Pregnancy-induced oxidative stress and inflammation are not associated with impaired maternal neuronal activity or memory function.

Pregnancy is associated with neural and behavioral plasticity, systemic inflammation, and oxidative stress. Yet, the impact of inflammation and oxidative stress on maternal neural and behavioral plasticity during pregnancy are unclear. We hypothesized that healthy pregnancy transiently reduces learning and memory, and these deficits are associated with pregnancy-induced elevations in inflammation and oxidative stress. Cognitive performance was tested using novel object recognition (recollective memory), Morris water maze (spatial memory), and open field (anxiety-like) behavior tasks in female Sprague-Dawley rats of varying reproductive states [non-pregnant (nulliparous), pregnant (near term), and 1-2 months post-pregnancy (primiparous); n = 7-8/group]. Plasma and CA1 proinflammatory cytokines were measured using a MILLIPLEX® magnetic bead assay. Plasma oxidative stress was measured via advanced oxidation protein products (AOPP) assay. CA1 markers of oxidative stress, neuronal activity, and apoptosis were quantified via western blotting. Our results demonstrate CA1 oxidative stress-associated markers were elevated in pregnant compared to nulliparous rats (p ≤ 0.017) but were equivalent levels in pregnant and primiparous rats. In contrast, reproductive state did not impact CA1 inflammatory cytokines, neuronal activity, or apoptosis. Likewise, there was no effect of reproductive state on recollective or spatial memory. Even so, spatial learning was impaired (p ≤ 0.007) while anxiety-like behavior (p ≤ 0.034) was reduced in primiparous rats. Overall, our data suggest maternal hippocampal CA1 is protected from systemic inflammation but vulnerable to peripartum oxidative stress. Peripartum oxidative stress elevations, such as in pregnancy complications, may contribute to peripartum neural and behavioral plasticity.

Oxidative stress induces release of mitochondrial DNA into the extracellular space in human placental villous trophoblast BeWo cells.

Circulating cell-free mitochondrial DNA (ccf-mtDNA) is an indicator of cell death, inflammation, and oxidative stress. ccf-mtDNA differs in pregnancies with placental dysfunction from healthy pregnancies and the direction of this difference depends on gestational age and method of mtDNA quantification. Reactive oxygen species (ROS) trigger release of mtDNA from non-placental cells; yet it is unknown whether trophoblast cells release mtDNA in response to oxidative stress, a common feature of pregnancies with placental pathology. We hypothesized that oxidative stress would induce cell death and release of mtDNA from trophoblast cells. BeWo cells were treated with antimycin A (10-320 mM) or rotenone (0.2-50 mM) to induce oxidative stress. A multiplex real-time quantitative PCR (qPCR) assay was used to quantify mtDNA and nuclear DNA in membrane bound, non-membrane bound, and vesicular-bound forms in cell culture supernatants and cell lysates. Treatment with antimycin A increased ROS (p<0.0001), induced cell necrosis (p=0.0004) but not apoptosis (p=0.6471) and was positively associated with release of membrane-bound and non-membrane bound mtDNA (p<0.0001). Antimycin A increased mtDNA content in exosome-like extracellular vesicles (vesicular-bound form; p=0.0019) and reduced autophagy marker expression (LC3A/B, p=0.0002; p62, p<0.001). Rotenone treatment did not influence mtDNA release or cell death (p>0.05). Oxidative stress induces release of mtDNA into the extracellular space and causes non-apoptotic cell death and a reduction in autophagy markers in BeWo cells, an established in vitro model of human trophoblast cells. Intersection between autophagy and necrosis may mediate the release of mtDNA from the placenta in pregnancies exposed to oxidative stress.

Sex-dependent effects of chronic intermittent hypoxia: implication for obstructive sleep apnea.

BACKGROUND: Obstructive sleep apnea (OSA) affects 10-26% of adults in the United States with known sex differences in prevalence and severity. OSA is characterized by elevated inflammation, oxidative stress (OS), and cognitive dysfunction. However, there is a paucity of data regarding the role of sex in the OSA phenotype. Prior findings suggest women exhibit different OSA phenotypes than men, which could result in under-reported OSA prevalence in women. To examine the relationship between OSA and sex, we used chronic intermittent hypoxia (CIH) to model OSA in rats. We hypothesized that CIH would produce sex-dependent phenotypes of inflammation, OS, and cognitive dysfunction, and these sex differences would be dependent on mitochondrial oxidative stress (mtOS). METHODS: Adult male and female Sprague Dawley rats were exposed to CIH or normoxia for 14 days to examine the impact of sex on CIH-associated circulating inflammation (IL-1ß, IL-6, IL-10, TNF-α), circulating steroid hormones, circulating OS, and behavior (recollective and spatial memory; gross and fine motor function; anxiety-like behaviors; and compulsive behaviors). Rats were implanted with osmotic minipumps containing either a mitochondria-targeting antioxidant (MitoTEMPOL) or saline vehicle 1 week prior to CIH initiation to examine how inhibiting mtOS would affect the CIH phenotype. RESULTS: Sex-specific differences in CIH-induced inflammation, OS, motor function, and compulsive behavior were observed. In female rats, CIH increased inflammation (plasma IL-6 and IL-6/IL-10 ratio) and impaired fine motor function. Conversely, CIH elevated circulating OS and compulsivity in males. These sex-dependent effects of CIH were blocked by inhibiting mtOS. Interestingly, CIH impaired recollective memory in both sexes but these effects were not mediated by mtOS. No effects of CIH were observed on spatial memory, gross motor function, or anxiety-like behavior, regardless of sex. CONCLUSIONS: Our results indicate that the impact of CIH is dependent on sex, such as an inflammatory response and OS response in females and males, respectively, that are mediated by mtOS. Interestingly, there was no effect of sex or mtOS in CIH-induced impairment of recollective memory. These results indicate that mtOS is involved in the sex differences observed in CIH, but a different mechanism underlies CIH-induced memory impairments.

Sleep apnea is a common sleeping condition in adults with a wide range of symptoms that include inflammation, oxidative stress, memory problems, anxiety, and compulsivity. Men are diagnosed with sleep apnea more often than women. Although there is limited information on how sleep apnea affects men and women differently, previous studies suggest that women may exhibit different sleep apnea symptoms than men. To examine the impact of male and female sex on common sleep apnea symptoms, we exposed adult male and female rats to a model of sleep apnea called chronic intermittent hypoxia (CIH). We found that many effects of CIH were different in males and females. CIH females had increased inflammation and motor problems, whereas CIH males had increased oxidative stress and compulsivity. To investigate the reason for these CIH sex differences, we blocked mitochondrial oxidative stress. Blocking mitochondrial oxidative stress decreased CIH associated sex differences. However, blocking mitochondrial oxidative stress had no impact on CIH-induced memory impairment that was observed in male and female rats. Our findings support previous reports that suggest that women exhibit different sleep apnea symptoms than men. Further, we extend these findings by showing that mitochondrial oxidative stress is involved in these sex differences. Clinically, patients diagnosed with sleep apnea are typically treated with continuous positive airway pressure (CPAP) machines, which have high rates of non-compliance (15­40%). Therefore, understanding why sleep apnea is causing these symptoms will be important in developing therapeutics.

Chronic intermittent hypoxia-induced hypertension: the impact of sex hormones.

Obstructive sleep apnea, a common form of sleep-disordered breathing, is characterized by intermittent cessations of breathing that reduce blood oxygen levels and contribute to the development of hypertension. Hypertension is a major complication of obstructive sleep apnea that elevates the risk of end-organ damage. Premenopausal women have a lower prevalence of obstructive sleep apnea and cardiovascular disease than men and postmenopausal women, suggesting that sex hormones play a role in the pathophysiology of sleep apnea-related hypertension. The lack of protection in men and postmenopausal women implicates estrogen and progesterone as protective agents but testosterone as a permissive agent in sleep apnea-induced hypertension. A better understanding of how sex hormones contribute to the pathophysiology of sleep apnea-induced hypertension is important for future research and possible hormone-based interventions. The effect of sex on the pathophysiology of sleep apnea and associated intermittent hypoxia-induced hypertension is of important consideration in the screening, diagnosis, and treatment of the disease and its cardiovascular complications. This review summarizes our current understanding of the impact of sex hormones on blood pressure regulation in sleep apnea with a focus on sex differences.

Sex-dependent effects of chronic intermittent hypoxia: Implication for obstructive sleep apnea.

Background Obstructive sleep apnea (OSA) affects 10-26% of adults in the United States with known sex differences in prevalence and severity. OSA is characterized by elevated inflammation, oxidative stress (OS), and cognitive dysfunction. However, there is a paucity of data regarding the role of sex in the OSA phenotype. Prior findings suggest women exhibit different OSA phenotypes than men, which could result in under-reported OSA prevalence in women. To examine the relationship between OSA and sex, we used chronic intermittent hypoxia (CIH) to model OSA in rats. We hypothesized that CIH would produce sex-dependent phenotypes of inflammation, OS, and cognitive dysfunction, and these sex differences would be dependent on mitochondrial oxidative stress (mtOS). Methods Adult male and female Sprague Dawley rats were exposed to CIH or normoxia for 14 days to examine the impact of sex on CIH-associated circulating inflammation (IL-1ß, IL-4, IL-6, IL-10, TNF-α), circulating OS, and behavior (recollective and spatial memory; gross and fine motor function; anxiety-like behaviors; and compulsive behaviors). A subset of rats was implanted with osmotic minipumps containing either a mitochondria-targeting antioxidant (MitoTEMPOL) or saline vehicle 1 week prior to CIH initiation to examine how inhibiting mtOS would affect the CIH phenotype. Results Sex-specific differences in CIH-induced inflammation, OS, motor function, and compulsive behavior were observed. In female rats, CIH increased inflammation (plasma IL-6 and IL-6/IL-10 ratio) and impaired fine motor function. Conversely, CIH elevated circulating OS and compulsivity in males. These sex-dependent effects of CIH were blocked by inhibiting mtOS. Interestingly, CIH impaired recollective memory in both sexes but these effects were not mediated by mtOS. No effects of CIH were observed on spatial memory, gross motor function, or anxiety-like behavior, regardless of sex. Conclusions Our results indicate that the impact of CIH is dependent on sex, such as an inflammatory response and OS response in females and males, respectively, that are mediated by mtOS. Interestingly, there was no effect of sex or mtOS in CIH-induced impairment of recollective memory. These results indicate that mtOS is involved in the sex differences observed in CIH, but a different mechanism underlies CIH-induced memory impairments.

Oxidative stress induces release of mitochondrial DNA into the extracellular space in human placental villous trophoblast BeWo cells.

Circulating cell-free mitochondrial DNA (ccf-mtDNA) is an indicator of cell death, inflammation, and oxidative stress. ccf-mtDNA differs in pregnancies with placental dysfunction from healthy pregnancies and the direction of this difference depends on gestational age and method of mtDNA quantification. Reactive oxygen species (ROS) trigger release of mtDNA from non-placental cells; yet it is unknown whether trophoblast cells release mtDNA in response to oxidative stress, a common feature of pregnancies with placental pathology. We hypothesized that oxidative stress would induce cell death and release of mtDNA from trophoblast cells. BeWo cells were treated with antimycin A (10-320 µM) or rotenone (0.2-50 µM) to induce oxidative stress. A multiplex real-time quantitative PCR (qPCR) assay was used to quantify mtDNA and nuclear DNA in membrane bound, non-membrane bound, and vesicular-bound forms in cell culture supernatants and cell lysates. Treatment with antimycin A increased ROS (p<0.0001), induced cell necrosis (p=0.0004) but not apoptosis (p=0.6471) and was positively associated with release of membrane-bound and non-membrane bound mtDNA (p<0.0001). Antimycin A increased mtDNA content in exosome-like extracellular vesicles (vesicular-bound form; p=0.0019) and reduced autophagy marker expression (LC3A/B, p=0.0002; p62, p<0.001). Rotenone treatment did not influence mtDNA release or cell death (p>0.05). Oxidative stress induces release of mtDNA into the extracellular space and causes non-apoptotic cell death and a reduction in autophagy markers in BeWo cells, an established in vitro model of human trophoblast cells. Intersection between autophagy and necrosis may mediate the release of mtDNA from the placenta in pregnancies exposed to oxidative stress. NEW & NOTEWORTHY: This is the first study to test whether trophoblast cells release mitochondrial DNA in response to oxidative stress and to identify mechanisms of release and biological forms of mtDNA from this cellular type. This research identifies potential cellular mechanisms that can be used in future investigations to establish the source and biomarker potential of circulating mitochondrial DNA in preclinical experimental models and humans.

Pregnancy-associated oxidative stress and inflammation are not associated with impaired maternal neuronal activity or memory function.

Pregnancy is associated with neural and behavioral plasticity, systemic inflammation, and oxidative stress. Yet, the impact of systemic inflammation and oxidative stress on maternal neural and behavioral plasticity during pregnancy are unclear. We hypothesized that the maternal hippocampal CA1, a brain region associated with cognition, would be protected from pregnancy-associated systemic elevations in inflammation and oxidative stress, mediating stable peripartum cognitive performance. Cognitive performance was tested using novel object recognition (recollective memory), Morris water maze (spatial memory), and open field (anxiety-like) behavior tasks in female Sprague-Dawley rats of varying reproductive states [non-pregnant (nulliparous), pregnant (near term), and two months post-pregnancy (primiparous); n = 7-8/group]. Plasma and CA1 proinflammatory cytokines were measured using a MILLIPLEX® magnetic bead assay. Plasma oxidative stress was measured via advanced oxidation protein products (AOPP) assay. CA1 markers of oxidative stress, neuronal activity, and apoptosis were quantified via western blotting. Our results demonstrate CA1 oxidative stress-associated markers were elevated in pregnant compared to nulliparous rats ( p ≤ 0.017) but were equivalent levels in pregnant and primiparous rats. In contrast, reproductive state did not impact CA1 inflammatory cytokines, neuronal activity, or apoptosis. Likewise, there was no effect of reproductive state on recollective or spatial memory. Even so, spatial learning was impaired ( p ≤ 0.007) while anxiety-like behavior ( p ≤ 0.034) was reduced in primiparous rats. Overall, our data suggest maternal hippocampal CA1 is protected from systemic inflammation but vulnerable to peripartum oxidative stress. Thus, peripartum oxidative stress elevations, such as in pregnancy complications, may contribute to peripartum neural and behavioral plasticity.

Sex and age differences in social and cognitive function in offspring exposed to late gestational hypoxia.

BACKGROUND: Gestational sleep apnea is a hypoxic sleep disorder that affects 8-26% of pregnancies and increases the risk for central nervous system dysfunction in offspring. Specifically, there are sex differences in the sensitivity of the fetal hippocampus to hypoxic insults, and hippocampal impairments are associated with social dysfunction, repetitive behaviors, anxiety, and cognitive impairment. Yet, it is unclear whether gestational sleep apnea impacts these hippocampal-associated functions and if sex and age modify these effects. To examine the relationship between gestational sleep apnea and hippocampal-associated behaviors, we used chronic intermittent hypoxia (CIH) to model late gestational sleep apnea in pregnant rats. We hypothesized that late gestational CIH would produce sex- and age-specific social, anxiety-like, repetitive, and cognitive impairments in offspring. METHODS: Timed pregnant Long-Evans rats were exposed to CIH or room air normoxia from GD 15-19. Behavioral testing of offspring occurred during either puberty or young adulthood. To examine gestational hypoxia-induced behavioral phenotypes, we quantified hippocampal-associated behaviors (social function, repetitive behaviors, anxiety-like behaviors, and spatial memory and learning), hippocampal neuronal activity (glutamatergic NMDA receptors, dopamine transporter, monoamine oxidase-A, early growth response protein 1, and doublecortin), and circulating hormones in offspring. RESULTS: Late gestational CIH induced sex- and age-specific differences in social, repetitive, and memory functions in offspring. In female pubertal offspring, CIH impaired social function, increased repetitive behaviors, and elevated circulating corticosterone levels but did not impact memory. In contrast, CIH transiently induced spatial memory dysfunction in pubertal male offspring but did not impact social or repetitive functions. Long-term effects of gestational CIH on social behaviors were only observed in female offspring, wherein CIH induced social disengagement and suppression of circulating corticosterone levels in young adulthood. No effects of gestational CIH were observed in anxiety-like behaviors, hippocampal neuronal activity, or circulating testosterone and estradiol levels, regardless of sex or age of offspring. CONCLUSIONS: Our results indicate that hypoxia-associated pregnancy complications during late gestation can increase the risk for behavioral and physiological outcomes in offspring, such as social dysfunction, repetitive behaviors, and cognitive impairment, that are dependent on sex and age.

Sleep apnea during late pregnancy is a common pregnancy complication that can impact the brain development of children born to mothers with sleep apnea. Children with impaired brain development may present with decreased social skills, memory issues, anxiety, and compulsivity. It is unclear if there is a cause and effect relationship between sleep apnea during late pregnancy and behavioral changes in offspring. Additionally, it is unknown whether male or female sex or age of the offspring affects these relationships. In this study, we exposed pregnant rats to a model of sleep apnea called chronic intermittent hypoxia (CIH) within late gestation and examined the behavior of the offspring and brain activity during puberty and young adulthood. We found that CIH during late pregnancy had long-term effects in the offspring that were different in males and females. Notably, female offspring displayed social impairments in response to late gestation CIH, whereas male offspring displayed cognitive dysfunction.

Sex and age differences in social and cognitive function in offspring exposed to late gestational hypoxia.

Background: Gestational sleep apnea affects 8-26% of pregnancies and can increase the risk for autism spectrum disorder (ASD) in offspring. ASD is a neurodevelopmental disorder associated with social dysfunction, repetitive behaviors, anxiety, and cognitive impairment. To examine the relationship between gestational sleep apnea and ASD-associated behaviors, we used a chronic intermittent hypoxia (CIH) protocol between gestational days (GD) 15-19 in pregnant rats to model late gestational sleep apnea. We hypothesized that late gestational CIH would produce sex- and age-specific social, mood, and cognitive impairments in offspring. Methods: Timed pregnant Long-Evans rats were exposed to CIH or room air normoxia from GD 15-19. Behavioral testing of offspring occurred during either puberty or young adulthood. To examine ASD-associated phenotypes, we quantified ASD-associated behaviors (social function, repetitive behaviors, anxiety-like behaviors, and spatial memory and learning), hippocampal activity (glutamatergic NMDA receptors, dopamine transporter, monoamine oxidase-A, EGR-1, and doublecortin), and circulating hormones in offspring. Results: Late gestational CIH induced sex- and age-specific differences in social, repetitive and memory functions in offspring. These effects were mostly transient and present during puberty. In female pubertal offspring, CIH impaired social function, increased repetitive behaviors, and increased circulating corticosterone levels, but did not impact memory. In contrast, CIH transiently induced spatial memory dysfunction in pubertal male offspring but did not impact social or repetitive functions. Long-term effects of gestational CIH were only observed in female offspring, wherein CIH induced social disengagement and suppression of circulating corticosterone levels in young adulthood. No effects of gestational CIH were observed on anxiety-like behaviors, hippocampal activity, circulating testosterone levels, or circulating estradiol levels, regardless of sex or age of offspring. Conclusions: Our results indicate that hypoxia-associated pregnancy complications during late gestation can increase the risk for ASD-associated behavioral and physiological outcomes, such as pubertal social dysfunction, corticosterone dysregulation, and memory impairments.

Exposure to unmethylated CpG oligonucleotides disrupts blood pressure circadian rhythms and placental clock gene network in pregnant rats.

Bacterial infections and impaired circulating mitochondrial DNA dynamics are associated with adverse pregnancy outcomes. Unmethylated cytosine-guanine dinucleotide (CpG) motifs are common in bacterial and mitochondrial DNA and act as potent immunostimulators. We tested the hypothesis that exposure to CpG oligonucleotides (ODN) during pregnancy would disrupt blood pressure circadian rhythms and placental molecular clock network, mediating aberrant fetoplacental growth dynamics. Rats were repeatedly treated with CpG ODN in the third trimester [gestational days (GD) 14, 16, and 18] and euthanized on GD20 (near term) or treated with a single dose of CpG ODN on GD14 and euthanized 4 h after treatment. Hemodynamic circadian rhythms were analyzed via Lomb-Scargle periodogram analysis on 24-h raw data collected continuously via radiotelemetry. A P value ≥ 0.05 indicates the absence of a circadian rhythm. Following the first treatment with CpG ODN, maternal systolic and diastolic blood pressure circadian rhythms were lost (P ≥ 0.05). Blood pressure circadian rhythm was restored by GD16 and remained unaffected after the second treatment with CpG ODN (P < 0.0001). Diastolic blood pressure circadian rhythm was again lost after the last treatment on GD18 (P ≥ 0.05). CpG ODN increased placental expression of Per2, Per3, and Tnfα (P ≤ 0.05) and affected fetoplacental growth dynamics. Reduced fetal and placental weights were disproportionately associated with increases in the number of resorptions in ODN-treated dams compared with controls. In conclusion, gestational exposure to unmethylated CpG ODN dysregulates the placental molecular clock network and fetoplacental growth dynamics and disrupts blood pressure circadian rhythms.NEW & NOTEWORTHY Gestational exposure to unmethylated CpG ODN dysregulates placental molecular clock network and fetoplacental growth dynamics and disrupts blood pressure circadian rhythms. These findings provide novel insights into the relationship between circadian rhythms and immune responses in pregnancy and propose new mechanisms by which maternal responses to immune triggers could dictate circadian rhythms of cardiovascular processes and placental clock machinery function to determine fetal growth trajectories.

Gestational exposure to unmethylated CpG oligonucleotides dysregulates placental molecular clock network and fetoplacental growth dynamics, and disrupts maternal blood pressure circadian rhythms in rats.

Bacterial infections and impaired mitochondrial DNA dynamics are associated with adverse pregnancy outcomes. Unmethylated cytosine-guanine dinucleotide (CpG) motifs are common in bacterial and mitochondrial DNA and act as potent immunostimulators. Here, we tested the hypothesis that exposure to CpG oligonucleotides (ODN) during pregnancy would disrupt blood pressure circadian rhythms and the placental molecular clock machinery, mediating aberrant fetoplacental growth dynamics. Rats were repeatedly treated with CpG ODN in the 3 rd trimester (gestational day, GD, 14, 16, 18) and euthanized on GD20 (near term) or with a single dose of CpG ODN and euthanized 4 hours after treatment on GD14. Hemodynamic circadian rhythms were analyzed via Lomb-Scargle periodogram analysis on 24-h raw data collected continuously via radiotelemetry. A p -value ≥ 0.05 indicates the absence of a circadian rhythm. Following the first treatment with CpG ODN, maternal systolic and diastolic blood pressure circadian rhythms were lost ( p ≥ 0.05). Blood pressure circadian rhythm was restored by GD16 and remained unaffected after the second treatment with CpG ODN ( p < 0.0001). Diastolic blood pressure circadian rhythm was again lost after the last treatment on GD18 ( p ≥ 0.05). CpG ODN increased placental expression of Per2 and Per3 and Tnfα ( p ≤ 0.05) and affected fetoplacental growth dynamics, such as reduced fetal and placental weights were disproportionately associated with increases in the number of resorptions in ODN-treated dams compared to controls. In conclusion, gestational exposure to unmethylated CpG DNA dysregulates placental molecular clock network and fetoplacental growth dynamics and disrupts blood pressure circadian rhythms.

Gestational hypoxia in late pregnancy differentially programs subcortical brain maturation in male and female rat offspring.

BACKGROUND: Hypoxia is associated with pregnancy complications, such as preeclampsia, placental abruption, and gestational sleep apnea. Hypoxic insults during gestation can impact the brain maturation of cortical and subcortical pathways, such as the nigrostriatal pathway. However, the long-term effects of in utero hypoxic stress exposure on brain maturation in offspring are unclear, especially exposure during late gestation. The purpose of this study was to determine the impact of gestational hypoxia in late pregnancy on developmental programming of subcortical brain maturation by focusing on the nigrostriatal pathway. METHODS: Timed pregnant Long-Evans rats were exposed to chronic intermittent hypoxia or room air normoxia from gestational day (GD) 15-19 (term 22-23 days). Male and female offspring were assessed during two critical periods: puberty from postnatal day (PND) 40-45 or young adulthood (PND 60-65). Brain maturation was quantified by examining (1) the structural development of the nigrostriatal pathway via analysis of locomotor behaviors and the substantia nigra dopaminergic neuronal cell bodies and (2) the refinement of the nigrostriatal pathway by quantifying ultrasonic vocalizations (USVs). RESULTS: The major findings of this study are gestational hypoxia has age- and sex-dependent effects on subcortical brain maturation in offspring by adversely impacting the refinement of the nigrostriatal pathway in the absence of any effects on the structural development of the pathway. During puberty, female offspring were impacted more than male offspring, as evidenced by decreased USV call frequency, chirp USV call duration, and simple call frequency. In contrast, male offspring were impacted more than female offspring during young adulthood, as evidenced by increased latency to first USV, decreased simple USV call intensity, and increased harmonic USV call bandwidth. No effects of gestational hypoxia on the structural development of the nigrostriatal pathway were observed. CONCLUSIONS: These novel findings demonstrate hypoxic insults during pregnancy mediate developmental programming of the cortical and subcortical pathways, in which male offspring exhibit long-term adverse effects compared to female offspring. Impairment of cortical and subcortical pathways maturation, such as the nigrostriatal pathway, may increase risk for neuropsychiatric disorders (e.g., mood disorders, cognitive dysfunction, brain connectivity dysfunction).

Exosomes facilitate intercellular communication between uterine perivascular adipose tissue and vascular smooth muscle cells in pregnant rats.

Perivascular adipose tissue (PVAT) is distinct from other adipose depots, as it has differential gene and protein profiles and vasoactive functions. We have shown that pregnancy affects the morphology of PVAT surrounding the uterine arteries (utPVAT) differentially than the morphology of nonperivascular reproductive adipose depots (i.e., periovarian adipose tissue, OVAT). Here, we hypothesized that pregnancy modifies the profile (size and molecular mass) of exosome-like extracellular vesicles released by utPVAT (Exo-utPVAT) compared with exosome-like extracellular vesicles released by OVAT (Exo-OVAT) and that primary uterine vascular smooth muscle cells (utVSMCs) can internalize Exo-utPVAT. Our findings indicate that utPVAT from pregnant and nonpregnant rats secrete exosome-like vesicles. Exo-utPVAT from pregnant rats were smaller (i.e., molecular size) and heavier (i.e., molecular mass) than those from nonpregnant rats, whereas pregnancy did not affect the size of Exo-OVAT. Immunocytochemistry and confocal microscopy showed that primary utVSMCs internalized Exo-utPVAT (both tissues from the same pregnant rat) labeled by the lipophilic tracer DiO. Treatment of isolated uterine arteries with Exo-utPVAT did not affect relaxation responses to acetylcholine in pregnant or nonpregnant rats. Collectively, these findings demonstrate a novel type of intercellular communication between Exo-utPVAT and utVSMCs and indicate pregnancy modulates the morphology and cargo of Exo-utPVAT.NEW & NOTEWORTHY Uterine perivascular adipose tissue secretes exosome-like vesicles, which are internalized by their adjacent uterine vascular smooth muscle cells. Consideration of the exosomal communication between adipose tissue and vascular smooth muscle cells in the uterine circulation in mathematical models and experimental designs may help us to improve understanding of mechanisms underlying uterine artery adaptive responses to a healthy pregnancy and during pregnancy complications.