Intracellular generation of free radicals and modifications of detoxifying enzymes in cultured neurons from the developing rat forebrain in response to transient hypoxia.

To address the influence of oxidative stress and defense capacities in the effects of transient hypoxia in the immature brain, the time course of reactive oxygen species generation was monitored by flow cytometry using dihydrorhodamine 123 and 2',7'-dichlorofluorescein-diacetate in cultured neurons issued from the fetal rat forebrain and subjected to hypoxia/reoxygenation (6 h/96 h). Parallel transcriptional and activity changes of superoxide dismutases, glutathione peroxidase and catalase were analyzed, in line with cell outcome. The study confirmed hypoxia-induced delayed apoptotic death, and depicted increased mitochondrial and cytosolic productions of free radicals (+30%) occurring over the 48-h period after the restoration of oxygen supply, with sequential stimulations of superoxide dismutases. Whereas catalase mRNA levels and activity were augmented by cell reoxygenation, glutathione peroxidase activity was transiently repressed (-24%), along with reduced glutathione reductase activity (-27%) and intracellular glutathione depletion (-19%). Coupled with the neuroprotective effects of the glutathione precursor N-acetyl-cysteine (50 microM), these data suggest that hypoxia/reoxygenation-induced production of reactive oxygen species can overwhelm glutathione-dependent antioxidant capacity, and thus may contribute to the resulting neuronal apoptosis.

Neonatal intensive care and birth weight-specific perinatal mortality in Michigan and Lorraine.

This study investigated the factors influencing use of neonatal intensive care and perinatal mortality in regions of the United States and France, two countries with similar health care systems but different approaches to health financing. The study employed birth certificates from Michigan and a birth registry from Lorraine in 1984. The study showed that geographic access and socioeconomic status were important in determining use of neonatal intensive care in both regions. Socioeconomic factors in perinatal mortality were also shown for both regions, after controlling for gestational age, birth weight, and neonatal intensive care use. In Michigan, infants of mothers with low education had higher mortality rates and in Lorraine residents of low income areas had higher mortality rates. A higher proportion of Michigan women delivered in hospitals with neonatal intensive care than in Lorraine, in all weight/gestation categories. Perinatal mortality rates were also lower in Michigan than in Lorraine, overall and within birth weight categories.

Effect of chronic exposure to methylxanthines on diazepam cerebral binding in female rats and their offsprings.

Caffeine, theophylline or saline were injected daily into female rats during the gestation and lactation periods. Crude synaptosomal membranes were isolated from the brains of offsprings at various stages of development and their ability to specifically bind [3H]diazepam was tested. An other approach consisted of injecting [3H]diazepam into offsprings and cerebral specifically bound diazepam was measured. It was shown that methylxanthines were able to inhibit [3H]diazepam binding by reducing total number of binding sites in the brain of 5- and 15-day-old rats born from treated mothers, with a total recovery of control values at 25 days of age. Moreover, in vivo percentage of cerebral bound diazepam dramatically fell when rats were exposed to methylxanthines in utero and through the mother's milk. Since caffeine and theophylline displace diazepam binding not necessarily in a competitive manner, it is suggested that they could interfere with diazepam as adenosine antagonists.

Comparative effects of acute and chronic administration of caffeine on local cerebral glucose utilization in the conscious rat.

The quantitative 2-[14C]deoxyglucose autoradiographic method was used to compare the effects of acute and chronic administration of caffeine on rat brain energy metabolism. The acute intravenous administration of caffeine (10 mg/kg) to naive rats induced widespread increases in glucose utilization in 20 of 62 structures, mainly in striatal and related areas as well as in the 2 raphe nuclei and the locus coeruleus. After 2 weeks' chronic intraperitoneal injection of caffeine (10 mg/kg), increases in glucose utilization were seen in 6 of 62 structures: the substantia nigra, pars compacta, dorsal raphe, locus coeruleus and the 3 parts of the caudate nucleus. An acute caffeine injection (10 mg/kg) to these chronically caffeine-treated rats induced a further increase in glucose utilization in 9 additional structures but there was no significant difference in the effects of an acute administration of caffeine whether the rats had been chronically pretreated with caffeine or saline. The results of the present study show that brain energy metabolism seems to be subject to only partial tolerance to central stimulation by caffeine.

Effects of bilirubin infusion on local cerebral glucose utilization in the immature rat.

The clinical features of kernicterus have been extensively described. However, there are still no data available on a possible correlation between the areas which appear to preferentially accumulate bilirubin and regional changes in cerebral functional activity. Therefore, we applied the quantitative autoradiographic [14C]2-deoxyglucose method to the measurement of local cerebral metabolic rates for glucose (LCMRglc) in immature rats receiving a bilirubin infusion. A loading dose of 160 mg/kg bilirubin in a buffered serum albumin solution was first given to the rats over 15 min. Thereafter, bilirubin was infused at a reduced rate, 64 mg/kg/h. Bilirubin infusion lasted from 2 to 3 h according to the age of the animal, in order to obtain a plasma concentration of bilirubin ranging from 200 to 300 mumol/l over the experimental period. Bilirubin entered the brain without any sign of blood-brain barrier alteration. The [14C]2-deoxyglucose was injected to the animals 45 min before the end of bilirubin infusion. Rats were studied at 3 postnatal ages, 10 (P10), 14 (P14) and 21 days (P21). Hyperbilirubinemia induced widespread decreases in LCMRglc's in all brain areas and at all ages. These decreases were mostly prominent in sensory areas, auditory and visual, as well as in hypothalamic and thalamic regions. Especially at P10, the distribution of LCMRglc's was strikingly heterogeneous in both cerebral cortex and caudate nucleus, appearing as alternate dark and white columns or as alternate dark and light dots, respectively. The data of the present study are in agreement with clinical observations reporting that bilirubin mostly accumulates in the striatum and cranial nerves and that the neurological sequelae of kernicterus are very often hearing loss as well as motor problems.

Regional cerebral metabolic consequences of bilirubin in rat depend upon post-gestational age at the time of hyperbilirubinemia.

While the accumulation of bilirubin in specific brain regions has been well characterized at autopsy in kernicteric infants, data on the regional effects of early cerebral bilirubin intoxication are still missing. Therefore, the quantitative autoradiographic [14C]2-deoxyglucose technique was applied to the measurement of the effects of a bilirubin infusion on local cerebral metabolic rates for glucose (LCMRglc) in immature rats. A loading dose of 80 mg/kg bilirubin was first administered to the animals over 15 min. Thereafter, the velocity of the infusion was reduced to 32 mg/kg/h and the infusion was continued for 105 min. The animals were studied at two ages, postnatal day 10 (P10) and P21. The [14C]2-deoxyglucose was injected to the animals 45 min before the end of the infusion. Bilirubin infusion led to plasma concentrations ranging from 100 to 200 mumol/l at both ages and to brain amounts of 10-16 nmol/g at P10 while bilirubin was not detectable in brain at P21. Hyperbilirubinemia induced widespread decreases in LCMRglcs at P10 and had rather limited consequences on cerebral glucose utilization at P21. At P10, decreases in LCMRglcs were mostly prominent in regions that have been shown to preferentially accumulate bilirubin in kernicteric infants. In conclusion, there appears to be a good correlation between these metabolic data and regional brain permeability to bilirubin.

An experimental model of generalized seizures for the measurement of local cerebral glucose utilization in the immature rat. I. Behavioral characterization and determination of lumped constant.

An experimental model of status epilepticus has been developed in the immature rat by administration of pentylenetetrazol (PTZ) using repetitive, timed intraperitoneal injections of subconvulsive doses. The pattern of behavioral signs has been well characterized in each age group, i.e. 10 (P10), 14 (P14), 17 (P17) and 21 postnatal days (P21). In this model, the dose of convulsant could be adjusted as a function of interindividual sensitivity and status epilepticus lated for quite a long duration to allow the measurement of local cerebral metabolic rates for glucose (LCMRglc) by means of the [14C]2-deoxyglucose method [J. Neurochem., 28 (1977) 897-916]. To estimate LCMRglc during status epilepticus, the lumped constant (LC) was re-calculated in controls and PTZ-treated rats. The control LC was 0.54 at P10 and 0.50-0.51 at the three older ages studied (P14, P17 and P21). During status epilepticus, it increased to 0.64 in P10 rats and decreased to 0.42 and 0.40, respectively, in P17 and P21 animals. At P14, LC was not affected by seizures. The measurements of brain lactate levels showed a large 4.5-10-fold increase in PTZ-treated rats as compared to controls at all ages. The results of the present study show that the immature brain responds to sustained seizure activity in a specific way according to its postnatal age. Moreover, our results underscore the necessity of re-calculation of LC to the quantification of LCMRglc in such pathological states, particularly in immature animals.

An experimental model of generalized seizures for the measurement of local cerebral glucose utilization in the immature rat. II. Mapping of brain metabolism using the quantitative [14C]2-deoxyglucose technique.

The quantitative autoradiographic [14C]2-deoxyglucose technique (2DG) was applied to measure the effects of pentylenetetrazol (PTZ)-induced status epilepticus (SE) on local cerebral metabolic rates for glucose (LCMRglc) in 10 (P10)-, 14 (P14)-, 17 (P17)- and 21 (P21)-day-old rats. To produce long-lasting SE (55 min), the animals received repetitive, timed intraperitoneal injections of subconvulsive doses of PTZ until SE was reached. At P10 and P14, SE induced a marked increase in LCMRglc which affected 66 of the 76 structures studied. Increases were especially high (200-400%) in limbic and motor cortices at P10 and in some brainstem areas at these 2 ages. At P17 and P21, average brain glucose utilization was similar in seizing and control rats, but in PTZ-treated rats reflected a redistribution in local metabolic rates with increases in brainstem, midbrain, hypothalamus and septum, decreases in cortex, hippocampus, some sensory areas and white matter and no change in many motor and limbic structures. In a few cerebral regions, such as hippocampus, dentate gyrus and mammillary body, LCMRglc did not increase at P10 and P14 and decreased at P17 and P21 in PTZ- vs. saline-treated rats. The results of the present study show that the immature brain responds to sustained seizure activity in a specific way according to its maturational state. Moreover, these data allow the mapping of the vulnerability of cerebral structures to seizures, according to their metabolic response to convulsions.

Acute hypoxia induces specific changes in local cerebral glucose utilization at different postnatal ages in the rat.

The quantitative autoradiographic 2-[14C]-deoxyglucose technique (2-DG) was applied to measure the effects of an acute hypoxic exposure on local cerebral metabolic rates for glucose (LCMRglcs) in the 10 (P10)-, 14 (P14)-, and 21 (P21)-day-old rat. The animals were exposed to hypoxic (7% O2/93% N2) or control gas mixture (21% O2/79% N2) for 20 min before the initiation and for the duration of the 2-DG procedure. Lumped constants were not affected by hypoxia at any age. At P10, the exposure to the hypoxic gas mixture induced a generalized increase in LCMRglc which affected 41 structures of the 45 studied. At P14, average cerebral glucose utilization was similar in hypoxic and control rats. LCMRglc increased in 5 areas and decreased in 11 regions, mainly brainstem and respiratory areas in hypoxic rats. Finally, at P21, LCMRglc decreased in 11 structures of hypoxic rats. The increase in LCMRglc in the hypoxic 10-day-old rat likely reflects stimulation of anaerobic glycolysis. Conversely, at P14 and P21, when the brain has become more dependent upon oxygen supply for its energy metabolism, levels of LCMRglc are similar in both groups of animals or decreased in a few structures of hypoxic compared to normoxic rats. The results of the present study show that the immature brain responds to an acute hypoxic insult in a specific way according to its maturational state. They are also in good accordance with the higher resistance of the immature animal to oxygen deprivation.

Mapping of the consequences of bilirubin exposure in the immature rat: local cerebral metabolic rates for glucose during moderate and severe hyperbilirubinemia.

The regional cerebral metabolic consequences of bilirubin intoxication are not well known. With the quantitative autoradiographic [14C]2-deoxyglucose (2DG), we studied the effect of moderate or severe bilirubin infusion on local cerebral metabolic rates for glucose utilization (LCMRglcs) in 10 (P10) and 21 day-old (P21) rats. After an 80 or 160 mg/kg loading dose of bilirubin administered over 15 min, the speed of bilirubin infusion was reduced to 32 or 64 mg/kg/h for the following 105 min, for moderate or severe intoxication, respectively. This infusion protocol led to plasma bilirubin concentrations of 100-200 nmol/ml (moderate intoxication) or 200-300 nmol/ml (severe intoxication). Cerebral bilirubin concentration was 10 nmol/g at P10 and undetectable at P21 in moderate hyperbilirubinemia while it reached 22-34 nmol/g at both ages during severe hyperbilirubinemia. At P10, bilirubin infusion, moderate or severe, induced significant decreases in LCMRglcs in 17 and 15 brain regions of the 24 studied, respectively. At P21, moderate hyperbilirubinemia induced a decrease in LCMRglcs in only 2 regions, auditory cortex and auditory nerve. Conversely, at that age, severe bilirubin intoxication led to significant decreases in LCMRglcs in all regions studied. These results demonstrate that metabolic changes induced by bilirubin are directly correlated to its entry into the brain which occurs without any alteration in the blood-brain barrier. Indeed, the effects of the dye are quite discrete during moderate hyperbilirubinemia at P21 when no bilirubin is detectable in the brain while they are massive during severe hyperbilirubinemia at P21 and at both levels of intoxication at P10 when bilirubin has entered the brain in measurable amounts.

The management of bronchopulmonary dysplasia.

After presentation of the actual knowledge concerning the pathophysiology of bronchopulmonary dysplasia, the prevention and the management of the disease are discussed. Techniques of ventilation, weaning procedures and prescription of drugs are also analyzed. The importance of a slow reduction of oxygen supply and the possibility of discharge with oxygen therapy is emphasized.

Neonatal apnea and apneic syndromes.

The physiologic factors that predispose premature infants to apnea are reviewed in this article. Management and treatment of "idiopathic apnea" are discussed. "Symptomatic apnea" should be treated according to its primary cause.

Detection of immune complexes in the sera of human newborns suspected of neonatal infection.

The special characteristics and relative immaturity of the immune system in human neonates favour the constitution and persistence of circulating immune complexes after any antigenic invasion so that their detection in serum during the first week of life could be of paticular interest for the early diagnosis of a neonatal infection. In this study, using a technique based on the inhibition of a latex agglutination, we detected circulating immune complexes in sixty-four neonates suspected of infection, at a significantly higher titre than in eleven newborns considered to be completely devoid of any clinical abnormalities. The level of those circulating immune complexes was related to the severity of a clinically considered as a high risk of infection. On the other hand, no significant correlation was found between the infection score and the level of fibrinogen in the blood or the percentage of nonsegmented neutrophils. Moreover, no correlation was demonstrated between each of these two classical biological tests and the level of circulating immune complexes.

Neonatal necrotizing enterocolitis: a retrospective and multicentric review of 331 cases.

The authors retrospectively review 331 cases of necrotizing enterocolitis from 13 different departments of pediatric surgery; 184 cases were treated only medically at the acute stage (47 of whom developed intestinal stricture) and 147 cases were operated on at the acute stage. The different procedures of acute surgical intervention are reported and the results of two surgical procedures are compared. The first is a classical one comprising a laparotomy with exploration of the intestinal tract and resection of the necrotic segments followed by enterostomy above the resected area. The other procedure comprises a minimal laparotomy in the right lower quadrant with ileostomy above necrotic areas, without resection of necrotic segments associated with peritoneal drainage. The results are assessed using the postoperative mortality rate and the number of secondary intestinal strictures. Mortality during the first postoperative month occurred in 27.9% of cases, and intestinal strictures were noted in 31.3% of cases after acute surgical procedure.

[Physiopathology of neonatal hyperbilirubinemia]. / Physiopathologie de l'hyperbilirubinémie néonatale.

The most important steps of bilirubin metabolism involved in the pathophysiology of neonatal hyperbilirubinemia are: 1) hemoglobin degradation by heme oxygenase; 2) bilirubin binding to serum albumin; 3) bilirubin conjugation to acid glucoronic by glucoronyl transferase. Progress in the knowledge of these metabolic steps allows to understanding of why massive hemolysis, infections, hypoxia and prematurity increase the risk of kernicterus and therefore justify adapted preventive and therapeutic measures.

[The toxicity of bilirubin to the central nervous system]. / La toxicité de la bilirubine sur le système nerveux central.

Whatever the causes of its accumulation, excessive production and/or insufficient elimination, unconjugated bilirubin when it reaches a certain concentration threshold, is responsible for neurones and astrocytes death. In this paper the mechanisms involved in this process of cellular death, from hemolysis to oxydation in neurons and neuroglial cells, are reviewed.

[Efficacy and limits of rescue high-frequency oscillatory ventilation in the treatment of hyaline membrane disease in preterm newborns]. / Efficacité et limites de la ventilation par oscillations à haute fréquence après échec de la ventilation conventionnelle chez des prématurés atteints de maladie des membranes hyalines sévère.

UNLABELLED: Conflicting reports of high-frequency oscillatory ventilation (HFOV) use as an alternative to conventional mechanical ventilation have been published. This retrospective study has evaluated the efficacy and safety of rescue HFOV in preterm infants with severe hyaline membrane disease (HMD) after the failure of conventional mechanical ventilation (CMV). POPULATION AND METHODS: All newborns hospitalized in our neonatal intensive care unit (NICU) from 10.1.1993 to 15.4.1995 with CMV failure, defined as the need for more than 55% FiO2 without any improvement for at least six hours, have been retrospectively studied. The infants were shared according to the absence (Gr I) or the presence (Gr II) of persistent pulmonary hypertension of neonate (PPHN) in addition to HMD before HFOV. RESULTS: Gestational age (GA) was 29.2 +/- 3.7 weeks (mean +/- SD) in Gr I and 30.3 +/- 2.8 in Gr II. Birth weight was 1379 +/- 750 g and 1471 +/- 612 g, respectively. As soon as three hours after the onset of HFOV in both groups, a dramatic improvement was observed with a FiO2 drop from 82 +/- 20% to 64.8 +/- 25.5% (P < 0.01). Among the infants, 62% survived without any major disability and 28% died (46% in Gr II vs 12% in Gr I, P < 0.01). A trend towards a worsening of pre-existing brain lesions has been noticed. An increased risk of mortality was observed when a secondary worsening in O2 requirements occurred 24 hours after the onset of HFOV, despite an initial significant improvement. SGA was also associated with a poor prognosis (46% of the deaths vs 29% for AGA infants, P < 0.05). CONCLUSION: HFOV has been successfully used in premature infants with severe respiratory disease and failure of CMV. Criteria of poor prognosis were PPHN and SGA, or a secondary worsening in oxygen requirements after initial improvement. A trend towards aggravation of pre-existing brain lesions has been noticed after HFOV. This aggravation is more frequent when PPHN is associated with HMD. This observation suggests caution for HFOV use when these conditions are present in premature infants.

[Handicaps in the perinatal period. I. Perinatal pathology and difficulties in school]. / Handicaps et périnatalité. I. Pathologie périnatale et difficultés scolaires.

BACKGROUND: Major handicaps are closely related to perinatal events. However, relationship of these events with other moderate disabilities such as school problems are still unclear. POPULATION AND METHODS: A representative sample of 1,408 children at school age and born in Lorraine in 1984 was studied. Two hundred and fifty children with problems were compared with 602 controls without school problems, using a multivariate analysis. RESULTS: The incidence of school difficulties among 6-year old children (in the last year of nursery school) is 17.8%. Children are at higher risk of school problems if their parents are not (odds ratio [OR] = 7.9) or are poor school graduate (OR = 2.7), if they are boys (OR = 2.0), if they are born at the end of the year (OR = 1.1) and also if they are preterm (OR = 2.7) or small for gestational age (OR = 2.5). Preterm delivery and intra-uterine growth retardation accounted for 9.6% of school difficulties. CONCLUSION: The relationship between perinatal events and school difficulties warrants to continue with prevention during pregnancy, especially among groups with multiple risk factors.

[Handicaps and the perinatal period. II. Perinatal pathology and severe deficiencies]. / Handicaps et périnatalité. II. Pathologie périnatale et déficiences graves.

BACKGROUND: In the last two decades, the infant mortality rate has dramatically declined. But improved management of newborns may induce an increased prevalence of neurodevelopmental handicaps. The aim of this paper is to estimate the rate of major disabilities and their relationships to perinatal events. POPULATION AND METHODS: Three hundred and sixteen children born in 1984 and registered by the "Commission Départementale de l'Education Spéciale" (CDES) were included in the study. Among these, 97 had either cerebral palsy, blindness, deafness, or mental retardation. These 97 children were compared to 602 school age controls using a multivariate analysis (logistic regression). RESULTS: The rate of major disabilities among the 6-year old children is 3.4/1000. It was 5/1000 in 1972 and 4.3/1000 in 1976. This frequency is higher in the children who were preterm (odds ratio--OR = 4.8), small for gestational age (OR = 3.3) or suffered from perinatal asphyxia (OR = 32.8). These three factors accounted for 37.4% of major disabilities. CONCLUSION: This study emphasizes the relationships between perinatal events and some major handicaps but also shows that antenatal factors may be involved in neurodevelopmental problems.

[Influence of the transfer mode on short-term outcome in neonates with high perinatal risk]. / Influence du mode de transfert sur le devenir à court terme des enfants à haut risque périnatal.

BACKGROUND: Transferring in utero children with high perinatal risk has been widely recommended in France over the last few years. The purpose of this study is to describe the different transfer modes for children less than 32 weeks GA or less than 1,500 g birthweight and analyse their impact on death and severe neurological lesion. POPULATION AND METHODS: This retrospective study concerned live births in a definite geographic area (Lorraine, France). Four hundred and twenty-seven children born alive between 1989 and 1992 and hospitalized in eight neonatology units were recruited. Multivariate analysis (logistic regressions) were performed to assess the influence of transfer mode on death or severe neurological lesion. RESULTS: Sixty-two percent of the children were born in the level 3 maternity, 19% in a level 1 maternity and 19% in a level 2 maternity. One hundred and twenty-one children (28%) were transferred in utero and 116 (27%) were transferred extra muros. Thirty children died during the hospital stay. Multivariate analysis does show that neonatal extra muros transfer plays a significative role on neonatal death after adjustment for other risks factors (OR = 3.3 (1.1-9.91). Thirty-one children presented a severe neurological lesion. In comparison with neonates born in the level 3 maternity without transfer, extra muros transfer appears to be a very significant risk factor (P = 0.0008): OR for transfer from level 2 and level 1 maternity is 15.8 (3.8-66.5), and 5.6 (1.3-24) respectively. There is no significant increased risk for children born after maternal transfer or born in a level 2 maternity without transfer. CONCLUSION: These data are consistent with data from the literature and confirm the risk related to extra muros transfer in premature babies less than 32 weeks GA or less than 1,500 g birthweight.