Expression Of Steroid Receptor Coactivator-1 In The Brain Of Female Rats And Its Role In The Regulation Of Hippocampal Postsynaptic Density Protein-95

Steroids have profound impact on nervous system, and accumulated evidences have shown that decreased circulating steroids with aging are associated with an increase in cognitive, neurological and psychiatric disorders, but the mechanisms are still unclear. So far as we have known that from fetal life, estrogens are crucial in determining central gender dimorphism, and an estrogen-induced synaptic plasticity is well evident during puberty and seasonal changes as well as during the ovarian cycle. Estrogen and nuclear ERs have been implied in the regulation of synaptic plasticity, learning and memory, and these actions are believed to be mediated through the coactivators of nuclear receptor.Steroid receptor coactivator-1 (SRC-1) is one of the coactivators of nuclear receptors, which has been demonstrated to dramatically enhance the transcriptional activity of a variety of nuclear receptors, including ER and progestin receptor (PR), in a ligand-dependent manner. Studies have reported that SRC-1 transcript was extensively expressed in the nerve system of embryonic and adult animals. Functional studies showed that SRC-1 was required for normal neuronal development, reduced SRC-1 protein interfered with the defeminizing actions of estrogen in the neonatal rat brain and resulted in a significantly smaller sexually dimorphic nucleus of the hypothalamus. The studies of knock-out mice models also found that adult SRC-1-/- mice exhibited moderate motor dysfunction and delayed development of Purkinje cells, loss of skeletal response to estrogen, and altered hypothalamic-pituitary-adrenal axis function. However, the expression of SRC-1 in young brain and whether normal aging has impact on that has not been fully defined. Thus, we detected the immunoreactivity of SRC-1 in the young and old female rat brain by immunohistochemistry to clarify how aging affects the expression of SRC-1 in specific brain regions.Hippocampus is one of the most important targets of estrogen, and studies have shown that hippocampal synaptic plasticity could be regulated by estrogen. In the hippocampus, the mRNAs and proteins of steroids receptors such as ERαand ERβare present, and these nuclear receptors function to regulate hippocampal neuronal excitability, synaptogenesis and plasticity as well as hippocampus-based learning and memory. SRC-1 transcripts in the hippocampus were detected in E14.5 mice and in adult rats. SRC-1 protein was detected in the CA1–CA4 pyramidal cell layer and the granular cell layer of the dentate gyrus by immunohistochemistry. The developmental profile of SRC-1 mRNA studied by in situ hybridization in mice from postnatal day 1 (P1) to adult has been reported, but the temporal and spatial profile of SRC-1 protein in the developmental especially in the aged hippocampus are currently unknown. More importantly, synaptic plasticity was hypothesized to be crucial in hippocampal learning and memory, high expression of SRC-1 mRNA and protein was detected in the hippocampus, but whether SRC-1 plays a role in the regulation of synaptic plasticity is still unknown.To address these questions above, we further used immunohistochemical (IHC) peroxidase staining and Western blot technique to examine the expression profile of SRC-1 and postsynaptic density-95 (PSD-95) in postnatal developmental hippocampus. PSD-95 is a cytoskeletal protein localized in postsynaptic density (PSD) and has been reported as marker for PSD. It is involved in several important functions such as synaptogenesis, synaptic plasticity, learning and memory performance for the mammalian braint. Then, ovariectomy was applied to detect whether the age-related decline of hippocampal SRC-1 was affected by ovary female hormones. Finally, a pool of dsRNA against SRC-1 was employed to knockdown SRC-1 expression in cultured hippocampal neurons to test its impact on PSD-95.Main results:1. SRC-1 immunoreactivities were extensively distributed in the brain, striking age-related decreases of SRC-1 were noticed in those regions related to central regulation of motor (striatum, substantia nigra, pontine nuclei, lateral reticular nucleus and Purkinje cells, etc.), learning and memory (olfactory bulb, hippocampus, basal forebrain, Purkinje cells, etc.), and neural stem cell (olfactory, dentate gyrus, cerebral cortex, etc.).2. Suprisingly, SRC-1 immunopositive materials were mainly detected in the nuclear, but also detected in the extranuclear in some regions, especially in those sub-regions related to motor-regulation. 3. In the hippocampus, SRC-1 expression was increased gradually during the postnatal development. At P0, it was nearly undetectable,then approached the peak at P14, and decreased lightly when adult, but it was dramatically reduced at old.4. PSD-95 has the similar developmental expression profile with SRC-1. What is different is that it approached the peak at P30, a little later than SRC-1.5. The level of PSD-95 expression in the hippocampus was significantly decreased when a dsRNA pool against SRC-1 was applied (p<0.05). SRC-1 RNAi also affected the growth and morphological character of cultured neurons, include less and shoter dendrites, and fewer connection between cells.6. It seems that the expression of SRC-1 in the hippocampus was not affected by ovariectomy. But the expression of PSD-95 was down-regulated in the first week after OVX, although it returned to the normal level two weeks later.Main conclusion:1. The extensively expression of SRC-1 suggests it has multifunctional roles in the brain. The age-related decrease of SRC-1 in specific motor, neural stem cells, learning and memory nuclei indicates its potential roles in neurodegenerative disorders, which may be one of the underlying mechanisms of the vulnerability of the aged brain.2. The extra-nuclear localization of SRC-1 immunoreactivities strongly implys that it may function through a novel non-genomic pathway.3. The high correlation between PSD-95 and SRC-1 expression profile at postnatal development suggests SRC-1 plays a crucial role in synaptogenesis and synaptic platisity. Age-related functional impairment of memory and cognition may result from decrease of SRC-1 in hippocampus.4. SRC-1 RNA interference followed by a down-regulation of PSD-95 in cultured hippocampal neurons. Such phenomenon demonstrates the potential roles of SRC-1 in synaptogenesis and synaptic platisity, which may be involved in recognition function and neurodegenerative disorders.5. Ovariectomy has little effect on expression of SRC-1 in hippocampus indicates that hippocampal synaptic plastisity may be regulated by estrogen synthesized in hippocampus location but not influenced by estrogen from circulation.In summary, we used immunohistochemistry to investigate the locations and age-related changes of SRC-1 in female rat brain; then we studied the developmental expression profile of SRC-1 and the effects of SRC-1 on PSD-95 through OVX modle and RNA interference techniques. Our studies demonstrated the potential funtions of SRC-1 on synaptogenesis and synaptic plasiticy, and provided new clues and evidences for further study on neurodegenerative diseases.