Sexually dimorphic organization of open field behavior following moderate prenatal alcohol exposure.

BACKGROUND: Prenatal alcohol exposure (PAE) can produce deficits in a wide range of cognitive functions but is especially detrimental to behaviors requiring accurate spatial information processing. In open field environments, spatial behavior is organized such that animals establish "home bases" marked by long stops focused around one location. Progressions away from the home base are circuitous and slow, while progressions directed toward the home base are non-circuitous and fast. The impact of PAE on the organization of open field behavior has not been experimentally investigated. METHODS: In the present study, adult female and male rats with moderate PAE or saccharin exposure locomoted a circular high walled open field for 30 minutes under lighted conditions. RESULTS: The findings indicate that PAE and sex influence the organization of open field behavior. Consistent with previous literature, PAE rats exhibited greater locomotion in the open field. Novel findings from the current study indicate that PAE and sex also impact open field measures specific to spatial orientation. While all rats established a home base on the periphery of the open field, PAE rats, particularly males, exhibited significantly less clustered home base stopping with smaller changes in heading between stops. PAE also impaired progression measures specific to distance estimation, while sex alone impacted progression measures specific to direction estimation. CONCLUSIONS: These findings support the conclusion that adult male rats have an increased susceptibility to the effects of PAE on the organization of open field behavior.

The LFA-1 antagonist BIRT377 reverses neuropathic pain in prenatal alcohol-exposed female rats via actions on peripheral and central neuroimmune function in discrete pain-relevant tissue regions.

Previous reports show that moderate prenatal alcohol exposure (PAE) poses a risk factor for developing neuropathic pain following adult-onset peripheral nerve injury in male rats. Recently, evidence suggests that immune-related mechanisms underlying neuropathic pain in females are different compared to males despite the fact that both sexes develop neuropathy of similar magnitude and duration following chronic constriction injury (CCI) of the sciatic nerve. Data suggest that the actions of peripheral T cells play a greater role in mediating neuropathy in females. The goal of the current study is to identify specificity of immune cell and cytokine changes between PAE and non-PAE neuropathic females by utilizing a well-characterized rodent model of sciatic nerve damage, in an effort to unmask unique signatures of immune-related factors underlying the risk of neuropathy from PAE. Cytokines typically associated with myeloid cell actions such as interleukin (IL)-1ß, tumor necrosis factor (TNF), IL-6, IL-4 and IL-10 as well as the neutrophil chemoattractant CXCL1, are examined. In addition, transcription factors and cytokines associated with various differentiated T cell subtypes are examined (anti-inflammatory FOXP3, proinflammatory IL-17A, IL-21, ROR-γt, interferon (IFN)-γ and T-bet). Lymphocyte function associated antigen 1 (LFA-1) is an adhesion molecule expressed on peripheral immune cells including T cells, and regulates T cell activation and extravasation into inflamed tissue regions. A potential therapeutic approach was explored with the goal of controlling proinflammatory responses in neuroanatomical regions critical for CCI-induced allodynia by blocking LFA-1 actions using BIRT377. The data show profound development of hindpaw allodynia in adult non-PAE control females following standard CCI, but not following minor CCI, while minor CCI generated allodynia in PAE females. The data also show substantial increases in T cell-associated proinflammatory cytokine mRNA and proteins, along with evidence of augmented myeloid/glial activation (mRNA) and induction of myeloid/glial-related proinflammatory cytokines, CCL2, IL-1ß and TNF in discrete regions along the pain pathway (damaged sciatic nerve, dorsal root ganglia; DRG, and spinal cord). Interestingly, the characteristic anti-inflammatory IL-10 protein response to nerve damage is blunted in neuropathic PAE females. Moreover, T cell profiles are predominantly proinflammatory in neuropathic Sac and PAE females, augmented levels of Th17-specific proinflammatory cytokines IL-17A and IL-21, as well as the Th1-specific factor, T-bet, are observed. Similarly, the expression of RORγt, a critical transcription factor for Th17 cells, is detected in the spinal cord of neuropathic females. Blocking peripheral LFA-1 actions with intravenous (i.v.) BIRT377 reverses allodynia in Sac and PAE rats, dampens myeloid (IL-1ß, TNF, CXCL1)- and T cell-associated proinflammatory factors (IL-17A and RORγt) and spinal glial activation. Moreover, i.v. BIRT377 treatment reverses the blunted IL-10 response to CCI observed only in neuropathic PAE rats and elevates FOXP3 in pain-reversed Sac rats. Unexpectedly, intrathecal BIRT377 treatment is unable to alter allodynia in either Sac or PAE neuropathic females. Together, these data provide evidence that: 1) fully differentiated proinflammatory Th17 cells recruited at the sciatic nerve, DRGs and lumbar spinal cord may interact with the local environment to shape the immune responses underlying neuropathy in female rats, and, 2) PAE primes peripheral and spinal immune responses in adult females. PAE is a risk factor in females for developing peripheral neuropathy after minor nerve injury.

Prenatal alcohol exposure is a risk factor for adult neuropathic pain via aberrant neuroimmune function.

BACKGROUND: Clinical studies show that prenatal alcohol exposure (PAE) results in effects that persist into adulthood. Experimental animal models of moderate PAE demonstrate that young adults with PAE display potentiated sensitivity to light touch, clinically termed allodynia, following sciatic nerve chronic constriction injury (CCI) that coincides with heightened spinal glial, spinal macrophage, and peripheral immune responses. However, basal touch sensitivity and corresponding glial and leukocyte activation are unaltered. Therefore, the current study explored whether the enduring pathological consequences of moderate PAE on sensory processing are unmasked only following secondary neural insult. METHODS: In middle-aged (1 year) Long Evans rats that underwent either prenatal saccharin exposure (control) or moderate PAE, we modified the well-characterized model of sciatic neuropathy, CCI, to study the effects of PAE on neuro-immune responses in adult offspring. Standard CCI manipulation required 4 chromic gut sutures, while a mild version applied a single suture loosely ligated around one sciatic nerve. Spinal glial immunoreactivity was examined using immunohistochemistry. The characterization and functional responses of leukocyte populations were studied using flow cytometry and cell stimulation assays followed by quantification of the proinflammatory cytokines interleukin-1beta (IL-1ß) and tumor necrosis factor-alpha (TNF-α). Data were statistically analyzed by ANOVA and unpaired t tests. RESULTS: The current report demonstrates that mild CCI generates robust allodynia only in PAE rats, while the pathological effects of PAE following the application of a standard CCI are revealed by enhanced allodynia and elevated spinal glial activation. Additionally, mild CCI increases spinal astrocyte activation but not microglia, suggesting astrocytes play a larger role in PAE-induced susceptibility to aberrant sensory processing. Leukocyte populations from PAE are altered under basal conditions (i.e., prior to secondary insult), as the distribution of leukocyte populations in lymphoid organs and other regions are different from those of controls. Lastly, following in vitro leukocyte stimulation, only PAE augments the immune response to antigen stimulation as assessed by heightened production of TNF-α and IL-1ß. CONCLUSIONS: These studies demonstrate PAE may prime spinal astrocytes and peripheral leukocytes that contribute to enduring susceptibility to adult-onset neuropathic pain that is not apparent until a secondary insult later in life.

Prenatal alcohol exposure potentiates chronic neuropathic pain, spinal glial and immune cell activation and alters sciatic nerve and DRG cytokine levels.

A growing body of evidence indicates that prenatal alcohol exposure (PAE) may predispose individuals to secondary medical disabilities later in life. Animal models of PAE reveal neuroimmune sequelae such as elevated brain astrocyte and microglial activation with corresponding region-specific changes in immune signaling molecules such as cytokines and chemokines. The aim of this study was to evaluate the effects of moderate PAE on the development and maintenance of allodynia induced by chronic constriction injury (CCI) of the sciatic nerve in adult male rat offspring. Because CCI allodynia requires the actions of glial cytokines, we analyzed lumbar spinal cord glial and immune cell surface markers indicative of their activation levels, as well as sciatic nerve and dorsal root ganglia (DRG) cytokines in PAE offspring in adulthood. While PAE did not alter basal sensory thresholds before or after sham manipulations, PAE significantly potentiated adult onset and maintenance of allodynia. Microscopic analysis revealed exaggerated astrocyte and microglial activation, while flow cytometry data demonstrated increased proportions of immune cells with cell surface major histocompatibility complex II (MHCII) and ß-integrin adhesion molecules, which are indicative of PAE-induced immune cell activation. Sciatic nerves from CCI rats revealed that PAE potentiated the proinflammatory cytokines interleukin (IL)-1ß, IL-6 and tumor necrosis factor-alpha (TNFα) protein levels with a simultaneous robust suppression of the anti-inflammatory cytokine, IL-10. A profound reduction in IL-10 expression in the DRG of PAE neuropathic rats was also observed. Taken together, our results provide novel insights into the vulnerability that PAE produces for adult-onset central nervous system (CNS) pathological conditions from peripheral nerve injury.

Moderate Prenatal Alcohol Exposure Alters Functional Connectivity in the Adult Rat Brain.

BACKGROUND: Past studies of moderate prenatal alcohol exposure (PAE) have focused on specific brain regions, neurotransmitter systems, and behaviors. However, the effects of PAE on brain function and behavior are complex and not limited to discrete brain regions. Thus, there is a critical need to understand the global effects of moderate PAE on neural function. A primary aim of this research was to explore the functional relationships in neural activity of spatially distinct areas by applying a widely used computational algorithm-group-independent component analysis (gICA)-to resting-state functional magnetic resonance imaging data from rats exposed to either an alcohol or saccharin control solution via maternal consumption during pregnancy. METHODS: Long-Evans rat dams consumed either 5% (v/v) alcohol or a saccharin control solution throughout gestation. Adult offspring from each prenatal treatment group were anesthetized for functional, structural, and perfusion magnetic resonance-based image acquisition sequences. gICA was applied to the functional data to extract components. To determine connectivity, component time-course correlations were computed and compared. Additionally, spectral power analyses were utilized as an additional measure of functional connectivity. Finally, blood perfusion-assessed by arterial spin labeling-and whole-brain volumetric analyses were evaluated. RESULTS: Analyses revealed 17 components in several brain regions such as the cortex, hippocampus, and thalamus. PAE was associated with reductions in coordinated activity between components, especially in males. PAE was also associated with reductions in low-frequency spectral power, an effect that was more robust in females. Brain volumetric analyses revealed sex-dependent reductions in females while blood flow analyses revealed sex-dependent reductions in males. CONCLUSIONS: Moderate PAE leads to persistent changes in functional connectivity in the absence of whole-brain volume or blood flow measures. Future studies will investigate the relationships between alterations in functional network connectivity and behavior.

Alcohol Use During Pregnancy is Associated with Specific Alterations in MicroRNA Levels in Maternal Serum.

BACKGROUND: Given the challenges of confirming prenatal alcohol exposure (PAE) during pregnancy using currently established biomarkers of alcohol consumption, we examined whether serum microRNAs (miRNAs) may serve as stable biomarkers for PAE. Alterations in the levels of specific circulating miRNAs have been associated with various disease states and in animal models of fetal alcohol spectrum disorder. METHODS: Pregnant women in this prospective study were recruited from substance abuse and general maternity clinics affiliated with the University of New Mexico. Serum was collected at the time of admission for delivery from 14 subjects who reported ≥1 binge-drinking episode or ≥3 drinks/wk during pregnancy and 16 subjects who reported abstinence during pregnancy and tested negative for 5 ethanol biomarkers. Total RNA was isolated from serum and used for microarray analysis. RESULTS: False discovery rate-corrected analyses of covariance revealed that 55 miRNAs were significantly altered between the 2 groups. Hierarchical clustering using only the significantly altered miRNAs grouped samples into alcohol-consuming and non-alcohol-consuming individuals. Discriminant analysis then identified miRs-122*, -126, -216b, -221*, -3119, -3942-5p, -4704-3p, -4743, -514-5p, and -602 as the top 10 discriminators between the 2 groups. Ingenuity Pathway Analysis of putative miRNA targets illustrated that miRNAs identified in this study are involved in biological pathways that mediate the effects of alcohol, such as brain-derived neurotrophic factor, ERK1/2, and PI3K/AKT signaling. CONCLUSIONS: This is the first report of alterations in serum miRNA expression that are associated with alcohol use during human pregnancy. These results suggest that serum miRNAs could be useful as biomarkers of alcohol exposure.

High incidence of ErbB3, ErbB4, and MET expression in ovarian cancer.

Ovarian cancer is the leading cause of death from gynecologic cancers in the United States. Failure may be due to variable expression and/or complex interactions of growth factor receptors in individual tumors. As ErbB3-MET cooperativity is implicated in solid tumor resistance to EGFR/ErbB2 inhibitors, we evaluated expression of MET and all 4 ErbB family members in ovarian cancers. Tissue arrays were prepared from archival formalin-fixed paraffin-embedded tumor samples, including 202 ovarian carcinomas (Stage I-IV) and controls. Of 202 patient samples, only 25% were positive for EGFR and 35% for ErbB2 expression. ErbB3, ErbB4, and MET showed marked expression in 76%, 98%, and 96% of cases. Consistent with high incidence, there was no significant correlation for expression of ErbB3, ErbB4, or MET with outcome. On the basis of their high expression in the majority of cases, inhibitors targeting ErbB3, ErbB4, and/or MET may be broadly applicable as therapeutic agents in this disease.

The histamine H3 receptor inverse agonist SAR-152954 reverses deficits in long-term potentiation associated with moderate prenatal alcohol exposure.

Prenatal alcohol exposure can have persistent effects on learning, memory, and synaptic plasticity. Previous work from our group demonstrated deficits in long-term potentiation (LTP) of excitatory synapses on dentate gyrus granule cells in adult offspring of rat dams that consumed moderate levels of alcohol during pregnancy. At present, there are no pharmacotherapeutic agents approved for these deficits. Prior work established that systemic administration of the histaminergic H3R inverse agonist ABT-239 reversed deficits in LTP observed following moderate PAE. The present study examines the effect of a second H3R inverse agonist, SAR-152954, on LTP deficits following moderate PAE. We demonstrate that systemic administration of 1 mg/kg of SAR-152954 reverses deficits in potentiation of field excitatory post-synaptic potentials (fEPSPs) in adult male rats exposed to moderate PAE. Time-frequency analyses of evoked responses revealed PAE-related reductions in power during the fEPSP, and increased power during later components of evoked responses which are associated with feedback circuitry that are typically not assessed with traditional amplitude-based measures. Both effects were reversed by SAR-152954. These findings provide further evidence that H3R inverse agonism is a potential therapeutic strategy to address deficits in synaptic plasticity associated with PAE.

Organization of spontaneous spatial behaviors under dark conditions is unaffected in adult male and female long-Evans rats after moderate prenatal alcohol exposure.

Prenatal alcohol exposure can produce disruptions in a wide range of cognitive functions, but it is especially detrimental to spatial navigation. In open environments, rodents organize their spatial behaviors around centralized locations, termed home bases, from which they make circuitous and slow locomotor trips (progressions) into the rest of the environment. Open-field behaviors are organized even under darkened test conditions, suggesting a role for self-motion cues (vestibular, motor, etc.). The impact of moderate prenatal alcohol exposure (mPAE) on the organization of spontaneous open-field behaviors under darkened conditions has not been investigated. Here we tested adult female and male rats with mPAE or saccharin control exposure in a circular open field for 30 min in a testing room that was made completely dark. While general locomotion, as measured by reductions in travel distance and increased stop duration, decreased across the test session, the organization of these behaviors, as measured by stop duration, home base establishment, home base stability, progression accuracy, and scaling of peak speeds with progression length, did not differ between mPAE and saccharin control rats. Together, the findings strongly suggest that spontaneous movement organization in relation to self-motion cues remains intact in adult mPAE rats. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Sex-specific alterations in cognitive control following moderate prenatal alcohol exposure and transient systemic hypoxia ischemia in the rat.

BACKGROUND: Prenatal alcohol exposure (PAE) continues to be a worldwide problem. Affected offspring display impaired neurodevelopment, including difficulties with executive control. Although PAE has also been associated with decreased blood flow to fetuses, the relationship between PAE and altered blood flow is not well understood. METHODS: We used preclinical models of PAE, transient systemic hypoxia ischemia (TSHI), and PAE + TSHI combined to assess the effects on neurodevelopmental outcomes using translationally relevant touchscreen operant platform testing. Twenty-eight Long-Evans (Blue Spruce, Strain HsdBlu:LE) dams were randomly assigned to one of four experimental groups: Saccharin Control (Sham), 5% Ethanol (PAE), TSHI, or 5% Ethanol and TSHI (PAE + TSHI). Dams consumed either saccharin or 5% ethanol during gestation. TSHI was induced on Embryonic Day 19 (E19) during an open laparotomy where the uterine arteries were transiently occluded for 1 h. Pups were born normally and, after weaning, were separated by sex. A total of 80 offspring, 40 males and 40 females, were tested on the 5-Choice Continuous Performance paradigm (5C-CPT). RESULTS: Female offspring were significantly impacted by TSHI, but not PAE, with an increase in false alarms and a decrease in hit rates, omissions, accuracy, and correct choice latencies. In contrast, male offspring were mildly affected by PAE, but not TSHI, showing decreases in premature responses and increases in accuracy. No significant interactions between PAE and TSHI were detected on any measure. CONCLUSION: Transient systemic hypoxia ischemia impaired performance on the 5C-CPT in females, leading to a bias toward stimulus responsivity regardless of stimulus type. In contrast, TSHI did not affect male offspring, and only slight effects of PAE were seen. Together, these data suggest that TSHI in females may cause alterations in cortical structures that override alterations caused by moderate PAE.

Impact of two different rodent diets on maternal ethanol consumption, serum ethanol concentration and pregnancy outcome measures.

Recent studies report varying levels of ethanol consumption by rodents maintained on different commercially available laboratory diets. As varied ethanol consumption by dams may impact offspring outcome measures in prenatal ethanol exposure paradigms, we compared ethanol consumption by rats maintained on the Envigo 2920 diet, used in our vivarium, with an isocalorically equivalent PicoLab 5L0D diet used in some alcohol consumption studies. Compared to 5L0D diet, female rats maintained on 2920 diet consumed 14% less ethanol during daily 4-h drinking sessions prior to pregnancy and 28% less ethanol during gestation. Rat dams consuming 5L0D diet gained significantly less weight during pregnancy. However, their pup birth weights were significantly higher. A subsequent study revealed that hourly ethanol consumption was not different between diets during the first 2 h, but was significantly lower on 2920 diet at the end of the third and fourth hours. The mean serum ethanol concentration in 5L0D dams after the first 2 h of drinking was 46 mg/dL compared to 25 mg/dL in 2920 dams. Further, ethanol consumption at the 2-h blood sampling time point was more variable in 2920 dams compared to 5L0D dams. An in vitro analysis mixing each powdered diet with 5% ethanol in acidified saline revealed that a 2920 diet suspension adsorbed more aqueous medium than the 5L0D diet suspension. The total ethanol remaining in aqueous supernatant of 5L0D mixtures was nearly twice the amount of ethanol in supernatants of the 2920 mixtures. These results suggest that the 2920 diet expands to a greater extent in aqueous medium than the 5L0D diet. We speculate that increasing adsorption of water and ethanol by the 2920 diet may reduce or delay the amount of ethanol absorbed and may decrease serum ethanol concentration to a greater extent than would be predicted from the amount of ethanol consumed.

Effect of moderate prenatal ethanol exposure on the differential expression of two histamine H3 receptor isoforms in different brain regions of adult rat offspring.

We have reported that prenatal alcohol exposure (PAE) elevates histamine H3 receptor (H3R) agonist-mediated inhibition of glutamatergic neurotransmission in the dentate gyrus. Here, we hypothesized that PAE alters the expression of two prominent H3R isoforms namely, the rH3A and rH3C isoforms, which have differing intrinsic activities for H3R agonists, in a manner that may contribute to heightened H3R function in PAE rats. In contrast to our predictions, we found different effects of sex and PAE in various brain regions with significant interactions between sex and PAE in dentate gyrus and entorhinal cortex for both isoforms. Subsequently, to confirm the PAE-and sex-induced differences on H3R isoform mRNA expression, we developed a polyclonal antibody selective for the rH3A inform. Western blots of rH3A mRNA-transfected HEK-293 cells identified a ~ 48 kDa band of binding consistent with the molecular weight of rH3A, thus confirming antibody sensitivity for rH3A protein. In parallel, we also established a pan-H3R knockout mice line to confirm antibody specificity in rodent brain membranes. Both qRT-PCR and H3R agonist-stimulated [35S]-GTPγS binding confirmed the absence of mH3A mRNA and H3 receptor-effector coupling in H3R knockout (KO) mice. Subsequent western blotting studies in both rat and mouse brain membranes were unable to detect rH3A antibody binding at ~48 kDa. Rather, the H3RA antibody bound to a ~ 55 kDa band in both rat and mouse membranes, including H3R KO mice, suggesting H3RA binding was not specific for H3Rs in rodent membranes. Subsequent LC/MS analysis of the ~55 kDa band in frontal cortical membranes identified the highly abundant beta subunit of ATPase in both WT and KO mice. Finally, LC/MS analysis of the ~48 kDa band from rH3A mRNA-transfected HEK-293 cell membranes was able to detect rH3A protein, but its presence was below the limits of quantitative reliability. We conclude that PAE alters rH3A and rH3C mRNA expression in some of the same brain regions where we have previously reported PAE-induced alterations in H3R-effector coupling. However, interpreting the functional consequences of altered H3R isoform expression was limited given the technical challenges of measuring the relatively low abundance of rH3A protein in native membrane preparations.

Prenatal cannabinoid exposure: why expecting individuals should take a pregnancy pause from using cannabinoid products.

Cannabinoid use in all populations is increasing as legalization across the United States continues. Concerningly, there is a lack of caution provided by medical providers to pregnant individuals as to the impact the use of cannabinoids could have on the developing fetus. Research continues in both the preclinical and clinical areas, and is severely needed, as the potency of delta-9-tetrahydrocannabinol (THC), the primary psychoactive component of cannabis, has increased dramatically since the initial studies were completed. Thus far, clinical studies raise compelling evidence for short term memory deficits, impulse control issues, and attention deficiencies following prenatal cannabinoid exposure (PCE). These changes may be mediated through epigenetic modifications that not only impact the current offspring but could carry forward to future generations. While additional studies are needed, a pregnancy pause from cannabinoid products should be strongly recommended by providers to ensure the optimal health and well-being of our future generations.

Prenatal alcohol exposure dysregulates spinal and circulating immune cell circular RNA expression in adult female rats with chronic sciatic neuropathy.

Alcohol consumption during pregnancy is associated with Fetal Alcohol Spectrum Disorders (FASD) that results in a continuum of central nervous system (CNS) deficits. Emerging evidence from both preclinical and clinical studies indicate that the biological vulnerability to chronic CNS disease in FASD populations is driven by aberrant neuroimmune actions. Our prior studies suggest that, following minor nerve injury, prenatal alcohol exposure (PAE) is a risk factor for developing adult-onset chronic pathological touch sensitivity or allodynia. Allodynia in PAE rats occurs concurrently with heightened proinflammatory peripheral and spinal glial-immune activation. However, minor nerve-injured control rats remain non-allodynic, and corresponding proinflammatory factors are unaltered. A comprehensive molecular understanding of the mechanism(s) that underlie PAE-induced proinflammatory bias during adulthood remains elusive. Non-coding circular RNAs (circRNAs) are emerging as novel modulators of gene expression. Here, we hypothesized that PAE induces dysregulation of circRNAs that are linked to immune function under basal and nerve-injured conditions during adulthood. Utilizing a microarray platform, we carried out the first systematic profiling of circRNAs in adult PAE rats, prior to and after minor nerve injury. The results demonstrate a unique circRNA profile in adult PAE rats without injury; 18 circRNAs in blood and 32 spinal circRNAs were differentially regulated. Following minor nerve injury, more than 100 differentially regulated spinal circRNAs were observed in allodynic PAE rats. Bioinformatic analysis identified that the parental genes of these circRNAs are linked to the NF-κB complex, a central transcription factor for pain-relevant proinflammatory cytokines. Quantitative real-time PCR was employed to measure levels of selected circRNAs and linear mRNA isoforms. We have validated that circVopp1 was significantly downregulated in blood leukocytes in PAE rats, concurrent with downregulation of Vopp1 mRNA levels. Spinal circVopp1 levels were upregulated in PAE rats, regardless of nerve injury. Additionally, PAE downregulated levels of circItch and circRps6ka3, which are linked to immune regulation. These results demonstrate that PAE exerts long-lasting dysregulation of circRNA expression in blood leukocytes and the spinal cord. Moreover, the spinal circRNA expression profile following peripheral nerve injury is differentially modulated by PAE, potentially contributing to PAE-induced neuroimmune dysregulation.

Global expression analysis of endometrial cancer cells in response to progesterone identifies new therapeutic targets.

Progesterone prevents development of endometrial cancers through its receptor (PR) although the molecular mechanisms have yet to be fully characterized. In this study, we performed a global analysis of gene regulation by progesterone using human endometrial cancer cells that expressed PR endogenously or exogenously. We found progesterone strongly inhibits multiple components of the platelet derived growth factor receptor (PDGFR), Janus kinase (JAK), signal transducer and activator of transcription (STAT) pathway through PR. The PDGFR/JAK/STAT pathway signals to control numerous downstream targets including AP-1 transcription factors Fos and Jun. Treatment with inhibitors of the PDGFR/JAK/STAT pathway significantly blocked proliferation in multiple novel patient-derived organoid models of endometrial cancer, and activation of this pathway was found to be a poor prognostic signal for the survival of patients with endometrial cancer from The Cancer Genome Atlas. Our study identifies this pathway as central to the growth-limiting effects of progesterone in endometrial cancer and suggests that inhibitors of PDGFR/JAK/STAT should be considered for future therapeutic interventions.

Detection of prenatal alcohol exposure using machine learning classification of resting-state functional network connectivity data.

Fetal Alcohol Spectrum Disorder (FASD), a wide range of physical and neurobehavioral abnormalities associated with prenatal alcohol exposure (PAE), is recognized as a significant public health concern. Advancements in the diagnosis of FASD have been hindered by a lack of consensus in diagnostic criteria and limited use of objective biomarkers. Previous research from our group utilized resting-state functional magnetic resonance imaging (fMRI) to measure functional network connectivity (FNC), which revealed several sex- and region-dependent alterations in FNC as a result of moderate PAE relative to controls. Considering that FNC is sensitive to moderate PAE, this study explored the use of FNC data and machine learning methods to detect PAE among a sample of rodents exposed to alcohol prenatally and controls. We utilized previously acquired resting state fMRI data collected from adult rats exposed to moderate levels of prenatal alcohol (PAE) or a saccharin control solution (SAC) to assess FNC of resting state networks extracted by spatial group independent component analysis (GICA). FNC data were subjected to binary classification using support vector machine (SVM) -based algorithms and leave-one-out-cross validation (LOOCV) in an aggregated sample of males and females (n = 48; 12 male PAE, 12 female PAE, 12 male SAC, 12 female SAC), a males-only sample (n = 24; 12 PAE, 12 SAC), and a females-only sample (n = 24; 12 PAE, 12 SAC). Results revealed that a quadratic SVM (QSVM) kernel was significantly effective for PAE detection in females. QSVM kernel-based classification resulted in accuracy rates of 62.5% for all animals, 58.3% for males, and 79.2% for females. Additionally, qualitative evaluation of QSVM weights implicates an overarching theme of several hippocampal and cortical networks in contributing to the formation of correct classification decisions by QSVM. Our results suggest that binary classification using QSVM and adult female FNC data is a potential candidate for the translational development of novel and non-invasive techniques for the identification of FASD.

Prenatal Alcohol Exposure and Chorioamnionitis Results in Microstructural Brain Injury in a Preclinical Investigation.

BACKGROUND: Prenatal Alcohol Exposure (PAE) impacts 2% to 5% of infants born in the United States yearly. Women who consume alcohol during pregnancy have a five-fold increased rate of Chorioamnionitis (CHORIO). Both PAE and CHORIO cause microstructural injury to multiple brain regions including major white matter tracts. OBJECTIVE: Utilizing two previously established animal models, we hypothesized that the combination of PAE+CHORIO would result in greater deficits in myelination and structural integrity than PAE alone. MATERIAL AND METHODS: Pregnant Long-Evans rats voluntarily drank 5% ethanol or saccharin until Gestational Day 19 (GD). On GD19, CHORIO was induced in one group of PAE dams by a 30 min uterine artery occlusion and injection of Lipopolysaccharide (LPS) into each amniotic sac. The remaining PAE dams and saccharin controls underwent sham surgery. Pups were born on GD22 and weaned on Postnatal Day 24 (PD). On PD28, offspring were sacrificed, and their brains examined using ex-vivo Diffusion Tensor Imaging (DTI). RESULTS: Compared to control, PAE alone did not affect offspring birth weights, mortality or any DTI measures. In contrast, PAE+CHORIO significantly reduced offspring survival and, in surviving pups, increased Radial Diffusivity (RD) in medial frontal cortex and decreased Fractional Anisotropy (FA) in medial and ventral frontal cortex and within capsular regions. CONCLUSION: The combination of moderate PAE+CHORIO results in an increased mortality, concomitant with diffuse microstructural brain injury noted in young adolescent offspring at PD28. Future studies should examine the extent to which PAE exacerbates the damage caused by CHORIO alone and whether these deficits persist into adulthood.

Moderate prenatal alcohol exposure impairs performance by adult male rats in an object-place paired-associate task.

Memory impairments, including spatial and object processing, are often observed in individuals with Fetal Alcohol Spectrum Disorders. The neurobiological basis of memory deficits after prenatal alcohol exposure (PAE) is often linked to structural and functional alterations in the medial temporal lobe, including the hippocampus. Recent evidence suggests that the medial temporal lobe plays a critical role in processing high-order sensory stimuli such as complex objects and their associated locations in space. In the first experiment, we tested male rat offspring with moderate PAE in a medial temporal-dependent object-place paired-associate (OPPA) task. The OPPA task requires a conditional discrimination between an identical pair of objects presented at two spatial locations 180° opposite arms of a radial arm maze. Food reinforcement is contingent upon selecting the correct object of the pair for a given spatial location. Adult rats were given a total of 10 trials per day over 14 consecutive days of training. PAE male rats made significantly more errors than male saccharin (SACC) control rats during acquisition of the OPPA task. In Experiment 2, rats performed an object-discrimination task in which a pair of objects were presented in a single arm of the maze. Moderate PAE and SACC control rats exhibited comparable performance. The results suggest that moderate PAE rats can learn to discriminate objects, but are impaired when required to discriminate between objects on the basis of spatial location in the environment.

The association between prenatal alcohol exposure and protein expression in human placenta.

BACKGROUND: The need for earlier recognition of children at risk for neurobehavioral problems associated with prenatal ethanol exposure (PAE) has prompted investigations of biomarkers prognostic for altered fetal development. Here, we examined whether PAE alters the expression of angiogenesis-related proteins and cytokines in human placenta in subjects from an Ethanol, Neurodevelopment, Infant and Child Health prospective cohort. METHODS: PAE was ascertained by screening questionnaires, Time-line Follow-back interviews and a panel of ethanol biomarkers at two study visits. After delivery, placental tissue samples were collected for protein analysis. RESULTS: No significant differences in the prevalence of substance use, demographic or medical characteristics were observed between the No PAE and PAE groups. PAE was associated with significant reductions in placental expression of VEGFR2 and annexin-A4, while the levels of VEGFR1 and CCM-3 trended downward. A trend toward higher expression of the cytokines TNF-α and IL-13 was also observed in the PAE group. Receiver operating characteristic analyses of the data demonstrated a moderate-to-high degree of diagnostic accuracy for individual placental proteins. Combinations of proteins substantially increased their ability to differentiate between PAE and No PAE subjects. CONCLUSIONS: These results establish the feasibility of harvesting placental tissue for protein analyses of PAE in a prospective manner. In addition, given the importance of vascular remodeling in both placenta and developing brain, the role of angiogenic and cytokine proteins in this process warrants further investigation for their utility for predicting alterations in brain development, as well as their mechanistic role in PAE-induced pathology.

Impact of moderate prenatal alcohol exposure on histaminergic neurons, histidine decarboxylase levels and histamine H2 receptors in adult rat offspring.

We have reported that moderate prenatal alcohol exposure (PAE) elevates histamine H3 receptor-mediated inhibition of glutamatergic neurotransmission in dentate gyrus (DG), and that the H3 receptor antagonist ABT-239 ameliorates PAE-induced deficits in DG long-term potentiation. Here, we investigated whether PAE alters other markers of histaminergic neurotransmission. Long-Evans rat dams voluntarily consumed either a 0% or a 5% ethanol solution 4 h each day throughout gestation. Young adult female offspring from each prenatal treatment group were used in histidine decarboxylase (HDC) immunohistochemical studies of histamine neuron number in ventral hypothalamus, quantitative Western blotting studies of HDC expression in multiple brain regions, radiohistochemical studies of H2 receptor density in multiple brain regions, and in biochemical studies of H2 receptor-effector coupling in dentate gyrus. Rat dams consumed a mean of 1.90 g of ethanol/kg/day during pregnancy. This level of consumption did not affect maternal weight gain, offspring birth weight, or litter size. PAE did not affect the number of HDC-positive neurons in ventral hypothalamus. However, HDC expression was reduced in frontal cortex, dentate gyrus, and cerebellum of PAE rats compared to controls. Specific [125I]-iodoaminopotentidine binding to H2 receptors was not altered in any of the brain regions measured, nor was basal or H2 receptor agonist-stimulated cAMP accumulation in DG altered in PAE rats compared to controls. These results suggest that not all markers of histaminergic neurotransmission are altered by PAE. However, the observation that HDC levels were reduced in the same brain regions where elevated H3 receptor-effector coupling was observed previously raises the question of whether a cause-effect relationship exists between HDC expression and H3 receptor function in affected brain regions of PAE rats. This relationship, along with the question of why these effects occur in some, but not all brain regions, requires more-detailed investigation.