Placental gas exchange during amniotic carbon dioxide insufflation in sheep.

OBJECTIVE: Insufflation of the amniotic cavity with carbon dioxide (CO2 ) is used clinically to improve visibility during complex fetoscopic surgery. Insufflation with heated, humidified CO2 has recently been shown to reduce fetal hypercapnia and acidosis in sheep, compared with use of cold and dry CO2 , but the underlying mechanisms are unclear. The aim of this study was to investigate whether differences in placental CO2 and oxygen (O2 ) exchange during insufflation with heated and humidified vs cold and dry CO2 could explain these findings. METHODS: Thirteen fetal lambs at 105 days of gestation (term, 146 days) were exteriorized partially, via a midline laparotomy and hysterotomy, and instrumented with an umbilical artery catheter, an umbilical vein catheter and a common umbilical vein flow probe. Arterial and venous catheters and flow probes were also inserted into the maternal uterine circulation. Six ewes were insufflated with cold, dry CO2 (22°C; 0-5% humidity) and seven with heated, humidified CO2 (40°C; 95-100% humidity) at 15 mmHg for 180 min. Blood-flow recordings and paired arterial and venous blood gases were sampled from uterine and umbilical vessels. Rates of placental CO2 and O2 exchange were calculated. RESULTS: After 180 min of insufflation, fetal survival was 33% (2/6) using cold, dry CO2 and 71% (5/7) using heated, humidified CO2 . By 120 min, fetuses insufflated with heated, humidified CO2 had lower arterial CO2 levels and higher arterial pH compared to those insufflated with cold, dry gas. Insufflation decreased significantly placental gas exchange in both groups, as measured by rates of both (i) fetal CO2 clearance and O2 uptake and (ii) maternal O2 delivery and CO2 uptake from the fetal compartment. CONCLUSIONS: Lower arterial CO2 and higher pH levels in fetuses insufflated with heated and humidified, compared to cold and dry, CO2 could not be explained by differences in placental gas exchange. Instead, heated and humidified insufflation appeared to reduce fetal CO2 absorption from the uterus, supporting its use in preference to cold, dry CO2 . © 2019 International Society of Ultrasound in Obstetrics and Gynecology.

Antenatal sildenafil treatment improves neonatal pulmonary hemodynamics and gas exchange in lambs with diaphragmatic hernia.

OBJECTIVES: Infants with congenital diaphragmatic hernia (CDH) are predisposed to pulmonary hypertension after birth, owing to lung hypoplasia that impairs fetal pulmonary vascular development. Antenatal sildenafil treatment attenuates abnormal pulmonary vascular and alveolar development in rabbit and rodent CDH models, but whether this translates to functional improvements after birth remains unknown. We aimed to evaluate the effect of antenatal sildenafil on neonatal pulmonary hemodynamics and lung function in lambs with diaphragmatic hernia (DH). METHODS: DH was surgically induced at approximately 80 days' gestation in 16 lamb fetuses (term in lambs is approximately 147 days). From 105 days' gestation, ewes received either sildenafil (0.21 mg/kg/h intravenously) or saline infusion until delivery (n = 8 fetuses in each group). At approximately 138 days' gestation, all lambs were instrumented and then delivered via Cesarean section. The lambs were ventilated for 120 min with continuous recording of physiological (pulmonary and carotid artery blood flow and pressure; cerebral oxygenation) and ventilatory parameters, and regular assessment of arterial blood gas tensions. Only lambs that survived until delivery and with a confirmed diaphragmatic defect at postmortem examination were included in the analysis; these comprised six DH-sildenafil lambs and six DH-saline control lambs. RESULTS: Lung-to-body-weight ratio (0.016 ± 0.001 vs 0.013 ± 0.001; P = 0.06) and dynamic lung compliance (0.8 ± 0.2 vs 0.7 ± 0.2 mL/cmH2 O; P = 0.72) were similar in DH-sildenafil lambs and controls. Pulmonary vascular resistance decreased following lung aeration to a greater degree in DH-sildenafil lambs, and was 4-fold lower by 120 min after cord clamping than in controls (0.6 ± 0.1 vs 2.2 ± 0.6 mmHg/(mL/min); P = 0.002). Pulmonary arterial pressure was also lower (46 ± 2 vs 59 ± 2 mmHg; P = 0.048) and pulmonary blood flow higher (25 ± 3 vs 8 ± 2 mL/min/kg; P = 0.02) in DH-sildenafil than in DH-saline lambs at 120 min. Throughout the 120-min ventilation period, the partial pressure of arterial carbon dioxide tended to be lower in DH-sildenafil lambs than in controls (63 ± 8 vs 87 ± 8 mmHg; P = 0.057), and there was no significant difference in partial pressure of arterial oxygen between the two groups. CONCLUSIONS: Sustained maternal antenatal sildenafil infusion reduced pulmonary arterial pressure and increased pulmonary blood flow in DH lambs for the first 120 min after birth. These findings of improved pulmonary vascular function are consistent with improved pulmonary vascular structure seen in two previous animal models. The data support the rationale for a clinical trial investigating the effect of antenatal sildenafil in reducing the risk of neonatal pulmonary hypertension in infants with CDH. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Physiological effects of partial amniotic carbon dioxide insufflation with cold, dry vs heated, humidified gas in a sheep model.

OBJECTIVE: Partial amniotic carbon dioxide (CO2 ) insufflation (PACI) is used to improve visualization and facilitate complex fetoscopic surgery. However, there are concerns about fetal hypercapnic acidosis and postoperative fetal membrane inflammation. We assessed whether using heated and humidified, rather than cold and dry, CO2 might reduce the impact of PACI on the fetus and fetal membranes in sheep. METHODS: Twelve fetal lambs of 105 days' gestational age (term = 145 days) were exteriorized partially, via a midline laparotomy and hysterotomy, and arterial catheters and flow probes were inserted surgically. The 10 surviving fetuses were returned to the uterus, which was then closed and insufflated with cold, dry (22 °C at 0-5% humidity, n = 5) or heated, humidified (40 °C at 100% humidity, n = 5) CO2 at 15 mmHg for 180 min. Fetal membranes were collected immediately after insufflation for histological analysis. Physiological data and membrane leukocyte counts, suggestive of membrane inflammation, were compared between the two groups. RESULTS: After 180 min of insufflation, fetal survival was 0% in the group which underwent PACI with cold, dry CO2 , and 60% (n = 3) in the group which received heated, humidified gas. While all insufflated fetuses became progressively hypercapnic (PaCO2  > 68 mmHg), this was considerably less pronounced in those in which heated, humidified gas was used: after 120 min of insufflation, compared with those receiving cold, dry gas (n = 3), fetuses undergoing heated, humidified PACI (n = 5) had lower arterial partial pressure of CO2 (mean ± standard error of the mean, 82.7 ± 9.1 mmHg for heated, humidified CO2 vs 170.5 ± 28.5 for cold, dry CO2 during PACI, P < 0.01), lower lactate levels (1.4 ± 0.4 vs 8.5 ± 0.9 mmol/L, P < 0.01) and higher pH (pH, 7.10 ± 0.04 vs 6.75 ± 0.04, P < 0.01). There was also a non-significant trend for fetal carotid artery pressure to be higher following PACI with heated, humidified compared with cold, dry CO2 (30.5 ± 1.3 vs 8.7 ± 5.5 mmHg, P = 0.22). Additionally, the median (interquartile range) number of leukocytes in the chorion was significantly lower in the group undergoing PACI with heated, humidified CO2 compared with the group receiving cold, dry CO2 (0.7 × 10-5 (0.5 × 10-5 ) vs 3.2 × 10-5 (1.8 × 10-5 ) cells per square micron, P = 0.02). CONCLUSIONS: PACI with cold, dry CO2 causes hypercapnia, acidosis, hypotension and fetal membrane inflammation in fetal sheep, raising potential concerns for its use in humans. It seems that using heated, humidified CO2 for insufflation partially mitigates these effects and this may be a suitable alternative for reducing the risk of fetal acid-base disturbances during, and fetal membrane inflammation following, complex fetoscopic surgery. © 2018 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of the International Society of Ultrasound in Obstetrics and Gynecology.

Exerted force on the face mask in preterm infants at birth is associated with apnoea and bradycardia.

BACKGROUND: During stabilisation of preterm infants at birth, a face mask is used to provide respiratory support. However, application of these masks may activate cutaneous stretch receptors of the trigeminal nerve, causing apnoea and bradycardia. This study investigated the amount of force exerted on the face mask during non-invasive ventilation of preterm infants at birth and whether the amount of exerted force is associated with apnoea and bradycardia. METHODS: A prospective observational study was performed in preterm infants born <32 weeks of gestation who were stabilised at birth. During the first 10 minutes of respiratory support, we measured breathing and heart rate as well as the amount of force exerted on a face mask using a custom-made pressure sensor placed on top of the face mask. RESULTS: Thirty infants were included (median (IQR) gestational age(GA) 28+3 (27+0-30+0) weeks, birthweight 1104 (878-1275) grams). The median exerted force measured was 297 (198-377) grams, ranging from 0 to 1455 grams. Significantly more force was exerted on the face mask during positive pressure ventilation when compared to CPAP (410 (256-556) vs 286 (190-373) grams, p = 0.009). In a binary logistic regression model, higher forces were associated with an increased risk of apnoea (OR = 1.607 (1.556-1.661), p < 0.001) and bradycardia (OR = 1.140 (1.102-1.180), p < 0.001) during the first 10 minutes of respiratory support at birth. CONCLUSION: During mask ventilation, the median exerted force on a face mask was 297 grams with a maximum of 1455 grams. Higher exerted forces were associated apnoea and bradycardia during the first 10 minutes of respiratory support at birth.

Adverse respiratory patterns in near-term spontaneously breathing newborn lambs with elevated airway liquid volumes at birth.

Introduction: Recent evidence indicates that respiratory distress (RD) in near-term infants is caused by elevated airway liquid (EL) volume at the beginning of air-breathing after birth. While the adverse effects EL volumes on newborn lung function are known, the effects on respiratory control and breathing patterns shortly after birth (<4 h) are unknown. We investigated the effects of EL volumes on cardiorespiratory function and breathing patterns in spontaneously breathing near-term newborn lambs in the first hours after birth. Methods: At 137-8 days gestation (2-3 days prior to delivery; term ∼147 days), sterile surgery was performed on fetal sheep (n = 17) to implant catheters and blood flow probes. At 140 days, lambs were delivered via caesarean section under spinal anaesthesia. Airway liquid volumes were adjusted to mimic the level expected following vaginal delivery (∼10 ml/kg; Controls; n = 7), or elective caesarean section (∼30 ml/kg; elevated airway liquid group; EL; n = 10). Spontaneous breathing and cardiorespiratory parameters were recorded over four hours after birth. Non-invasive respiratory support with supplemental oxygen was provided if required. Results: EL lambs required higher inspired oxygen levels (p = 0.0002), were less active (p = 0.026), fed less (p = 0.008) and had higher respiratory morbidity scores than Controls (p < 0.0001). EL lambs also displayed higher rates of breathing patterns associated with RD, such as expiratory braking and tachypnoea. These patterns were particularly evident in male EL lambs who displayed higher levels of severe respiratory morbidity (e.g., expiratory braking) than female EL lambs. Conclusion: The study demonstrates that EL volumes at birth trigger respiratory behaviour and breathing patterns that resemble clinically recognised features of RD in term infants.

Assessing lung aeration using ultrasound after birth in near-term lambs at risk of respiratory distress.

Background: Optimizing respiratory support after birth requires real-time feedback on lung aeration. We hypothesized that lung ultrasound (LUS) can accurately monitor the extent and progression of lung aeration after birth and is closely associated with oxygenation. Methods: Near-term (140 days gestation, term ∼147 days), spontaneously breathing lambs with normal (controls; n = 10) or elevated lung liquid levels (EL; n= 9) were delivered by Caesarean section and monitored for four hours after birth. LUS (Phillips CX50, L3-12 transducer) images and arterial blood gases were taken every 5-20 min. LUS images were analyzed both qualitatively (grading) and quantitatively (using the coefficient of variation of pixel intensity (CoV) to estimate the degree of lung aeration), which was correlated with the oxygen exchange capacity of the lungs (Alveolar-arterial difference in oxygen; AaDO2). Results: Lung aeration, measured using LUS, and the AaDO2 improved over the first 4 h after birth. The increase in lung aeration measured using CoV of pixel intensity, but not LUS grade, was significantly reduced in EL lambs compared to controls (p = 0.02). The gradual decrease in AaDO2 after birth was significantly correlated with increased lung aeration in both control (grade, r2 = 0.60, p < 0.0001; CoV, r2 = 0.54, p < 0.0001) and EL lambs (grade, r2 = 0.51, p < 0.0001; CoV, r2 = 0.44, p < 0.0001). Conclusions: LUS can monitor lung aeration and liquid clearance after birth in spontaneously breathing near-term lambs. Image analysis techniques (CoV) may be able detect small to moderate differences in lung aeration in conditions with lung liquid retention which are not readily identified using qualitative LUS grading.

Respiratory patterns in spontaneously breathing near-term lambs delivered by caesarean section under spinal anaesthesia.

Introduction: The transition to newborn life has typically been studied in intubated and mechanically ventilated newborn lambs delivered via caesarean section (CS) under general anaesthesia. As a result, little is known of the spontaneous breathing patterns in lambs at birth, particularly those at risk of developing respiratory distress (RD). We have developed a method for delivering spontaneously breathing near-term lambs to characterise their breathing patterns in the immediate newborn period. Methods: At 137-8 days gestation (2-3 days prior to delivery; term ∼147 days), fetal lambs (n = 7) were partially exteriorised for instrumentation (insertion of catheters and flow probes) before they were returned to the uterus. At 140 days, lambs were delivered via CS under light maternal sedation and spinal anaesthesia. Lambs were physically stimulated and when continuous breathing was established, the umbilical cord was clamped. Breathing patterns were assessed by measuring intrapleural and upper-tracheal pressures during the first four hours after birth. Results: Newborn lambs display significant heterogeneity in respiratory patterns in the immediate newborn period that change with time after birth. Seven distinct breathing patterns were identified including: (i) quiet (tidal) breathing, (ii) breathing during active periods, (iii) breathing during oral feeding, (iv) tachypnoea, (v) expiratory braking manoeuvres, (vi) expiratory pauses or holding, and (vii) step changes in ventilation. Conclusions: We have described normal respiratory behaviour in newborn lambs, in order to identify respiratory behaviours that are indicative of RD in term newborn infants.

Accurate measures of changes in regional lung air volumes from chest x-rays of small animals.

Objective. To develop a robust technique for calculating regional volume changes within the lung from x-ray radiograph sequences captured during ventilation, without the use of computed tomography (CT).Approach. This technique is based on the change in transmitted x-ray intensity that occurs for each lung region as air displaces the attenuating lung tissue.Main results. Lung air volumes calculated from x-ray intensity changes showed a strong correlation (R2= 0.98) against the true volumes, measured from high-resolution CT. This correlation enables us to accurately convert projected intensity data into relative changes in lung air volume. We have applied this technique to measure changes in regional lung volumes from x-ray image sequences of mechanically ventilated, recently-deceased newborn rabbits, without the use of CT.Significance. This method is suitable for biomedical research studies,enabling quantitative regional measurement of relative lung air volumes at high temporal resolution, and shows great potential for future clinical application.

Establishment of functional residual capacity at birth: Observational study of 821 neonatal resuscitations.

AIM OF THE STUDY: Establishing functional residual capacity (FRC) during positive pressure ventilation (PPV) of apnoeic neonates is critical for survival. This may be difficult due to liquid-filled airways contributing to low lung compliance. The objectives were to describe initial PPV, changes in lung compliance and establishment of FRC in near-term/term neonates ≥36 weeks gestation at birth. METHODS: Observational study of all neonatal resuscitations between 01.07.13 and 30.06.18 in a Tanzanian referral hospital. Perinatal events and characteristics were observed and recorded by trained research assistants. PPV were performed using self-inflating bag-masks without positive end-expiratory pressure (PEEP). Ventilation signals (pressure/flow), expired CO2 (ECO2) and heart rate were recorded by resuscitation monitors. RESULTS: 19,587 neonates were born, 1451 received PPV, of these 821 of median (p25, p75) birthweight 3180 (2844, 3500) grams and gestation 38 (37, 40) weeks had ≥20 ventilations and complete datasets. There was a significant increase in expired volume (from 3.3 to 6.0 ml/kg), ECO2 (0.3-2.4%), lung compliance (0.13-0.19 ml/kg/mbar) and heart rate (109-138 beats/min) over the first 20 PPVs. Inflation volume, time, and peak inflation pressure (PIP) were stable around 12-13 ml/kg, 0.45 s, and 36 mbar, respectively. CONCLUSIONS: The combination of increasing expired volumes, ECO2, and heart rate with decreasing inflation/expired volume ratios and constant PIP, suggests establishment of FRC during the first 20 PPVs in near-term/term neonates using a self-inflating bag-mask without PEEP, the most common device worldwide for ventilating non-breathing neonates. Initial lung compliance is low, and with short inflation times, higher than recommended PIP seem necessary to deliver adequate tidal volumes.

Time to achieve desired fraction of inspired oxygen using a T-piece ventilator during resuscitation of preterm infants at birth.

AIM: To determine the time between adjustment of FiO2 at the oxygen blender and the desired FiO2 reaching the preterm infant during respiratory support at birth. METHODS: This observational study was performed using a Neopuff™ T-piece Resuscitator attached to either a test lung (during initial bench tests) or a face mask during the stabilization of infants at birth. FiO2 was titrated following resuscitation guidelines. The duration for the desired FiO2 to reach either the test lung or face mask was recorded, both with and without leakage. A respiratory function monitor was used to record FiO2 and amount of leak. RESULTS: In bench tests, the median (IQR) time taken to achieve a desired FiO2 was 34.2 (21.8-69.1) s. This duration was positively associated with the desired FiO2 difference, the direction of titration (upwards) and the occurrence of no leak (R2 0.863, F 65.016, p < 0.001). During stabilization of infants (median (IQR) gestational age 29+0 (28+2-30+0) weeks, birthweight 1290 (1240-1488) g), the duration (19.0 (0.0-57.0) s) required to reach a desired FiO2 was less, but still evident. In 27/55 (49%) titrations, the desired FiO2 was not achieved before the FiO2 levels were again changed. CONCLUSION: There is a clear delay before a desired FiO2 is achieved at the distal end of the T-piece resuscitator. This delay is clinically relevant as this delay could easily lead to over- and under titration of oxygen, which might result in an increased risk for both hypoxia and hyperoxia.

Dynamic measures of regional lung air volume using phase contrast x-ray imaging.

Phase contrast x-ray imaging can provide detailed images of lung morphology with sufficient spatial resolution to observe the terminal airways (alveoli). We demonstrate that quantitative functional and anatomical imaging of lung ventilation can be achieved in vivo using two-dimensional phase contrast x-ray images with high contrast and spatial resolution (<100 microm) in near real time. Changes in lung air volume as small as 25 microL were calculated from the images of term and preterm rabbit pup lungs (n = 28) using a single-image phase retrieval algorithm. Comparisons with plethysmography and computed tomography showed that the technique provided an accurate and robust method of measuring total lung air volumes. Furthermore, regional ventilation was measured by partitioning the phase contrast images, which revealed differences in aeration for different ventilation strategies.

Assessment of airway response distribution and paradoxical airway dilation in mice during methacholine challenge.

Detailed information on the distribution of airway diameters during bronchoconstriction in situ is required to understand the regional response of the lungs. Imaging studies using computed tomography (CT) have previously measured airway diameters and changes in response to bronchoconstricting agents, but the manual measurements used have severely limited the number of airways measured per subject. Hence, the detailed distribution and heterogeneity of airway responses are unknown. We have developed and applied dynamic imaging and advanced image-processing methods to quantify and compare hundreds of airways in vivo. The method, based on CT, was applied to house dust-mite-sensitized and control mice during intravenous methacholine (MCh) infusion. Airway diameters were measured pre- and post-MCh challenge, and the results compared demonstrate the distribution of airway response throughout the lungs during mechanical ventilation. Forced oscillation testing was used to measure the global response in lung mechanics. We found marked heterogeneity in the response, with paradoxical dilation of airways present at all airway sizes. The probability of paradoxical dilation decreased with decreasing baseline airway diameter and was not affected by pre-existing inflammation. The results confirm the importance of considering the lung as an entire interconnected system rather than a collection of independent units. It is hoped that the response distribution measurements can help to elucidate the mechanisms that lead to heterogeneous airway response in vivo.NEW & NOTEWORTHY Information on the distribution of airway diameters during bronchoconstriction in situ is critical for understanding the regional response of the lungs. We have developed an imaging method to quantify and compare the size of hundreds of airways in vivo during bronchoconstriction in mice. The results demonstrate large heterogeneity with both constriction and paradoxical dilation of airways, confirming the importance of considering the lung as an interconnected system rather than a collection of independent units.

Does growth restriction increase the vulnerability to acute ventilation-induced brain injury in newborn lambs? Implications for future health and disease.

Fetal growth restriction (FGR) and preterm birth are frequent co-morbidities, both are independent risks for brain injury. However, few studies have examined the mechanisms by which preterm FGR increases the risk of adverse neurological outcomes. We aimed to determine the effects of prematurity and mechanical ventilation (VENT) on the brain of FGR and appropriately grown (AG, control) lambs. We hypothesized that FGR preterm lambs are more vulnerable to ventilation-induced acute brain injury. FGR was surgically induced in fetal sheep (0.7 gestation) by ligation of a single umbilical artery. After 4 weeks, preterm lambs were euthanized at delivery or delivered and ventilated for 2 h before euthanasia. Brains and cerebrospinal fluid (CSF) were collected for analysis of molecular and structural indices of early brain injury. FGRVENT lambs had increased oxidative cell damage and brain injury marker S100B levels compared with all other groups. Mechanical ventilation increased inflammatory marker IL-8 within the brain of FGRVENT and AGVENT lambs. Abnormalities in the neurovascular unit and increased blood-brain barrier permeability were observed in FGRVENT lambs, as well as an altered density of vascular tight junctions markers. FGR and AG preterm lambs have different responses to acute injurious mechanical ventilation, changes which appear to have been developmentally programmed in utero.

Dynamic imaging of the lungs using x-ray phase contrast.

High quality real-time imaging of lungs in vivo presents considerable challenges. We demonstrate here that phase contrast x-ray imaging is capable of dynamically imaging the lungs. It retains many of the advantages of simple x-ray imaging, whilst also being able to map weakly absorbing soft tissues based on refractive index differences. Preliminary results reported herein show that this novel imaging technique can identify and locate airway liquid and allows lung aeration in newborn rabbit pups to be dynamically visualized.

Phase contrast X-ray imaging of mice and rabbit lungs: a comparative study.

The significant degree of X-ray phase contrast created by air-tissue interfaces, coupled with the poor radiographic contrast of conventional chest radiographs, makes the inflated lung an ideal candidate for investigating the potential diagnostic improvement afforded by phase contrast X-ray imaging. In small animals these methods highlight the lung airways and lobe boundaries and reveal the lung tissue as a speckled intensity pattern not seen in other soft tissues. We have compared analyser-based and propagation-based phase contrast imaging modalities, together with conventional radiographic imaging, to ascertain which technique shows the greatest image enhancement for various lung sizes. The conventional radiographic image of a mouse was obtained on a Siemens Nova 3000 mammography system, whilst phase contrast images of mice and rabbit chests were acquired at the medical imaging beamline (20B2) at the SPring-8 synchrotron radiation research facility in Japan. For mice aged 1 day, 1 week and 1 month old it was determined that analyser-based imaging showed the greatest overall image contrast, however, for an adult rabbit both techniques yielded excellent contrast. The success of these methods in creating high quality images for rabbit lungs raises the possibility of improving human lung imaging using phase contrast techniques.

Umbilical blood flow patterns directly after birth before delayed cord clamping.

BACKGROUND: Delayed umbilical cord clamping (DCC) affects the cardiopulmonary transition and blood volume in neonates immediately after birth. However, little is known of blood flow in the umbilical vessels immediately after birth during DCC. The objective is to describe the duration and patterns of blood flow through the umbilical vessels during DCC. METHODS: Arterial and venous umbilical blood flow was measured during DCC using Doppler ultrasound in uncomplicated term vaginal deliveries. Immediately after birth, the probe was placed in the middle of the umbilical cord, pattern and duration of flow in vein and arteries were evaluated until cord clamping. RESULTS: Thirty infants were studied. Venous flow: In 10% no flow was present, in 57% flow stopped at 4:34 (3:03-7:31) (median (IQR) min:sec) after birth, before the cord was clamped. In 33%, flow continued until cord clamping at 5:13 (2:56-9:15) min:sec. Initially, venous flow was intermittent, increasing markedly during large breaths or stopping and reversing during crying, but then became continuous. Arterial flow: In 17% no flow was present, in 40% flow stopped at 4:22 (2:29-7:17) min:sec, while cord pulsations were still palpable. In 43% flow continued until the cord was clamped at 5:16 (3:32-10:10) min:sec. Arterial flow was pulsatile, unidirectional towards placenta or bidirectional to/from placenta. In 40% flow became continuous towards placenta later on. CONCLUSIONS: During delayed umbilical cord clamping, venous and arterial umbilical flow occurs for longer than previously described. Net placental transfusion is probably the result of several factors of which breathing could play a major role. Umbilical flow is unrelated to cessation of pulsations.

The effect of oxygen content during an initial sustained inflation on heart rate in asphyxiated near-term lambs.

OBJECTIVE: At birth, an initial sustained inflation (SI) uniformly aerates the lungs, increases arterial oxygenation and rapidly improves circulatory recovery in asphyxiated newborns. We hypothesised that lung aeration, in the absence of an increase in arterial oxygenation, can increase heart rate (HR) in asphyxiated near-term lambs. INTERVENTIONS: Lambs were delivered and instrumented at 139±2 days of gestation. Asphyxia was induced by umbilical cord clamping and then delaying the onset of ventilation until mean carotid arterial pressures (CAPs) had decreased <20 mm Hg. Lambs then received a single 30-s SI using nitrogen (N2; n=6), 5% oxygen (O2; n=6), 21% O2 (n=6) or 100% O2 (n=6) followed by ventilation in air for 30 min. MAIN OUTCOME MEASURES: HR, CAP and pulmonary blood flow (PBF) were continuously recorded. RESULTS: HR and PBF increased more quickly in lambs resuscitated with 100% and 21% O2 than with 5% O2 or N2. HR and PBF recovery in the 5% O2 group was delayed relative to all other oxygen SI groups. HR in 5%, 21% and 100% O2 groups reached 100 bpm before the SI was complete. HR and PBF in the N2 group did not increase until 10 s after the SI was completed and ventilation was initiated with air. CAP tended to increase quicker in all O2 groups than in N2 group. CONCLUSIONS: Oxygen content during an SI is important for circulatory recovery in asphyxiated lambs. This increase in HR is likely driven by the increase in PBF and venous return to the heart.

Oxytocin, oxytocin-associated neurophysin, and prostaglandin F2 alpha concentrations in the utero-ovarian vein of pregnant and nonpregnant sheep.

Oxytocin, oxytocin-associated neurophysin (neurophysin), prostaglandin F2 alpha (PGF2 alpha), and progesterone concentrations were measured in the utero-ovarian vein (UOV) of sheep during the estrous cycle and early pregnancy. On days 13-16 of the cycle, large pulses of PGF2 alpha, oxytocin, and neurophysin were measured in samples collected at hourly intervals from the UOV draining a corpus luteum (UOV/CL). Most of the PGF2 alpha pulses (96.5%) coincided with a pulse of both oxytocin and neurophysin, whereas only 55.6% of oxytocin pulses coincided with a pulse of PGF2 alpha. Therefore, during luteolysis in sheep, uterine PGF2 alpha release is closely associated with ovarian oxytocin release, and oxytocin release is unlikely to be dependent upon a uterine PGF2 alpha stimulus. During frequent sampling, coincident oxytocin pulses were measured in 1) both UOVs when a CL was present in both ovaries and 2) the jugular vein, carotid artery, and UOV/CL, with a significantly higher oxytocin pulse concentration occurring in jugular venous compared with carotid arterial plasma. Pituitary and luteal release of oxytocin may, therefore, occur simultaneously and be controlled by a circulating factor in sheep. Compared to days 13-16 of the cycle, significantly (P less than 0.001) fewer pulses of PGF2 alpha, which were significantly (P less than 0.001) smaller in amplitude, were measured in UOV samples collected frequently during early pregnancy. The frequency of oxytocin pulses observed in the UOV/CL of pregnant sheep was not significantly (P greater than 0.1) different from that observed in cyclic ewes, although most (86.4%) oxytocin pulses occurred in the absence of a PGF2 alpha pulse. In contrast, when a pulse of PGF2 alpha was observed in the UOV/CL of pregnant ewes, it usually coincided with a pulse of oxytocin. The suppression of uterine PGF2 alpha release during early pregnancy is not considered to result from a lack of stimulation by oxytocin.

Prolonged hypoxia induced by the reduction of maternal uterine blood flow alters insulin-like growth factor-binding protein-1 (IGFBP-1) and IGFBP-2 gene expression in the ovine fetus.

Insulin-like growth factors (IGF-I and IGF-II) are potent mitogenic and differentiating peptides which are synthesized by many fetal tissues. In the circulation and tissue fluids, IGFs are bound to binding proteins (BPs) which not only function as carrier proteins, but also inhibit or modulate the biological actions of IGFs. We have previously shown that prolonged hypoxia in the ovine fetus induced by the reduction of maternal uterine blood flow for 24 h causes a reduction in the DNA synthesis rate in selected fetal tissues. To determine if this effect is due to alterations in the local synthesis of tissue IGFs and their binding proteins or to changes in systemic concentrations of IGFs and IGFBPs, we have investigated the abundance of mRNAs encoding IGFs and IGFBPs in selected tissues and changes in plasma IGFs and IGFBPs. Ovine fetuses (115-120 days gestation; n = 6) underwent 24 h of hypoxia by the reduction of maternal uterine blood flow (RUBF). Controls (n = 6) underwent the same surgical procedure without RUBF. Serial plasma samples were collected before, during, and after the experiment, and tissues were collected at the end of 24 h. Mean plasma IGF-I and IGF-II concentrations tended to be lower in hypoxic fetuses than in controls during the course of hypoxia, but these differences were not statistically significant. Tissue mRNA levels for IGF-I and IGF-II in lung, muscle, thymus, and kidney were similar in control and hypoxic fetuses after 24 h of hypoxia. The relative abundance of liver IGF-I and IGF-II mRNAs was lower in hypoxic fetuses, but only IGF-I mRNA levels were significantly different from the control values (P < 0.05). Compared to control fetuses, IGFBP-1 mRNA levels in the liver of hypoxic fetuses were increased 3- to 7-fold, and IGFBP-1 mRNA expression was induced in kidneys of some hypoxic fetuses (two of six). In addition, IGFBP-2 mRNA levels were decreased in the liver (50%) and kidney (30%) of hypoxic fetuses. The increase in liver IGFBP-1 mRNA abundance and the decrease in liver and kidney IGFBP-2 mRNA abundance were accompanied by an increase in IGFBP-1 levels and a decrease in IGFBP-2 levels in fetal plasma. No changes were observed in either plasma levels or tissue mRNA abundance for IGFBP-3. Analysis of the time course of changes in plasma revealed that the changes in IGFBP-1 and IGFBP-2 occurred within 4 h of hypoxia.(ABSTRACT TRUNCATED AT 400 WORDS)

Catecholamines stimulate the synthesis and release of insulin-like growth factor binding protein-1 (IGFBP-1) by fetal sheep liver in vivo.

In fetal sheep, prolonged hypoxia (for 24 h) induced by a reduction in maternal uterine artery blood flow, increases insulin-like growth factor binding protein-1 (IGFBP-1) levels and decreases IGFBP-2 levels in the plasma, with corresponding changes in messenger RNA (mRNA) levels in the liver. Since IGFBP-1 synthesis in liver cells in vitro is stimulated by compounds that increase intracellular cAMP concentrations, we hypothesized that the increased IGFBP-1 synthesis during prolonged hypoxemia may be induced by circulating catecholamines, that are released during hypoxia, and that elevate fetal liver cAMP levels. Our aim was to determine the effect of 24-h catecholamine infusions on the synthesis and release of IGFBP-1 and IGFBP-2 in fetal sheep. Vascular catheters were implanted into fetuses at 110-115 days gestation in 14 pregnant ewes. After a 5-day recovery period, fetuses received a 24-h infusion of either norepinephrine (1 micrograms/kg.min, n = 5), epinephrine (0.25 micrograms/kg.min, n = 5), or vehicle (normal saline, n = 4). Fetal carotid arterial samples were collected at specified intervals throughout the infusion for the determination of blood glucose concentrations, plasma catecholamine concentrations by HPLC, insulin, and glucagon concentrations by RIA, and IGFBP levels by Western ligand blotting. After 24 h, the ewe and fetus were killed and selected fetal tissues (liver and kidney) were collected, and analyzed for IGFBP mRNA levels by northern blotting followed by laser densitometric quantification. Plasma catecholamine concentrations were increased in treated fetuses to levels that may be expected in fetuses subjected to prolonged hypoxia. In epinephrine and norepinephrine infused fetuses, blood glucose and plasma glucagon concentrations were increased significantly, whereas plasma insulin concentrations were decreased significantly. Norepinephrine and epinephrine infusions increased IGFBP-1 levels significantly (2- to 5-fold) in fetal plasma within 8-12 h, and the time course pattern of elevation of plasma IGFBP-1 levels was similar to that observed in prolonged hypoxia. After 24 h of either norepinephrine or epinephrine infusion, IGFBP-1 mRNA levels in the liver of fetuses were increased significantly (5- to 7-fold) compared to those of vehicle infused fetuses. IGFBP-2, -3, and -4 levels in fetal plasma were not affected by either infusion, nor were IGFBP-2 mRNA levels in fetal liver and kidney.(ABSTRACT TRUNCATED AT 400 WORDS)