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Fetal heart rate variability (FHRV) is a key index of antenatal and intrapartum fetal well-being. FHRV is well established to be mediated by both arms of the autonomic nervous system, but it remains unknown whether higher centers in the forebrain contribute to FHRV. We tested the hypothesis that selective forebrain ischemia would impair the generation of FHRV. Sixteen chronically instrumented near-term fetal sheep were subjected to either forebrain ischemia induced by bilateral carotid occlusion or sham-ischemia for 30 min. Time, frequency, and nonlinear measures of FHRV were assessed during and for seven days after ischemia. Ischemia was associated with profound suppression of electroencephalographic (EEG) power, which remained suppressed throughout the recovery period ( P < 0.001). During the first 5 min of ischemia, multiple time and frequency domain measures were increased (all P < 0.05) before returning back to sham levels. A delayed increase in sample entropy was observed during ischemia ( P < 0.05). For the first 3 h after ischemia, there was moderate suppression of two measures of FHRV (very-low frequency power and the standard deviation of RR-intervals, both P < 0.05) and increased sample entropy ( P < 0.05). Thereafter, all measures of FHRV returned to control levels. In conclusion, profound forebrain ischemia sufficient to lead to severe neural injury had only transient effect on multiple measures of FHRV. These findings suggest that the forebrain makes a limited contribution to FHRV. FHRV therefore primarily originates in the hindbrain and is unlikely to provide meaningful information on forebrain neurodevelopment or metabolism.

Background: Sensitive biomarkers are needed to rapidly identify high-risk infants after hypoxia-ischemia for neuroprotective treatment. Hypotension is a key determinant of hypoxic-ischemic neural injury, and a potent stimulus of humoral pressors including angiotensin-II and arginine vasopressin. We therefore aimed to quantify the relationship between vasopressin and angiotensin-II levels in the latent phase after hypoxia-ischemia induced by umbilical cord occlusion (UCO) with both the severity of preceding hypotension and subsequent neuronal injury., Methods: Chronically instrumented near-term fetal sheep underwent sham-UCO or UCO for either 15 min or until mean arterial pressure was <8 mmHg. Neuronal injury was assessed after 72 h recovery., Results: Umbilical cord occlusion was associated with severe hypotension that recovered after UCO; two fetuses developed profound secondary hypotension within 6 h and died. Vasopressin levels but not angiotensin-II were significantly elevated 1-3 h after UCO and were closely associated with the severity of hypotension during UCO and the subsequent severity of neuronal loss in the parasagittal and lateral cortex, caudate nucleus and putamen. The Youden cut-point for vasopressin at 1 h was 180.0 pmol/L, with sensitivity 100% and specificity 92.3% for severe neuronal injury or death., Conclusion: Vasopressin levels shortly after moderate-severe hypoxia-ischemia may be a useful early biomarker to guide the timely implementation of neuroprotective treatment., Impact: It can be difficuIt to rapidly identify infants who might benefit from therapeutic hypothermia. We investigated whether increases in plasma pressor hormones early after hypoxia-ischemia were biomarkers for neonatal hypoxic-ischemic encephalopathy using near-term fetal sheep. Arginine vasopressin levels were elevated at 1-3 h after hypoxia-ischemia and were predictive of the severity of preceding hypotension and subsequent risk of severe neuronal injury or death after hypoxia-ischemia. Arginine vasopressin may help identify neonates at high risk of hypoxic-ischemic encephalopathy early within the therapeutic window for hypothermia.

Fetal heart rate variability (FHRV) is a widely used index of intrapartum well being. Both arms of the autonomic system regulate FHRV under normoxic conditions in the antenatal period. However, autonomic control of FHRV during labor when the fetus is exposed to repeated, brief hypoxemia during uterine contractions is poorly understood. We have previously shown that the sympathetic nervous system (SNS) does not regulate FHRV during labor-like hypoxia. We therefore investigated the hypothesis that the parasympathetic system is the main mediator of intrapartum FHRV. Twenty-six chronically instrumented fetal sheep at 0.85 of gestation received either bilateral cervical vagotomy ( n = 7), atropine sulfate ( n = 7), or sham treatment (control, n = 12), followed by three 1-min complete umbilical cord occlusions (UCOs) separated by 4-min reperfusion periods. Parasympathetic blockade reduced three measures of FHRV before UCOs (all P < 0.01). Between UCOs, atropine and vagotomy were associated with marked tachycardia (both P < 0.005), suppressed measures of FHRV (all P < 0.01), and abolished FHRV on visual inspection compared with the control group. Tachycardia in the atropine and vagotomy groups resolved over the first 10 min after the final UCO, in association with evidence that the SNS contribution to FHRV progressively returned during this time. Our findings support that SNS control of FHRV is acutely suppressed for at least 4 min after a deep intrapartum deceleration and takes 5-10 min to recover. The parasympathetic system is therefore likely to be the key mediator of FHRV once frequent FHR decelerations are established during labor.

Premature babies are at high risk of serious neurodevelopmental disabilities, which in many cases are related to perinatal hypoxic-ischemic encephalopathy (HIE). Studies of neuroprotection in animal models consistently suggest that treatment must be started as early as possible in the first 6 h after hypoxia-ischemia (HI), the so-called latent phase before secondary deterioration, to improve outcomes. We have shown in preterm sheep that EEG biomarkers of injury, in the form of high-frequency micro-scale spike transients, develop and evolve in this critical latent phase after severe asphyxia. Real-time automatic identification of such events is important for the early and accurate detection of HI injury, so that the right treatment can be implemented at the right time. We have previously reported successful strategies for accurate identification of EEG patterns after HI. In this study, we report an alternative high-performance approach based on the fusion of spectral Fourier analysis and Type-I fuzzy classifiers (FFT-Type-I-FLC). We assessed its performance in over 2520 min of latent phase EEG recordings from seven asphyxiated in utero preterm fetal sheep exposed to a range of different occlusion periods. The FFT-Type-I-FLC classifier demonstrated 98.9 ± 1.0% accuracy for identification of high-frequency spike transients in the gamma frequency band (namely 80-120 Hz) post-HI. The spectral-based approach (FFT-Type-I-FLC classifier) has similar accuracy to our previous reverse biorthogonal wavelets rbio2.8 basis function and type-1 fuzzy classifier (rbio-WT-Type-1-FLC), providing competitive performance (within the margin of error: 0.89%), but it is computationally simpler and would be readily adapted to identify other potentially relevant EEG waveforms.

Perinatal Hypoxic-Ischemia Encephalopathy (HIE) in newborn infants, due to birth-related circumstances such as oxygen deprivation in brain cells, is caused by the disruption in blood flow through the umbilical cord. Subcortical neuronal loss due to the HIE can lead to cerebral palsy and other chronic neurological conditions. Pre-clinical EEG studies using in utero sheep have demonstrated that particular micro-scale HI transients emerge along a suppressed EEG background during a latent phase of 3-6 hours, after a severe HI insult. Whilst the nature of these micro-scale transients is not well understood, it has been hypothesized that such transients may be signatures of the evolving hypoxic-ischemic brain injury, possessing the potential to be served as the diagnosis biomarkers for the injury. Cerebral hypothermia is optimally neuroprotective only if administered within the first 2-3 hours post HI insult. Using data from a cohort of in utero preterm fetal sheep (n=5, at 0.7 of gestational age), this paper indicates how the number of automatically quantified micro-scale sharp wave transients from asphyxiated preterm fetal sheep, statistically correlate to the amount of NeuN-positive neurons measured in caudate nucleus of striatum. Different temporal window sizes of 2hrs, 1hr, ½hr and 10mins within the early phase of the latent phase are examined using our developed Wavelet Type-2 Fuzzy classifier for sharp detection. Analyses were narrowed down to 10min intervals to assess where exactly in time the occurrence of the HI micro-scale sharp waves demonstrate a significant correlation. Signal processing wise, results from the sub-windows indicate a timing trend that highlights a positive correlation, between the number of automatic quantifications and the amount of surviving neurons in the preterm brain, permitting the possibility of a point of care (POC) intervention to stop the spread of injury before it becomes irreversible.

Basal ganglia injury at term remains a major cause of disability, such as cerebral palsy. In this study we tested the hypotheses that blockade of astrocytic connexin hemichannels with a mimetic peptide would improve survival of striatal phenotypic neurons after global cerebral ischaemia in term-equivalent fetal sheep, and that neuronal survival would be associated with electrophysiological recovery. Fetal sheep (0.85 gestation) were randomly assigned to receive a short or long (1 or 25 h) intracerebroventricular infusion of a mimetic peptide or vehicle, starting 90 minutes after 30 minutes of cerebral ischaemia. Sheep were killed 7 days after ischaemia. Cerebral ischaemia was associated with reduced numbers of calbindin-28k, calretinin, parvalbumin and GAD positive striatal neurons (P < 0.05 ischaemia + vehicle, n = 6 vs. sham ischaemia, n = 6) but not ChAT or nNOS positive neurons. Short infusion of peptide (n = 6) did not significantly improve survival of any striatal phenotype. Long infusion of peptide (n = 6) was associated with increased survival of calbindin-28k, calretinin, parvalbumin and GAD positive neurons (P < 0.05 vs. ischaemia + vehicle). Neurophysiological recovery was associated with improved survival of calbindin-28k, calretinin and parvalbumin positive striatal neurons (P < 0.05 for all). In conclusion, connexin hemichannel blockade after cerebral ischaemia in term-equivalent fetal sheep improves survival of striatal GABA-ergic neurons.

Subclinical (shallow) heart rate decelerations occur during neonatal sepsis, but there is limited information on their relationship with hypotension or whether they occur before birth. We examined whether subclinical decelerations, a fall in fetal heart rate (FHR) that remained above 100 bpm, were associated with hypotension in preterm fetal sheep exposed to lipopolysaccharide (LPS). Chronically-instrumented fetal sheep at 0.7 gestation received continuous low-dose LPS infusions (n = 15, 100 ng/kg over 24 h, followed by 250 ng/kg/24 h for 96 h) or saline (n = 8). Boluses of 1 μg LPS or saline were given at 48 and 72 h. FHR variability (FHRV) was calculated, and sample asymmetry was used to assess the severity and frequency of decelerations. Low-dose LPS infusion did not affect FHR. After the first LPS bolus, 7 fetuses remained normotensive, while 8 developed hypotension (a fall in mean arterial blood pressure of ≥5 mmHg). Developing hypotension was associated with subclinical decelerations, with a corresponding increase in sample asymmetry and FHRV (p < 0.05). The second LPS bolus was associated with similar but attenuated changes in FHR and blood pressure (p < 0.05). In conclusion, subclinical decelerations are not consistently seen during prenatal exposure to LPS, but may be a useful marker of developing inflammation-related hypotension before birth.

Power spectral analysis of fetal heart rate variability has been proposed to provide a non-invasive estimate of autonomic balance. However, there are few systematic data before birth. We therefore examined developmental changes in the frequency power spectrum at very low (0-0.04 Hz), low (0.04-0.15 Hz) and high frequencies (0.15-0.4 Hz), as well as the ratio of low- to high-frequency power (LF/HF), in chronically catheterized, healthy fetal sheep at 0.6 (n = 8), 0.7 (n = 7) and 0.8 gestational age (ga; n = 11). In a second study, 0.8 ga fetuses received either atropine (4.8 mg bolus, then 4.8 mg h(-1) for 30 min, n = 6) or 6-hydroxydopamine (20 mg ml(-1) at 2.5 ml h(-1) for 3 h; n = 9). Data were analysed by sleep state, defined by low-voltage-high-frequency (LV) or high-voltage-low-frequency (HV) EEG. Total spectral power increased with gestational age (P < 0.05), while LF/HF decreased from 0.6 to 0.7 ga. At 0.8 ga, heart rate and LF/HF were significantly higher during HV than LV sleep (P < 0.05). Consistent with this, although total spectral power was not significantly greater during HV sleep, there was a significant interaction between sleep state and frequency band (P = 0.02). Both atropine (P = 0.05) and 6-hydroxydopamine (P < 0.05) were associated with an overall reduction in spectral power but no significant effect on the LF/HF ratio. This study does not support substantial, consistent differences between the frequencies of sympathetic and parasympathetic activity in late-gestation fetal sheep.

Spontaneous antenatal hypoxia is associated with high risk of adverse outcomes, however, there is little information on neural adaptation to labor-like insults. Chronically instrumented near-term sheep fetuses (125 ± 3 days, mean ± SEM) with baseline PaO2 < 17 mmHg (hypoxic group: n = 8) or > 17 mmHg (normoxic group: n = 8) received 1-minute umbilical cord occlusions repeated every 5 minutes for a total of 4 hours, or until mean arterial blood pressure (MAP) fell below 20 mmHg for two successive occlusions. 5/8 fetuses with pre-existing hypoxia were unable to complete the full series of occlusions (vs. 0/8 normoxic fetuses). Pre-existing hypoxia was associated with progressive metabolic acidosis (nadir: pH 7.08 ± 0.04 vs. 7.33 ± 0.02, p<0.01), hypotension during occlusions (nadir: 24.7 ± 1.8 vs. 51.4 ± 3.2 mmHg, p<0.01), lower carotid blood flow during occlusions (23.6 ± 6.1 vs. 63.0 ± 4.8 mL/min, p<0.01), greater suppression of EEG activity during, between, and after occlusions (p<0.01) and slower resolution of cortical impedance, an index of cytotoxic edema. No normoxic fetuses, but 4/8 hypoxic fetuses developed seizures 148 ± 45 minutes after the start of occlusions, with a seizure burden of 26 ± 6 sec during the inter-occlusion period, and 15.1 ± 3.4 min/h in the first 6 hours of recovery. In conclusion, in fetuses with pre-existing hypoxia, repeated brief asphyxia at a rate consistent with early labor is associated with hypotension, cephalic hypoperfusion, greater EEG suppression, inter-occlusion seizures, and more sustained cytotoxic edema, consistent with early onset of neural injury.

Progenitor cell proliferation is ubiquitous in the subventricular zone (SVZ) and subgranular zone (SGZ) of adult mammalian brains, however, the abundance and distribution of proliferation are surprisingly heterogeneous between species. In rodents, proliferation is high in both the SVZ and SGZ, while in humans proliferation is prominent in the SVZ but limited in the SGZ. To accurately study proliferation and how it changes in human disease, we should focus on animals in which the patterns of proliferation are consistent with the human brain. In this study, we characterized the neurogenic niches of the adult sheep, an animal model with a longer lifespan than rodents and a highly gyrencephalic brain, using 5-bromo-2'-deoxyuridine (BrdU) as a mitotic marker and neuronal nuclear antigen to identify neuronal lineage cells. Our study demonstrates that the sheep SVZ is organized into the same distinct layers that are comparable to what has been described in humans. The rate of maturation of new neurons was slower in sheep than in previous reports in rodents, with only 20% of BrdU-positive cells showing neuronal phenotype after 4 months survival following BrdU administration. Most importantly, as in the human, there was much greater proliferation in the sheep SVZ than in the SGZ. These results suggest that the sheep is a better basis for comparisons with human SVZ and SGZ neurogenesis than rodents., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

The sympathetic nervous system (SNS) is an important mediator of fetal adaptation to life-threatening in utero challenges, such as asphyxia. Although the SNS is active well before term, SNS responses mature significantly over the last third of gestation, and its functional contribution to adaptation to asphyxia over this critical period of life remains unclear. Therefore, we examined the hypotheses that increased renal sympathetic nerve activity (RSNA) is the primary mediator of decreased renal vascular conductance (RVC) during complete umbilical cord occlusion in preterm fetal sheep (101 ± 1 days; term 147 days) and that near-term fetuses (119 ± 0 days) would have a more rapid initial vasomotor response, with a greater increase in RSNA. Causality of the relationship of RSNA and RVC was investigated using surgical (preterm) and chemical (near-term) denervation. All fetal sheep showed a significant increase in RSNA with occlusion, which was more sustained but not significantly greater near-term. The initial fall in RVC was more rapid in near-term than preterm fetal sheep and preceded the large increase in RSNA. These data suggest that although RSNA can increase as early as 0.7 gestation, it is not the primary determinant of RVC. This finding was supported by denervation studies. Interestingly, chemical denervation in near-term fetal sheep was associated with an initial fall in blood pressure, suggesting that by 0.8 gestation sympathetic innervation of nonrenal vascular beds is critical to maintain arterial blood pressure during the rapid initial adaptation to asphyxia.

Unlabelled: Fetal susceptibility to hypoxic brain injury increases over the last third of gestation. This study examined the hypothesis that this is associated with impaired mitochondrial adaptation, as measured by more rapid oxidation of cytochrome oxidase (CytOx) during profound asphyxia., Methods: Chronically instrumented fetal sheep at 0.6, 0.7, and 0.85 gestation were subjected to either 30 min (0.6 gestational age (ga), n = 6), 25 min (0.7 ga, n = 27) or 15 min (0.85 ga, n = 17) of complete umbilical cord occlusion. Fetal EEG, cerebral impedance (to measure brain swelling) and near-infrared spectroscopy-derived intra-cerebral oxygenation (ΔHb = HbO(2) - Hb), total hemoglobin (THb) and CytOx redox state were monitored continuously. Occlusion was associated with profound, rapid fall in ΔHb in all groups to a plateau from 6 min, greatest at 0.85 ga compared to 0.6 and 0.7 ga (p<0.05). THb initially increased at all ages, with the greatest rise at 0.85 ga (p<0.05), followed by a progressive fall from 7 min in all groups. CytOx initially increased in all groups with the greatest rise at 0.85 ga (p<0.05), followed by a further, delayed increase in preterm fetuses, but a striking fall in the 0.85 group after 6 min of occlusion. Cerebral impedance (a measure of cytotoxic edema) increased earlier and more rapidly with greater gestation. In conclusion, the more rapid rise in CytOx and cortical impedance during profound asphyxia with greater maturation is consistent with increasing dependence on oxidative metabolism leading to earlier onset of neural energy failure before the onset of systemic hypotension.

Moderate cerebral hypothermia significantly improves survival without disability from perinatal hypoxia-ischemia. However, protection is partial. Insulin-like growth factor 1 (IGF-1) plays a key role in oligodendrocyte survival and myelination. The purpose of this study was to test the hypothesis that the combination of IGF-1 plus hypothermia could reduce postischemic white matter damage compared with hypothermia alone. Unanesthetized near-term fetal sheep received 30 min of cerebral ischemia, followed by either an infusion of 3 μg of IGF-1 intracerebroventricularly from 4.5 to 5.5 h plus cooling from 5.5 to 72 h (IGF-1 + hypothermia; n = 8), vehicle infusion plus cooling from 5.5 to 72 h (vehicle + hypothermia; n = 12), sham cooling plus sham infusion (ischemia control; n = 12) or sham ischemia (n = 5). The fetal extradural temperature was reduced from 39.4 ± 0.1°C to between 30 and 33°C. White matter was assessed after 5 days. Ischemia was associated with severe loss of CNPase-positive oligodendrocytes in white matter compared with sham ischemia (380 ± 138 vs. 1,180 ± 152 cells/field; mean ± SD; p < 0.001). Delayed hypothermia reduced cell loss (847 ± 297 cells/field, p < 0.01, vs. ischemia control), but there was no significant difference between vehicle + hypothermia and IGF-1 + hypothermia (1,015 ± 211 cells/field; NS). Ischemia was associated with increased caspase 3 expression in white matter (216 ± 41 vs. 19 ± 18 cells/field; p < 0.001). Hypothermia reduced numbers of activated caspase 3-positive cells (116 ± 81 cells/field; p < 0.05), with no significant difference between vehicle + hypothermia and IGF-1 + hypothermia (91 ± 27 cells/field; NS). In conclusion, delayed cotreatment with IGF-1 plus hypothermia after ischemia was associated with an improvement in white matter damage similar to that achieved by hypothermia alone., (Copyright © 2011 S. Karger AG, Basel.)

Perinatal hypoxia remains a significant cause of brain damage. Currently there are no biomarkers to detect the at risk brain. Recent research, however, suggests that the appearance of epileptiform transients in the first 6-8 hours after hypoxia (the latent phase of injury) are predictive of neural outcome. To quantify this further a key need is to automate EEG signal analysis to aid clinical staff with the vast amounts of complex data to review. In this study, we present a semi-automated method for spike detection in the fetal sheep EEG. The method utilizes the short time Fourier transform and peak separation to extract spikes. The performance of the method was found to be high in sensitivity and selectivity over 3 distinct time points.

Objective: To develop a model to study the fetal effects of intrapleural infusion of OK-432 (Picibanil), a pleurodesis agent derived from killed Gram-positive streptococci., Methods: OK-432 (0.1 mg, n = 5), or normal saline (n = 5) were infused over 20 min into the pleural space of chronically instrumented preterm fetal sheep at 0.7 gestation. Fetal physiological parameters, including breathing and nuchal activity were monitored in utero from 6 h before infusion until 12 h afterward, and fetuses were killed after 7 days recovery., Results: OK-432 was associated with transient suppression of fetal EEG activity, breathing and body movements from 3-6 h after infusion. Hypotension and hypoxia did not occur. At postmortem, local pleural adhesions were seen around the site of OK-432 infusion but not in saline treated fetuses., Conclusions: Intrapleural administration of OK-432 is associated with marked but transient fetal behavioral effects. This model will enable preclinical investigation of the neural and cardiovascular safety of OK-432 at a clinical relevant stage of development., (Copyright 2009 S. Karger AG, Basel.)

There is evidence that preterm fetuses have blunted chemoreflex-mediated responses to hypoxia. However, the preterm fetus has much lower aerobic requirements than at term, and so moderate hypoxia may not be sufficient to elicit maximal chemoreflex responses; there are only limited quantitative data on the ontogeny of chemoreflex and hemodynamic responses to severe asphyxia. Chronically instrumented fetal sheep at 0.6 (n = 12), 0.7 (n = 12), and 0.85 (n = 8) of gestational age (GA; term = 147 days) were exposed to 30, 25, or 15 min of complete umbilical cord occlusion, respectively. At all ages, occlusion was associated with early onset of bradycardia, profoundly reduced femoral blood flow and conductance, and hypertension. The 0.6-GA fetuses showed a significantly slower and lesser fall in femoral blood flow and conductance compared with the 0.85-GA group, with a correspondingly reduced relative rise in mean arterial blood pressure. As occlusion continued, the initial adaptation was followed by loss of peripheral vasoconstriction and progressive development of hypotension in all groups. The 0.85-GA fetuses showed significantly more sustained reduction in femoral conductance but also more rapid onset of hypotension than either of the younger groups. Electroencephalographic (EEG) activity was suppressed during occlusion in all groups, but the degree of suppression was less at 0.6 GA than at term. In conclusion, the near-midgestation fetus shows attenuated initial (chemoreflex) peripheral vasomotor responses to severe asphyxia compared with more mature fetuses but more sustained hemodynamic adaptation and reduced suppression of EEG activity during continued occlusion of the umbilical cord.

After exposure to asphyxia, infants may develop both prolonged, clinically evident seizures and shorter, clinically silent seizures; however, their effect on cerebral tissue oxygenation is unclear. We therefore examined the hypothesis that the increase in oxygen delivery during postasphyxial seizures might be insufficient to meet the needs of increased metabolism, thus causing a fall in tissue oxygenation, in unanesthetized near-term fetal sheep in utero (gestational age 125+/-1 days). Fetuses were administered an infusion of the specific adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine, followed by 10 mins of asphyxia induced by complete umbilical cord occlusion. The fetuses then recovered for 3 days. Sixty-one episodes of electrophysiologically defined seizures were identified in five fetuses. Tissue PO(2) (tPO(2)) did not change significantly during short seizures (<3.5 mins), 5.2+/-0.2 versus baseline 5.6+/-0.1 mm Hg (NS), but fell to 2.2+/-0.2 mm Hg during seizures lasting more than 3.5 mins (P<0.001). During prolonged seizures, cortical blood flow did not begin to increase until tPO(2) had begun to fall, and then rose more slowly than the increase in metabolism, with a widening of the brain to blood temperature gradient. In conclusion, in the immature brain, during prolonged, but not short seizures, there is a transient mismatch between cerebral blood flow and metabolism leading to significant cerebral deoxygenation.

Secondary amino acid disturbances from circulatory responses during hypoxia may cause problems in interpreting plasma amino acid profiles of sick babies investigated for possible inherited defects. Systematic studies to characterise them are difficult in man. We investigated the effects of hypoxia on plasma amino acids by studying 9 late gestation fetal sheep in utero during 11 one hour episodes of moderately severe isocapnic hypoxia. In 6 experiments, maternal plasma amino acids were also monitored. Fourteen fetal plasma amino acids increased significantly, with the largest proportionate changes in alanine, valine, leucine, isoleucine, phenylalanine, tyrosine, ornithine and lysine. Maternal amino acids did not increase. Probable explanations were reflex peripheral vasoconstriction in skeletal muscle beds and decreased hepatic blood flow. The findings extend our knowledge of the fetal response to hypoxic stress, demonstrate the importance of skeletal muscle in branched-chain amino acid metabolism, and should help with interpretation of postnatal plasma amino acid disturbances.

We measured mean arterial blood pressure (MAP), fetal heart rate (FHR), the baroreflex, weight, blood gases, blood glucose and lactate in chronically instrumented fetal sheep for 14 days from Day 105 to Day 109 gestation, with or without daily administration of 1 h of acute isocapnic hypoxaemia (arterial O2 partial pressure, PaO2, 11-13 mm Hg; 1 mm Hg = 133 Pa). Fetuses subjected to hypoxia showed no significant differences in MAP or FHR v. control fetuses. However, examination of all fetuses together revealed that there were two distinct groups: those showing a rise in MAP over the 14 days ('pressure up' group, PU), and those in which blood pressure did not increase or showed only a slight decrease ('pressure down' group, PD). PU fetuses were proportionately larger than PD fetuses. In contrast to PU fetuses, PD fetuses had lower blood glucose concentration, arterial O2 saturation (SaO2), PaO2, total haemoglobin, haematocrit and oxygen content, and higher lactate concentration, pH and PaCO2. PU fetuses showed a shift of the baroreflex MAP/R-R interval curve to the right, however, the PD group showed a shift upwards from Day 1 to Day 14. The PD group responded to hypoxia with a greater increase in MAP than the PU group. Thus, repeated acute moderate isocapnic hypoxia does not affect development of MAP or FHR in late gestation fetal sheep. However, MAP follows different trajectories in individual fetuses, related to fetal size and the availability of oxygen and/or glucose. Cardiovascular chemoreflexes and baroreflexes are also different, depending on the MAP trajectory. These data indicate an important association between growth and blood pressure development, and also show that differences in growth are associated with changes in cardiovascular control.

1. The respiratory effects of corticotrophin-releasing factor (CRF) and the CRF antagonists alpha-helical CRF 9-41 (alpha hCRF) and [DPhe 12, Nle 21-38] rCRF (12-41) (DPhe CRF) have been studied in unanaesthetized fetal lambs of 125-140 days gestation. 2. CRF when given as a 10 micrograms bolus followed by a 5 micrograms h-1 infusion into a lateral cerebral ventricle caused prolonged continuous fetal breathing movements which were stimulated in both amplitude and frequency but which did not persist during hypoxia. 3. Lower doses of CRF (20 ng bolus followed by 10 ng h-1) increased the amplitude but not the frequency of fetal breathing movements which did not become continuous. 4. At higher doses (20 micrograms bolus followed by 10-15 micrograms h-1) CRF induced cerebral convulsions which were also associated with fetal breathing movements of increased amplitude and frequency. 5. The CRF antagonists alpha hCRF and DPhe CRF both inhibited fetal breathing movements and induced a prolonged apnoea which was resistant to the stimulatory effects of 5-6% hypercapnia. 6. We conclude that CRF stimulates breathing movements in the fetal lamb. The finding that administration of the CRF antagonists alone cause apnoea suggests that CRF may have a tonic role in the regulation of fetal breathing movements.

Objective: Deceleration area (DA) and capacity (DC) of the fetal heart rate can help predict risk of intrapartum fetal compromise. However, their predictive value in higher risk pregnancies is unclear. We investigated whether they can predict the onset of hypotension during brief hypoxaemia repeated at a rate consistent with early labour in fetal sheep with pre‐existing hypoxaemia. Design: Prospective, controlled study. Setting: Laboratory. Sample Chronically instrumented, unanaesthetised near‐term fetal sheep. Methods One‐minute complete umbilical cord occlusions (UCOs) were performed every 5 minutes in fetal sheep with baseline paO2 <17 mmHg (hypoxaemic, n = 8) and >17 mmHg (normoxic, n = 11) for 4 hours or until arterial pressure fell <20 mmHg. Main outcome measures: DA, DC and arterial pressure. Results: Normoxic fetuses showed effective cardiovascular adaptation without hypotension and mild acidaemia (lowest arterial pressure 40.7 ± 2.8 mmHg, pH 7.35 ± 0.03). Hypoxaemic fetuses developed hypotension (lowest arterial pressure 20.8 ± 1.9 mmHg, P < 0.001) and acidaemia (final pH 7.07 ± 0.05). In hypoxaemic fetuses, decelerations showed faster falls in FHR over the first 40 seconds of UCOs but the final deceleration depth was not different to normoxic fetuses. DC was modestly higher in hypoxaemic fetuses during the penultimate (P = 0.04) and final (P = 0.012) 20 minutes of UCOs. DA was not different between groups. Conclusion: Chronically hypoxaemic fetuses had early onset of cardiovascular compromise during labour‐like brief repeated UCOs. DA was unable to identify developing hypotension in this setting, while DC only showed modest differences between groups. These findings highlight that DA and DC thresholds need to be adjusted for antenatal risk factors, potentially limiting their clinical utility. [ABSTRACT FROM AUTHOR]

Activation of the hypothalamic-pituitary-adrenal (HPA) axis is a critical response to perinatal hypoxia. Recent data show that adenosine appears to inhibit baseline levels of fetal cortisol and to restrict the increase in ACTH and cortisol during moderate hypoxia. Because adenosine increases substantially during profound asphyxia, it is possible, but untested, that counterintuitively it might restrict the HPA response to more severe insults. It is unclear which receptors mediate the effects of adenosine on the HPA axis; however, adenosine A1 receptor activation is important for adaptation to hypoxia. We therefore investigated whether adenosine i receptor blockade modulates ACTH and cortisol levels in fetal sheep at 118 to 126 days gestation, randomly allocated to receive an intravenous infusion of either vehicle (vehicle-occlusion, n = 7) or 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, an A1 receptor antagonist, DPCPX-occlusion, n = 7) infused 60 mm before and during 10 min of umbilical cord occlusion, or infusion of DPCPX for 70 mm without occlusion (DPCPX-sham, n = 6). Experiments were terminated after 72 h. Fetal ACTH levels increased significantly (P < 0.01) during occlusion, but not sham occlusion, and returned to baseline values by 60 min after occlusion. In the vehicle-occlusion group, fetal cortisol and cortisone plasma levels increased significantly (P < 0.05) 60 mm after the occlusion and returned to baseline values by 24 h. In contrast, there was a marked increase in both fetal cortisol and cortisone during DPCPX infusion before occlusion to a level greater even than the maximum rise seen after occlusion alone. This increase was sustained after occlusion, with increased cortisol levels compared with occlusion alone up to 72 h. In conclusion, fetal cortisol concentrations are suppressed by adenosine A1 receptor activity, largely though a direct adrenal mechanism. This suppression can be partially overcome by supraphysiological stimuli such as asphyxia. [ABSTRACT FROM AUTHOR]

Early onset cerebral hypoperfusion after birth is highly correlated with neurological injury in premature infants, but the relationship with the evolution of injury remains unclear. We studied changes in cerebral oxygenation, and cytochrome oxidase (CytOx) using near-infrared spectroscopy in preterm fetal sheep (103–104 days of gestation, term is 147 days) during recovery from a profound asphyxial insult ( n= 7) that we have shown produces severe subcortical injury, or sham asphyxia ( n= 7). From 1 h after asphyxia there was a significant secondary fall in carotid blood flow ( P < 0.001), and total cerebral blood volume, as reflected by total haemoglobin ( P < 0.005), which only partially recovered after 72 h. Intracerebral oxygenation (difference between oxygenated and deoxygenated haemoglobin concentrations) fell transiently at 3 and 4 h after asphyxia ( P < 0.01), followed by a substantial increase to well over sham control levels ( P < 0.001). CytOx levels were normal in the first hour after occlusion, was greater than sham control values at 2–3 h ( P < 0.05), but then progressively fell, and became significantly suppressed from 10 h onward ( P < 0.01). In the early hours after reperfusion the fetal EEG was highly suppressed, with a superimposed mixture of fast and slow epileptiform transients; overt seizures developed from 8 ± 0.5 h. These data strongly indicate that severe asphyxia leads to delayed, evolving loss of mitochondrial oxidative metabolism, accompanied by late seizures and relative luxury perfusion. In contrast, the combination of relative cerebral deoxygenation with evolving epileptiform transients in the early recovery phase raises the possibility that these early events accelerate or worsen the subsequent mitochondrial failure. [ABSTRACT FROM AUTHOR]

Secondary cerebral hypoperfusion is common following perinatal hypoxia–ischaemia. However, it remains unclear whether this represents a true failure to provide sufficient oxygen and nutrients to tissues, or whether it is simply a consequence of reduced cerebral metabolic demand. We therefore examined the hypothesis that cerebral oxygenation would be reduced during hypoperfusion after severe asphyxia, and further, that the greater neural injury associated with blockade of the adenosine A1 receptor during the insult would be associated with greater hypoperfusion and deoxygenation. Sixteen near-term fetal sheep received either vehicle or 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) for 1 h, followed by 10 min of severe asphyxia induced by complete occlusion of the umbilical cord. Infusions were discontinued at the end of the occlusion and data were analysed for the following 8 h. A transient, secondary fall in carotid artery blood flow and laser Doppler flow was seen from approximately 1–4 h after occlusion ( P < 0.001), with no significant differences between vehicle and DPCPX. Changes in laser Doppler blood flow were highly correlated with carotid blood flow ( r2= 0.81, P < 0.001). Cortical metabolism was suppressed, reaching a nadir 1 h after occlusion and then resolving. Cortical tissue was significantly increased at 1, 2 and 3 h after occlusion compared to baseline, and inversely correlated with carotid blood flow ( r2= 0.69, P < 0.001). In conclusion, contrary to our initial hypothesis, delayed posthypoxic hypoperfusion was associated with suppression of cerebral metabolism and increased tissue , and was not significantly affected by preceding adenosine A1 blockade. These data suggest that posthypoxic hypoperfusion is actively mediated and reflects suppressed cerebral metabolism. [ABSTRACT FROM AUTHOR]