Abstract

The ramifications of endocrine and neural senescence converge in the hippocampus, particularly with respect to glutamatergic synapses. In this review, we will focus on current literature suggesting that potential synaptic alterations induced by estrogen in the hippocampus are mediated through interactions between ER-α and NMDA receptors. In addition, we will examine the data suggesting that these interactions may be uncoupled with aging. These studies demonstrate that while estrogen helps retain a youthful synaptic phenotype by some measures, the aged synapse may differ from the young synapse in several key respects that impact plasticity in general, and endocrine influences on the synapse, in particular.

Recent evidence indicates that changes in sex steroid hormones affect not only the hypothalamic circuits directly involved in reproductive behavior and physiology, but also brain regions and circuits that mediate cognitive functions, such as the hippocampal formation. Estrogens regulate multiple aspects of synaptic plasticity in the hippocampus, and these effects may influence memory. Rats given estrogen replacement following an ovariectomy show an increase in the number of spines and synaptic boutons in the CA1 region of the hippocampus relative to ovariectomized animals that lack estrogen replacement (Gould et al., 1990; Woolley and McEwen, 1992, 1993; Woolley et al., 1996). In addition, fluctuations in the dendritic spine density appear during the estrous cycle of the rat: dendritic spine density on CA1 pyramidal neurons decreases on days when levels of estrogen are low, i.e. proestrus, as compared to days when estrogen levels are higher, i.e. estrus (Woolley et al., 1990; Woolley and McEwen, 1992).

Current neurobiological evidence suggests that one mechanism by which estrogen alters synaptic plasticity is through N-methyl-D-aspartate (NMDA)-type glutamate receptors. Glutamate receptors are the primary mediators of excitatory transmission in the central nervous system (Hollmann and Heinemann, 1994). They play an important role in learning and memory, and have the capacity to mediate cell death under certain conditions, implicating them in several devastating neurodegenerative disorders and age-related functional decline (Olney, 1983; Rothman, 1984; Morris et al., 1986; Choi, 1988; Bliss and Collingridge, 1993; Lipton and Rosenbeerg, 1995). Pharmacologically, glutamate receptors can be divided into several classes, one metabotropic (G-protein linked) and three classes of ionotropic: NMDA, alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA), and kainate (KA) receptors (Hollmann and Heinemann, 1994). They are each likely to function differently in mediating excitation, learning and memory, and excitotoxicity. Each of the defined classes of glutamate receptors are composed of multimeric assemblies of protein subunits (Hollmann and Heinemann, 1994), with NR1 an obligatory subunit (NMDAR1), and functional attributes of the receptor complex partially dependent on which NR2 subunits are present.

Estradiol appears to have a direct effect on hippocampal NMDA receptors. For example, it increases NMDA agonist binding in the CA1 region of the hippocampus, but has no effect on AMPA receptors (Weiland, 1992). In addition, NMDA receptor antagonists block the estrogen-induced increase in dendritic spine density, and NMDA receptor dependent long-term potentiation is enhanced on days of proestrus, i.e. when estrogen levels area high (Woolley and McEwen, 1994; Warren et al., 1995; Murphy and Segal, 1996). A study in our laboratory using confocal laser scanning microscopy, demonstrated a 30% increase in NMDAR1 fluorescence intensity in the dendrites of CA1 pyramidal neurons with estrogen replacement (Gazzaley et al., 1996).

In summary, these data demonstrate conclusively that estrogens regulate synaptic plasticity in the young hippocampus, reinforcing the fact that sex steroids such as estrogen can directly impact regions and circuits underlying cognitive function, such as the hippocampal formation. The data summarized above are all from young rats, however, what role does estrogen play in age-related cognitive impairment? While many studies have addressed the effects of estrogen deprivation and replacement in hypothalamus in both young and aged animals (Wise and Ratner, 1980; Steger et al., 1983; Gee et al., 1984; Mobbs et al., 1984; Rubin et al., 1985; Caraty and Locatelli, 1988; Belisle et al., 1990; Hwang et al., 1990; Joshi et al., 1995), few studies have addressed the differential effects of estrogen and age on the hippocampus (Miranda et al., 1999). Thus, we proposed the following questions: (i) Does estrogen increase axospinous synapse density on CA1 pyramidal cells in aged rats in a manner similar to young animals? (ii) Is the dendritic increase in NMDAR1 manifested at the synapse? (iii) If so, is the synaptic effect equivalent in young and aged rats?

To answer these questions, we used quantitative ultrastructural approaches to reveal age-related differences in estrogen-induced synaptic plasticity in the CA1 region of the hippocampus [see Fig. 3 legend for animal ages and numbers (Adams et al., 2001)]. An analysis of axospinous synapse density revealed that while estrogen increases synapse density in the young hippocampus in a manner similar to that described by other investigators (Gould et al., 1990; Woolley et al., 1990; Woolley and McEwen, 1992, 1993; Woolley et al., 1996), it fails to increase it in the aged hippocampus (Adams et al., 2001). Moreover, there appears to be an overall loss of axospinous synapses that is occurring in the aged CA1. In order to determine whether NMDAR1 levels are shifting at the synapse we used post-embedding immunogold (Fig. 1A,B), and quantified the distribution of each gold particle using software developed in our laboratory (i.e. SYNBIN; Fig. 2), on the basis of principles regarding proximity to membranes articulated by Ottersen and colleagues (Blackstad et al., 1990; Ruud and Blackstad, 1999). Accordingly, the position of each gold particle (Fig. 2A) is determined as it relates to the post- and presynaptic membrane structures (Fig. 2B). The program analyzes the resulting data map (Fig. 2C) and objectively assigns each gold particle to a given bin (Fig. 2D), with bin sizes and targeted synaptic domains established prospectively. Through this process, a precise gold particle/bin density emerges that is an accurate reflection of gold particle distribution and density in different compartments of the synaptic complex (Adams et al., 2001). Only synapses that contained two or more gold particles associated with the postsynaptic density were considered in the current analyses. Moreover, minimal mitochondrial labeling was observed and controls omitting the primary antibody were also performed to assess the specificity of the secondary antibody.

We found that estrogen does not alter the expression of NMDAR1 per synapse in the young CA1 synapse, however, it does affect the aged CA1 synapse, increasing the amount of NMDAR1 per synapse in the aged animal to that of a young animal (Adams et al., 2001). Therefore, we concluded that the increase in dendritic NMDAR1 revealed with immunofluorescence (Gazzaley et al., 1996) serves to provide adequate NMDA receptors to the additional axospinous synapses induced by estrogen replacement therapy (ERT) in young female rats, but does not alter the NMDA receptor profile at the synapse in young animals. With age there is a loss of CA1 synapses that is not reversible by ERT, yet these synapses still respond to estrogen through increasing NMDA receptors at the synapse. Moreover, these changes in NMDA receptor levels are occurring in the absence of alterations in postsynaptic density lengths (Adams et al., 2001). Thus, while estrogen impacts the aged CA1 synapse in a manner that might help preserve hippocampal function, it does so in the context of a synaptic density compromised by age (Adams et al., 2001). In addition, an estrogen-induced increase in synaptic NMDA receptors may have implications for excitotoxicity that need to be explored further.

What is it about the aged CA1 spine that makes it less responsive to ERT? To this end we focused on the role of the estrogen receptor (ER). Recent evidence demonstrated that both estrogen receptor-alpha (ER-α) and estrogen receptor-beta (ER-β) are expressed in the hippocampus (Li et al., 1997; Shughrue et al., 1997; Weiland et al., 1997; Pau et al., 1998; Petersen et al., 1998; Register et al., 1998; Shughrue et al., 1998; Shughrue and Merchenthaler, 2000; Brinton, 2001). Additionally, Milner et al. (2001) found ultrastructural evidence for the localization of ER-α within dendritic spines in CA1. These data suggest that ER-α may act locally, as well as by regulating nuclear transcription. The potential local, non-genomic effects related to plasticity include regulating neurotransmitter release, signaling cascades, and mRNA translation (McEwen, 2002).

We performed a quantitative ultrastructural analysis of synaptic ER-α in the same animals that were analyzed for synapse density and NMDAR1 localization (Adams et al., 2002). Using postembedding immunogold we observed that the percentage of ER-α labeled synapses decreases with age, independent of estrogen status, by ∼50% (Adams et al., 2002). In addition, ERT led to a decrease in ER-α in both the terminal and the spine in young CA1, but did not change ER-α distribution in the aged spine. Thus, in young animals, peri-synaptic ER-α in CA1 is dynamic and responsive to circulating estrogen levels, but with aging, responsiveness to circulating estrogen decreases (Adams et al., 2002). These highly localized shifts in ER-α regulation and distribution may contribute to age-related decreases in synaptic plasticity. In addition, recent data suggests that the combined effects of aging and ovarian hormones can impact basal forebrain cholinergic function, and this may augment the functional impact of our observation in the aged animals (Gibbs, 2003).

While the study from our laboratory (Adams et al., 2002) confirms a previous study (Milner et al., 2001) demonstrating that ER-α-IR is present within the presynaptic terminal and spines of CA1 pyramidal neurons, and also that it is responsive to estrogen in young animals and decreases with aging, it is important to note that the distribution and nature of estrogen receptors in the hippocampal formation is far from conclusive. Interestingly, most of the ER-α-IR is within extra-synaptic membranes, the spine cytoplasm or associated with presynaptic vesicles, rather than within the post-synaptic density. It will be of great importance to determine the identity of proteins that are co-localized and potentially interact with the spinous ER-α-IR with greater precision. For example, it will be important to determine the degree to which the ER-α-IR is sequestered in membranous caveolae-like structures as has been hypothesized (Toran-Allerand, 2000), since such a localization would position ER-α to interact with the multiple signal transduction pathways that have been hypothesized to function within such an organelle in non-nervous system tissues (Schlegel et al., 1999). A critical role for ER-α in such caveolae was recently demonstrated in endothelial cells, in that caveolar ER-α was functionally and biochemically linked to nitric oxide synthase (NOS) in a critical signaling module capable of regulating the local calcium levels. Such a link between ER-α and NOS could have profound local effects on synaptic plasticity in CA1. However, the substrate for such a complex is hypothetical at present, since calveolin and calveolin-containing structures do not appear to be present in brain, though there may be a structural and functional homologue involving flotillin-anchored calveolae that may be a site for ER-α or a putative homologous ER (Kelly and Levin, 2001), that may be recognized by ER-α antibodies referred to by Toran-Allerand (2000) as ER-X. While such a scenario is compelling, given the rich environment that it would offer regarding local effects of ER-α or its homologue, its characterization in the nervous system in general and CA1 in particular will require extensive additional ultrastructural and biochemical analyses.

Conclusions

Our data suggests that there are potential synaptic alterations induced by estrogen in CA1 mediated through interactions between ER-α and NMDA receptors, and this interaction may be uncoupled with aging (Fig. 3A–D). In young animals, NMDAR1 levels per synapse are consistent across estrogen treatment groups (Fig. 3A,B). Thus, we interpret the increased levels of NMDAR1 in dendritic shafts (Gazzaley et al., 1996) as required to maintain normal levels of NMDAR1 in the new synapses. Furthermore, in the young animal, we hypothesize that only synapses that contain ER-α and NMDA receptors respond to estrogen through formation of multiple synaptic contacts. In contrast, in the aged animals, there is a dramatic decline in the number of spines/synapses that contain ER-α and thus a blunted capacity to form new synapses (Fig. 3C,D). In addition the aged synapse has less NMDAR1 per synapse in the absence of estrogen, yet retains a youthful NMDA receptor profile in the presence of estrogen (Fig. 3C,D). However, given the decrease in ER-α in the aged animal, we must consider that this NMDA receptor response to estrogen is mediated through some mechanism other than ER-α such as, perhaps, ER-β.

While these cellular and synaptic data are compelling with respect to estrogen’s effect on hippocampus and may be relevant to age-related cognitive decline, their relevance to hormone replacement in women is unclear at this time. First, endocrine senescence in rats is quite different from humans, with rodents entering into what is referred to as estropause with the cyclicity becoming irregular and then halted. Unlike humans, this estropause is not accompanied by a dramatic loss of estrogen (Huang HH et al., 1978; Lu et al., 1979; Wise and Ratner, 1980). Thus, the primate model may be a critical intermediate for relating the cellular and synaptic rodent data to human, though little data from non-human primates are presently available. First, like the human, the rhesus monkey has a 28 day menstrual cycle and experiences menopause with reproductive senescence, though it is later in life relative to humans. Recently our laboratory demonstrated that estrogen increased the spine density in the CA1 region of both young and aged female rhesus monkeys (Hao et al., 2002). Thus, unlike the rat, the CA1 region of the aged monkey hippocampus is responsive to estrogen replacement at the level of increased spine and synapse number.

In summary, estrogen impacts the aging synapse at the cellular and molecular level within cells and circuits that are involved in learning and memory. Additionally, estrogen affects performance on learning and memory tasks (Berry et al., 1997; Packard and Teather, 1997; Roberts et al., 1997; Sherwin, 1997; Stackman et al., 1997; Warren and Jurska, 1997; Rapp et al., 2003). Preliminary evidence suggests that primates and rodents may respond differently to ERT, but in both cases there is evidence to suggest that estrogen impacts the aging synapse. Thus, future studies need to be directed at integrating cellular/molecular data and behavior, and determining whether estrogen can help retain a youthful phenotype of plasticity in the aging synapse in both rodents and primates, and the impact of such synaptic effects on behavior. Such data will have important implications for ERT in humans particularly with respect to cognitive function.

Supported by NIH grant AG 16765.

Figure 1. Images of NMDAR1-positive asymmetric synapses between axon terminals and dendritic spines (*) in the CA1 subfield of the hippocampus (A, B). Immunogold particles, indicated by the arrowheads, were observed to be associated with the postsynaptic density. Immunogold particles were also observed within the head of the spine, which may represent a non-synaptic pool. Scale bar = 250 nm.

Figure 1. Images of NMDAR1-positive asymmetric synapses between axon terminals and dendritic spines (*) in the CA1 subfield of the hippocampus (A, B). Immunogold particles, indicated by the arrowheads, were observed to be associated with the postsynaptic density. Immunogold particles were also observed within the head of the spine, which may represent a non-synaptic pool. Scale bar = 250 nm.

Figure 2. Schematic images that illustrate the software developed in our laboratory for quantification of the distribution of gold particles associated with the synaptic complex. (A) Schematic diagram illustrates the possible distribution of gold particles within a synapse. (B) The user outlines the relevant synaptic components [i.e. the length of the postsynaptic density (red line), the length of the presynaptic membrane that corresponds to the length of the postsynaptic density (green line), and the location of the gold particles (yellow closed circles)]. (C) The computer program provides a data map of the distribution of the gold particles relative to these membranes and postsynaptic compartments. (D) Table representing the different synaptic compartments or bins that are associated with the synaptic complex.

Figure 2. Schematic images that illustrate the software developed in our laboratory for quantification of the distribution of gold particles associated with the synaptic complex. (A) Schematic diagram illustrates the possible distribution of gold particles within a synapse. (B) The user outlines the relevant synaptic components [i.e. the length of the postsynaptic density (red line), the length of the presynaptic membrane that corresponds to the length of the postsynaptic density (green line), and the location of the gold particles (yellow closed circles)]. (C) The computer program provides a data map of the distribution of the gold particles relative to these membranes and postsynaptic compartments. (D) Table representing the different synaptic compartments or bins that are associated with the synaptic complex.

Figure 3. Schematic diagram illustrating the potential sites of synaptic alterations induced by estrogen in the CA1 region of young [A, B; 3- to 4-month-old female Sprague–Dawley rats; n = 4–5 per group (Adams et al., 2001)] and aged animals [C, D; 24-month-old female Sprague–Dawley rats; n = 4–5 per group (Adams et al., 2001)]. Estrogen treatment increases NMDAR1 expression per synapse in aged hippocampus, whereas it increases spine number but not synaptic NMDAR1 in young female rat hippocampus. Estrogen treatment either decreases the amount of ER-α per synapse in young animals or leads to a redistribution of the receptor resulting from the formation of new dendritic spines and alterations in presynaptic terminal shape. Presynaptic terminal numbers remain unchanged in the young animal that is given estrogen replacement and they form multiple synapse boutons with new spines (Woolley et al., 1996; Yankova et al., 2001). Moreover, the presynaptic terminals become irregular in shape and less spherical (Woolley et al., 1996). ER-α levels are unchanged in aged animals in both presynaptic and postsynaptic compartments, but there is an overall decrease in the percentage of terminals that express ER-α. The events depicted in this schematic are occurring in the absence of changes in the lengths of the postsynaptic densities [average length for all animals approximately 250 nm (Adams et al., 2001)]. Small black dots represent immunogold particles labeling NMDAR1 associated with postsynaptic Bin 1, small red dots represent immunogold particles labeling ER-α, open circles represent synaptic vesicles, and the gray zones represent the postsynaptic density. OVX = ovariectomized; Veh = vehicle-treated; E2 = estrogen-replaced.

Figure 3. Schematic diagram illustrating the potential sites of synaptic alterations induced by estrogen in the CA1 region of young [A, B; 3- to 4-month-old female Sprague–Dawley rats; n = 4–5 per group (Adams et al., 2001)] and aged animals [C, D; 24-month-old female Sprague–Dawley rats; n = 4–5 per group (Adams et al., 2001)]. Estrogen treatment increases NMDAR1 expression per synapse in aged hippocampus, whereas it increases spine number but not synaptic NMDAR1 in young female rat hippocampus. Estrogen treatment either decreases the amount of ER-α per synapse in young animals or leads to a redistribution of the receptor resulting from the formation of new dendritic spines and alterations in presynaptic terminal shape. Presynaptic terminal numbers remain unchanged in the young animal that is given estrogen replacement and they form multiple synapse boutons with new spines (Woolley et al., 1996; Yankova et al., 2001). Moreover, the presynaptic terminals become irregular in shape and less spherical (Woolley et al., 1996). ER-α levels are unchanged in aged animals in both presynaptic and postsynaptic compartments, but there is an overall decrease in the percentage of terminals that express ER-α. The events depicted in this schematic are occurring in the absence of changes in the lengths of the postsynaptic densities [average length for all animals approximately 250 nm (Adams et al., 2001)]. Small black dots represent immunogold particles labeling NMDAR1 associated with postsynaptic Bin 1, small red dots represent immunogold particles labeling ER-α, open circles represent synaptic vesicles, and the gray zones represent the postsynaptic density. OVX = ovariectomized; Veh = vehicle-treated; E2 = estrogen-replaced.

References

Adams MM, Shah RA, Janssen WGM, Morrison JH (
2001
) Different modes of hippocampal plasticity in response to estrogen in young and aged female rats.
Proc Natl Acad Sci USA
 
98
:
8071–
8076.
Adams MM, Fink SE, Shah RA, Janssen WGM, Hayashi S, Milner TA, McEwen BS, Morrison JH (
2002
) Estrogen and aging affect the subcellular distribution of estrogen receptor-alpha in the hippocampus of female rats.
J Neurosci
 
22
:
3608–
3614.
Belisle S, Bellabarba D, Lehoux J-G (
1990
) Hypothalamic–pituitary axis during reproductive aging in mice.
Mech Ageing Dev
 
52
:
207–
217.
Berry B, McMahan R, Gallagher M (
1997
) Spatial learning and memory at defined points of the estrous cycle: effects on performance of a hippocampal-dependent task.
Behav Neurosci
 
111
:
267–
274.
Blackstad TW, Karagülle T, Ottersen OP (
1990
) MORFOREL, a computer program for two-dimensional analysis of micrographs of biological specimens, with emphasis on immungold preparations.
Comput Biol Med
 
20
:
15–
34.
Bliss TVP, Collingridge GL (
1993
) A synaptic model of memory: long-term potentiation in the hippocampus.
Nature
 
361
:
31–
39.
Brinton RD (
2001
) Cellular and molecular mechanisms of estrogen regulation of memory function and neuroprotection against Alzheimer’s disease: recent insights and remaining challenges.
Learn Mem
 
8
:
121–
133.
Caraty A, Locatelli A (
1988
) Effect of time after castration on secretion of LHRH and LH in the ram.
Reprod Fertil
 
82
:
263–
269.
Choi DW (
1988
) Glutamate neurotoxicity and diseases of the nervous system.
Neuron
 
1
:
623–
634.
Gazzaley AH, Weiland NG, McEwen BS, Morrison JH (
1996
) Differential regulation of NMDAR1 mRNA and protein by estradiol in the rat hippocampus.
J Neurosci
 
16
:
6830–
6838.
Gee DM, Flurkey K, Mobbs CV, Sinha YN, Finch CE (
1984
) The regulation of luteinizing hormone and prolactin in C57BL/6J mice: effects of estradiol implant size, duration of ovariectomy, and aging.
Endocrinology
 
114
:
685–
693.
Gibbs RB (
2003
) Effects of ageing and long-term hormone replacement on cholinergic neurones in the medial septum and nucleus basalis magnocellularis of ovariectomized rats.
J Neuroendocrinol
 
15
:
477–
485.
Gould E, Woolley CS, Frankfurt M, McEwen BS (
1990
) Gonadal steroids regulate dendritic spine density in hippocampal pyramidal cells in adulthood.
J Neurosci
 
4
:
1286–
1291.
Hao J, Jansse WGM, Tang Y, Roberts J, Hof PR, Morrison JH (
2002
) Estrogen induces formation of dendritic spines in hippocampus of young and aged female rhesus monkeys.
Soc Neurosci Abstr
  368.12.
Hollmann M, Heinemann S (
1994
) Cloned glutamate receptors.
Annu Rev Neurosci
 
17
:
31–
108.
Huang HH SR, Bruni JF, Meites J (
1978
) Patterns of sex steroid and gonadotropin secretion in aging female rats.
Endocrinology
 
103
:
1855–
1859.
Hwang C, Pu H-F, Hwang C-Y, Liu J-Y, Yao H-C, Tung Y-F, Wang PS (
1990
) Age-related differences in the release of luteinizing hormone and gonadotropin-releasing hormone in ovariectomized rats.
Neuroendocrinology
 
52
:
127–
132.
Joshi D, Billiar RB, Miller MM (
1995
) Luteinizing hormone response to N-methyl-D,L-aspartic acid in the presence of physiological estradiol concentrations: influence of age and the ovary.
Proc Soc Exp Biol Med
 
209
:
237–
244.
Kelly MJ, Levin ER (
2001
) Rapid actions of plasma membrane estrogen receptors.
Trends Endocrinol Metab
 
12
:
152–
156.
Li X, Schwartz PE, Rissman EF (
1997
) Distribution of estrogen receptor-β-like immunoreactivity in rat forebrain.
Neuroendocrinology
 
66
:
63–
67.
Lipton SA, Rosenbeerg PA (
1995
) Excitatory amino acids as a final common pathway for neuorlogic disorders. New England
J Med
 
330
:
613–
622.
Lu KH, Hopper BR, Vargo TM, Yen SSC (
1979
) Chronolgical changes in sex steroid, gonadotropin and prolactin secretion in aging female rats displaying different reproductive states.
Biol Reprod
 
21
:
193–
203.
McEwen BS (
2002
) Estrogen actions throughout the brain.
Recent Prog Horm Res
 
57
:
357–
384.
Milner TA, McEwen BS, Hayashi S, Li CJ, Reagan LP, Alves SE (
2001
) Ultrastructural evidence that hippocampal alpha estrogen receptors are located at extranuclear sites.
J Comp Neurol
 
429
:
355–
371.
Miranda P, Williams CL, Einstein G (
1999
) Granule cells in aging rats are sexually dimorphic in their response to estradiol.
J Neurosci
 
19
:
3316–
3325.
Mobbs CV, Gee DM, Finch CE (
1984
) Reproductive senescence in female C57BL/6J mice: ovarian impairments and neuroendocrine impairments that are partially reversible and delayable by ovariectomy.
Endocrinology
 
115
:
1653–
1662.
Morris RGM, Anderson E, Lynch GS, Baudry M (
1986
) Selective impairment of learning and blockade of long-term potentiation by an N-methyl-D-aspartate receptor antagonist, AP5.
Nature
 
319
:
774–
776.
Murphy DD, Segal M (
1996
) Regulation of dendritic spine density in cultured rat hippocampal neurons by steroid hormones.
J Neurosci
 
16
:
4059–
4068.
Olney JW (
1983
) Brain lesions, obesity and other disturbances in mice treated with monosodium glutamate.
Science
 
164
:
719–
721.
Packard MG, Teather LA (
1997
) Intra-hippocampal estradiol infusion enhances memory in ovariectomized rats.
Neuroreport
 
8
:
3009–
3013.
Pau CY, Pau K-YF, Spies HG (
1998
) Putative estrogen receptor β and α mRNA expression in male and female rhesus macaques.
Mol Cell Endocrinol
 
146
:
59–
68.
Petersen DN, Tkalcevic GT, Koza-Taylor PH, Turi TG, Brown TA (
1998
) Identification of estrogen receptor β2, a functional variant of estrogen receptor β expressed in normal rat tissues.
Endocrinology
 
139
:
1082–
1092.
Rapp PR, Morrison JH, Roberts J (
2003
) Cyclic estrogen replacement improves cognitve function in aged ovariectomized rhesus monkeys.
J Neurosci
 
23
:
5708
–5714.
Register TC, Shively CA, Lewis CE (
1998
) Expression of estrogen receptor α and β transcripts in female monkey hippocampus and hypothalamus.
Brain Res
 
788
:
320–
322.
Roberts JA, Gilardi KVK, Lasley B, Rapp PR (
1997
) Reproductive senescence predicts cognitive decline in aged female monkeys.
Neuroreport
 
8
:
2047–
2051.
Rothman S (
1984
) Synaptic activity mediates death of hypoxic neurons.
Science
 
220
:
536–
537.
Rubin BS, Elkind-Hirsch K, Bridges RS (
1985
) Hypothalamic LHRH in aging rats: effects of ovariectomy and steroid replacement.
Neurobiol Aging
 
6
:
309–
315.
Ruud HK, Blackstad TW (
1999
) PALIREL, a compter program for analyzing particle-to-membrane relations, with emphasis on electron micrographs of immunocytochemical preparations and gold labeled molecules.
Comput Biomed Res
 
32
:
93–
122.
Schlegel A, Wang C, Katzenellenbogen BS, Pestell RG, Lisanti MP (
1999
) Caveolin-1 potentiates estrogen receptor alpha (ERalpha ) signaling. Caveolin-1 drives ligand-independent nuclear translocation and activation of ERalpha.
J Biol Chem
 
274
:
33551–
33556.
Sherwin BB (
1997
) Estrogen effects on cognition in menopausal women.
Neurology
 
48
:S21-S26.
Shughrue PJ, Merchenthaler I (
2000
) Evidence for novel estrogen binding sites in the rat hippocampus.
Neuroscience
 
99
:
605–
612.
Shughrue PJ, Lane MV, Merchenthaler I (
1997
) Comparative distribution of estrogen receptor-α and -β mRNA in the rat central nervous system.
J Comp Neurol
 
388
:
507–
525.
Shughrue PJ, Scrimo PJ, Merchenthaler I (
1998
) Evidence for the colocalization of estrogen receptor-β mRNA and estrogen receptor-α immunoreactivity in neurons of the rat forebrain.
Endocrinology
 
139
:
5267–
5270.
Stackman RW, Blasberg ME, Langan CJ, Clark AS (
1997
) Stability of spatial working memory across the estrous cycle of Long-Evans rats.
Neurobiol Learn Mem
 
67
:
167–
171.
Steger RW, Sonntag WE, Peluso JJ, Meites J (
1983
) Effects of ovariectomy and steroid replacement on hypothalamic LHRH content in aging female rats.
Neurobiol Aging
 
4
:
53–
57.
Toran-Allerand CD (
2000
) Novel sites and mechanisms of oestrogen action in the brain. In: Neural and cognitive effects of oestrogens (Novartis Foundation Symposium 230), pp.
56–
73. Chichester: Wiley.
Warren SG, Juraska JM (
1997
) Spatial and nonspatial learning across the rat estrous cycle.
Behav Neurosci
 
111
:
259–
266.
Warren SG, Humphreys AG, Juraska JM, Greenough WT (
1995
) LTP varies across the estrous cycle: enhanced synaptic plasticity in proestrous rats.
Brain Res
 
703
:
26–
30.
Weiland NG (
1992
) Estradiol selectively regulates agonist binding sites on the N-methyl-D-aspartate receptor complex in the CA1 region of the hippocampus.
Endocrinology
 
131
:
662–
668.
Weiland NG, Orikasa C, Hayashi S, McEwen BS (
1997
) Distribution and hormone regulation of estrogen receptor immunoreactive cells in the hippocampus of male and female rats.
J Comp Neurol
 
388
:
603–
612.
Wise PM, Ratner A (
1980
) Effect of ovariectomy on plasma LH, FSH, estradiol, and progesterone and medial basal hypothalamic LHRH concentrations in old and young rats.
Neuroendocrinology
 
30
:
15–
19.
Woolley CS, McEwen BS (
1992
) Estradiol mediates fluctuations in hippocampal synapse density during the estrous cycle in the adult rat.
J Neurosci
 
12
:
2549–
2554.
Woolley CS, McEwen BS (
1993
) Roles of estradiol and progesterone in regulation of hippocampal dendritic spine density during the estrous cycle in the rat.
J Comp Neurol
 
336
:
293–
306.
Woolley CS, McEwen BS (
1994
) Estradiol regulates hippocampal dendritic spine density via an N-methyl-D-aspartate receptor-dependent mechanism.
J Neurosci
 
14
:
7680–
7687.
Woolley CS, Wenzel HJ, Schwartzkroin PA (
1996
) Estradiol increases the frequency of multiple synapse boutons in the hippocampal CA1 region of the adult female rat.
J Comp Neurol
 
373
:
108–
117.
Woolley CW, Gould E, Frankfurt M, McEwen BS (
1990
) Naturally occurring fluctuations in dendritic spine density on adult hippocampal pyramidal neurons.
J Neurosci
 
10
:
4035–
4039.
Yankova M, Hart SA, Woolley CS (
2001
) Estrogen increases synaptic connectivity between single presynaptic inputs and multiple postsynaptic CA1 pyramidal cells: a serial electron-microscopic study.
Proc Natl Acad Sci USA
 
98
:
3525–
3530.