Vincent Kirkbride

Delayed ("secondary") cerebral energy failure after acute hypoxia-ischemia in the newborn piglet: continuous 48-hour studies by phosphorus magnetic resonance spectroscopy

ABSTRACT: Phosphorus (31P) spectra from the brains of severely birth-asphyxiated human infants are commonly normal on the first day of life. Later, cerebral energy failure develops, which carries a serious prognosis. The main purpose of this study was to test the hypothesis that this delayed (“secondary”) energy failure could be reproduced in the newborn piglet after a severe acute reversed cerebral hypoxicischemic insult. Twelve piglets were subjected to temporary occlusion of the common carotid arteries and hypoxemia [mean arterial Po2 3.1 (SD 0.6) kPa]. Mean cerebral phosphocreatine concentration [PCr]/inorganic orthophosphate concentration [Pi] decreased from 1.40 (SD 0.29) to 0.01 (SD 0.02), and nucleotide triphosphate concentration [NTP]/exchangeable phosphate pool concentration [EPP] decreased from 0.19 (SD 0.02) to 0.06 (SD 0.04) (p<0.001 for each decrease). On reperfusion and reoxygenation of the brain, mean [PCr]/[Pi] and [NTP]/[EPP] returned to baseline. Observations continuing for the next 48 h showed that [PCr]/[Pi] again decreased, in spite of normal arterial Po2, mean arterial blood pressure, and blood glucose, to 0.62 (SD 0.61) at 24 h (p<0.01) and 0.49 (SD 0.37) at 48 h (p<0.001). [NTP]/[EPP] also decreased, but to a lesser degree. Intracellular pH remained unchanged. These findings appeared identical with those seen in birth-asphyxiated human infants. No changes in cerebral metabolite concentrations took place in six control piglets. The severity of secondary energy failure, as judged by the lowest [PCr]/[Pi] recorded at 24-48 h, was directly related to the extent of acute energy depletion, obtained as the time integral of reduction in [NTP]/[EPP] (p<0.0001). This animal model of secondary energy failure may prove useful for testing cerebroprotective strategies.

Mild hypothermia after severe transient hypoxia-ischemia ameliorates delayed cerebral energy failure in the newborn piglet

ABSTRACT: Severely birth-asphyxiated human infants develop delayed (“secondary”) cerebral energy failure, which carries a poor prognosis, during the first few days of life. This study tested the hypothesis that mild hypothermia after severe transient cerebral hypoxia-ischemia decreases the severity of delayed energy failure in the newborn piglet. Six piglets underwent temporary occlusion of the common carotid arteries and hypoxemia. Resuscitation was started when cerebral [phosphocreatine (PCr)]/ [inorganic phosphate (Pi)] as determined by phosphorus magnetic resonance spectroscopy had fallen almost to zero and [nucleotide triphosphate (NTP)]/[exchangeable phosphate pool (EPP)] had fallen below about 30% of baseline. Rectal and tympanic temperatures were then reduced to 35°C for 12 h after which normothermia (38.5°C) was resumed. Spectroscopy results over the next 64 h were compared with previously established data from 12 piglets similarly subjected to transient cerebral hypoxia-ischemia, but maintained normothermic, and six sham-operated controls.The mean severity of the primary insult (judged by the time integral of depletion of [NTP]/[EPP]) was similar in the hypothermic and normothermic groups. In the normothermic group, [PCr]/[Pi] and [NTP]/[EPP] recovered after the acute insult and then fell again. Minimum values for these variables observed between 24 and 48 h were significantly higher in the hypothermic group and not significantly different from the control values (p < 0.05, analysis of variance). A large reduction in secondary energy failure relative to the extent of the primary insult was shown and no further fall in either [PCr]/[Pi] or [NTP]/[EPP] took place up to 64 h in the hypothermic piglets. We conclude that mild hypothermia after a severe acute cerebral hypoxicischemic insult ameliorated delayed energy failure.

Proton Magnetic Resonance Spectroscopy of the Brain during Acute Hypoxia-Ischemia and Delayed Cerebral Energy Failure in the Newborn Piglet

Studies of the brains of severely birth-asphyxiated infants using proton(1H) magnetic resonance spectroscopy (MRS) have shown changes indicating a rise in cerebral lactate (Lac) and a fall in N- acetylaspartate (Naa). The aim of this study was to test two hypotheses: 1) that these changes can be reproduced in the newborn piglet after transient reversed cerebral hypoxia-ischemia, and their time course determined; and 2) that changes in Lac peak-area ratios are related to changes in phosphorylation potential as determined by phosphorus(31P) MRS. Eighteen piglets aged <24 h were anesthetized and ventilated. Twelve underwent temporary occlusion of the carotid arteries and hypoxemia, and six served as sham-operated controls. 1H and 31P spectra were acquired alternately, both during the insult and for the next 48 h, using a 7-tesla spectrometer. During hypoxia-ischemia, the median Lac/total creatine (Cr) peak-area ratio rose from a baseline of 0.14 (interquartile range 0.07-0.27), to a maximum of 4.34 (3.33-7.45). After resuscitation, Lac/Cr fell to 0.75 (0.45-1.64) by 2 h, and then increased again to 2.43(1.13-3.08) by 48 h. At all stages after resuscitation Lac/Cr remained significantly above baseline and control values. Naa/Cr was significantly reduced below baseline and control values by 48 h after resuscitation. The increases in the Lac peak-area ratios were concomitant with the falls in the[phosphocreatine (PCr)*]/[inorganic phosphate (Pi)] ratio, during both acute hypoxia-ischemia and delayed energy failure. The maximum Lac/Naa during delayed energy failure correlated strongly with the minimum[nucleotide triphosphate (NTP)]/[exchangeable phosphate pool (EPP)](r = -0.94, p < 0.0001). We conclude that both hypotheses have been confirmed.

Increased apoptosis in the cingulate sulcus of newborn piglets following transient hypoxia-ischaemia is related to the degree of high energy phosphate depletion during the insult

An increase in the number of cells undergoing apoptosis was observed in the cingulate sulcus of newborn piglets 48 h after a global hypoxic-ischaemic insult. Apoptotic death was identified morphologically (by light and electron microscopy) and by DNA fragmentation, detected by in situ end labelling. The number of apoptotic cells was directly related to the degree of high-energy phosphate depletion during hypoxia-ischaemia, measured using continuous 31P magnetic resonance spectroscopy. These results may have implications for the understanding and treatment of perinatal hypoxic-ischaemic brain injury.

Brain-metabolite transverse relaxation times in magnetic resonance spectroscopy increase as adenosine triphosphate depletes during secondary energy failure following acute hypoxia-ischaemia in the newborn piglet.

The adenosine triphosphate (ATP)-dependent sodium/potassium pump extrudes intracellular sodium in exchange for extracellular potassium. Low ATP causes pump dysfunction increasing both intracellular sodium and water thereby enhancing metabolite mobility. This should be detectable by proton magnetic resonance spectroscopy (MRS) as increased metabolite transverse relaxation times (T2s). During secondary cerebral energy failure in the newborn piglet, proton and phosphorus MRS showed large increases in the T2s of choline, creatine, N-acetylaspartate, and lactate that correlated with ATP depletion. These results provide insight into factors affecting metabolite T2s and show that T2s may be useful for studying cellular oedema.

Follow-up by questionnaire?

Recently, proposals have been made to include larger numbers of infants and reduce the cost of obtaining follow-up information pertaining to modern perinatal management. These proposals have been made in response to requests from purchasers and providers of health care as well as the obstetricians and neonatologists actively engaged in delivery of the service. These initiatives are welcome, but care must be taken to provide objective, meaningful data. In addition to standardised recording including by questionnaire, standardised data collection designed to identify relevant impairments must be the primary objective; the nature and extent of disability at particular ages can then be assigned but it is misleading to regard disability as the principle outcome measure.

A comparision of linear-array and mechanical-sector cranial ultrasound scanning techniques to predict neurodevelopmental outcome at 8 years in preterm newborn infants

Two methods of neonatal cranial ultrasound (US) scanning, linear-array and mechanical-sector, were compared for their accuracy in predicting neurodevelopmental outcome in a cohort of 854, of whom 782 (92%) infants, all born less than 33 weeks of gestation and cared for on the Neonatal Intensive Care Unit at University College Hospital, London between 1979 and 1988, were included in the analysis. A total of 205 infants were studied by linear-array and 577 infants by mechanical-sector scan. Ultrasound findings were grouped into three risk categories on the basis of the US diagnosis. Outcome was assessed at 8 years of age. The probability estimates for neurologically disabling and nondisabling impairments, extra education and mean IQ were compared for the two US methods. There was no significant difference between the two methods in the accuracy of prediction of neurodevelopmental outcome.

220 DO PERIVENTRICULAR FLARES (PVF) PREDICT OUTCOME

Interpretation and significance of PVF seen with ultrasound (US) in the brains of newborn infants remain controversial. To find out if PVF predict adverse neurodevelopment, follow up data for PVF without periventricular haemorrhage were analysed from a prospectively scanned cohort (n=725) of very preterm (<33 w) survivors born 1983-89. PVF were defined as non-haemorrhagic echodensities in the periventricular region. Probabilities (p%, 95% CI) were calculated for disabling impairments and for total impairments, with and without disability, identified by neurological and developmental assessments at 1 year of corrected age. Values are given for normal US scans (n=355), PVF without later cyst formation (n=50), PVF with cystic periventricular leucomalacia <n=7, all parieto-occipital) (PVF+PVL) and PVL cysts not preceded by PVF (n=7; 5 frontal, 1 parietal, 1 parieto-occipital), of which 6 were noted aged 1-4 days, and thus were antenatal in origin.We conclude a) only PVF followed by cyst formation caused a significant excess of neurodevelopmental impairments at 1 year b) cysts aquired before birth were predominantly frontal and not associated with neuromotor or sensory impairments.

134 CEREBRAL LACTATE AND N-ACETYLASPARTATE/CHOLINE RATIOS AND DELAYED ENERGY FAILURE FOLLOWING ACUTE HYPOXIA-ISCHAEMIA IN THE NEWBORN PIG

Falling [phosphocreatine(PCr)]/[inorganic phosphate(Pi)] following perinatal asphyxia indicates a bad prognosis, as may increases in lactate(lac)/choline(cho) peak area ratios. The aim of this study was to define the relation between these ratios in the newborn pig. 11 pigs less than 24h old were studied continuously by magnetic resonance spectroscopy (MRS) at 7T for 48h: 7 were given an acute reversed cerebral hypoxic-iscbaemic insult and 4 were sham-operated controls. During the insult [PCr]/[Pi] fell from 1.32(0.21) (mean (SD)) to 0.01(0.01), and lac/cho increased from 0.04(0.01) to 1.62(0.34) (p<0.001 for both). These values returned to 1.12(0.31) and 0.31(0.13) after resuscitation. During subsequent delayed (‘secondary’) energy failure [PCr]/Pi] decreased to 0.27(0.24), and lac/cho rose gradually to 0.73(0.30) (p<0.01 for both). N-acetylaspartate(Naa)/cho fell during the insult from 1.05(0.24) to 0.83(0.16)(p<0.05), did not rise on resuscitation, and fell variably over 48 hours to 0.50(0.22) (p<0.01). All values in control pigs were unaltered. We conclude that the MRS abnormalities previously seen in the asphyxiated human infant can be reproduced in the piglet, and that increased lac/cho and reduced Naa/cho were related to falling [PCr]/[Pi].

Does Neonatal Ultrasound (US) Predict Mri Findings in Adolescence? 129

Hypothesis: in very preterm (VPT) subjects at 14 years a) serious neonatal US lesions persist b) MRI identifies more lesions than US.