The protocols involved in this project are as follows: 00-M-0103, 03-M-0138, 03-M-0161, 04-M-0139, 05-M-0059, 81-M-0126, 90-M-0088, 92-M-0174, and 95-M-0097. This report includes work arising from the following protocols: NCT00005011, NCT00056901, NCT00059228, NCT00082043, NCT00100360, NCT00001177, NCT00001259, and NCT00001481. In our studies we have developed experimental models for the triggering of symptoms in premenstrual dysphoric disorder (PMDD) and postpartum depression that we employ in our efforts to identify both the underlying biology of these conditions. We have extended and replicated our earlier findings by demonstrating that women with PMDD (n = 35), who respond to gonadotropin releasing hormone (GnRH) agonist-induced ovarian suppression experience a recurrence of PMDD after the initial re-exposure to combined estradiol and progesterone. However, recurrent symptoms do not occur once hormone levels are stabilized over the subsequent two months of continuous ovarian steroid therapy. Thus the change in ovarian steroid level not the absolute level itself is the trigger for affective destabilization in these women. These observations are of both clinical and scientific importance, as they suggest the physiologic basis for the susceptibility to experience PMDD and will also provide alternative hormone-based therapies for women with this condition. In addition to our studies on the behavioral effects of changes in ovarian steroids across the menstrual cycle and during the postpartum, we employ methodologies to investigate the underlying biological mechanisms of these conditions including studies investigating positron emission tomography (PET), structural magnetic resonance imaging (MRI), and functional magnetic resonance imaging (fMRI), as well as both metabolomics platforms for evaluating neurosteroid synthesis, and functional genomics studies examining the effects of a change in ovarian steroid on cellular function. First, our neuroimaging studies have identified both a neural substrate of risk in PMDD as well as a brain-region specific response that could underlie the differential behavioral response to ovarian steroids in this condition. We performed O15 PET and fMRI studies in women who are participating in the GnRH agonist-induced hypogonadism study. In this study, cognitive activation was achieved using the N-back test which allows us to vary the cognitive load and the effort required to perform the task. We observed that women with PMDD show abnormal prefrontal recruitment, specifically greater activation than controls throughout the DLPFC bilaterally. These findings are robust and observable in both 015 PET and fMRI techniques. In both imaging modalities, significantly increased DLPFC activations in PMDD compared with controls occurred in all three hormone conditions and, therefore, were independent of hormone condition. Moreover, DLPFC activations in PMDD correlated negatively with measures of PMD functional impairment (i.e., global assessment of functional impairment GAF scores), age of onset of PMDD, and pre-treatment symptom severity measures: the greater the over-activation, the greater the disability, the earlier age of onset, and the more severe baseline symptomatology. These findings suggest an enduring, trait-like predisposition to this hormonally-triggered disorder. In a second completed study, in women with PMDD and controls who underwent resting state PET studies during each of the three hormone conditions in the Lupron paradigm, we observed differences in resting rCBF in women with PMDD compared with controls in the subgenual cingulate (BA25), medial orbital frontal cortex (mOFC), and right hippocampal/parahippocampal region. Higher resting state rCBF in PMDD was present during hypogonadism (when PMDD symptoms are in remission) compared with estradiol or progesterone replacement (when PMDD symptoms are at risk of recurring), whereas the opposite pattern (i.e., lower resting rCBF during hypogonadism compared with either estradiol or progesterone) was observed in controls who experience no change in mood across identical hormone conditions. These data demonstrate for the first time a differential pattern of rCBF within neuroanatomical loci implicated in the process of affective adaptation. Additionally, the fact that the changes in rCBF correspond in time to the hormone state-related changes in symptoms raises the possibility that the changes in rCBF may reflect the altered mood state rather than a direct effect of ovarian steroid hormonal exposure. One possible trigger for the abnormal ovarian steroid-triggered mood state in PMDD is an altered profile of neurosteroid metabolites during exposure to estradiol or progesterone. This possibility is also suggested by our demonstration of the therapeutic efficacy of dutasteride to mitigate symptoms of PMDD since dutasteride inhibits the rate limiting enzymatic step in neurosteroid synthesis. Thus, we next examined the profile of steroid and neurosteroid metabolites in women with PMDD using a metabolomics platform. Metabolomic studies (employing liquid chromatography-tandem mass spectroscopy) are performed in women with PMDD who respond to Lupron with elimination of symptoms and who experience return of symptoms during progesterone or estradiol replacement, as well as controls who experience no change in mood during the identical experimental paradigm. These data have been analyzed at the UC Davis metabolomics core lab and are now being analyzed for differences between women with PMDD and controls in the activity of neurosteroid synthetic pathways. Our metabolomic and neuroimaging studies are complemented by functional genomic studies, performed in collaboration with David Goldman's laboratory, in which we employ lymphoblastoid and induced pluripotent cells obtained from women with PMDD and controls who participate in our Lupron studies. These experiments allow us to explore the nature of the differential behavioral response by examining gene expression and changes in cellular behaviors associated with the exposure to physiologic levels of either estradiol or progesterone across the two different behavioral phenotypes (women with PMDD who experience a remission of PMDD during ovarian suppression and a recurrence of PMDD when ovarian steroids are added back, and controls who are asymptomatic during all three hormone conditions). Preliminary results show that women with PMDD express more estrogen receptor (ER) alpha (but not beta) compared with controls. Additionally, RNA sequencing studies show statistically significant, quantitative differences in the PMDD transcriptome compared to an asymptomatic control group. We identified significant differences in the expression of the chromatin modifying EzH2 complex, which is of particular interest as a potential candidate since it is regulated by estradiol and leads to genomic regions of transcriptional repression. Ten of the 13 constituent genes in this complex showed increased expression at baseline in women with PMDD compared with controls. These results were confirmed with quantitative RT-PCR and in a larger sample of women with PMDD and controls. Additionally, several of these genes including members of the sirtuin family (e.g., SIRT1) showed differential expression patterns in response to progesterone exposure of these cells in women with PMDD compared to controls. The mechanism by which these pathways converge and possibly alter CNS function sufficient to manifest in PMDD will be the focus of our future studies in induced neuronal and glial cells.
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