described a subregion of the hamster hypothalamic suprachiasmatic nucleus (SCN) containing cells that are immunopositive for the cytosolic calcium-binding protein, Calbindin-D28K (CaBP). Several lines of evidence indicate that this region may constitute the site of the pacemaker cells that are responsible for the regulation of circadian locomotor rhythms. First, 79% of the CaBP-immunoreactive (ir) neurons express Fos in response to photic stimulation, indicating that they are close to or part of the input pathway to pacemakers. Second, at the light microscopy level, retinal terminals innervate the CaBP subnucleus. Finally, destruction of this subnucleus renders animals arrhythmic in locomotor activity. In this study, the authors examined the ultrastructural relationship between cholera toxin (CTbeta) labeled retinal fibers and the CaBP-ir subregion within the hamster SCN. CTbeta-ir retinal terminals make primarily axo-somatic, symmetric, synaptic contacts with CaBP-ir perikarya. In addition, retinal terminals form synapses with CaBP processes as well as with unidentified profiles. There are also complex interactions between retinal terminals, CaBP perikarya, and unidentified profiles. Given that axo-somatic synaptic input has a more potent influence on a cell's electrical activity than does axo-dendritic synaptic input, cells of the CaBP subregion of the SCN are ideally suited to respond rapidly to photic stimulation to reset circadian pacemakers.

The effects of active and passive stimulation on chemosensory event-related potentials.

During normal olfaction, stimulation is dependent upon nasal inhalation. When collecting chemosensory event-related potentials (CSERPs), inhalations may produce artifacts such as respiration-related brain potentials that confound interpretation of the data. To avoid this type of artifact, CSERPs have often been collected using stimulation that is independent of respiratory cycle. It is possible, however, that inspirations prime the olfactory tract for odor input, and traditional CSERP data acquisition techniques, obscure this neural preparation. To investigate this question, twelve subjects were tested using two different approaches to stimulation. Odorants (butanol 2% and 4%) were introduced into a warmed and humidified air stream and delivered to one nostril through a cannula. In one condition, subjects mouth-breathed and stimuli occurred asynchronously with respiratory cycle (passive). In the other condition, subjects inhaled through the nose and stimuli were delivered synchronously with nasal inhalations (active). CSERP data were collected from thirty scalp sites for 3 s following stimulation. Data were corrected for eye movements, smoothed, and averaged. Maximal amplitudes for an early negativity (N1) and later positivity (P2) were determined and submitted to separate analyses of variance. These analyses indicated that administration technique interacted with both odor concentration and recording site. Additionally, amplitude of P2 was greater in the passive condition. Such results suggest that the two administration techniques produce different neural processing of olfactory stimuli and that the passive technique may be better suited for determination of the integrity of the olfactory tract for single subjects because of its greater amplitude.