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Pregnancy diet based on ancestral patterns increases growth in subcortical fetal brain regions.

Evidence on the biological basis for maternal nutrition effects on fetal and newborn neurodevelopment remains limited. This randomized controlled trial in Ecuador tested a maternal dietary pattern-derived from empirical studies of nutrition in human evolution and adapted locally-on offspring growth and brain development. Pregnant women (n = 215) in their first trimester were randomized to: 1) control (n = 104); or 2) Mikhuna ("nourish" in Kichwa) intervention (n = 111). The intervention, from 12 wk gestation to birth, consisted of a weekly food delivery (8 eggs, 500 g fish, and a variety of sustainably sourced fruits and vegetables) and a behavior change communication strategy encouraging diet diversity and limiting highly processed foods. Longitudinal data collection occurred at 12 wk, 21 wk, 35 wk gestation, and 2 wk postpartum, and included ultrasound imaging of fetal bone and brain parameters, maternal dietary intakes, anthropometry, socioeconomic and demographic variables, and other biomarkers. At close of intervention, a significantly higher percentage of women met the minimum dietary diversity threshold in Mikhuna (74.5%) vs. control groups (55.8%) (P = 0.004). Generalized linear regression models showed significant differences in Mikhuna compared to control for: corpus callosum length 0.19 cm (95% CI [0.02, 0.35]), gangliothalamic ovoid height 0.15 cm (95% CI [0.03 to 0.26]), and femur length -0.10 cm (95% CI [-0.19, -0.02]) from 21 wk to 35 wk; and corpus callosum Z 0.56 (95% CI [0.03, 1.09]) and femur length Z -0.21 (95% CI [-0.42, 0.00]) at 35 wk. The Mikhuna intervention increased the growth of subcortical fetal brain structures, which have established roles in motor control, cognition, and signal transmission.

Female

Maturation of the hypothalamo-neurohypophysial system. II. Neurophysin, vasopressin and oxytocin in the median eminence of the developing rat brain.

Transverse sections of the median eminence from fetal and neonatal rats were examined by the immunoperoxidase technique to detect the presence of oxytocin, vasopressin and neurophysin. Neurophysin was observed in the 18-day fetus. Vasopressin and oxytocin were not detected until after birth, on the 4th and 8th days respectively. There was an accumulation of material crossreactive with neurophysin and vasopressin antibodies in the palisade layer of the median eminence between the 4th and 9th days after birth. This distribution of immunoreactive material in the palisade layer was suggestive of neurosecretory substances localized in two fibre tracts on either side of the median eminence. The data are consistent with the accumulation of corticotropin releasing factor and an associated neurophysin in this area. It is suggested that the accumulation of material occurs because of the relative immaturity of the capillary loops that constitute the primary plexus of the hypophysial portal system.

Animals

Maturation of the hypothalamo-neurohypophysial system. I. Localization of neurophysin, oxytocin and vasopressin in the hypothalamus and neural lobe of the developing rat brain.

Sections of the hypothalamus, median eminence and pituitary from fetal and neonatal rats were examined with the immunoperoxidase staining technique and light microscopy. Purified antisera raised against vasopressin and oxytocin, and antisera cross-reactive with rat neurophysin were used to localize these antigens in the hypothalamo-neurohypophysial system (HNS). Neurophysin was detected throughout the HNS of the 18-day fetal rat. Vasopressin was present in the hypothalamus and pituitary of the 19-day fetus, and in the median eminence of the 4-day neonate. Oxytocin was not detected in the pituitary until 1--2 days after birth, in the hypothalamus after 4 days, and in the median eminence after 8 days. During the first days after birth the supraoptic nucleus was more mature than the paraventricular nucleus. The HNS did not approach maturity until at least 7 days after birth. The relative maturity of the supraoptic nucleus compared with the paraventricular nucleus, and the detection of vasopressin before oxytocin are evidence for the one-neuron-one-hormone theory. The data do not exclude the possibility that the fetal hypothalamo-neurohypophysial system, and perhaps the fetal hormone, vasotocin, affect the initiation and course of parturition.

Animals

Isoenzymic forms of NAD-linked glycerol-3-phosphate dehydrogenase from rabbit brain.

Two major enzyme forms of cytosolic NAD-linked glycerol-3-phosphate dehydrogenase in rabbit brain have been purified to apparent homogeneity. One major enzyme form designated I6.5 exhibits an iso-electric point at pH 6.5, and is indistinguishable from the major form I6.5 found in other tissues. The other major form, designated I5.9, has an isolectric point at pH 5.9, and by amino acid analysis is shown to be a true isoenzyme distinct from form I6.5. Form I5.9 appears to be closely related to or identical with the major enzyme characteristic of heart. Neither the brain enzyme form I5.9 nor the major heart isoenzyme are inhibited by antiserum to the muscle enzyme. Because of the high apparent Km for NADH, it is postulated that the brain isoenzyme I5.9 serves to maintain glycolysis when NADH levels rise under relatively anaerobic conditions especially during fetal and neonatal development.

Amino Acids

The derivation and characterization of neuronal cell lines from rat and mouse brain.

This study shows that permanent cell lines can be established from rat and mouse brain by direct tissue culture methodology without the aid of exogenous chemical or viral transforming agents. These cells were derived from specific areas of the brain, such as the cerebellum and hippocampus, at chosen times during fetal and neonatal development. Success in establishing neuronal cell lines was dependent upon the use of selection pressures designed to keep the background of glial cells and fibroblasts at a minimum. These manipulations included care in the choice and processing of the original tissue, utilization of cytotoxic anti-glial sera, and continuous manual isolation of cells with neuronal morphology. Slow-growing nerve cells were thus allowed to adapt spontaneously to culture with a minimum of competition from faster-adapting cell types. Many of these cell lines are judged to be neuronal on the basis of their electrical excitability and their characteristic surface antigens. The cells respond positively in a sodium flux assay which has been shown to correlate well with the ability to generate an action potential, and also express one or more of three antigens previously found to be specific for nerve cells.

Animals

Taurine in development and nutrition.

Taurine is an amino acid that is widely distributed in the fluids and tissues of man. In mammals, taurine is a major end-product of methionine metabolism. Taurine is found in most mammalian tissues but is only present in trace amounts in many plants. During fetal development of the brain in man and other mammals taurine is present in high concentrations and declines to lower, adult concentrations during neonatal life. However, during this time there is a net accumulation of taurine when the amount per brain rather than per gram of tissue is calculated. In man, taurine is apparently an essential nutrient, unlike in other animals which have a much greater capacity to synthesize this compound. The human infant, is particular, needs a dietary supply of taurine to synthesize the bile salt taurocholate. Thus, taurine appears to be an important component of the developing brain and must be supplied to man in the diet.

Animals

Thyrotropin-releasing hormone is not required for thyrotropin secretion in the perinatal rat.

To determine the role of thyrotropin-releasing hormone (TRH) in the regulation of thyroid-stimulating hormone (TSH) secretion in the perinatal period, a physiological approach of neutralizing circulating TRH in the fetal and early neonatal rat was employed. TRH-antiserum (TRH-AS) raised in rabbits and administered daily to low iodine-propylthiouracil (LID-PTU)-fed pregnant rats from days 12 to 19 of gestation markedly impaired the rise in serum TSH to LID-PTU when compared with normal rabbit serum-treated controls. In contrast, fetal serum TSH was unaffected by TRH-AS. The binding capacity of TRH-AS in the fetal serum (111 ng/ml) far exceeded circulating TRH in the fetus. Similarly, acute TRH-AS administration to the pregnant rat fed LID-PTU markedly decreased the serum TSH concentration in the mother, but not in the fetus, 60 min after TRH-AS administration. Chronic TRH-AS administration to neonatal rats whose nursing mothers were fed LID-PTU was in-effective in decreasing the elevated serum TSH in the neonate through day 8 of life, whereas a slight but significant decrease in serum TSH was observed on day 10. Chronic daily TRH-AS administration to neonatal rats through day 10 of life had no effect on the later development of the hypothalamic-pituitary-thyroid axis. These findings suggest that TRH does not participate in TSH regulation during the perinatal life in the rat and that thyroid hormones are probably the main regulators of TSH secretion during this period. Placental TRH is not important in regulating TSH secretion in the fetal rat. Furthermore, TRH "deprivation" during neonatal life does not prevent normal later development of the hypothalamic-pituitary-thyroid axis.

Animals

Biochemical differentiation of mechanically dissociated mammalian brain in aggregating cell culture.

Mouse and rat brain cells were dissociated by a simple mechanical sieving technique and studied in culture for the formation of aggregates and the activities of choline acetyltransferase, acetylcholinesterase, glutamic acid decarboxylase, tyrosine 3-monooxygenase, aromatic L-amino acid decarboxylase, catechol methyltransferase, and monoamine oxidase. Cells from fetal and neonatal tissue formed aggregates but not cells from tissue older than two days after birth. The pattern of development of enzyme activities in these aggregates varied with the age of starting tissue. The highest levels of specific activity for the neuron-specific enzymes were found after 3-4 weeks in culture for aggregates of cells derived from relatively undeveloped brains.

Acetylcholinesterase

Galactocerebrosidase activity in canine globoid leukodystrophy.

In an investigation of canine globoid leukodystrophy, cerebroside beta-galactosidase activities were assayed in 24 brains from neonatal and older dogs and in 90 canine leukocyte pellets from nine samplings. The neonatal brains had significantly less enzyme activity than the brains of older dogs, which indicates a potential complication in making neonatal and fetal enzymatic diagnoses. For leukocytes, heterozygous activities averaged 51 percent and globoid leukodystrophy activities 18 percent of mean enzyme activity of normal leukocytes. Variability of leukocyte enzyme activities among the nine samplings was large, but within each sample, variability per genotypic category was moderate. A statistical model was developed to facilitate enzyme diagnosis in the dog and, by implication, in human globoid leukodystrophy and other sphingolipidoses.

Animals

Effects of maternal ethanol ingestion on amine uptake into synaptosomes of fetal and neonatal rat brain.

Pregnant rats were kept on an ethanol-containing (6.8% v/v) liquid diet either from the 13th or 18th day of gestation. In pups exposed to ethanol from the 13th day of gestation, synaptosomal uptake of 3H-tyramine and its conversion to 3H-octopamine were increased initially and exposure to ethanol from the 18th day of gestation produced increases in synaptosomal uptake and conversion for longer periods. These changes were not observed in adult rats administered ethanol. Brain tyrosin hydroxylase activity in the developing rats was unaltered by continuous exposure to ethanol. Withdrawal from ethanol either at 0, 3 or 5 days of age enhanced the tendancy toward increased synaptosomal uptake and conversion, but the magnitude of change depended upon the time of exposure and the age at which withdrawal was initiated. These results suggest that maternal ethanol ingestion can affect synaptic function in the developing central noradrenergic system, with consequent alterations in neurotransmitter uptake and storage.

Amines

The effects of hydrocortisone on the development of the amine systems in the fetal brain.

Human and animal studies suggest impaired central nervous system (CNS) development due to corticoid use in the perinatal period. In this study, hydrocortisone was given to pregnant rats and the development of the fetal dopamine (DA) and norepinephrine (NE) systems in the CNS was investigated. In the fetal rat brain DA and NE systems develop between days 12 and 17 of gestation. Hydrocortisone (HC), 57 mg/kg/day, or saline (SAL) was given intraperitoneally at day 12 or 15 of gestation. The offspring were studied at days 20 to 21 of gestation and days 12 to 13 in the neonatal period. Brain amine systems were visualized using a modified cryostat glyoxylic acid histofluorescence method, and DA and NE levels were determined in whole brains by means of a radioenzymatic assay. The visualized amine systems were evaluated semiquantitatively for distribution and fluorescence intensity without previous knowledge of the administered drug. The amine systems of the HC and SAL groups showed an equal maturation. In both groups cell bodies were demonstrable in areas A1 to A13 and axon terminals in all examined final regions. The distribution and the fluorescence intensity did not show consistent differences for the HC and SAL brains. The concentrations of DA and NE were similar in the offspring of the SAL- and HC-treated animals. The results indicate that HC given during pregnancy does not influence the proliferation of amine cell bodies or the arrival of axon terminals in the regions where the synapses form.

Animals

Lipofuscin formation in the developing nervous system of squirrel monkeys consequent to maternal dietary protein deficiency during gestation.

The formation of lipofuscin pigment under the extrinsic influence of maternal protein deprivation during most of the gestation period has been demonstrated in the cerebellum and motor cortex of the developing fetuses and neonates in squirrel monkeys. The controls maintained on high protein diets failed to show lipofuscin pigment in appreciable quantities. The dirty yellow to dark brown pigment bodies exist as homogeneous particulates as well as duplex structures, each with a thin, irregular, light brown cortex, studded with one or more darker granules, and a medulla filled with sharp yellow substance. The malnourished neonates, especially in the third and fifth laminae of motor cortex, show loose aggregations of lipofuscin bodies in the apical dendrite and occasionally in the body of the dendritic branches but not in the axon hillock or the body of the axon. Histochemically, the pigment is a complex of lipids, carbohydrates and proteins, showing strong reaction for non-specific esterase and acid phosphatase and mild activity of succinate and lactate dehydrogenase and monoamine oxidase, which puts the pigment observed in the same category as the typical aging pigment described in literature. It is clear that maternal protein deficiency during the critical period of the fetal brain development has an adverse effect on its biochemical maturation. The accumulation of lipofuscin pigment is probably one manifestation of the metabolic changes in the neurons, which may have the impact of hastening the aging process, if protein deprivation is continued into the postnatal period.

Aging

Ontogenetic changes in levels of phosphodiesterase for adenosine 3':5'-monophosphate and glucosine 3':5'-monophosphate in the lung, brain and heart from guinea pigs.

Changes in tissue levels of the low Km phosphodiesterase for adenosine 3':5'-monophosphate (cyclic AMP) and guanosine 3':5'-monophosphate (cyclc GMP) in the lung, liver, heart and brain from developing guinea pigs were studied. It was found that the contents of the soluble (cytosol) phosphodiesterase for both cyclic AMP and cyclic GMP were higher in the lung from the fetus than from the neonate and adult. The ontogenetic changes seen in the liver were qualitatively similar to thos in the lung with respect to cyclic GMP hydrolysis, while a reversed pattern of change was noted in the brain. The level of cyclic AMP phosphodiesterase was highest in the fetal heart. Throughout the fetal stage, the levels of the enzyme for cyclic GMP hydrolysis were higher than those for cyclic AMP in the lung. At or around birth, a reversal in the relative levels of the two enzymes took place; two days after birth, the level of the enzyme for cyclic AMP was 2-3times higher than thos for cyclic GMP. Kinetic analysis showed that phohphodiesterases from extracts of the lung from all developmental stages of guinea pigs had the same Km (2.6 muM) for cyclic AMP and the same Km (6.6 muM) for cyclic GMP. The relative values of V, based on assays using the same amount of enzyme protein, in decreasing order, were fetus greater than neonate greater than adult. The present findings suggest that metabolism of the two cyclic nucleotides may be closely related to developmental processes of the tissues. Moreover, the actions involving cyclic GMP may be more predominent in the fetal lung and adult brain.

3',5'-Cyclic-AMP Phosphodiesterases

Computer interpreted fetal electroencephalogram: sharp wave detection and classification of infants for one year neurological outcome.

The presence of visually discernible sharp waves (SWs) in the fetal electroencephalogram (FEEG) has been found to be associated with abnormal neurological infant outcome, but no method of programmed SW detection for FEEG was available. In order to develop an algorithm for SW detection, the first and second derivatives for visually identified SWs and non-SWs were examined and five random variables chosen for discriminant function analysis (DFA). The resulting equation, incorporated into program logic along with logic for artifact rejection, produced classifications from 85% to 89% consistent with visual identifications, suggesting that the number of SWs/epoch (NSW) corresponds with visually identified SWs. In addition, in 61 cases using a threshold for NSW derived by DFA, computer recognized SWs were found to be significantly related to the overall visual interpretation of the tracings (P less than 0.005). Finally, NSW alone produced correct classification of 65.5% of infants for 1 year neurological outcome. The overall consistency was increased to as high as 80% using additional FEEG and neonatal data. These findings imply that some forms of brain damage are present before birth and can be detected during labor using FEEG.

Brain Diseases

Bovine fetal inoculations with calf rotavirus.

The serological and histopathological responses of bovine fetuses to in utero inoculation with virulent and attenuated strains of the calf rotavirus (reovirus-like agent of neonatal calf diarrhea) are described. Thirteen bovine fetuses, 63 to 190 days of gestation, were inoculated in utero with attenuated (three fetuses) or field strain virus (nine fetuses) or both (one fetus).Serum-neutralizing antibody titers ranging from 1:16 to > 1:256 were detected in six of eight fetuses tested, demonstrating the ability of the bovine fetus to respond immunologically to this agent. The youngest fetus in the series was inoculated at 63 days of gestation and developed a titer of 128 in 64 days. This represents the earliest stage of gestation at which a bovine fetus has been inoculated with a bovine virus and found to produce antibody to it. Serum neutralizing titers in six of the eight dams tested increased significantly following the inoculations of their fetuses in utero. Histological changes associated with viral replication and antigenic stimulation of the lymphoreticular system were observed. Pneumonic lesions consisting of both local and diffuse lymphoreticular proliferation were present in five of the nine fetuses that were alive at slaughter. Gliosis and perivascular cuffing were noted in the brains of two of these fetuses and meningitis was seen in one. No evidence of teratogenic change was found.

Animals

Depression of cytidine 3':5'-cyclic monophosphate phosphodiesterase activity in developing tissues of guinea pigs.

Changes in levels of the newly discovered cytidine 3':5'-cyclic monophosphate (cyclic CMP) phosphodiesterase in some representative tissues (cerebral cortex, kidney, intestine, liver, heart, and lung) of guinea pigs from various developmental stages (fetus, neonate, pup, and adult) were studied and compared with those of cyclic AMP and cyclic GMP phosphodiesterases in the same tissues. It was observed that the tissue levels of cyclic CMP phosphodiesterase were invariably the lowest in every one of the fetal tissues examined, the highest in the corresponding tissues from the pups and adult, with intermediate levels seen in some neonatal tissues. The patterns of the ontogenetic changes in levels of cyclic AMP and cyclic GMP phosphodiesterase activities, however, were variable and tissue specific. These findings suggest that the depressed cyclic CMP phosphodiesterase activity (hence, the elevated cyclic CMP concentration) is perhaps a common factor in developing tissues undergoing rapid cell proliferation. The data also suggest that metabolism of cyclic CMP is perhaps more closely related to cell proliferation than the metabolism of cyclic AMP or cyclic GMP.

Animals

Acute leukemia in pregnancy: transient neonatal myelosuppression after combination chemotherapy in the mother.

An 18-year-old primagravida received combination chemotherapy with vincristine, prednisone, L-asparaginase, cyclophosphamide, daunomycin, 6-mercaptopurine and central nervous system (CNS) prophylaxis with intrathecal methotrexate and whole-brain irradiation for acute lymphoblastic leukemia (ALL) beginning in the 12th week of pregnancy. Therapy resulted in sustained complete remission of the leukemia and delivery of a normally developed female infant whose immediate neonatal course was complicated by transient severe bone marrow hypoplasia. Our experience confirms the reports of others that intensive chemotherapy can be administered in the last two trimesters of pregnancy without serious teratogenic complications. However, we conclude that such therapy may cause significant myelosuppression in the newborn.

Antineoplastic Agents

Experimental congenital infection with cytomegalovirus: a guinea pig model.

An animal model permitting study of congenital infections with cytomegalovirus (CMV) has been developed in guinea pigs. Fifteen Hartley strain guinea pigs in the latter half of pregnancy were inoculated intraperitoneally with 10(5.5) 50% tissue culture infective doses of guinea pig CMV. Forty percent of infected mothers delivered litters containing at least one infected newborn, as defined by a positive culture of lung, spleen, or brain. All tissues were cultured by an explant technique. The three mothers who had no detectable complement-fixing antibody to CMV prior to experimental infection delivered infected litters, whereas three of 12 immune mothers delivered infected litters (P less than 0.01). A low-passage, tissue culture-adapted virus produced neonatal infection as frequently as did salivary gland-passaged virus. No congenital abnormalities were found in any of the seven infected newborns. CMV was isolated from lung, spleen, or brain in the four newborns of nonimmune mothers; CMV was isolated from lung only in the three newborns of immune mothers. These preliminary experiments demonstrate that the guinea pig is a suitable animal for further study of maternal-fetal CMV infections.

Animals