Osmotic threshold and sensitivity for vasopressin release and fos expression by hypertonic NaCl in ovine fetus

In adults, hyperosmolality stimulates central osmoreceptors, resulting in arginine vasopressin (AVP) secretion. Near-term fetal sheep have also developed mechanisms to respond to intravascular hypertonicity with stimulation of in utero AVP release. However, prior studies demonstrating fetal AVP secretion have utilized plasma tonicity changes greater than those required for adult osmotically induced AVP stimulation. We sought to examine near-term fetal plasma osmolality threshold and sensitivity for stimulation of AVP secretion and to correlate plasma hormone levels with central neuronal responsiveness. Chronically instrumented ovine fetuses (130 +/- 2 days) and maternal ewes simultaneously received either isotonic or hypertonic intravascular NaCl infusions. Maternal and fetal plasma AVP and angiotensin II (ANG II) levels were examined at progressively increasing levels of plasma hypertonicity. Intravenous hypertonic NaCl gradually elevated plasma osmolality and sodium levels. Both maternal and fetal plasma AVP increased during hypertonicity, whereas ANG II levels were not changed. Maternal AVP levels significantly increased with a 3% increase in plasma osmolality, whereas fetal plasma AVP significantly increased only at higher plasma osmolality levels (over 6%). Thus the slope of the regression of AVP vs. osmolality was greater for ewes than for fetuses (0.232 vs. 0.064), despite similar maternal and fetal plasma osmolality thresholds for AVP secretion (302 vs. 304 mosmol/kg). Hyperosmolality induced Fos immunoreactivity (FOS-ir) in the circumventricular organs of the fetal brain. FOS-ir was also demonstrated in the fetal supraoptic and paraventricular nuclei (SON and PVN), and double labeling demonstrated that AVP-containing neurons in the SON and PVN expressed Fos in response to intravenous NaCl. These results demonstrate that, in the ovine fetus at 130 days of gestation, neuroendocrine responses to cellular dehydration are functional, although they evidence a relatively reduced sensitivity for AVP secretion compared with the adult.     

Maternal dehydration: impact on ovine amniotic fluid volume and composition

Maternal dehydration consistent with mild water deprivation or moderate exercise results in maternal and fetal plasma hyperosmolality and increased plasma arginine vasopressin (AVP). Previous studies have demonstrated a reduction in fetal urine and lung fluid production in response to maternal dehydration or exogenous fetal AVP. As fetal urine and perhaps lung liquid combine to produce amniotic fluid, maternal dehydration may affect the amniotic fluid volume and/or composition. In the present study, six chronically-prepared pregnant ewes with singleton fetuses (128 +/- 1 day) were water deprived for 54 h to determine the effect on amniotic fluid. Maternal plasma osmolality (306.5 +/- 0.9 to 315.6 +/- 1.9 mOsm/kg) and AVP (1.9 +/- 0.2 to 22.2 +/- 3.2 pg/ml) significantly increased during dehydration. Similarly, fetal plasma osmolality (300.0 +/- 0.9 to 312.7 +/- 1.7 mOsm/kg) and AVP (1.4 +/- 0.1 to 10.4 +/- 2.4 pg/ml) increased in parallel to maternal values. Amniotic fluid osmolality (276.8 +/- 5.7 to 311.6 +/- 6.5 mOsm/kg) and sodium (139.8 +/- 4.8 to 154.0 +/- 5.4 mEq/l) and potassium (9.1 +/- 1.3 to 13.9 +/- 2.4 mEq/l) concentrations increased while a significant (35%) reduction in amniotic fluid volume occurred (871 +/- 106 to 520 +/- 107 ml). These results indicate that maternal dehydration may have marked effects on maternal-fetal-amniotic fluid dynamics, possibly contributing to the development of oligohydramnios.     

Maternal DDAVP-induced hyponatremia preserves fetal urine flow during acute fetal hemorrhage

Maternal administration of DDAVP induces maternal and fetal plasma hyponatremia, accentuates fetal urine flow, and increases amniotic fluid volume. Fetal hemorrhage represents an acute stress that results in fetal AVP secretion and reduced urine flow rate. In view of the potential therapeutic use of DDAVP for pregnancies with reduced amniotic fluid volume, we sought to examine the impact of maternal hypotonicity during acute fetal hemorrhage. Chronically catheterized pregnant ewes (130 +/- 2 days) were allocated to control or to DDAVP-induced hyponatremia groups. In the latter group, tap water (2,000 ml) was administered intragastrically to the ewe followed by DDAVP (20 microg bolus, 4 microg/h) and a maintenance intravenous infusion of 5% dextrose water for 4 h to achieve maternal hyponatremia of 10-12 meq/l. Thereafter, ovine fetuses from both groups were continuously hemorrhaged to 30% of estimated blood volume over a 60-min period. DDAVP caused similar degree of reductions in plasma sodium and osmolality in pregnant ewes and their fetuses. In response to hemorrhage, DDAVP fetuses showed greater reduction in hematocrit than control fetuses (14 vs. 10%). Both groups of fetuses demonstrated similar increases in plasma AVP concentration. However, the AVP-hemorrhage threshold was greater in DDAVP fetuses (22.5%) than in control (17.5%). Hemorrhage had no significant impact on plasma osmolality, electrolyte levels, or cardiovascular responses in either group of fetuses. Despite similar increases in plasma AVP, DDAVP fetuses preserved fetal urine flow rates, with values threefold those of control fetuses. These results suggest that under conditions of acute fetal stress of hemorrhage, maternal DDAVP may preserve fetal urine flow and amniotic fluid volume.     

Tissue-specific regulation of angiotensinogen mRNA accumulation by dexamethasone

The regulation of angiotensinogen gene expression in response to adrenalectomy and dexamethasone treatment was examined in multiple rat tissues. Angiotensinogen mRNA as quantitated by slot blot hybridization utilizing an angiotensinogen cRNA probe was most abundant in the liver with levels in the brain, kidney, and adrenal of 50, 25, and 10%, respectively. No angiotensinogen mRNA was detected in testes or heart. Although no change in the quantity of angiotensinogen mRNA was found following adrenalectomy and maintenance on 0.9% saline, dexamethasone treatment of both normal and adrenalectomized rats resulted in a time-dependent and tissue-specific accumulation of angiotensinogen mRNA. In normal animals, the hepatic response to treatment was a 4.5-fold increase in angiotensinogen mRNA by 8 h which remained 2.4-fold above basal levels by 24 h. Angiotensinogen mRNA levels in the brains of normal rats treated with dexamethasone increased only 60% by 6 h and returned to basal levels by 24 h. In contrast to the increases seen in brain and liver, angiotensinogen mRNA derived from kidney did not significantly change following dexamethasone treatment. In adrenalectomized animals, the hepatic response to dexamethasone was similar to normal animals with a 3.7-fold increase by 6 h. The accumulation in brain was greater in these animals compared to normals and increased 3-fold by 8 h. Finally, dexamethasone did not significantly increase levels in the kidney. These results clearly demonstrate glucocorticoid regulation of angiotensinogen mRNA levels in liver and brain. In contrast, the kidney, an organ known to contain glucocorticoid receptors, does not respond with increased angiotensinogen mRNA levels following glucocorticoid stimulation. These studies provide the first evidence for tissue-specific differences in the control of angiotensinogen mRNA.     

Prolonged prenatal hypernatremia alters neuroendocrine and electrolyte homeostasis in neonatal sheep

Arginine vasopressin (AVP) is a neuroendocrine hormone synthesized in the hypothalamus, and is stored and secreted by the posterior pituitary gland in response to stimuli such as plasma hypertonicity and hypotension. The primary physiologic roles of AVP include plasma osmolality and blood pressure regulation. We have previously demonstrated that chronic prenatal plasma hypertonicity alters the AVP regulatory pathway in newborn lambs. The objectives of the present study were to evaluate prolonged effects of antenatal plasma hypertonicity on neonatal plasma osmoregulation. Pregnant ewes at 119 +/- 3 days of gestation were water restricted to achieve and maintain hypertonicity until normal-term delivery. After delivery, ewes were provided food and water ad libitum and lambs were allowed maternal nursing. At the age of 28 days, blood samples were obtained for the analysis of plasma osmolality, electrolytes, and AVP levels from study (n= 5) and age-matched control (n= 6) lambs. Subsequently, lambs were euthanized, and the pituitary and hypothalamus were processed for the determination of pituitary AVP content by radioimmunoassay, and AVP gene expression by Northern analysis. In response to water restriction, maternal plasma osmolality significantly increased (306 +/- 1.1 to 326 +/- 1.2 mOsm/kg, P< 0.001). At the age of 28 days, plasma sodium level was higher in study (prenatally dehydrated) than control lambs (144.6 +/- 0.4 vs 142.6 +/- 0.3,P< 0.05). Study lambs had higher plasma AVP concentrations than the control lambs (4.1 +/- 0.4 vs 1.7 +/- 0.4 pg/ml,P< 0.05). Similarly, total pituitary AVP content was higher in thein utero hypertonic lambs than in the control lambs (6.5 +/- 1.0 vs 2.8 +/-1.2 microg, P< 0.05). However, there was no difference in hypothalamic AVP mRNA levels between the two groups. The present study demonstrates that chronic maternal and fetal plasma hypertonicity has prolonged effects on pituitary and plasma AVP, as well as plasma sodium in neonatal lambs, providing further evidence suggesting prenatal imprinting of osmoregulation through at least 1 month of age.     

Acute effects of ethanol on brain,plasma and adrenal monoamine concentrations in virgin and pregnant rats and their fetuses

The dose-response relationship in brain, plasma, and adrenal monoamine changes after acute oral ethanol administration (1, 2, 4 g/kg body wt) was studied in virgin rats to determine whether the response to the highest dose differed in 21-day pregnant animals, and to assess the potential consequences of ethanol on the neurotransmitter systems of their fetuses. Blood ethanol and acetaldehyde concentrations in blood increased progressively with the ethanol dose in virgin rats, and values in pregnant animals were very similar. Ethanol concentration in fetal blood and amniotic fluid did not differ from that in mother's blood whereas fetal acetaldehyde concentrations were negligible. In a dose-related manner, ethanol decreased brain DA, DOPAC and 5HT concentrations did not affect those of NA and 5HIAA, or adrenal A and NA concentrations, whereas it enhanced plasma NA levels. Basal levels of monoamines and their changes after ethanol intake did not differ in pregnant and virgin rats. Monoamine and metabolite concentrations were much lower in fetal than in maternal brains whereas plasma and adrenal catecholamine concentrations were very similar and maternal ethanol intake did not modify these fetal parameters in the fetus. Results are in agreement with the known similar metabolic response to ethanol in fed pregnant and virgin rats. The lack of fetal monoamine response to maternal ethanol intake may be a consequence of the incapacity of fetal liver to form acetaldehyde and the ability of the placenta to oxidize maternal acetaldehyde which protects the fetus from maternal alcohol intake at late gestation.     

Insulin-like growth factors and binding proteins in the fetal rat: Alterations during maternal starvation and effects in fetal brain cell culture

Maternal malnutrition adversely affects fetal body and brain growth during late gestation. We utilized a fetal brain cell culture model to examine whether alternations in circulating factors may contribute to reduce brain growth during maternal starvation; we then used specific immunoassay and western blotting techniques, and purified peptides to investigate the potential role that altered levels of insulin-like growth factors (IGFs) and IGF binding proteins (IGFBPs) may play in impaired growth during maternal nutritional restriction.Fetal, body, liver, and brain weight were reduced after 72 hr maternal starvation, and plasma from starved fetuses were less potent than fed fetal plasma in stimulating brain cell growth. Circulating levels of IGF-I were reduced in starved compared to fed fetuses, while levels of IGF-II were similar in both groups. In contrast, [¹²⁵I]-IGF-I binding assay demonstrated an increase in the availability of plasma IGFBPs following starvation. Western ligand blotting and densitometry indicated that levels of 32 Kd IGFBPs were 2-fold higher in starved compared to fed fetal plasma. Immunoblotting and immunoprecipitation with antiserum against rat IGFBP-1 confirmed that heightened levels of immunoreactive IGFBP-1 accounted for the increase in 32 Kd IGFBPs in starved plasma. Levels of 34 Kd BPs, representing IGFBP-2, were unaffected by starvation. Reconstitution experiments in cell culture showed that IGF-I promoted fetal brain cell growth, and that when they were supplemented with IGF-I, the growth promoting activity of starved fetal plasma was restored to fed levels. These changes were measured using MTT to assess mitochondrial reductase activity. Conversely, addition of physiological amounts of rat IGFBP-1 inhibited the effects of fed fetal plasma on brain cell growth, and bioactivity was reduced even further with higher concentrations of IGFBP-1. Based on these results, we conclude that reciprocal changes in circulating levels of IGFBP-1 (increased) and IGF-I (decreased) may combine to reduce the availability of IGF-I to this tissue and limit fetal brain cell growth when maternal nutrition is impaired.     

Iron deficiency in the rat: biochemical studies of fetal metabolism

The effects of dietary-induced iron deficiency on fetal and maternal metabolism were studied in the rat. Concentrations of phenylalanine, but not tyrosine, were significantly elevated in plasma from iron-deficient maternal and fetal rats at day 20 of gestation with individual fetal plasma levels of phenylalanine as high as 10 mg per 100 ml. Concentrations of total 5-hydroxyindole compounds were significantly decreased in brain tissue from iron-deficient fetuses (day 20 of gestation), suggesting that synthesis of the compounds may be inhibited by iron deficiency. Mitochondrial NADH oxidase activity was markedly decreased (60%) in homogenates of fetuses at day 14 of gestation and may account for the high fetal resorption rate and small fetal size observed in the rat in iron deficiency.     

Rat alpha-fetoprotein in experimental utero-placental ischaemia.

Retardation of growth and death of fetal rats were produced after uteroplacental ischaemia was induced by surgical ligation of the uterine arteries. Changes in maternal plasma levels of alpha-fetoprotein (AFP) were measured by radioimmunoassay. In rats in which the uterine blood supply was totally occluded, the resultant increase in maternal plasma AFP was due to resorption of fetal elements, because AFP levels in maternal rat plasma did not increase following hysterectomy in a control group. Maternal plasma AFP levels in rats with a partly occluded blood supply (and therefore some dead and some live fetuses) paralleled those of sham-operated rats, suggesting that increased placental transfer of AFP to maternal plasma may have offset the anticipated decline of AFP due to a decreased number of live fetuses.     

Endogenous angiotensin II regulates hepatic angiotensinogen production

We have investigated the effects of endogenous angiotensin II (ANG II) on hepatic angiotensinogen mRNA levels in rats. Changes in endogenous ANG II were induced by various sodium intakes (standard-, low-, and high-sodium) or by enalapril treatment. In a low sodium state for 2 weeks, angiotensinogen mRNA levels and plasma ANG II concentration increased 1.3-fold and 1.6-fold compared to those in standard sodium state, respectively. In a high sodium state, angiotensinogen mRNA levels and plasma ANG II concentration decreased by 42% and 56% compared to the standard sodierm state, respectively. Four hours after treatment with enalapril (3 mg/kg), angiotensinogen mRNA level and plasma ANG II concentration decreased by 25% and 12% compared to the standard sodium state, respectively. There was a close correlation between angiotensinogen mRNA level and plasma ANG II concentration (r = 0.79, P less than 0.01). These results suggest that endogenous ANG II may play an important role in the regulation of hepatic angiotensinogen synthesis.     

Dominant role of glucagon in the initial induction of phosphoenolpyruvate carboxykinase mRNA in cultured hepatocytes from fetal rats.

The injection of streptozotocin to 18-day-old rat fetuses induced, 2 days later, a 50% fall in plasma insulin and a twofold increase in plasma glucagon concentrations and liver cAMP levels. Phosphoenolpyruvate carboxykinase mRNA that were undetectable in the fetal rat liver, accumulated 48 h after streptozotocin injection, their concentration being 30% of that found in the liver of 1-day-old newborn rats in whom liver phosphoenolpyruvate carboxykinase gene expression is maximal. Physiological concentrations of glucagon (0.7 +/- 0.2 nM) induced, within 2 h, phosphoenolpyruvate carboxykinase mRNA accumulation in cultured hepatocytes from 20-day-old fetuses. The addition of insulin (0.01-100 nM) inhibits, by no more than 30%, the glucagon-induced phosphoenolpyruvate carboxykinase mRNA accumulation. Exposure of fetal hepatocytes to insulin for 24 h did not change the glucagon dose/response curve and did not lead to a more efficient inhibition of the glucagon-induced phosphoenolpyruvate carboxykinase mRNA accumulation, despite a clear stimulatory effect on the rate of lipogenesis. In contrast, when hepatocytes were cultured in the presence of dexamethasone, the glucagon-induced phosphoenolpyruvate carboxykinase mRNA accumulation can be totally inhibited by pharmacological concentrations of insulin (10 nM). From these in-vivo and in-vitro studies, it is concluded that, under physiological conditions, the postnatal rise in plasma glucagon concentration is more important than the fall in the plasma insulin concentration for the primary induction of liver phosphoenolpyruvate carboxykinase gene expression.     

Effects of 7-day amino acid infusion on renal growth, function, and renin-angiotensin system in fetal sheep

These experiments examined whether renal growth and the fetal renin-angiotensin system could be stimulated by infusion of amino acids and whether chronic amino acid infusions restored glomerulotubular balance, which had been disrupted during 4-h infusions. Five fetal sheep aged 122 +/- 1 days gestation received an infusion of alanine, glycine, proline and serine in 0.15 M saline at 0.22 mmol/min for 7 days. Six control fetuses were given saline at the same rate (5 ml/h). Kidney wet weights after amino acid infusion were 28% larger than control fetuses (P < 0.05), and renal angiotensinogen mRNA levels were approximately 2.6-fold higher (P < 0.005). Circulating renin levels and renal renin mRNA levels were suppressed (P < 0.05), and renal renin protein levels tended to be lower. Arterial pressure was increased, and there was a marked, sustained natriuresis and diuresis. Glomerular filtration rate and filtered sodium were approximately two-fold higher throughout infusion (P < 0.05). Fractional proximal sodium reabsorption, suppressed at 4 h (from 73.4 +/- 6.5 to 53.7 +/- 10.2%), did not return to control levels (36.1 +/- 3.4% on day 7, P < 0.05). Distal sodium reabsorption was markedly increased (from 79 +/- 25 to 261 +/- 75 mumol/min by day 7, P < 0.005), but this was not sufficient to restore glomerulotubular balance. The resultant high rates of sodium excretion led to hyponatremia and polyhydramnios. In conclusion, long-term amino acid infusions increased renal angiotensinogen gene expression, kidney weight, and distal nephron sodium reabsorptive capacity but failed to restore proximal and total glomerulotubular balance.     

The effect of maternal fluid intake on the volume and composition of fetal urine

The effects on fetal renal function of restricting maternal water intake to 1 l/day for 6 days was investigated in 7 chronically-catheterized fetuses (gestation age 118-131 days). Restriction of water intake caused a significant decrease in maternal urine flow rate and significant increases in maternal plasma and urinary osmolality. Fetal renal function was investigated on the third and sixth days of the period of restricted maternal intake of water. Urine flow rate from the fetus was depressed significantly, and urinary osmolality increased significantly. The glomerular filtration rate remained unchanged, and free water clearance was decreased. These changes indicate increased water reabsorption in the distal parts of the nephron, probably consequent upon increased circulating levels of antidiuretic hormone. In 3 fetuses whose mothers subsequently had free access to water, these changes in urine flow rate and free water clearance that occurred during water restriction were reversed. There was an inverse correlation between maternal plasma osmolality and fetal free water clearance corrected for glomerular filtration rate. It is concluded that when water intake by a pregnant animal is restricted, the availability of water to the fetus is reduced and fetal sheep respond by producing a concentrated urine.     

The renal response to the ingestion of fluid by the fetal sheep

To see if the variability in fetal urine flow and sodium excretion was related to fetal drinking activity, renal function was investigated in two groups of oesophageally-ligated fetuses and one group of non-ligated fetuses. There was no significant difference in urine flow, sodium excretion or glomerular filtration rate in the ligated fetuses compared with the non-ligated fetuses. Furthermore, oesophageal ligation had no effect on the variability in urine flow and sodium excretion rate. The response of fetal kidney to ingestion of fluid was investigaeed in 2 groups of oesophageally-ligated fetuses. In one group it was shown that ingestion of 20 ml/kg of amniotic fluid by the fetus had no consistent effect on fetal renal function. In the other group it was shown that the ingestion of 200 ml water also had no consistent effect on fetal renal function. The water load caused a rise in fetal blood pressure and a fall in plasma osmolality. Since there was no significant increase in free water clearance and fetal plasma osmolality decreased then rose towards control levels, it is concluded that the oral water load was absorbed from the fetal gastrointestinal tract and diffused out of the fetal compartment across the placenta. These experiments show that fetal drinking is probably not responsible for the variability often seen in fetal urine flow and sodium excretion rate.     

Maternal diets enriched in olive oil regulate lipid metabolism and levels of PPARs and their coactivators in the fetal liver in a rat model of gestational diabetes mellitus

In a rat model of gestational diabetes mellitus (GDM) programmed in the offspring of neonatal streptozotocin-induced (nSTZ) diabetic rats, lipids are accumulated in the fetal liver in a sex-dependent way. Here, we evaluated whether maternal diets enriched in olive oil in rats that will develop GDM ameliorate lipid metabolic impairments in the fetal livers. Pregnant offspring of control and nSTZ diabetic rats (F0) were fed a 6% olive oil-supplemented diet throughout the F1 gestation. We evaluated maternal metabolic parameters as well as lipid content, expression of lipid metabolizing enzymes and protein expression of PLIN2, PPARs and PPAR coactivators in the fetal livers. The offspring of nSTZ diabetic rats developed GDM regardless of the maternal treatment. Hypertriglyceridemia in GDM rats was prevented by the olive oil-enriched maternal treatment. In the livers of male fetuses of GDM rats, the maternal olive oil-supplemented diet prevented lipid overaccumulation and prevented the increase in PPARγ and PPARδ levels. In the livers of female fetuses of GDM rats, the maternal olive oil supplementation prevented the increase in PPARδ levels and the reduction in PGC1α levels, but did not prevent the reduced lipid content. Control and GDM rats showed a reduction of lipid metabolic enzymes in the fetal livers, which was associated with reduced levels of the PPAR coactivators PGC-1α and SRC-1 in males and of SRC-1 in females. These results suggest powerful effects of a maternal olive oil-supplemented diet in the fetal liver, possibly providing benefits in the fetuses and offspring from GDM rats.     

Enhanced central response to dehydration in mice lacking angiotensin AT(1a) receptors

The objective was to determine the central nervous system (CNS) responses to dehydration (c-Fos and vasopressin mRNA) in mice lacking the ANG AT(1a) receptor [ANG AT(1a) knockout (KO)]. Control and AT(1a) KO mice were dehydrated for 24 or 48 h. Baseline plasma vasopressin (VP) was not different between the groups; however, the response to dehydration was attenuated in AT(1a) KO (24 +/- 11 vs. 10.6 +/- 2.7 pg/ml). Dehydration produced similar increases in plasma osmolality and depletion of posterior pituitary VP content. Neuronal activation was observed as increases in c-Fos protein and VP mRNA. The supraoptic responses were not different between groups. In the paraventricular nucleus (PVN), c-Fos-positive neurons (57.4 +/- 10.7 vs. 98.4 +/- 7.4 c-Fos cells/PVN, control vs. AT(1a) KO) and VP mRNA levels (1.0 +/- 0.1 vs. 1.4 +/- 0.1 microCi, control vs. AT(1a) KO) were increased with greater responses in AT(1a) KO. A comparison of 1- to 2-day water deprivation showed that plasma VP, brain c-Fos, and VP mRNA returned toward control on day 2, although plasma osmolality remained high. Data demonstrate that AT(1a) KO mice show a dichotomous response to dehydration, reduced for plasma VP and enhanced for PVN c-Fos protein and VP mRNA. The results illustrate the importance of ANG AT(1a) receptors in the regulation of osmotic and endocrine balance.     

Isocaloric maternal low-protein diet alters IGF-I,IGFBPs, and hepatocyte proliferation in the fetal rat

We investigated the effect of an isocaloric maternal low-protein diet during pregnancy in rats on the proliferative capacity of cultured fetal hepatocytes. The potential roles of these changes on the IGF-IGF-binding protein (IGFBP) axis, and the role of insulin and glucocorticoids in liver growth retardation, were also evaluated. Pregnant Wistar rats were fed a control (C) diet (20% protein) or a low-protein (LP) diet (8%) throughout gestation. In primary culture, the DNA synthesis of hepatocytes derived from LP fetuses was decreased by approximately 30% compared with control hepatocytes (P < 0.05). In parallel, in vivo moderate protein restriction in the dam reduced the fetal liver weight and IGF-I level in fetal plasma (P < 0.01) and augmented the abundance of 29- to 32-kDa IGFBPs in fetal plasma (P < 0.01) and fetal liver (P < 0.01). By contrast, the abundance of IGF-II mRNA in liver of LP fetuses was unaffected by the LP diet. In vitro, the LP-derived hepatocytes produced less IGF-I (P < 0.01) and more 29- to 32-kDa IGFBPs (P < 0.01) than hepatocytes derived from control fetuses. These alterations still appeared after 3-4 days of culture, indicating some persistence in programming. Dexamethasone treatment of control-derived hepatocytes decreased cell proliferation (54 +/- 2.3%, P < 0.01) and stimulated 29- to 32-kDa IGFBPs, whereas insulin promoted fetal hepatocyte growth (127 +/- 5.5%, P < 0.01) and inhibited 29- to 32-kDa IGFBPs. These results show that liver growth and cell proliferation in association with IGF-I and IGFBP levels are affected in utero by fetal undernutrition. It also suggests that glucocorticoids and insulin may modulate these effects.     

Effects of dehydration on plasma osmolality, thirst-related behavior, and plasma and brain angiotensin concentrations in Couch's spadefoot toad, Scaphiopus couchii

Under dehydrating conditions, many terrestrial vertebrates species exhibit increases in plasma osmolality and their drinking behavior. Under some circumstances, this behavioral change is accompanied by changes in plasma and central angiotensin concentrations, and it has been proposed that these changes in angiotensin levels induce the thirst-related behaviors. In response to dehydration, the spadefoot toad, Scaphiopus couchii, exhibits thirst-related behavior in the form of cutaneous drinking. This behavior has been termed water absorption response (WR) behavior. Spadefoot toads live in harsh desert environments and are subject annually to dehydrating conditions that may induce thirst-related behavior. We tested the hypothesis that an increase in WR behavior is associated with both an increase in plasma osmolality and an increase in plasma and brain angiotensin concentrations. First, we determined the degree of dehydration that was necessary to initiate WR behavior. Animals dehydrated to 85% of their standard bladder-empty weight via deprivation of water exhibited WR behavior more frequently than control toads left in home containers with water available. Next, using the same dehydration methods, we determined the plasma osmolality and sodium concentrations of dehydrated toads. Toads dehydrated to 85% standard weight also had a significant increase in plasma osmolality, but exhibited no overall change in plasma sodium concentrations, indicating that while an overall increase in plasma osmolality appears to be associated with WR behavior in S. couchii, changes in sodium concentrations alone are not sufficient to induce the behavior. Finally, plasma and brain angiotensin concentrations were measured in control toads and toads dehydrated to 85% standard weight. Plasma and brain angiotensin concentrations did not increase in dehydrated toads, indicating that dehydration-induced WR behavior that is associated with changes in plasma osmolality may not be induced by changes in endogenous angiotensin concentrations in S. couchii.