Obese mice have decreased uterine contractility and altered energy metabolism in the uterus at term gestation†.

Over 35% of reproductive-age women in the USA have obesity, putting them at increased risk for numerous obstetric complications due to abnormal labor. While the association between maternal obesity and abnormal labor has been well documented, the mechanisms responsible for this remain understudied. The uterine smooth muscle, myometrium, has high energy needs in order to fuel regular uterine contractions during parturition. However, the precise mechanisms by which the myometrium meets its energy demands has not been defined. Here, our objective was to define the effects of obesity on energy utilization in the myometrium during labor. We generated a mouse model of maternal diet-induced obesity and found that these mice had a higher rate of dystocia than control chow-fed mice. Moreover, compared to control chow-fed mice, DIO mice at term, both before and during labor had lower in vivo spontaneous uterine contractility. Untargeted transcriptomic and metabolomic analyses suggest that diet-induced obesity is associated with elevated long-chain fatty acid uptake and utilization in the uterus, but also an accumulation of medium-chain fatty acids. Diet-induced obesity uteri also had an increase in the abundance of long chain-specific beta-oxidation enzymes, which may be responsible for the observed increase in long-chain fatty acid utilization. This altered energy substrate utilization may be a contributor to the observed contractile dysfunction.

BKCa channels are involved in spontaneous and lipopolysaccharide-stimulated uterine contraction in late gestation mice†.

The large-conductance, voltage-gated, calcium (Ca2+)-activated potassium channel (BKCa) is one of the most abundant potassium channels in the myometrium. Previous work conducted by our group has identified a link between inflammation, BKCa channels and excitability of myometrial smooth muscle cells. Here, we investigate the role of BKCa channels in spontaneous and lipopolysaccharide (LPS)-stimulated uterine contraction to gain a better understanding of the relationship between the BKCa channel and uterine contraction in basal and inflammatory states. Uteri of C57BL/6 J mice on gestational day 18.5 (GD18.5) were obtained and either fixed in formalin or used immediately for tension recording or isolation of primary myocytes for patch-clamp. Paraffin sections were used for immunofluorescenctdetection of BKCa and Toll-like receptor (TLR4). For tension recordings, LPS was administered to determine its effect on uterine contractions. Paxilline, a BKCa inhibitor, was used to dissect the role of BKCa in uterine contraction in basal and inflammatory states. Finally, patch-clamp recordings were performed to investigate the relationship between LPS, the BKCa channel and membrane currents in mouse myometrial smooth muscle cells (mMSMCs). We confirmed the expression of BKCa and TLR4 in the myometrium of GD18.5 mice and found that inhibiting BKCa channels with paxilline suppressed both spontaneous and LPS-stimulated uterine contractions. Furthermore, application of BKCa inhibitors (paxilline or iberiotoxin) after LPS inhibited BKCa channel activity in mMSMCs. Moreover, pretreatment with BKCa inhibitor or the TLR4 inhibitor suppressed LPS-activated BKCa currents. Our study demonstrates that BKCa channels are involved in both basal and LPS-stimulated uterine contraction in pregnant mice.

Sleep and circadian rhythms during pregnancy, social disadvantage, and alterations in brain development in neonates.

Pregnant women in poverty may be especially likely to experience sleep and circadian rhythm disturbances, which may have downstream effects on fetal neurodevelopment. However, the associations between sleep and circadian rhythm disturbances, social disadvantage during pregnancy, and neonatal brain structure remains poorly understood. The current study explored the association between maternal sleep and circadian rhythm disturbances during pregnancy and neonatal brain outcomes, examining sleep and circadian rhythm disturbances as a mediator of the effect of social disadvantage during pregnancy on infant structural brain outcomes. The study included 148 mother-infant dyads, recruited during early pregnancy, who had both actigraphy and neuroimaging data. Mothers' sleep was assessed throughout their pregnancy using actigraphy, and neonates underwent brain magnetic resonance imaging in the first weeks of life. Neonatal structural brain outcomes included cortical gray matter, subcortical gray matter, and white matter volumes along with a measure of the total surface area of the cortex. Neonates of mothers who experienced greater inter-daily deviations in sleep duration had smaller total cortical gray and white matter volumes and reduced cortical surface areas. Neonates of mothers who had higher levels of circadian misalignment and later sleep timing during pregnancy showed smaller subcortical gray matter volumes. Inter-daily deviations in sleep duration during pregnancy mediated the association between maternal social disadvantage and neonatal structural brain outcomes. Findings highlight the importance of regularity and rhythmicity in sleep schedules during pregnancy and bring to light the role of chronodisruption as a potential mechanism underlying the deleterious neurodevelopmental effects of prenatal adversity. RESEARCH HIGHLIGHTS: Social disadvantage was associated with sleep and circadian rhythm disturbances during pregnancy, including later sleep schedules, increased variability in sleep duration, circadian misalignment, and a higher proportion of the sleep period spent awake. Maternal sleep and circadian rhythm disturbances during pregnancy were associated with decreased brain volume and reduced cortical surface area in neonates. Maternal inter-daily deviations in sleep duration during pregnancy mediated the association between social disadvantage and neonatal brain volume and cortical surface area.

The Association between Mode of Transportation Support and Research Study Visit Attendance among Pregnant Patients.

OBJECTIVE: This study aimed to examine the association between transportation assistance and study visits, and explore differences by transportation modality. STUDY DESIGN: This was a secondary analysis of prospective cohort study. We identified patients requesting transportation support for research ultrasound visits and identified controls (1:2 ratio) who did not request support matched for age, race, and insurance type. Conditional logistic regression examined the association between transportation support and mode of transportation with study visit attendance. RESULTS: Transportation support was requested by 57/1,184 (4.8%) participants. Participants that requested transportation support were three times more likely to attend visits than their matched controls (adjusted odds ratio [aOR] = 3.16, 95% confidence interval [CI]: 1.76-5.68). Among visits with transportation support, those supported by a ridesharing service had five-fold higher odds of attendance than visits supported with taxi service (aOR = 5.06, 95% CI: 1.50-16.98). CONCLUSION: Transportation support, especially a ridesharing service, is associated with improved attendance at research study visits in a sample of predominantly low-income, Black, pregnant participants. Implementing transportation support may be a promising strategy to improve engagement in research studies. KEY POINTS: · Participants utilizing transportation assistance were more likely to attend study appointments.. · Participants using ridesharing had higher likelihood of attendance than those using taxi service.. · Transportation assistance may improve research engagement for historically marginalized people..

The association between maternal sleep and circadian rhythms during pregnancy and infant sleep and socioemotional outcomes.

Studies have established that maternal sleep and circadian rhythm disturbances during pregnancy are associated with poor prenatal and perinatal outcomes for mothers and offspring. However, little work has explored its effects on infant sleep or socioemotional outcomes. The current study examined the relationship between maternal sleep and circadian rhythm disturbances during pregnancy and infant sleep and socioemotional outcomes in a diverse sample of N = 193 mothers and their infants (51% White; 52% Female; Mage = 11.95 months). Maternal sleep and circadian rhythms during pregnancy were assessed using self-reports and actigraphy. Mothers reported on infants' sleep and socioemotional outcomes when infants were one year old. When controlling for infant sex, age, gestational age at birth, family income-to-needs ratios, and maternal depression, mothers who reported more sleep problems during pregnancy had infants with more sleep disturbances when they were one year old. Moreover, mothers who had later sleep timing (i.e., went to bed and woke up later, measured via actigraphy) during pregnancy had infants with more dysregulation (e.g., increased feeding difficulties, sensory sensitivities) and externalizing problems, and mothers with increased intra-daily variability in rest-activity rhythms (as measured via actigraphy) had infants with more externalizing problems. Findings suggest that maternal sleep and circadian rhythm disturbances during pregnancy may be a risk factor for infant sleep problems and socioemotional difficulties.

Pharmacological chaperones for the oxytocin receptor increase oxytocin responsiveness in myometrial cells.

Oxytocin is a potent uterotonic agent administered to nearly all patients during childbirth in the United States. Inadequate oxytocin response can necessitate Cesarean delivery or lead to uterine atony and postpartum hemorrhage. Thus, it may be clinically useful to identify patients at risk for poor oxytocin response and develop strategies to sensitize the uterus to oxytocin. Previously, we showed that the V281M variant in the oxytocin receptor (OXTR) gene impairs OXTR trafficking to the cell surface, leading to a decreased oxytocin response in cells. Here, we sought to identify pharmacological chaperones that increased oxytocin response in cells expressing WT or V281M OXTR. We screened nine small-molecule agonists and antagonists of the oxytocin/vasopressin receptor family and identified two, SR49059 and L371,257, that restored both OXTR trafficking and oxytocin response in HEK293T cells transfected with V281M OXTR. In hTERT-immortalized human myometrial cells, which endogenously express WT OXTR, treatment with SR49059 and L371,257 increased the amount of OXTR on the cell surface by two- to fourfold. Furthermore, SR49059 and L371,257 increased the endogenous oxytocin response in hTERT-immortalized human myometrial cells by 35% and induced robust oxytocin responses in primary myometrial cells obtained from patients at the time of Cesarean section. If future studies demonstrate that these pharmacological chaperones or related compounds function similarly in vivo, we propose that they could potentially be used to enhance clinical response to oxytocin.

Blocking the BKCa channel induces NF-κB nuclear translocation by increasing nuclear calcium concentration†.

Nuclear factor kappa B (NF-κB) transcriptionally regulates several genes involved in initiating uterine contractions. A key factor controlling NF-κB activity is its translocation to the nucleus. In myometrial smooth muscle cells (MSMCs), this translocation can be stimulated by the inflammatory molecule lipopolysaccharide (LPS) or by blocking the potassium calcium-activated channel subfamily M alpha 1 (KCNMA1 or BKCa) with paxilline (PAX). Here, we sought to determine the mechanism by which blocking BKCa causes NF-κB-p65 translocation to the nucleus in MSMCs. We show that LPS- and PAX-induced NF-κB-p65 translocation are similar in that neither depends on several mitogen-activated protein kinase pathways, but both require increased intracellular calcium (Ca2+). However, the nuclear transport inhibitor wheat germ agglutinin prevented NF-κB-p65 nuclear translocation in response to LPS but not in response to PAX. Blocking BKCa located on the plasma membrane resulted in a transient NF-κB-p65 nuclear translocation that was not sufficient to induce expression of its transcriptional target, suggesting a role for intracellular BKCa. We report that BKCa also localizes to the nucleus and that blocking nuclear BKCa results in an increase in nuclear Ca2+ in MSMCs. Together, these data suggest that BKCa localized on the nuclear membrane plays a key role in regulating nuclear Ca2+ and NF-κB-p65 nuclear translocation in MSMCs.

Association between Maternal Serum Lipids and Intrapartum Oxytocin Requirements during Labor Induction and Augmentation.

OBJECTIVE: This study aimed to investigate the relationship between maternal serum lipid parameters and oxytocin requirements among women with term vaginal deliveries. STUDY DESIGN: In this secondary analysis of a prospective cohort study, women who presented for delivery at ≥37 weeks' gestation and received oxytocin during their labor were included. Maternal serum was collected intrapartum. The cohort was stratified into two groups based on maximum oxytocin infusion dose during labor. Primary outcomes were maternal total cholesterol, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and triglyceride levels. Generalized linear regression models were used to assess the association between lipid parameters and maximum oxytocin dose requirements while controlling for potential confounders. For secondary analyses, the cohort was stratified by HDL-C into two groups. Multivariable logistic regression was used to evaluate the relationship between low maternal HDL-C and additional intrapartum oxytocin parameters. RESULTS: There were no differences in maternal total cholesterol, LDL-C, or triglyceride values between high and low maximum oxytocin groups. Median serum HDL-C was significantly lower among women in the high oxytocin group compared with those in the low oxytocin group (56 vs. 62 mg/dL, p < 0.01). For every 0.26 mg/dL lower HDL-C, women had 1 mU/min higher maximum oxytocin infusion dose during labor. Women with low serum HDL-C were also more likely to require maximum oxytocin doses above the 75th percentile (adjusted odds ratio [aOR]: 1.99, 95% confidence interval [CI]: 1.06-3.75) and above the 90th percentile (aOR: 2.47, 95% CI: 1.10-5.54). Among women undergoing induction of labor, low serum HDL-C was also associated with longer duration of oxytocin infusion (aOR: 2.07, 95% CI: 1.02-4.20). CONCLUSION: Low maternal HDL-C levels at term are associated with higher maximum oxytocin infusion doses among women undergoing labor induction or augmentation. Given the growing prevalence of metabolic syndrome in the United States and persistently high rates of cesarean delivery, HDL-C or its components may present a new target for predicting and improving labor outcomes. KEY POINTS: · Serum HDL-C at term is inversely correlated with oxytocin infusion doses at term.. · Low maternal serum HDL-C is associated with higher oxytocin requirements during labor induction or augmentation.. · No association between maternal serum total cholesterol, LDL-C, or triglyceride levels and oxytocin requirements in labor..

Guidelines for the measurement of vascular function and structure in isolated arteries and veins.

The measurement of vascular function in isolated vessels has revealed important insights into the structural, functional, and biomechanical features of the normal and diseased cardiovascular system and has provided a molecular understanding of the cells that constitutes arteries and veins and their interaction. Further, this approach has allowed the discovery of vital pharmacological treatments for cardiovascular diseases. However, the expansion of the vascular physiology field has also brought new concerns over scientific rigor and reproducibility. Therefore, it is appropriate to set guidelines for the best practices of evaluating vascular function in isolated vessels. These guidelines are a comprehensive document detailing the best practices and pitfalls for the assessment of function in large and small arteries and veins. Herein, we bring together experts in the field of vascular physiology with the purpose of developing guidelines for evaluating ex vivo vascular function. By using this document, vascular physiologists will have consistency among methodological approaches, producing more reliable and reproducible results.

The impact of physical activity during pregnancy on labor and delivery.

BACKGROUND: Physical activity in pregnancy is associated with decreased risks of adverse pregnancy outcomes such as gestational diabetes and preeclampsia. However, the relationship between the amount and type of physical activity during pregnancy and subsequent labor outcomes remains unclear. OBJECTIVE: This study aimed to test the hypothesis that higher levels of physical activity across different lifestyle domains in pregnancy are associated with a shorter duration of labor. STUDY DESIGN: This study is a secondary analysis of a prospective cohort study in which patients with singleton pregnancies without a major fetal anomaly were administered the Kaiser Physical Activity Survey in each trimester. The Kaiser Physical Activity Survey was designed specifically to quantify various types of physical activities in women and includes 4 summative indices-housework/caregiving, active living habits, sports, and occupation. The study included women at full-term gestations admitted for induction of labor or spontaneous labor. The primary outcome of this analysis was duration of the second stage of labor. Secondary outcomes were duration of the active stage, prolonged first and second stage, mode of delivery, rates of second-stage cesarean delivery, operative vaginal delivery, severe perineal lacerations, and postpartum hemorrhage. These outcomes were compared between patients with and without high physical activity levels, defined as overall Kaiser Physical Activity Survey score ≥75th percentile in the third trimester. Multivariable logistic regression was used to adjust for obesity and epidural use. In addition, a subgroup analysis of nulliparous patients was performed. RESULTS: A total of 811 patients with complete Kaiser Physical Activity Survey data in the third trimester were included in this analysis. The median Kaiser Physical Activity Survey score was 9.5 (8.2-10.8). Of the 811 patients, 203 (25%) had higher levels of physical activity in pregnancy. There was no difference in the duration of the second stage of labor between patients with and without higher physical activity levels (1.29±2.94 vs 0.97±2.08 hours; P=.15). The duration of active labor was significantly shorter in patients with higher levels of physical activity (5.77±4.97 vs 7.43±6.29 hours; P=.01). Patients with higher physical activity levels were significantly less likely to have a prolonged first stage (9.8% vs 19.4%; P<.01; adjusted relative risk, 0.55; 95% confidence interval, 0.34-0.83). However, rates of prolonged second-stage cesarean delivery, operative vaginal deliveries, and perineal lacerations were similar between the 2 groups. CONCLUSION: Patients who are more physically active during pregnancy have a shorter duration of active labor.

Progesterone and estrogen regulate NALCN expression in human myometrial smooth muscle cells.

During pregnancy, the uterus transitions from a quiescent state to an excitable, highly contractile state to deliver the fetus. Two important contributors essential for this transition are hormones and ion channels, both of which modulate myometrial smooth muscle cell (MSMC) excitability. Recently, the sodium (Na+) leak channel, nonselective (NALCN), was shown to contribute to a Na+ leak current in human MSMCs, and mice lacking NALCN in the uterus had dysfunctional labor. Microarray data suggested that the proquiescent hormone progesterone (P4) and the procontractile hormone estrogen (E2) regulated this channel. Here, we sought to determine whether P4 and E2 directly regulate NALCN. In human MSMCs, we found that NALCN mRNA expression decreased by 2.3-fold in the presence of E2 and increased by 5.6-fold in the presence of P4. Similarly, E2 treatment decreased, and P4 treatment restored NALCN protein expression. Additionally, E2 significantly inhibited, and P4 significantly enhanced an NALCN-dependent leak current in MSMCs. Finally, we identified estrogen response and progesterone response elements (EREs and PREs) in the NALCN promoter. With the use of luciferase assays, we showed that the PREs, but not the ERE, contributed to regulation of NALCN expression. Our findings reveal a new mechanism by which NALCN is regulated in the myometrium and suggest a novel role for NALCN in pregnancy.

Microelectrode array analysis of mouse uterine smooth muscle electrical activity†.

Uterine contractions are important for various functions of the female reproductive cycle. Contractions are generated, in part, by electrical coupling of smooth muscle cells of the myometrium, the main muscle layer of the uterus. Aberrant myometrial electrical activity can lead to uterine dysfunction. To better understand and treat conditions associated with aberrant activity, it is crucial to understand the mechanisms that underlie normal activity. Here, we used microelectrode array (MEA) to simultaneously record and characterize myometrial electrical activities at high spatial and temporal resolution. Mouse myometrial longitudinal muscle tissue was isolated at different stages throughout the estrous cycle and placed on an 8×8 MEA. Electrical activity was recorded for 10 min at a sampling rate of 12.5 kHz. We used a spike-tracking algorithm to independently analyze each channel and developed a pipeline to quantify the amplitude, duration, frequency, and synchronicity of the electrical activities. Electrical activities in estrous were more synchronous, and had shorter duration, higher frequency, and lower amplitude than electrical activities in non-estrous. We conclude that MEA can be used to detect differential patterns of myometrial electrical activity in distinct estrous cycle stages. In the future, this methodology can be used to assess different physiological and pathological states and evaluate therapeutic agents that regulate uterine function.

Sodium-activated potassium channels moderate excitability in vascular smooth muscle.

KEY POINTS: We report that a sodium-activated potassium current, IKNa , has been inadvertently overlooked in both conduit and resistance arterial smooth muscle cells. IKNa is a major K+ resting conductance and is absent in cells of IKNa knockout (KO) mice. The phenotype of the IKNa KO is mild hypertension, although KO mice react more strongly than wild-type with raised blood pressure when challenged with vasoconstrictive agents. IKNa is negatively regulated by angiotensin II acting through Gαq protein-coupled receptors. In current clamp, KO arterial smooth muscle cells have easily evoked Ca2+ -dependent action potentials. ABSTRACT: Although several potassium currents have been reported to play a role in arterial smooth muscle (ASM), we find that one of the largest contributors to membrane conductance in both conduit and resistance ASMs has been inadvertently overlooked. In the present study, we show that IKNa , a sodium-activated potassium current, contributes a major portion of macroscopic outward current in a critical physiological voltage range that determines intrinsic cell excitability; IKNa is the largest contributor to ASM cell resting conductance. A genetic knockout (KO) mouse strain lacking KNa channels (KCNT1 and KCNT2) shows only a modest hypertensive phenotype. However, acute administration of vasoconstrictive agents such as angiotensin II (Ang II) and phenylephrine results in an abnormally large increase in blood pressure in the KO animals. In wild-type animals Ang II acting through Gαq protein-coupled receptors down-regulates IKNa , which increases the excitability of the ASMs. The complete genetic removal of IKNa in KO mice makes the mutant animal more vulnerable to vasoconstrictive agents, thus producing a paroxysmal-hypertensive phenotype. This may result from the lowering of cell resting K+ conductance allowing the cells to depolarize more readily to a variety of excitable stimuli. Thus, the sodium-activated potassium current may serve to moderate blood pressure in instances of heightened stress. IKNa may represent a new therapeutic target for hypertension and stroke.

Oxytocin can regulate myometrial smooth muscle excitability by inhibiting the Na+ -activated K+ channel, Slo2.1.

KEY POINTS: At the end of pregnancy, the uterus transitions from a quiescent state to a highly contractile state. This transition requires that the uterine (myometrial) smooth muscle cells increase their excitability, although how this occurs is not fully understood. We identified SLO2.1, a potassium channel previously unknown in uterine smooth muscle, as a potential significant contributor to the electrical excitability of myometrial smooth muscle cells. We found that activity of the SLO2.1 channel is negatively regulated by oxytocin via Gαq-protein-coupled receptor activation of protein kinase C. This results in depolarization of the uterine smooth muscle cells and calcium entry, which may contribute to uterine contraction. These findings provide novel insights into a previously unknown mechanism by which oxytocin may act to modulate myometrial smooth muscle cell excitability. Our findings also reveal a new potential pharmacological target for modulating uterine excitability. ABSTRACT: During pregnancy, the uterus transitions from a quiescent state to a more excitable contractile state. This is considered to be at least partly a result of changes in the myometrial smooth muscle cell (MSMC) resting membrane potential. However, the ion channels controlling the myometrial resting membrane potential and the mechanism of transition to a more excitable state have not been fully clarified. In the present study, we show that the sodium-activated, high-conductance, potassium leak channel, SLO2.1, is expressed and active at the resting membrane potential in MSMCs. Additionally, we report that SLO2.1 is inhibited by oxytocin binding to the oxytocin receptor. Inhibition of SLO2.1 leads to membrane depolarization and activation of voltage-dependent calcium channels, resulting in calcium influx. The results of the present study reveal that oxytocin may modulate MSMC electrical activity by inhibiting SLO2.1 potassium channels.

BKCa channel regulates calcium oscillations induced by alpha-2-macroglobulin in human myometrial smooth muscle cells.

The large-conductance, voltage-gated, calcium (Ca(2+))-activated potassium channel (BKCa) plays an important role in regulating Ca(2+)signaling and is implicated in the maintenance of uterine quiescence during pregnancy. We used immunopurification and mass spectrometry to identify proteins that interact with BKCain myometrium samples from term pregnant (≥37 wk gestation) women. From this screen, we identified alpha-2-macroglobulin (α2M). We then used immunoprecipitation followed by immunoblot and the proximity ligation assay to confirm the interaction between BKCaand both α2M and its receptor, low-density lipoprotein receptor-related protein 1 (LRP1), in cultured primary human myometrial smooth muscle cells (hMSMCs). Single-channel electrophysiological recordings in the cell-attached configuration demonstrated that activated α2M (α2M*) increased the open probability of BKCain an oscillatory pattern in hMSMCs. Furthermore, α2M* caused intracellular levels of Ca(2+)to oscillate in oxytocin-primed hMSMCs. The initiation of oscillations required an interaction between α2M* and LRP1. By using Ca(2+)-free medium and inhibitors of various Ca(2+)signaling pathways, we demonstrated that the oscillations required entry of extracellular Ca(2+)through store-operated Ca(2+)channels. Finally, we found that the specific BKCablocker paxilline inhibited the oscillations, whereas the channel opener NS11021 increased the rate of these oscillations. These data demonstrate that α2M* and LRP1 modulate the BKCachannel in human myometrium and that BKCaand its immunomodulatory interacting partners regulate Ca(2+)dynamics in hMSMCs during pregnancy.

The large-conductance voltage- and Ca2+ -activated K+ channel and its γ1-subunit modulate mouse uterine artery function during pregnancy.

KEY POINTS: The uterine artery (UA) markedly vasodilates during pregnancy to direct blood flow to the developing fetus. Inadequate UA vasodilatation leads to intrauterine growth restriction and fetal death. The large-conductance voltage- and Ca2+ -activated K+ (BKCa ) channel promotes UA vasodilatation during pregnancy. We report that BKCa channel activation increases the UA diameter at late pregnancy stages in mice. Additionally, a BKCa channel auxiliary subunit, γ1, participates in this process by increasing channel activation and inducing UA vasodilatation at late pregnancy stages. Our results highlight the importance of the BKCa channel and its γ1-subunit for UA functional changes during pregnancy. ABSTRACT: Insufficient vasodilatation of the uterine artery (UA) during pregnancy leads to poor utero-placental perfusion, contributing to intrauterine growth restriction and fetal loss. Activity of the large-conductance Ca2+ -activated K+ (BKCa ) channel increases in the UA during pregnancy, and its inhibition reduces uterine blood flow, highlighting a role of this channel in UA adaptation to pregnancy. The auxiliary γ1-subunit increases BKCa activation in vascular smooth muscle, but its role in pregnancy-associated UA remodelling is unknown. We explored whether the BKCa and its Î³1-subunit contribute to UA remodelling during pregnancy. Doppler imaging revealed that, compared to UAs from wild-type (WT) mice, UAs from BKCa knockout (BKCa-/- ) mice had lower resistance at pregnancy day 14 (P14) but not at P18. Lumen diameters were twofold larger in pressurized UAs from P18 WT mice than in those from non-pregnant mice, but this difference was not seen in UAs from BKCa-/- mice. UAs from pregnant WT mice constricted 20-50% in response to the BKCa blocker iberiotoxin (IbTX), whereas UAs from non-pregnant WT mice only constricted 15%. Patch-clamp analysis of WT UA smooth muscle cells confirmed that BKCa activity increased over pregnancy, showing three distinct voltage sensitivities. The γ1-subunit transcript increased 7- to 10-fold during pregnancy. Furthermore, γ1-subunit knockdown reduced IbTX sensitivity in UAs from pregnant mice, whereas γ1-subunit overexpression increased IbTX sensitivity in UAs from non-pregnant mice. Finally, at P18, γ1-knockout (γ1-/- ) mice had smaller UA diameters than WT mice, and IbTX-mediated vasoconstriction was prevented in UAs from γ1-/- mice. Our results suggest that the γ1-subunit increases BKCa activation in UAs during pregnancy.

Na+-Leak Channel, Non-Selective (NALCN) Regulates Myometrial Excitability and Facilitates Successful Parturition.

BACKGROUND/AIMS: Uterine contractility is controlled by electrical signals generated by myometrial smooth muscle cells. Because aberrant electrical signaling may cause inefficient uterine contractions and poor reproductive outcomes, there is great interest in defining the ion channels that regulate uterine excitability. In human myometrium, the Na+ leak channel, non-selective (NALCN) contributes to a gadolinium-sensitive, Na+-dependent leak current. The aim of this study was to determine the role of NALCN in regulating uterine excitability and examine its involvement in parturition. METHODS: Wildtype C57BL/6J mice underwent timed-mating and NALCN uterine expression was measured at several time points across pregnancy including pregnancy days 7, 10, 14, 18 and 19. Sharp electrode current clamp was used to measure uterine excitability at these same time points. To determine NALCN's contribution to myometrial excitability and pregnancy outcomes, we created smooth-muscle-specific NALCN knockout mice by crossing NALCNfx/fx mice with myosin heavy chain Cre (MHCCreeGFP) mice. Parturition outcomes were assessed by observation via surveillance video recording cre control, flox control, smNALCN+/-, and smNALCN-/- mice. Myometrial excitability was compared between pregnancy day 19 flox controls and smNALCN-/- mice. RESULTS: We found that in the mouse uterus, NALCN protein levels were high early in pregnancy, decreased in mid and late pregnancy, and then increased in labor and postpartum. Sharp electrode current clamp recordings of mouse longitudinal myometrial samples from pregnancy days 7, 10, 14, 18, and 19 revealed day-dependent increases in burst duration and interval and decreases in spike density. NALCN smooth muscle knockout mice had reduced myometrial excitability exemplified by shortened action potential bursts, and an increased rate of abnormal labor, including prolonged and dysfunctional labor. CONCLUSIONS: Together, our findings demonstrate that the Na+ conducting channel NALCN contributes to the myometrial action potential waveform and is important for successful labor outcomes.

Mouse models of preterm birth: suggested assessment and reporting guidelines.

Preterm birth affects approximately 1 out of every 10 births in the United States, leading to high rates of mortality and long-term negative health consequences. To investigate the mechanisms leading to preterm birth so as to develop prevention strategies, researchers have developed numerous mouse models of preterm birth. However, the lack of standard definitions for preterm birth in mice limits our field's ability to compare models and make inferences about preterm birth in humans. In this review, we discuss numerous mouse preterm birth models, propose guidelines for experiments and reporting, and suggest markers that can be used to assess whether pups are premature or mature. We argue that adoption of these recommendations will enhance the utility of mice as models for preterm birth.