Refractory glossopharyngeal neuralgia successfully treated with onabotulinumtoxinA: A case report.

BACKGROUND: Glossopharyngeal neuralgia is a rare but severe and disabling pain condition often caused by vascular compression of the glossopharyngeal nerve. Treatment is similar to that of trigeminal neuralgia, but some patients may be refractory to both medical and surgical approaches. Here we present a case of refractory glossopharyngeal neuralgia that responded well to onabotulinumtoxinA (BTX-A). CASE: We report a case of a 65-year-old man with well-controlled human immunodeficiency virus disease with glossopharyngeal neuralgia symptoms since 2015. He had partial response to medications but was limited by side-effects. He underwent microvascular decompression twice with initial relief both times, but experienced recurrence of attacks 1-3 years after each surgery. He was treated with BTX-A using the chronic migraine PREEMPT protocol (i.e., 31-39 injection sites in head and neck muscles), which led to significant relief of his glossopharyngeal neuralgia pain. CONCLUSIONS: This is the first case to our knowledge of glossopharyngeal neuralgia treated with BTX-A. BTX-A can be an effective treatment for glossopharyngeal neuralgia, even when injections are not administered directly over the sensory distribution of the glossopharyngeal nerve.

The genomic regulatory elements for estrogen receptor alpha transactivation-function-1 regulated genes.

Estrogen receptor alpha (ERα) is a ligand-dependent transcription regulator, containing two transactivation functional domains, AF-1 and AF-2. The selective estrogen receptor modulators (SERMs), including 4-hydroxytamoxifen (4OHT), activate AF-1 preferentially rather than AF-2. However, it is unclear whether this specific function is related to the tissue-selective functionality of SERMs. Moreover, there is no information determining AF-1-dependent estrogenic-genes existing in tissues. We sought to identify AF-1-dependent estrogenic-genes using the AF-2 mutated knock-in (KI) mouse model, AF2ERKI. AF2ER is an AF-2 disrupted estradiol (E2)-insensitive mutant ERα, but AF-1-dependent transcription can be activated by the estrogen-antagonists, fulvestrant (ICI) and 4OHT. Gene profiling and ChIP-Seq analysis identified Klk1b21 as an ICI-inducible gene in AF2ERKI uterus. The regulatory activity was analyzed further using a cell-based reporter assay. The 5'-flanking 0.4k bp region of Klk1b21 gene responded as an ERα AF-1-dependent estrogen-responsive promoter. The 150 bp minimum ERα binding element (EBE) consists of three direct repeats. These three half-site sequences were essential for the ERα-dependent transactivation and were differentially recognized by E2 and 4OHT for the gene activation. This response was impaired when the minimum EBE was fused with a thymidine-kinase promoter but could be restored by fusion with the 100 bp minimum transcription initiation element (TIE) of Klk1b21, suggesting that the cooperative function of EBE and TIE is essential for mediating AF-1-dependent transactivation. These findings provide the first in vivo evidence that endogenous ERα AF-1 dominant estrogenic-genes exist in estrogen-responsive organs. Such findings will aid in understanding the mechanism of ERα-dependent tissue-selective activity of SERMs.

DNA methylation and transcriptome aberrations mediated by ERα in mouse seminal vesicles following developmental DES exposure.

Early transient developmental exposure to an endocrine active compound, diethylstilbestrol (DES), a synthetic estrogen, causes late-stage effects in the reproductive tract of adult mice. Estrogen receptor alpha (ERα) plays a role in mediating these developmental effects. However, the developmental mechanism is not well known in male tissues. Here, we present genome-wide transcriptome and DNA methylation profiling of the seminal vesicles (SVs) during normal development and after DES exposure. ERα mediates aberrations of the mRNA transcriptome in SVs of adult mice following neonatal DES exposure. This developmental exposure impacts differential diseases between male (SVs) and female (uterus) tissues when mice reach adulthood due to most DES-altered genes that appear to be tissue specific during mouse development. Certain estrogen-responsive gene changes in SVs are cell-type specific. DNA methylation dynamically changes during development in the SVs of wild-type (WT) and ERα-knockout (αERKO) mice, which increases both the loss and gain of differentially methylated regions (DMRs). There are more gains of DMRs in αERKO compared with WT. Interestingly, the methylation changes between the two genotypes are in different genomic loci. Additionally, the expression levels of a subset of DES-altered genes are associated with their DNA methylation status following developmental DES exposure. Taken together, these findings provide an important basis for understanding the molecular and cellular mechanism of endocrine-disrupting chemicals (EDCs), such as DES, during development in the male mouse tissues. This unique evidence contributes to our understanding of developmental actions of EDCs in human health.

A distal super enhancer mediates estrogen-dependent mouse uterine-specific gene transcription of Igf1 (insulin-like growth factor 1).

Insulin-like growth factor 1 (IGF1) is primarily synthesized in and secreted from the liver; however, estrogen (E2), through E2 receptor α (ERα), increases uterine Igf1 mRNA levels. Previous ChIP-seq analyses of the murine uterus have revealed a potential enhancer region distal from the Igf1 transcription start site (TSS) with multiple E2-dependent ERα-binding regions. Here, we show E2-dependent super enhancer-associated characteristics and suggest contact between the distal enhancer and the Igf1 TSS. We hypothesized that this distal super-enhancer region controls E2-responsive induction of uterine Igf1 transcripts. We deleted 430 bp, encompassing one of the ERα-binding sites, thereby disrupting interactions of the enhancer with gene-regulatory factors. As a result, E2-mediated induction of mouse uterine Igf1 mRNA is completely eliminated, whereas hepatic Igf1 expression remains unaffected. This highlights the central role of a distal enhancer in the assembly of the factors necessary for E2-dependent interaction with the Igf1 TSS and induction of uterus-specific Igf1 transcription. Of note, loss of the enhancer did not affect fertility or uterine growth responses. Deletion of uterine Igf1 in a PgrCre;Igf1f/f model decreased female fertility but did not impact the E2-induced uterine growth response. Moreover, E2-dependent activation of uterine IGF1 signaling was not impaired by disrupting the distal enhancer or by deleting the coding transcript. This indicated a role for systemic IGF1, suggested that other growth mediators drive uterine response to E2, and suggested that uterine-derived IGF1 is essential for reproductive success. Our findings elucidate the role of a super enhancer in Igf1 regulation and uterine growth.

Dysregulation of hypothalamic-pituitary estrogen receptor α-mediated signaling causes episodic LH secretion and cystic ovary.

Polycystic ovary syndrome (PCOS) is a hypothalamic-pituitary-gonadal (HPG) axis disorder. PCOS symptoms most likely result from a disturbance in the complex feedback regulation system of the HPG axis, which involves gonadotrophic hormones and ovarian steroid hormones. However, the nature of this complex and interconnecting feedback regulation makes it difficult to dissect the molecular mechanisms responsible for PCOS phenotypes. Global estrogen receptor α (ERα) knockout (KO) mice exhibit a disruption of the HPG axis, resulting in hormonal dysregulation in which female ERα KO mice have elevated levels of serum estradiol (E2), testosterone, and LH. The ERα KO females are anovulatory and develop cystic hemorrhagic ovaries that are thought to be due to persistently high circulating levels of LH from the pituitary. However, the role of ERα in the pituitary is still controversial because of the varied phenotypes reported in pituitary-specific ERα KO mouse models. Therefore, we developed a mouse model where ERα is reintroduced to be exclusively expressed in the pituitary on the background of a global ERα-null (PitERtgKO) mouse. Serum E2 and LH levels were normalized in PitERtgKO females and were comparable to wild-type serum levels. However, the ovaries of PitERtgKO adult mice displayed a more overt cystic and hemorrhagic phenotype when compared with ERα KO littermates. We determined that anomalous sporadic LH secretion caused the severe ovarian phenotype of PitERtgKO females. Our observations suggest that pituitary ERα is involved in the estrogen negative feedback regulation, whereas hypothalamic ERα is necessary for the precise control of LH secretion. Uncontrolled, irregular LH secretion may be the root cause of the cystic ovarian phenotype with similarities to PCOS.-Arao, Y., Hamilton, K. J., Wu, S.-P., Tsai, M.-J., DeMayo, F. J., Korach, K. S. Dysregulation of hypothalamic-pituitary estrogen receptor α-mediated signaling causes episodic LH secretion and cystic ovary.

Negative elongation factor is essential for endometrial function.

Pausing of RNA polymerase II (Pol II) during early transcription, mediated by the negative elongation factor (NELF) complex, allows cells to coordinate and appropriately respond to signals by modulating the rate of transcriptional pause release. Promoter proximal enrichment of Pol II occurs at uterine genes relevant to reproductive biology; thus, we hypothesized that pausing might impact endometrial response by coordinating hormonal signals involved in establishing and maintaining pregnancy. We deleted the NELF-B subunit in the mouse uterus using PgrCre (NELF-B UtcKO). Resulting females were infertile. Uterine response to the initial decidual stimulus of NELF-B UtcKO was similar to that of control mice; however, subsequent full decidual response was not observed. Cultured NELF-B UtcKO stromal cells exhibited perturbances in extracellular matrix components and also expressed elevated levels of the decidual prolactin Prl8a2, as well as altered levels of transcripts encoding enzymes involved in prostaglandin synthesis and metabolism. Because endometrial stromal cell decidualization is also critical to human reproductive health and fertility, we used small interfering to suppress NELF-B or NELF-E subunits in cultured human endometrial stromal cells, which inhibited decidualization, as reflected by the impaired induction of decidual markers PRL and IGFBP1. Overall, our study indicates NELF-mediated pausing is essential to coordinate endometrial responses and that disruption impairs uterine decidual development during pregnancy.-Hewitt, S. C., Li, R., Adams, N., Winuthayanon, W., Hamilton, K. J., Donoghue, L. J., Lierz, S. L., Garcia, M., Lydon, J. P., DeMayo, F. J., Adelman, K., Korach, K. S. Negative elongation factor is essential for endometrial function.

Assessment of Progressive Muscle Relaxation (PMR) as a Stress-Reducing Technique for First-Year Veterinary Students.

The veterinary profession continually strives to address wellness issues such as compassion fatigue, burnout, stress, anxiety, and depression. Wellness issues may begin during the professional curriculum when students experience intense academic, clinical, social, and personal demands on their time. The purpose of this article was to assess the use of progressive muscle relaxation (PMR) as a simple, non-invasive stress reduction technique for first-year veterinary students (n = 101) at a US veterinary college. Students completed a 38-item questionnaire, the Smith Relaxation States Inventory 3 (SRSI3), both before and after performing PMR. Scores for the categories of basic relaxation, mindfulness, positive energy, transcendence, and stress were assessed. Female students (n = 92) had significant (p < .05) improvement in basic relaxation, mindfulness, and stress after completing PMR. Male students (n = 9) had significant (p < .05) improvement in basic relaxation and stress after completing PMR. When grouped according to age, all students had significant (p < .05) improvement in the categories of basic relaxation and stress. Students in the 22-year-old (n = 31), 23-year-old (n = 29), 24-year-old (n = 15), and 25-year-old or greater (n = 17) groups also had significant improvement (p < .05) in mindfulness. Additionally, students in the 23-year-old group had significant (p < .05) improvement in positive energy. These results support the use of PMR as a potential self-care strategy for students to implement during their academic and professional careers.

Migraine Treatment in Pregnant Women Presenting to Acute Care: A Retrospective Observational Study.

OBJECTIVE: To assess the acute treatment of pregnant women presenting to a hospital with migraine. BACKGROUND: Migraine is a common problem in pregnancy; however, migraine treatment is challenging in pregnant women for fears of medication teratogenicity and lack of data in this population. To date, no study has directly explored physician practices for treatment of acute migraine in pregnant women. METHODS: We conducted a retrospective chart review of medication administration for pregnant women who presented to an acute care setting with a migraine attack and received neurology consultation between 2009 and 2014. RESULTS: We identified 72 pregnant women with migraine who were treated with pain medications. Fifty-one percent (37/72) were in the third trimester of pregnancy, 39% (28/72) in the second trimester, and 10% (7/72) in the first trimester. Thirty-two percent (23/72) had not tried any acute medications at home before coming to the hospital, and 47% (34/72) presented in status migrainosus. Patients received treatment in the hospital for a median of 23 hours (interquartile range = 5-45 hours). The most common medications prescribed were metoclopramide in 74% (53/72) of patients (95% confidence interval [CI] 62-82%) and acetaminophen in 69% (50/72) of patients (95% CI 58-79%). Metoclopramide was administered along with diphenhydramine in 81% (44/53) of patients (95% CI 71-91%). Acetaminophen was the most frequent medicine administered first (53%, 38/72). Patients were often treated with butalbital (35%, 25/72) or opioids (30%, 22/72), which were used as second- or third-line treatments in 29% of patients (20/72). Thirty-eight percent (27/72) received an intravenous (IV) fluid bolus, 24% received IV magnesium (17/72), and 6% (4/72) had peripheral nerve blocks performed. CONCLUSIONS: While the majority of pregnant women with acute migraine received medications considered relatively safe in pregnancy, there was variation in treatment choice and sequence. Some acute medications considered potentially hazardous for fetal health and less effective for migraine (opioids and butalbital) were used frequently, whereas other treatments that may have low teratogenic risk (nerve blocks, IV fluid boluses, and triptans) were used less or not at all. These results indicate a need for developing guidelines and protocols to standardize acute treatment of migraine in pregnancy.

Clinical report: A patient with a late diagnosis of cerebrotendinous xanthomatosis and a response to treatment.

Cerebrotendinous xanthomatosis (CTX) is a rare, autosomal recessive, inborn error of bile acid metabolism characterized by diarrhea in infancy, juvenile cataracts in childhood, tendon xanthomas developing in the second to third decades of life, and progressive neurologic dysfunction in adulthood. The condition is caused by mutations in the CYP27A1 gene that result in decreased production of chenodeoxycholic acid (CDCA) and elevated levels of cholestanol and bile alcohols. We present a 36-year-old male of Han ethnicity who developed xanthomas of his Achilles tendons and suffered neurocognitive declines and gait deterioration in his second decade. The diagnosis of CTX was confirmed by marked elevation of the serum cholestanol level. Sequencing of CYP27A1 showed a paternally inherited splice mutation, c.446 + 1G>T, and a maternally inherited nonsense mutation, c.808C>T, predicting p.(Arg270*). Despite the advanced disease in this patient, treatment with CDCA reduced the xanthoma size and improved his cognition and strength, and the patient made significant gains in his ambulation and coordination. We report this case to illustrate the potential benefits of therapy in patients with CTX who have advanced disease at the time of diagnosis.

The Development and Validation of the Rational and Intuitive Decision Styles Scale.

Decision styles reflect the typical manner by which individuals make decisions. The purpose of this research was to develop and validate a decision style scale that addresses conceptual and psychometric problems with current measures. The resulting 10-item scale captures a broad range of the rational and intuitive styles construct domain. Results from 5 independent samples provide initial support for the dimensionality and reliability of the new scale, as demonstrated by a clear factor structure and high internal consistency. In addition, our results show evidence of convergent and discriminant validity through expected patterns of correlations across decision-making individual differences and the International Personality Item Pool (IPIP) Big Five traits. Research domains that would benefit from incorporating the concept of decision styles are discussed.

Transactivating function (AF) 2-mediated AF-1 activity of estrogen receptor α is crucial to maintain male reproductive tract function.

Estrogen receptor alpha (ERα) is a ligand-dependent transcription factor containing two transcriptional activation function (AF) domains. AF-1 is in the N terminus of the receptor protein, and AF-2 activity is dependent on helix 12 of the C-terminal ligand-binding domain. We recently showed that two point mutations converting leucines 543 and 544 to alanines in helix 12 (AF2ER) minimized estrogen-dependent AF-2 transcriptional activation. A characteristic feature of AF2ER is that the estrogen antagonists ICI182780 and tamoxifen (TAM) act as agonists through intact AF-1, but not through mutated AF-2. Here we report the reproductive phenotype of male AF2ER knock-in (AF2ERKI) mice and demonstrate the involvement of ERα in male fertility. The AF2ERKI male homozygotes are infertile because of seminiferous tubular dysmorphogenesis in the testis, similar to ERα KO males. Sperm counts and motility did not differ at age 6 wk in AF2ERKI and WT mice, but a significant testis defect was observed in adult AF2ERKI male mice. The expression of efferent ductal genes involved in fluid reabsorption was significantly lower in AF2ERKI males. TAM treatment for 3 wk beginning at age 21 d activated AF-2-mutated ERα (AF2ER) and restored expression of efferent ductule genes. At the same time, the TAM treatment reversed AF2ERKI male infertility compared with the vehicle-treated group. These results indicate that the ERα AF-2 mutation results in male infertility, suggesting that the AF-1 is regulated in an AF-2-dependent manner in the male reproductive tract. Activation of ERα AF-1 is capable of rescuing AF2ERKI male infertility.

Estrogen receptor α L543A,L544A mutation changes antagonists to agonists, correlating with the ligand binding domain dimerization associated with DNA binding activity.

A ligand-dependent nuclear transcription factor, ERα has two transactivating functional domains (AF), AF-1 and AF-2. AF-1 is localized in the N-terminal region, and AF-2 is distributed in the C-terminal ligand-binding domain (LBD) of the ERα protein. Helix 12 (H12) in the LBD is a component of the AF-2, and the configuration of H12 is ligand-inducible to an active or inactive form. We demonstrated previously that the ERα mutant (AF2ER) possessing L543A,L544A mutations in H12 disrupts AF-2 function and reverses antagonists such as fulvestrant/ICI182780 (ICI) or 4-hydoxytamoxifen (OHT) into agonists in the AF2ER knock-in mouse. Our previous in vitro studies suggested that the mode of AF2ER activation is similar to the partial agonist activity of OHT for WT-ERα. However, it is still unclear how antagonists activate ERα. To understand the molecular mechanism of antagonist reversal activity, we analyzed the correlation between the ICI-dependent estrogen-responsive element-mediated transcription activity of AF2ER and AF2ER-LBD dimerization activity. We report here that ICI-dependent AF2ER activation correlated with the activity of AF2ER-LBD homodimerization. Prevention of dimerization impaired the ICI-dependent ERE binding and transcription activity of AF2ER. The dislocation of H12 caused ICI-dependent LBD homodimerization involving the F-domain, the adjoining region of H12. Furthermore, F-domain truncation also strongly depressed the dimerization of WT-ERα-LBD with antagonists but not with E2. AF2ER activation levels with ICI, OHT, and raloxifene were parallel with the degree of AF2ER-LBD homodimerization, supporting a mechanism that antagonist-dependent LBD homodimerization involving the F-domain results in antagonist reversal activity of H12-mutated ERα.

Estrogen receptor α AF-2 mutation results in antagonist reversal and reveals tissue selective function of estrogen receptor modulators.

The estrogen receptor (ER) is a ligand-dependent transcription factor containing two transcriptional activation domains. AF-1 is in the N terminus of the receptor protein and AF-2 activity is dependent on helix 12 of the C-terminal ligand-binding domain. Two point mutations of leucines 543 and 544 to alanines (L543A, L544A) in helix 12 minimized estrogen-dependent transcriptional activation and reversed the activity of the estrogen antagonists ICI182780 (ICI) and tamoxifen (TAM) into agonists in a similar manner that TAM activated WT ERα through AF-1 activation. To evaluate the physiological role of AF-1 and AF-2 for the tissue-selective function of TAM, we generated an AF-2-mutated ERα knock-in (AF2ERKI) mouse model. AF2ERKI homozygote female mice have hypoplastic uterine tissue and rudimentary mammary glands similar to ERα-KO mice. Female mice were infertile as a result of anovulation from hemorrhagic cystic ovaries and elevated serum LH and E2 levels, although the mutant ERα protein is expressed in the AF2ERKI model. The AF2ERKI phenotype suggests that AF-1 is not activated independently, even with high serum E2 levels. ICI and TAM induced uterotropic and ER-mediated gene responses in ovariectomized AF2ERKI female mice in the same manner as in TAM- and E2-treated WT mice. In contrast, ICI and TAM did not act as agonists to regulate negative feedback of serum LH or stimulate pituitary prolactin gene expression in a different manner than TAM- or E2-treated WT mice. The functionality of the mutant ERα in the pituitary appears to be different from that in the uterus, indicating that ERα uses AF-1 differently in the uterus and the pituitary for TAM action.

Effect of CGRP inhibitors on interictal cerebral hemodynamics in individuals with migraine.

Introduction: Calcitonin gene-related peptide (CGRP) plays an important role in cerebral vasodilation, so here we aim to quantify the impact of CGRP monoclonal antibody (mAb) therapy on cerebral hemodynamics. Methods: In 23 patients with chronic and episodic migraine, cerebral hemodynamic monitoring was performed (1) prior to and (2) 3-months into CGRP-mAb therapy. Transcranial Doppler monitored cerebral blood flow velocity (CBFv) in the middle cerebral artery (MCA) and posterior cerebral artery (PCA), from which cerebrovascular reactivity (CVR) and cerebral autoregulation (CA; Mx-index) were calculated. Results: CA was similar off and on treatment, in the MCA (p = 0.42) and PCA (p = 0.72). CVR was also unaffected by treatment, in the MCA (p = 0.38) and PCA (p = 0.92). CBFv and blood pressure were also unaffected. The subgroup of clinical responders (>50% reduction in migraine frequency) exhibited a small reduction in MCA-CBFv (6.0 cm/s; IQR: 1.1-12.4; p = 0.007) and PCA-CBFv (8.9 cm/s; IQR: 6.9-10.3; p = 0.04). Discussion: Dynamic measures of cerebrovascular physiology were preserved after 3 months of CGRP-mAb therapy, but a small reduction in CBFv was observed in patients who responded to treatment. Subgroup findings should be interpreted cautiously, but further investigation may clarify if CBFv is dependent on the degree of CGRP inhibition or may serve as a biomarker of drug sensitivity.

Multiple tissue-specific epigenetic alterations regulate persistent gene expression changes following developmental DES exposure in mouse reproductive tissues.

Clinically, developmental exposure to the endocrine disrupting chemical, diethylstilboestrol (DES), results in long-term male and female infertility. Experimentally, developmental exposure to DES results in abnormal reproductive tract phenotypes in male and female mice. Previously, we reported that neonatal DES exposure causes ERα-mediated aberrations in the transcriptome and in DNA methylation in seminal vesicles (SVs) of adult mice. However, only a subset of DES-altered genes could be explained by changes in DNA methylation. We hypothesized that alterations in histone modification may also contribute to the altered transcriptome during SV development. To test this idea, we performed a series of genome-wide analyses of mouse SVs at pubertal and adult developmental stages in control and DES-exposed wild-type and ERα knockout mice. Neonatal DES exposure altered ERα-mediated mRNA and lncRNA expression in adult SV, including genes encoding chromatin-modifying proteins that can impact histone H3K27ac modification. H3K27ac patterns, particularly at enhancers, and DNA methylation were reprogrammed over time during normal SV development and after DES exposure. Some of these reprogramming changes were ERα-dependent, but others were ERα-independent. A substantial number of DES-altered genes had differential H3K27ac peaks at nearby enhancers. Comparison of gene expression changes, H3K27ac marks and DNA methylation marks between adult SV and adult uterine tissue from ovariectomized mice neonatally exposed to DES revealed that most of the epigenetic changes and altered genes were distinct in the two tissues. These findings indicate that the effects of developmental DES exposure cause reprogramming of reproductive tract tissue differentiation through multiple epigenetic mechanisms.

Refractory Short-Lasting Unilateral Neuralgiform Headache Attacks With Conjunctival Injection and Tearing (SUNCT) Responding to Erenumab Adjuvant Therapy: A Case Report.

Many patients with short-lasting unilateral neuralgiform headache attacks with conjunctival injection and tearing (SUNCT) fail to respond to the first-line treatment of lamotrigine. Additionally, data for other treatments are limited in this rare headache disorder. SUNCT involves activation of the trigeminal nerve which uses the neuropeptide calcitonin gene-related peptide (CGRP); thus CGRP-targeted treatments may be beneficial in this disorder. We present a patient with SUNCT who failed to respond optimally to 10 medications and four surgical treatments. However, she had minimal attacks after erenumab 140 mg was added to carbamazepine 200 mg three times daily and pregabalin 75 mg twice daily. Decreasing any of these three medications worsened her attacks. Our case represents the second case report of a SUNCT patient responding to a CGRP monoclonal antibody, suggesting this treatment may be a consideration in refractory SUNCT.

A Novel Mouse Model to Analyze Non-Genomic ERα Physiological Actions.

Nongenomic effects of estrogen receptor α (ERα) signaling have been described for decades. Several distinct animal models have been generated previously to analyze the nongenomic ERα signaling (eg, membrane-only ER, and ERαC451A). However, the mechanisms and physiological processes resulting solely from nongenomic signaling are still poorly understood. Herein, we describe a novel mouse model for analyzing nongenomic ERα actions named H2NES knock-in (KI). H2NES ERα possesses a nuclear export signal (NES) in the hinge region of ERα protein resulting in exclusive cytoplasmic localization that involves only the nongenomic action but not nuclear genomic actions. We generated H2NESKI mice by homologous recombination method and have characterized the phenotypes. H2NESKI homozygote mice possess almost identical phenotypes with ERα null mice except for the vascular activity on reendothelialization. We conclude that ERα-mediated nongenomic estrogenic signaling alone is insufficient to control most estrogen-mediated endocrine physiological responses; however, there could be some physiological responses that are nongenomic action dominant. H2NESKI mice have been deposited in the repository at Jax (stock no. 032176). These mice should be useful for analyzing nongenomic estrogenic responses and could expand analysis along with other ERα mutant mice lacking membrane-bound ERα. We expect the H2NESKI mouse model to aid our understanding of ERα-mediated nongenomic physiological responses and serve as an in vivo model for evaluating the nongenomic action of various estrogenic agents.