Developmental programming of cardiovascular dysfunction by prenatal hypoxia and oxidative stress

Fetal hypoxia is a common complication of pregnancy. It has been shown to programme cardiac and endothelial dysfunction in the offspring in adult life. However, the mechanisms via which this occurs remain elusive, precluding the identification of potential therapy. Using an integrative approach at the isolated organ, cellular and molecular levels, we tested the hypothesis that oxidative stress in the fetal heart and vasculature underlies the molecular basis via which prenatal hypoxia programmes cardiovascular dysfunction in later life. In a longitudinal study, the effects of maternal treatment of hypoxic (13% O(2)) pregnancy with an antioxidant on the cardiovascular system of the offspring at the end of gestation and at adulthood were studied. On day 6 of pregnancy, rats (n = 20 per group) were exposed to normoxia or hypoxia ± vitamin C. At gestational day 20, tissues were collected from 1 male fetus per litter per group (n = 10). The remaining 10 litters per group were allowed to deliver. At 4 months, tissues from 1 male adult offspring per litter per group were either perfusion fixed, frozen, or dissected for isolated organ preparations. In the fetus, hypoxic pregnancy promoted aortic thickening with enhanced nitrotyrosine staining and an increase in cardiac HSP70 expression. By adulthood, offspring of hypoxic pregnancy had markedly impaired NO-dependent relaxation in femoral resistance arteries, and increased myocardial contractility with sympathetic dominance. Maternal vitamin C prevented these effects in fetal and adult offspring of hypoxic pregnancy. The data offer insight to mechanism and thereby possible targets for intervention against developmental origins of cardiac and peripheral vascular dysfunction in offspring of risky pregnancy.     

Maternal zinc deficiency impairs brain nestin expression in prenatal and postnatal mice

INTRODUCTIONZinc is essential for normal brain development,evidenced by the fact that zinc deficiency in lactating mothers is characterized by a high incidence ofneuroanatomical maiformatinns and functional abnormalities in suckling offspring[1-3]. By colltrast,relatively little is known about the relationship be{tween maternal zinc nutrition and fetal brain development[2, 4, 5]. Dvergsten et al[6-81 investigated theeffects of maternal zinc deficiency on postnatal development of the rat ce…     

Developmental programming: impact of prenatal testosterone excess on pre- and postnatal gonadotropin regulation in sheep

The goal of this study was to explore mechanisms that mediate hypersecretion of LH and progressive loss of cyclicity in female sheep exposed during fetal life to excess testosterone. Our working hypothesis was that prenatal testosterone excess, by its androgenic action, amplifies GnRH-induced LH (but not FSH) secretion and, thus, hypersecretion of LH in adulthood, and that this results from altered developmental gene expression of GnRH and estradiol (E2) receptors, gonadotropin subunits, and paracrine factors that differentially regulate LH and FSH synthesis. We observed that, relative to controls, females exposed during fetal life to excess testosterone, as well as the nor-aromatizable androgen dihydrotestosterone, exhibited enhanced LH but not FSH responses to intermittent delivery of GnRH boluses under conditions in which endogenous LH (GnRH) pulses were suppressed. Luteinizing hormone hypersecretion was more evident in adults than in prepubertal females, and it was associated with development of acyclicity. Measurement of pituitary mRNA concentrations revealed that prenatal testosterone excess induced developmental changes in gene expression of pituitary GnRH and E2 receptors and paracrine modulators of LH and FSH synthesis in a manner consistent with subsequent amplification of LH release. Together, this series of studies suggests that prenatal testosterone excess, by its androgenic action, amplifies GnRH-induced LH response, leading to LH hypersecretion and acyclicity in adulthood, and that this programming involves developmental changes in expression of pituitary genes involved in LH and FSH release.     

Adaptation of male and female rats to iodine deficiency

The response of the hypothalamic pituitary axis to chronic iodine deficiency was compared in male and female Sprague-Dawley rats. The animals were kept on a low iodine diet for 12 weeks. Blood samples as well as thyroid and pituitary weights were obtained every two weeks. Baseline values of thyroid weight and serum thyroxine (T4) were similar in both sexes. However, females had lower serum TSH and higher serum triiodothyronine (T3), pituitary weight and pituitary TSH content. After initiation of the low iodine diet, both sexes showed similar decreases in serum T4 and similar increases of serum TSH and thyroid weight. Serum T3, pituitary weight and TSH content remained higher in females throughout the study. Pituitary TSH was directly correlated with serum TSH in both sexes. When adjusted for pituitary TSH and analyzed by a stepwise regression analysis, serum TSH was lower in females suggesting a difference in TSH secretion between males and females. Our studies demonstrate significant sex differences in the regulation of TSH secretion and maintenance of serum T3 level in response to a chronic stimulus.     

Effect of prenatal stress upon parameters of the behavioral response induced by a tonic pain focus in male and female rats during postnatal ontogenesis

The long-term effects of prenatal stress (immobilization of females during the last week of pregnancy) were studied on pain sensitivity to a prolonged irritant in Wistar rats during prepubertal, pubertal periods of development and in adults. Pain sensitivity was evaluated from indices of the biphasic behavioral response in the classical formalin test--the intensity of patterns of flexes, shakes and time spent licking, and from duration of the first, acute and the second, tonic phases and from the duration of interphase. Consequences of the prenatal stress manifested themselves differently in the patterns of the response organized at the spinal and supraspinal levels, during tonic phase mainly, differently in females and males; there were peculiarities of effects of prenatal stress in each age period. The data obtained suggest that the consequences of the prenatal stress at the spinal level manifest themselves in activation of modulating descending facilitating and in suppression of inhibitory monoaminergic systems in prepubertal and pubertal rats, and on the contrary, in activation of descending inhibitory and suppression of facilitating systems in adults. Furthermore, considerable evidence is obtained corroborating the idea of inhibitory nature of the interphase and the mechanisms of modulation of acute and tonic phases in the formalin test during different age periods of the individual development.     

Prostaglandin synthetase activity in vascular tissue of male and female rats

1. Isolated perfused lungs from mature male rats show greater conversion of 14C Arachidonic Acid to cyclo-oxygenase products than females. 2. Following unlabelled arachidonate infusion, the male lungs release more 6-K-PGF1 alpha and TxB2 than females. 3. Aortic rings from male rats release more PGI2-like material and 6-K-PGF1 alpha than the females. 4. These data indicate an elevated PG synthetase activity in male rats as compared with females.     

LHRH and LH in peripubertal female rats following prenatal and/or postnatal ethanol exposure

The effects of pre- and postnatal exposure to ethanol (ETOH) on LHRH and LH were investigated. Pregnant and/or lactating dams were fed ETOH during: 1) gestation, 2) lactation, or 3) gestation-lactation. Female offspring were decapitated at 30 or 40 days-of-age; trunk blood was collected for plasma LH RIA; and hypothalamic tissues were collected for LHRH RIA. Hypothalamic LHRH content of all ETOH-exposed groups was less than that of non-ETOH-fed controls at 30 and 40 days-of-age (p less than 0.05). Plasma LH concentrations of all ETOH-exposed groups were less than those of non-ETOH-fed controls at 30 and 40 days-of-age (p less than 0.05). Also, at 30 and 40 days-of-age, the plasma LH concentrations of the animals exposed to ETOH during lactation and gestation-lactation were less than those of the animals exposed to ETOH during gestation (p less than 0.05). These data suggest that ETOH exposure during gestation and/or lactation negatively affects hypothalamic LHRH content of female rat offspring. Decreased hypothalamic LHRH content with corresponding lowered plasma LH concentration suggests that ETOH influences development or maturation of hypothalamic LHRH neurons by possibly decreasing their number or synthesizing capability.     

Developmental changes in organic osmolytes in prenatal and postnatal rat tissues

At high osmotic pressures, mammalian kidney medulla, heart, lens, and brain utilize organic osmolytes to regulate cell volume. However the types and proportions of these solutes vary among tissues in patterns and for non-osmotic roles not fully elucidated. To clarify these, we analyzed osmolyte-type solute contents in rat tissues at 7 and 2 days prenatal and at 0, 7, 14, 21 (weaning), 35 (juvenile) and 77 (adult) days postnatal. Placentas were dominated by betaine, taurine, and creatine, which decreased between the prenatal times. Fetuses were dominated by glutamate and taurine, which increased between the times. In cerebrum, hindbrain and diencephalon, taurine dominated at early stages, but dropped after postnatal day 7, while myo-inositol, glutamine, creatine and glutamate increased after birth, with the latter two dominating in adults. In olfactory bulb, taurine content declined gradually with age and was equal to glutamate in adults. In all brain regions, glycerophosphorylcholine (GPC) reached a peak in juveniles. In postnatal renal medulla, urea, sodium, GPC, betaine, and taurine increased sharply at day 21. Thereafter, most increased, but taurine decreased. In heart, taurine dominated, and increased with age along with creatine and glutamine, while glutamate decreased after postnatal day 7. In lens, taurine dominated and declined in adults. These patterns are discussed in light of hypotheses on non-osmotic and pathological roles of these solutes.     

Sex difference and postnatal change of maternal behavioral patterns in juvenile male and female rats

Juvenile rats are known to show certain elements of maternal behavior. In this experiment, to investigate sex difference and postnatal change of retrieving and pup-cleaning (licking) behaviors in juvenile rats, these behaviors were recorded using new observation method at 20, 30 and 45 days of age in female and male Wistar rats. At 20 days of age, maternal behavior was observed in a common plastic observation cage (test A) and then test B was performed. In the test B, observation was carried out using a cage with a wooden box that was open on one side, helping the juveniles to establish a nest. As the results of day 20, most rats in all groups showed licking behavior in both the test A and B. The incidence of retrieving behavior increased from the test A to the test B with the box in both sexes, especially in males (p<0.01). The box is thought to play a facilitative role in induction of retrieving. Moreover, the incidence in males was higher than that in females in the test B (p<0.001). At 30 and 45 days of age, only a test B with box was performed. The incidences of licking and retrieving behaviors at 30 days of age were decreased significantly compared to those at 20 days of age in both sexes(p<0.001). Further decrease from 30 days to 45 days was observed. These results suggest that in juvenile rat, incidence of retrieving behavior in males is higher than that in females but there is no sex difference in incidence of licking behavior. Potency to show these behaviors decreases acutely before puberty in rats.     

Developmental programming in skeletal muscle in response to overnourishment in the immediate postnatal life in rats

Overnourishment during the suckling period [small litter (SL)] results in the development of adult-onset obesity. To investigate the mechanisms that underlie the development of insulin resistance in the skeletal muscle of young and adult female SL rats, the litter size was reduced to 3 female pups/dam (SL) while the control litter had 12 pups/dam from the postnatal Day 3 until Day 21. Protein content, mRNA expression and methylation status of the promoter region of key components in the insulin signaling pathway were determined in the skeletal muscle of SL rats. Overnutrition during the suckling period resulted in increased body weight gains, hyperphagia and adult-onset obesity as well as increased levels of serum insulin, glucose and leptin in SL rats. No differences in the expression of total protein as well as tyrosine phosphorylation of insulin receptor β and glucose transporter 4 (Glut4) were observed in skeletal muscle between two groups at both ages. A significant decrease of total insulin receptor substrate 1 (IRS-1) and an increase in serine phosphorylation of IRS-1 were observed in skeletal muscle from adult SL rats. Hypermethylation of specific cytidyl-3',5'phospho-guanylyl (CpG) dinucleotides in the proximal promoter region was observed for the Irs1 and Glut4 genes, which correlated with the reduction in Irs1 and Glut4 mRNA levels in skeletal muscle of adult SL rats. Our results suggest that epigenetic modifications of the key genes involved in the insulin signaling pathway in skeletal muscle could result in the development of insulin resistance in SL female rats.     

The effect of late prenatal and/or early postnatal zinc deficiency on the development and some biochemical aspects of the cerebellum and hippocampus in rats

In rats, late prenatal and/or early postnatal zinc deficiency results in behavioural anomalies in adult animals, but not in overt dysmorphogenesis of the central nervous system. Cerebellar and hippocampal development occurs mainly in the first three weeks postnatally and zinc accumulates specifically in the mossy fibres of the hippocampus during this period.In the present investigation, rat pups were suckled by dams fed a zinc-deficient (<0.5 mg/kg) diet either from day 19 of pregnancy or from parturition. Control animals were restricted-fed the same diet supplemented with 100 mg zinc/kg. Studies were performed on pups either on day 18 postpartum in the case of animals fed the experimental diets from parturition, or on day 20 for pups which received treatment from day 19 of gestation.Cerebellar and hippocampal weights were lower in pups suckling from zinc-deficient dams but zinc levels were not affected in either organ, although histological evidence suggested less zinc in the hippocampal mossy fibres. Incorporation of H-thymidine into cerebellar and hippocampal DNA was not affected by maternal zinc status, nor was the activity of the zinc metalloenzyme alkaline phosphatase.The activity of the myelin-marker enzyme 2′, 3′-cyclic nucleotide 3′-phosphohydrolase was substantially lower in both regions of the brain in zinc deprived pups, especially in the hippocampus. Activity of the zinc metalloenzyme L-glutamic acid dehydrogenase was also diminished in both tissues from 20-day-old pups and in the hippocampi of 18-day-old animals.The data suggest that cerebellar and hippocampal DNA synthesis is not seriously affected by late prenatal and/or early postnatal zinc depletion, but that the activities of two enzymes associated with neural function are. The possibility is raised that these defects may be associated with the behavioural changes observed in rats subjected to zinc impoverishment during the period of maximal cerebellar and the hippocampal development.     

Early-in-life dietary zinc deficiency and supplementation and mammary tumor development in adulthood female rats

Zinc deficiency during pregnancy and postnatal life can adversely increase risk of developing human diseases at adulthood. The present study was designed to evaluate whether dietary zinc deficiency or supplementation during the pregnancy, lactation and juvenile stages interferes in the development of mammary tumors induced by 7,12-dimethylbenzanthracene (DMBA) in female Sprague–Dawley (SD) rats. Pregnant female SD rats were allocated into three groups: zinc-adequate diet (ZnA - 35-mg/kg chow), zinc-deficient diet (ZnD - 3-mg/kg chow) or zinc-supplemented diet (ZnS - 180-mg/kg chow) during gestational day 10 (GD 10) until the litters' weaning. Female offspring received the same diets as their dams until postnatal day (PND) 51. At PND 51, the animals received a single dose of DMBA (50 mg/kg, ig) and zinc-adequate diets. At PND 180, female were euthanized, and tumor samples were processed for histological evaluation and gene expression microarray analysis. The ZnD induced a significant reduction in female offspring body weight evolution and in mammary gland development. At late in life, the ZnD or ZnS did not alter the latency, incidence, multiplicity, volume or histological types of mammary tumors in relation to the ZnA group. However, the total tumor number in ZnS group was higher than in ZnA group, accompanied by distinct expression of 4 genes up- and 15 genes down-regulated. The present findings indicate that early-in-life dietary zinc supplementation, differently to zinc deficiency, has a potential to modify the susceptibility to the development of mammary tumors induced by DMBA.     

Effects of prenatal stress on lever-press acquisition with delayed reinforcement in male and female rats

The prenatally stressed (PS) rat shows enhanced conditioned fear and increased behavioral inhibition in response to footshock compared to control (CON) rats. It is unclear whether this facilitated learning will occur only with aversive stimulation, or if it will also be observed in the context of positive reinforcement. There are limited and inconsistent data regarding sex differences and the impact of prenatal stress on learning. The present study was designed to examine lever-press acquisition with a 10-s delay to food reinforcement in male and female PS and CON rats. Overall, twice as many PS male rats acquired the lever-press response than the PS female rats, CON male rats, and CON female rats. PS male rats also earned significantly more reinforcers and responded on the operative lever at a significantly greater rate than the other three rat groups. These findings suggest that PS rats exhibit altered learning with a task involving positive reinforcement, and this effect of PS is sex specific for male rats.     

The postnatal development of the genital system in male rats following the prenatal administration of corticosterone

Corticosterone administration to pregnant Wistar rats on days 16 and 18 of pregnancy leads to changes in genital system of male offspring during postnatal ontogenesis: reduction of ano-genital distance in two days old rats, increase of preputial glands' weight in 35 and 70 day old embryos, changes in nature of puberal increase in testosterone blood level from day 35 to day 70 of life. The obtained data suggest that the increase in the corticosteroid level in blood of pregnant females owing to any stress factor can affect the postnatal development of genital system of male offspring.     

Effect of prenatal stress and gonadal hormone condition on depressive behaviors of female and male rats

Whether prenatal stress (PNS) and gonadal hormones may influence depressive behavior of rats in the forced swim test was investigated. In Experiment I, adult diestrous female rats had increased immobility, which is indicative of depression, but did not show any significant difference in the duration of struggling compared to intact adult males. In Experiment 2, the behavior of adult intact, castrated, or castrated dihydrotestosterone (DHT)- or estrogen (E2)-replaced offspring of dams that were restrained under lights for 45 min on gestational day 18 (PNS) or were not subjected to gestational stress (non-PNS, control condition) were compared. There were no effects of PNS, but DHT and E2 produced anti-depressant effects on behavior of male rats. Castration decreased struggling and increased immobility compared to intact rats. DHT or E2 replacement was able to partially reinstate struggling and immobility behavior but not to levels of intact males. In Experiment 3, behavior of PNS or control rats that were in proestrus or were ovariectomized and DHT, E2, or vehicle-replaced were compared. Ovariectomy decreased struggling and increased immobility compared to that of proestrous rats. E2 or DHT to control females increased anti-depressant struggling behavior compared to ovariectomized control or PNS rats administered vehicle, which demonstrated greater duration of struggling than did E2-primed, PNS rats. E2 or DHT administration decreased immobility of PNS and control females. These findings suggest that E2 and DHT have some anti-depressant effects but that modest PNS may alter E2's ability to alleviate some depressive behavior in female, but not male rats.     

Thermogenesis in newborn rats after prenatal or postnatal hypoxia

Oxygenconsumption (O₂)was measured in normoxia as ambient temperature(T_a) was lowered from 40 to15°C, at the rate of 0.5°C/min (thermoneutrality ~33°C).In 2-day-old rats born in hypoxia after hypoxic gestation, theT_a-O₂relationship was as in controls; their interscapular brown adiposetissue (IBAT) was hypoplastic (less proteins and DNA), with lowerconcentration of the mitochondrial uncoupling proteinthermogenin. In 8-day-old rats exposed to hypoxiapostnatally (day 2 today 8), at anyT_a below thermoneutralityO₂ was higher than incontrols; also, in this group IBAT was hypoplastic with decreasedthermogenin. Additional measurements under variousexperimental conditions indicated that the increased thermogeniccapacity was not explained by the smaller body mass and increased bloodoxygen content or by the eventuality of intermittent cold stimuliduring the chronic hypoxia. On the other hand, chronic hypercapnia (3%CO₂ in normoxia, fromday 2 to day8) also resulted in increased normoxic thermogenesis. We conclude that chronic hypoxia in the perinatal period1) reduces IBAT mass andthermogenin concentration and2) can increase the newborn's thermogenic capacity because of stress-related mechanisms not specific to hypoxia.

     

Nociceptive sensitivity to a long-term stimulus in the formalin test in female and male rats in postnatal ontogenesis

In Wistar rats, a comparative study of pain sensitivity to the long-term stimulus in the formalin test was carried out at different age periods—the prepubertal (25 days), pubertal (40 days), and sex maturity (90 days) periods. The pain sensitivity was evaluated by standard indexes of the biphasic behavioral response (BBR)—patterns of flexing, shaking, licking of the leg injected by formalin, and by duration of the first and second response phases and of the interphase interval. It is found that with development of the central nervous system the pain sensitivity changed, with differences in the first, acute and the second, tonic BBR phases: in the tonic phase of response the pain sensitivity increased in males by the indexes organized at the spinal, while in females, at the supraspinal level, whereas in the acute phase it decreased essentially at the supraspinal level in individuals of both sexes. The greater number of age differences in the pain sensitivity is revealed in females by the response patterns organized at the supraspinal level. At the same level, essential readjustments in hormonal and neurotransmitter systems are reflected in the high BBR indexes. In adult individuals the sex dimorphism is detected in duration of the interphase intervals. Activity of the bulbospinal descending inhibitory monoaminergic systems is shown to continue increasing for the first three months of life, with predominance of this process in females. The obtained data allow concluding that the BBR characteristics depend on age and sex of individuals and are determined by the organization level in CNS of the response patterns characterizing the pain sensitivity in the formalin test.Translated from Zhurnal Evolyutsionnoi Biokhimii i Fiziologii, Vol. 41, No. 1, 2005, pp. 76–81.Original Russian Text Copyright © 2005 by Butkevich, Vershinina.     

Zinc uptake by blood cells of rats in zinc deficiency and inflammation

In zinc deficiency, the function of leukocytes is impaired. However, the results of studies on the zinc concentration of blood cells in zinc deficiency are conflicting, probably in part because of technical and analytical problems. The aim of this study was to investigate, under standard conditions, the uptake of⁶⁵Zn-labeled zinc by blood cells, taken from zinc-deficient rats and from rats in which an inflammation is induced. In both conditions, the serum zinc concentration is reduced. In clinical practice, this makes it difficult to determine whether the decrease in serum zinc is the result of a real or an apparent zinc deficiency. In stress, like an inflammatory disease, the decrease of zinc reflects an apparent zinc deficiency because of redistribution of serum zinc into the liver and because of decrease in serum albumin concentration. Over 70% of the serum zinc is bound to albumin. Blood cells from zinc-deficient and control rats were isolated using a discontinuous Percoll gradient and incubated under nearly physiological conditions in a⁶⁵Zn-containing medium. A significant increase in the in vitro uptake of⁶⁵Zn-labeled zinc by the blood cells of zinc-deficient rats was seen: erythrocytes 1.3, mononuclear cells 2.0, and polymorphonuclear cells 2.6 times the control values. During inflammation, no change in⁶⁵Zn-labeled zinc uptake by erythrocytes and mononuclear cells was demonstrated after 2 d, although the serum zinc and albumin concentrations were decreased, but a small but significant increase in zinc uptake by polymorphonuclear cells was observed. This study of⁶⁵Zn uptake in vitro under standard conditions may prove of value for distinguishing in patients real zinc deficiency from apparent zinc deficiency owing to, e.g., stress, although additional experiments should be performed. A part of this study has been presented at the Meeting of The American Gastroenterological Association on May 12–18, 1990, San Antonio, TX, and has been published in abstract inGastroenterology 98 suppl., A423.