Delayed fetal pulmonary maturation in a rabbit model of the diabetic pregnancy.

A rabbit model for the diabetic pregnancy was used to investigate the etiology of delayed pulmonary maturation observed in infants of diabetic mothers. Pregnant rabbit does were made glucose intolerant and insulinopenic by injection of alloxan, a pancreatic beta-cell cytotoxin. At 28 d (term approximately 31 d) fetuses of these animals were hyperglycemic, but were not hyperinsulinemic and did not demonstrate tissue overgrowth. Fetal pulmonary maturity was assessed by measurement of pressure-volume relationships on the fetal lungs. Fetuses of glucose-intolerant does demonstrated less retention of air on deflation. Phospholipid components of pulmonary surfactant were assayed on fluid obtained from lavage of the fetal lungs. Levels of disaturated phosphatidylcholine (per-cent total-lavage phospholipids) were diminished in fetuses of glucose-intolerant does compared to control fetuses (20.5 +/- 4.2 vs. 38.0 +/ 4.3%; P less than 0.01). Lecithin/sphingomyelin ratios were similar in both groups and phosphatidylglycerol was not detected in either group. There was a direct correlation between the percentage of alveolar disaturated phosphatidylcholine and retention of air on lung deflation. These findings suggest that in this model pulmonary instability was a result of diminished alveolar disaturated phosphatidylcholine, and this diminution did not result from fetal hyperinsulinemia.

Serum somatomedin-C concentrations in a rabbit model of diabetic pregnancy.

We have developed and validated a method for measuring immunoreactive somatomedin-C (Sm-C) in serum of rabbits, and have shown that during midgestation (11-26 days; gestation = 31 days) serum Sm-C concentrations are higher in normal pregnant animals than in pregnant diabetic animals. Sm-C concentrations in the serum of 28-day gestation fetuses of diabetic rabbits (3.14 +/- 0.25 U/ml) were significantly higher than in the fetuses of nondiabetic rabbits (2.31 +/- 0.23 U/ml; P less than 0.05). Fetuses from litters of the most severely hyperglycemic diabetic mothers (glucose greater than 250 mg/dl) had higher serum Sm-C (3.66 +/- 0.41 U/ml) than those of mothers who were mildly hyperglycemic (2.71 +/- 0.2 U/ml). Although these differences were not statistically significant, fetuses from the former litters accounted in great part for the difference between the fetuses of diabetic and normal pregnancy. The diabetes-related increment in Sm-C does not appear to be due to insulin, since fetal insulin concentrations were not different between the normal and diabetic litters (normal, 50.0 +/- 3.6 microU/ml versus diabetic, 49.6 +/- 7.6 microU/ml). Despite their elevation in serum Sm-C, fetuses from litters of diabetic rabbits were growth retarded in weight (26.8 +/- 6.9 g and 33.8 +/- 6.9 g, diabetic versus normal pregnancy; P less than 0.05) and in length (7.9 +/- 0.7 cm and 8.6 +/- 0.7 cm, diabetic versus normal pregnancy; P less than 0.025). We speculate that these discrepancies between growth and Sm-C might be secondary to the toxic effects of glucose on embryonic growth and that later in gestation, the excessive energy provided to the fetus might stimulate Sm-C synthesis.

A minicomputer-based perinatal/neonatal telecommunications network.

A minicomputer-based telecommunications network has been developed to link all of the state's nine tertiary neonatal intensive care units in order to provide information on availability of neonatal and maternal beds. The information is updated automatically on computer terminals located in each neonatal intensive care unit every two hours by a complete system poll (350 polls per month). This information has been found to be accurate 95% of the time. Through a computer simulation, this network is shown to provide a significant reduction (P less than .0005) in the number of referral telephone calls, and thus the time, required to locate a bed within the state for obstetric or pediatric patients needing transfer. In addition, this network has provided a data base to aid in local, regional, and statewide planning for perinatal facilities. A 6-month summary of the data indicated that for 16% of the system polls, less than two neonatal intensive care units could accept outside referrals, demonstrating frequent severe statewide restrictions for care of high-risk patients. The total operating budget for the most recent fiscal year was less than +25,000, which is far less than that of manual 24-hour central telephone operator systems providing similar services. This telecommunications network is a cost-effective answer to the common problem of limited regional perinatal resources.

Regulation of gene expression by hypoxia.

The present study was undertaken to determine if gene expression for tyrosine hydroxylase (TH), the rate limiting enzyme in the biosynthesis of catecholamines, is regulated in the carotid body, sympathetic ganglia and adrenal medulla by hypoxia. We found that a reduction in oxygen tension from 21% to 10% caused a substantial increase (200% at 1 hour and 500% at 6 hours exposure) in the concentration of TH mRNA in carotid body type I cells but not in either the sympathetic ganglia or adrenal gland. In addition, we found that hypercapnia, another natural stimulus of carotid body activity, failed to enhance TH mRNA in type I cells. Removal of the sensory and sympathetic innervation of the carotid body failed to prevent the induction of TH mRNA by hypoxia in type I cells. Our results show that TH gene expression is regulated by hypoxia in the carotid body but not in other peripheral catecholamine synthesizing tissue and that the regulatory mechanism is intrinsic to type I cells.

Brain stem extracellular fluid pH and respiratory drive during hypoxia in newborn pigs.

In response to moderate hypoxia many newborn animals are capable of increasing ventilation only transiently. To examine the hypothesis that changes in brain stem extracellular fluid (ECF) pH explain this transient ventilatory response, we measured brain stem ECF pH and respiratory drive during hypoxia in newborn pigs. The animals were anesthetized with alpha-chloralose-urethan, paralyzed, vagotomized, and mechanically ventilated with a servo-controlled ventilator to regulate end-tidal CO2. Hypoxic ventilation for 6 min was achieved by changing inspired gas from 100% to 10-15% O2. Respiration, measured as integrated phrenic nerve activity, showed a range of responses. In 13 trials increased phrenic activity early in the hypoxic period was sustained or further augmented for the duration of the period. In contrast, in eight other trials phrenic activity increased and then declined. Regardless of the respiratory response, ECF pH (measured with a flat-surface electrode) increased slightly (0.009 +/- 0.002 U) during the first 2.5 min of hypoxia and then declined 0.061 +/- 0.017 U by the 6th min. This acidotic shift in ECF pH is inconsistent with the hypothesis that an alkalotic shift causes the nonsustained respiratory response of newborn pigs.

Intracellular recordings of respiratory neurons in the lateral medulla of piglets.

Membrane potentials of respiratory neurons in the ventral respiratory group were recorded using intracellular techniques in the medulla of newborn piglets. Three types of neurons were demonstrated: inspiratory neurons with an augmenting pattern of spike activity during inspiration; postinspiratory neurons with a short decrementing firing pattern that started immediately after inspiration ended; and stage II expiratory neurons with an augmenting spiking pattern that began shortly after inspiratory termination and ended before onset of the next inspiration. When not firing, the membrane potential trajectories of each cell type revealed two complementary patterns of relative inhibition. This latter finding suggests arrival of inhibitory synaptic potentials during these periods. These findings suggest that the respiratory control mechanisms of the newborn piglet are organized in a three-phased manner similar to that of adult cats.

Respiratory neuronal activity during apnea and other breathing patterns induced by laryngeal stimulation.

Respiration cycles through three distinct phases (inspiration, postinspiration, and expiration) each having corresponding medullary cells that are excited during one phase and inhibited during the other two. Laryngeal stimulation is known to induce apnea in newborn animals, but the cellular mechanisms underlying this effect are not known. Intracellular recording of ventral respiratory group neurons was accomplished in intact anesthetized, paralyzed, and mechanically ventilated piglets. Apnea was induced by insufflation of the larynx with ammonia-saturated air, smoke, or water. Laryngeal insufflation induced phrenic nerve apnea, stimulation of postinspiratory neurons, and stable membrane potentials in inspiratory and expiratory cells consistent with postinspiratory inhibition. Usually the membrane potential of each neuronal type cycled through an expiratory level before onset of the first recovery breath. Variants of the apnea response, probably reflecting the aspiration reflex or sniffing, sneezing, coughing, and swallowing, were also observed. These latter patterns showed oscillation between inspiration and postinspiration without an apparent intervening stage II expiratory phase. However, stage II expiratory activity always preceded onset of the first ramp inspiration after such a pattern. These findings suggest that activation of postinspiratory mechanisms causes profound alterations in the respiratory pattern and that stage II expiration importantly modulates recovery of ramp inspiratory activity. The mechanism of this latter effect may be inhibition of early inspiratory neurons with consequent postinhibitory rebound.

Ontogeny of biogenic amines in respiratory nuclei of the rabbit brainstem.

The concentrations of the biogenic amines, norepinephrine, dopamine and serotonin, were determined in 5 respiratory-related brainstem regions by use of a micropunch technique and a sensitive enzymatic isotopic assay. Samples were taken from rabbits at different ages (term fetuses, 3, 7, 14, 21 days and 2-year-old adults). All brain regions contained norepinephrine, dopamine, and serotonin in measurable amounts, but the distribution was not uniform. Norepinephrine and dopamine remained relatively low (less than 5 ng/mg) in the fetus and during the first weeks of life. Norepinephrine achieved its highest levels in the nucleus tractus solitarius and the locus coeruleus of adult animals. Dopamine concentrations did not change significantly in any nuclear groups over the ages tested. Compared with the newborn values, high concentrations of serotonin were seen in all brainstem nuclei in the term fetus. Adult serotonin levels were not achieved during the neonatal period. These data demonstrate pre- and postnatal changes in neurotransmitter levels in brainstem nuclei which regulate respiratory control. The susceptibility of the newborn to abnormalities in respiratory control may be related to low serotonin levels observed during the postnatal period.

Turnover and synthesis of biogenic amines in discrete brainstem nuclei of the rabbit.

By use of a microtechnique and sensitive enzymatic isotopic assays norepinephrine (NE), dopamine (DA) and serotonin (5-HT) turnover rates were measured in 6 discrete brainstem regions. The results from young (3 days) and adult (1 year) rabbits were compared. In the dorsal raphe nucleus (dr) of the younger animals the slopes of disappearance of NE and 5-HT were significantly higher than in the adults. Whereas, in the dr the slope of the decline of DA was found to be significantly lower in younger animals. In the dr turnover rates of NE and 5-HT were higher in the young animals. However, higher turnover rates for DA were seen only in the LC-A6 region. The young animals had significantly lower turnover times for NE and 5-HT in the dr. The nts was the only nuclear group to reach significance for DA, and the young animals had longer turnover times when compared to adults. These results are indicative of the different roles the neurotransmitter systems play in maintaining homeostasis. The delicate balances in these systems in the brains of younger animals may contribute to their increased susceptibility to perturbations.

Effect of hypoxia on brainstem concentration of biogenic amines in postnatal rabbits.

The effects of hypoxia FiO2 = 0.10 on concentration of biogenic amines in specific brainstem nuclear groups were investigated in 3-and 21-day-old rabbits. The rabbit pups were confined to temperature-controlled water-jacketed chambers and exposed to 6h of 21% O2 or to one of 3 combinations of 21% O2 and 10% O2. These 3 combinations were either intermittent hypoxia, or 4 h of normoxia followed by 2 h constant hypoxia, or 2 h of hypoxia followed by recovery for 4 h in normoxia. Radioenzymatic assays were used to determine the concentration of dopamine, norepinephrine and serotonin in the following brainstem nuclei: substantia nigra, locus coeruleus, dorsal raphe and the nucleus reticularis pontis oralis. Compared with control, hypoxia did not affect dopamine levels at either age. The concentration of norepinephrine was inconsistently affected by hypoxia at either age. In contrast, in the 3-day-old rabbits serotonin was consistently reduced in each of the nuclei. In the 21-day-old rabbits, serotonin was either unchanged or increased following hypoxia. Our results show that hypoxia alters the concentration of serotonin in an age-specific manner. This change in serotonin concentration may reflect altered serotonin metabolism and suggests a possible mechanism by which hypoxia disrupts physiologic homeostasis in newborns.

Development of methionine-enkephalin in microdissected areas of the rabbit brain.

Microdissected areas of the rabbit brain were isolated at prenatal day E-29, postnatal days P-3, 7, 14, 21, 2 months and adults. Methionine-enkephalin (ME) was assayed by RIA and ME concentration [ME] was expressed relative to the protein content of the extracted brain tissues. In brain nuclei with important roles in respiratory control [ME] was higher in prenatal and early postnatal life than in adults. In contrast, the prenatal and early postnatal [ME] levels in other nuclei were lower than or equal to adult values. These data suggest an important and changing role for ME in respiratory control throughout development. Early high [ME] levels within brainstem respiratory control nuclei may contribute to the newborn's increased susceptibility to respiratory depression.

Immunocytochemical detection of serotonin content in raphe neurons of newborn and young adult rabbits before and after acute hypoxia.

The present immunocytochemical study demonstrates serotonin (5-HT) depletion in the dorsal raphe nucleus (DRN) of 3- and 21-day-old rabbits following exposure to mild (10% ambient partial pressure of oxygen) and severe hypoxia (5% ambient oxygen). Under the mild hypoxic condition, 5-HT immunoreactivity in cells and fibers of the DRN was decreased in 3-day-old as well as 21-day-old rabbits, as indicated by decreased intensity of the staining compared to age-matched controls. Although this decrease was more pronounced in the younger animals, recovery from mild hypoxia was seen in both age groups. Hypoxic effects were more striking in 3-day-old animals under the severe hypoxic condition, indicating a greater depletion of 5-HT than in the mildly hypoxic condition. However, little additional effect on the older age group was seen. Further, a decreased ability of the 3-day-old rabbits to recover following severe hypoxia suggests that protracted effects on the developing serotonergic system occur following severe hypoxia during the neonatal period. This was demonstrated by the long-lasting decrease in the number of stained cells and fibers of the DRN 4-hr after return to normal conditions (21% O2). We conclude that newborns have a decreased rate of 5-HT synthesis and/or metabolic turnover that results in rapid depletion of intracellular stores and protracted time to recover from a hypoxic challenge. Similar effects could occur in human fetuses, newborns or infants following birth trauma, apnea or other events associated with severe hypoxia.

Expression of messenger RNAs for peptides and tyrosine hydroxylase in primary sensory neurons that innervate arterial baroreceptors and chemoreceptors.

Retrograde fiber tracing and in situ hybridization were used to determine expression of mRNAs for preprotachykinin A (ppTA), calcitonin gene related peptide (CGRP), preproenkephalin A (ENK), neuropeptide tyrosine (NPY) and somatostatin (SOM) as well as tyrosine hydroxylase (TH) in the petrosal ganglia primary sensory neurons which innervate carotid sinus baroreceptors and carotid body chemoreceptors. Perfusion of the carotid sinus with the retrogradely transported dye (Fluoro-Gold) labeled primary sensory neurons in petrosal ganglion. Numerous somata in the petrosal ganglion labeled with dye contained mRNAs for all the above peptides, except SOM. Moreover, TH mRNA was found in a substantial number of retrogradely labeled cells in the petrosal ganglion. This study provides information concerning which of the numerous peptides identified in sensory neurons of petrosal ganglion may be involved in modulation of the arterial baroreceptor and chemoreceptor reflexes.

Interaction of salicylic acid with verrucae assessed by FT-Raman spectroscopy.

FT-Raman spectroscopy has been used to investigate treated verrucae (warts from the sole of the foot) with a local application of a salicylic acid paint. Differences in the molecular structure of the stratum corneum across the verruca sample were observed, and by comparison with normal and hyperkeratotic skin it was concluded that the tissue around the edges of the verrucae was typically hyperkeratotic skin. In the centre of the verruca, the molecular structure of the skin was altered showing evidence of the interaction with salicylic acid. Salicylic acid was not observed in its characteristic dimerised acid structure, but spectroscopic evidence suggested that fission of the intermolecular H-bonding essentially cleaved the dimer. Observed changes in the v(CCO) stretching mode of the carboxyl and hydroxyl groups indicate the inter H-bonds have broken. These spectral changes are believed to be more consistent with salicylic acid bonding within the human papillomavirus-containing verruca tissue rather than simple acid dissociation upon dissolution in water within the tissue. No evidence for the presence of the other paint components, lactic acid and flexible collodion, was found in the verrucae spectra. This Raman approach may help to elucidate the molecular basis for therapeutic agents interacting with diseased skin.

Developmental characteristics in the daily rhythm of norepinephrine concentration within rabbit brainstem regions.

To examine the development of daily variations in norepinephrine levels, norepinephrine concentrations were measured within five distinct brainstem regions in 3-day-old, 21-day-old, and adult rabbits at 6-h intervals throughout the day. Norepinephrine was measured by radioenzymatic assay, and norepinephrine concentration was expressed relative to wet tissue weight. The data suggest that daily variations for norepinephrine concentrations are established by the third day of life. In the brainstem as a whole, there was an early nocturnal peak (2130 hours) for 3-day-old animals in contrast to a late nocturnal peak (0330 hours) for 21-day-old animals. Adult animals showed a late diurnal (1530 hours) peak. These gross daily variations constitute the sum of distinct region-specific patterns in the development of daily variations in norepinephrine concentration. Norepinephrine is involved in cardiorespiratory regulation and in the regulation of sleep/wake cycles. The observed developmental patterns may relate to the maturation and integration of these physiologic processes.

Neonatal pneumopericardium: current management.

Neonatal pneumopericardium (PPC) is a frequently encountered complication of ventilator therapy. However, the appropriate management remains controversial. We describe seven infants who demonstrate the clinical spectrum of PPC. It is apparent that PPC can occur as an asymptomatic finding and may not require invasive therapy. PPC may present also with cardiac tamponade and require immediate diagnosis and therapy. Simple needle pericardiocentesis is appropriate therapy for most cases with tamponade, however a few babies with PPC uncontrolled by needle aspiration required placement of pericardial catheter for continuous drainage of the air. Mortality from PPC with tamponade (86% without therapy) should be much improved with modern management.

Fourier-transform Raman spectra of ivory. III: Identification of mammalian specimens.

The FT-Raman spectra of six mammalian ivories, other than elephant and mammoth, are presented and spectral differences formulated into a protocol for the identification of animal species from the ivory samples. In this study, sperm whale, walrus, wart hog, narwhal, hippopotamus and domestic pig are considered. The results, which are obtained non-destructively from a variety of specimens, suggest that FT-Raman spectroscopy provides a potentially useful method for the identification of mammalian ivory.

Thermally induced molecular disorder in human stratum corneum lipids compared with a model phospholipid system; FT-Raman spectroscopy.

The molecular basis of lipid packing in human stratum corneum and a model phospholipid system has been studied as a function of temperature using Fourier Transform (FT) Raman spectroscopy. Thermally induced molecular rearrangements of the model lipid system, dipalmitoylphosphatidyl choline (DPPC), and stratum corneum were investigated using FT Raman spectroscopy coupled to a heating chamber. Spectra were recorded for a range of sample temperatures and the results for the two systems were compared, producing previously unreported information of the thermal behaviour for the different systems. Discrete thermal events were recorded for both systems by plotting band separation of the lipid v(CH2) symmetric and asymmetric stretching modes against temperature. The main thermal events observed for DPPC included a 'pre-melting' between 37 and 39 degrees C, the main transition observed between 41 and 42 degrees C, a 'post-transition' between 42 and 43 degrees C and three minor transitions at 58-60, 65-70 and 75-80 degrees C. No evidence was found for the pre-transition of DPPC, previously observed at 34-35 degrees C. The main transitions for human stratum corneum were observed at 35-45, 55, 72 and 83 degrees C, measured from lipid CH2 stretching and bending vibrations. The keratin thermal transition at about 100 degrees C exerted little effect on the lipid bands and no characterisable structural changes were reflected in the keratotic bands.