Postnatal effects of intrauterine treatment of the growth-restricted ovine fetus with intra-amniotic insulin-like growth factor-1.

KEY POINTS: Fetal growth restriction increases the risk of fetal and neonatal mortality and morbidity, and contributes to increased risk of chronic disease later in life. Intra-amniotic insulin-like growth factor-1 (IGF1) treatment of the growth-restricted ovine fetus improves fetal growth, but postnatal effects are unknown. Here we report that intra-amniotic IGF1 treatment of the growth-restricted ovine fetus alters size at birth and mechanisms of early postnatal growth in a sex-specific manner. We also show that maternal plasma C-type natriuretic peptide (CNP) products are related to fetal oxygenation and size at birth, and hence may be useful for non-invasive monitoring of fetal growth restriction. Intrauterine IGF1 treatment in late gestation is a potentially clinically relevant intervention that may ameliorate the postnatal complications of fetal growth restriction. ABSTRACT: Placental insufficiency-mediated fetal growth restriction (FGR) is associated with altered postnatal growth and metabolism, which are, in turn, associated with increased risk of adult disease. Intra-amniotic insulin-like growth factor-1 (IGF1) treatment of ovine FGR increases growth rate in late gestation, but the effects on postnatal growth and metabolism are unknown. We investigated the effects of intra-amniotic IGF1 administration to ovine fetuses with uteroplacental embolisation-induced FGR on phenotypical and physiological characteristics in the 2  weeks after birth. We measured early postnatal growth velocity, amino-terminal propeptide of C-type natriuretic peptide (NTproCNP), body composition, tissue-specific mRNA expression, and milk intake in singleton lambs treated weekly with 360 µg intra-amniotic IGF1 (FGRI; n = 13 females, 19 males) or saline (FGRS; n = 18 females, 12 males) during gestation, and in controls (CON; n = 15 females, 22 males). There was a strong positive correlation between maternal NTproCNP and fetal oxygenation, and size at birth in FGR lambs. FGR lambs were ∼20% lighter at birth and demonstrated accelerated postnatal growth velocity. IGF1 treatment did not alter perinatal mortality, partially abrogated the reduction in newborn size in females, but not males, and reduced accelerated growth in both sexes. IGF1-mediated upregulation of somatotrophic genes in males during the early postnatal period could suggest that treatment effects are associated with delayed axis maturation, whilst treatment outcomes in females may rely on the reprogramming of nutrient-dependent mechanisms of growth. These data suggest that the growth-restricted fetus is responsive to intra-amniotic intervention with IGF1, and that sex-specific somatotrophic effects persist in the early postnatal period.

Effects of pre-eclampsia and fetal growth restriction on C-type natriuretic peptide.

OBJECTIVE: To determine changes in plasma C-type natriuretic peptide (CNP), a paracrine product of the vascular endothelium, in pregnancies with vascular disorders, and relate these to time of presentation and severity. DESIGN: Retrospective nested cases and controls. SETTING: Community study, Auckland New Zealand. POPULATION: Screening for Pregnancy Endpoints (SCOPE) data and bio-bank of maternal plasma. METHODS: Maternal plasma amino terminal proCNP (NTproCNP) was measured by radioimmunoassay in early (14-16 weeks of gestation, and again at 19-21 weeks of gestation) and late (34-36 weeks of gestation) pregnancy in three groups of women (20 per group): pre-eclampsia (pre-eclampsia); gestational hypertension (GHT) with small for gestational age (SGA); and uncomplicated pregnancy. MAIN OUTCOME MEASURES: Change in NTproCNP and associations with concurrent blood pressure, time of case presentation, severity, and infant birthweight. RESULTS: Plasma NTproCNP in early pregnancy in women with vascular disorders did not differ from those found in controls. In late pregnancy, levels in pre-eclampsia (28.8 ± 2.3 pM) and in GHT with SGA (28.6 ± 4.8 pM) were significantly increased (P = 0.01 and 0.027, respectively) compared with controls (21.3 ± 1 pM). In pre-eclampsia, levels were significantly higher (P < 0.03) at 14-16 weeks of gestation in women diagnosed prior to 34 weeks of gestation. Combining all three groups, associations of NTproCNP with concurrent diastolic and mean arterial pressure were found at 34-36 weeks of gestation (r = 0.46). No significant associations were identified with birthweight. CONCLUSIONS: CNP secretion during gestation is responsive to vascular stress. Plasma NTproCNP measurements may have clinical application in late pregnancy in defining the different phenotypes associated with pre-eclampsia.

C-type natriuretic peptide in Parkinson's disease: reduced secretion and response to deprenyl.

C-type natriuretic peptide (CNP) is a neurotrophic factor widely expressed in the central nervous system including the basal ganglia, limbic system and hypothalamus. Nothing is known of CNP's role in the human brain but in rodents CNP promotes axon growth and branching, and interacts with dopaminergic function in models of addiction. Because preliminary evidence showed reduced levels in Parkinson's disease (PD), we examined concentrations of CNP peptides in cerebrospinal fluid (CSF) in 146 PD patients from the DATATOP study to determine changes over time in relation to medication status and cognitive function. CNP and an aminoterminal product of proCNP (NTproCNP) were measured in extracts from stored CSF by radioimmunoassay. CSF samples were obtained twice-at enrolment and at the study's endpoint (requirement for levodopa treatment) after treatment with placebo or deprenyl. At enrolment, median baseline concentration of CSF NTproCNP (776 pmol/L, n = 146) was significantly lower than that in a reference group without neurological disorder (1,010 pmol/L, p < 0.001). Concentrations declined significantly during placebo (p = 0.02) and lower values at enrolment were associated with more rapid functional decline (p < 0.01). In contrast, deprenyl-a treatment which delayed the need for levodopa-nullified the time-dependent decline in CSF NTproCNP. In conclusion subnormal CSF NTproCNP which declines with time and associates with increasing functional disability implicates CNP in PD. Concordant clinical and peptide responses to deprenyl suggest that some of the benefits of monoamine oxidase inhibitors in PD are mediated by preserving tissue CNP activity.

Impact of age, phenotype and cardio-renal function on plasma C-type and B-type natriuretic peptide forms in an adult population.

CONTEXT: In contrast to the cardiac hormones, atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP), variations in plasma concentrations of C-type natriuretic peptide (CNP) in healthy adults are ill-defined, limiting their clinical application. OBJECTIVE: Our objective was to define the effect of age, phenotype (gender, height, BMI), and cardiac and renal function on plasma CNPs in an adults population without renal or cardiovascular disease. DESIGN AND SETTING: This was a prospective cross-sectional observational study of adult volunteers, aged 21-80 years, randomly selected from the electoral roll. SUBJECTS AND METHODS: Plasma CNP and its associated aminoterminal propeptide (NTproCNP) were measured in 258 subjects and related to age, gender, height and plasma creatinine. Subgroup analyses seeking associations with cardiac function (plasma BNP and NTproBNP) and bone turnover bone-specific alkaline phosphatase (bALP) were also determined. RESULTS: Plasma concentrations of CNPs in men continued to decline from adolescent values to reach a nadir in the 5th decade after which values increased. Similar but less marked changes occurred in women. In both sexes, NTproCNP was inversely and independently correlated with height. In contrast to B-type natriuretic peptides (BNPs), NTproCNP was higher in men, significantly related to creatinine and positively related to bALP. CONCLUSIONS: Gender- and age-specific changes affect CNPs in adults. Inverse associations of NTproCNP with adult height, positive correlation with creatinine - and in contrast to CNP - no association with BNP are further unique findings distinguishing NTproCNP, which need to be considered in future studies.

Plasma C-type natriuretic peptide forms and thyroid status in prepubertal children with acquired thyroid disease.

OBJECTIVE: C-type natriuretic peptide (CNP) and thyroid hormone (TH) are essential for normal skeletal growth. Plasma CNP peptides correlate with growth velocity, but the relationship between thyroid status and CNP production is unknown. This study examined the impact of restoring normal TH levels on CNP and height velocity (HV) in children with acquired hypo- and hyperthyroidism. DESIGN: We performed a prospective, observational study in prepubertal children with acquired hypothyroidism (n = 15) and hyperthyroidism (n = 12). MEASUREMENTS: Blood levels of CNP, amino-terminal proCNP (NTproCNP), bone-specific alkaline phosphatase (BSAP), IGF-I and TH levels were measured before and during the first 6 months of standard treatment for hypo- and hyperthyroidism, and correlations were determined. RESULTS: At baseline, HV, CNP, NTproCNP and BSAP were significantly higher in hyper- than in hypothyroid subjects. Changes in TH after treatment were closely coupled to change in CNP and NTproCNP in hyperthyroid, but not in hypothyroid, children. In addition, a positive association of HV with CNP peptides was found during treatment of hyperthyroidism. Normalizing TH did not correlate with changes in BSAP or IGF-I in either group. CONCLUSIONS: Plasma CNP peptides are higher in children with hyperthyroidism than in those with hypothyroidism at diagnosis and, in hyperthyroid children, change concordantly with TH and HV during treatment. Differential responses of CNP in the two groups suggest CNP production is dependent on growth plate activity and not a direct effect of TH on CNP gene expression. Our findings suggest novel mechanisms underlying changes in skeletal response during treatment in children with acquired thyroid disease.

Caloric restriction, but not caloric loading, affects circulating fetal and maternal C-type natriuretic peptide concentrations in late ovine gestation.

The factors regulating the greatly elevated concentrations of maternal plasma C-type natriuretic peptide (CNP) forms in ruminant pregnancy are largely unknown, but nutrient status is likely to be important. Previous work has shown that increases in maternal plasma CNP, sourced from the placenta, occur in response to caloric restriction in late gestation. Whether oversupply of nutrients also regulates CNP secretion in pregnancy has not been studied. Hypothesising that CNP in fetal and maternal tissues will be responsive to both deficiency and excess, we studied changes in CNP and a cosecreted fragment, namely N-terminal pro-CNP (NTproCNP), during short-term periods of caloric restriction (CR) and loading (CL). Twin-bearing ewes received CR (fasted Days 121-124), CL (Days 110-124) or control maintenance diets. During CR, fetal plasma CNP forms, insulin-like growth factor (IGF)-1 and liveweight all fell, and maternal plasma NTproCNP increased. During CL, fetal IGF-1 increased, whereas CNP forms and liveweight were unchanged, as were maternal concentrations of CNP forms. The high abundance of CNP peptides in placental tissues was unaffected by these short-term changes in nutrient supply. We conclude that CNP in the fetal-maternal unit is acutely responsive to undernutrition, but is unaffected by oversupply in late gestation.

Antibody binding of insulin in diabetic ketoacidosis.

This study investigates the effects of insulin antibody binding on free insulin levels measured in patients with acute diabetic ketoacidosis receiving insulin by constant infusion. In spite of antibody binding ranging from 10 to 90 per cent of the total circulating insulin, the steady state concentrations of free insulin were similar to those observed in individuals on identical infusion rates but without insulin-binding antibodies. However, the levels of free insulin in two patients were substantially lower than expected for the rate of insulin infusion, even though levels of bound insulin were not greatly elevated. An infusion rate of at least 4 U. per hour produced satisfactory rate of fall of plasma glucose, whereas lower dose regimens (2 U. per hour)--producing steady state free insulin concentrations ranging from 28 to 49 mU. per liter in different subjects--were unreliable in controlling the metabolic abnormalities of diabetic ketoacidosis.

Plasma N-terminal pro-brain natriuretic peptide and adrenomedullin: prognostic utility and prediction of benefit from carvedilol in chronic ischemic left ventricular dysfunction. Australia-New Zealand Heart Failure Group.

OBJECTIVES: We sought to assess plasma concentrations of the amino (N)-terminal portion of pro-brain natriuretic peptide (N-BNP) and adrenomedullin for prediction of adverse outcomes and responses to treatment in 297 patients with ischemic left ventricular (LV) dysfunction who were randomly assigned to receive carvedilol or placebo. BACKGROUND: Although neurohormonal status has known prognostic significance in heart failure, the predictive power of either N-BNP or adrenomedullin in chronic ischemic LV dysfunction has not been previously reported. METHODS: Plasma N-BNP and adrenomedullin were measured in 297 patients with chronic ischemic (LV) dysfunction before randomization to carvedilol or placebo, added to established treatment with a converting enzyme inhibitor and loop diuretic (with or without digoxin). The patients' clinical outcomes, induding mortality and heart failure events, were recorded for 18 months. RESULTS: Above-median N-BNP and adrenomedullin levels conferred increased risks (all p < 0.001) of mortality (risk ratios [95% confidence intervals]: 4.67 [2-10.9] and 3.92 [1.76-8.7], respectively) and hospital admission with heart failure (4.7 [2.2-10.3] and 2.4 [1.3-4.5], respectively). Both of these predicted death or heart failure independent of age, New York Heart Association functional class, LV ejection fraction, previous myocardial infarction or previous admission with heart failure. Carvedilol reduced the risk of death or heart failure in patients with above-median levels of N-BNP or adrenomedullin, or both, to rates not significantly different from those observed in patients with levels below the median value. CONCLUSIONS: In patients with established ischemic LV dysfunction, plasma N-BNP and adrenomedullin are independent predictors of mortality and heart failure. Carvedilol reduced mortality and heart failure in patients with higher pre-treatment plasma N-BNP and adrenomedullin.

Evidence for activation of brain atrial natriuretic factor during acute plasma volume expansion in sheep.

Atrial Natriuretic Factor (ANF) has been demonstrated within the central nervous system (CNS), where it has been implicated in the regulation of blood pressure, fluid and electrolyte balance. To explore the effect of acute plasma vol expansion on the activation of CNS ANF, changes in ANF levels of CSF drawn from the cisterna magna of anesthetized sheep were measured. Intravenous infusions of Dextran (10 ml/kg body wt, n = 7) or control (total vol 7.5 ml Dextran, n = 7) were administered over 30 min. Compared to control experiments, plasma vol expansion resulted in a 3-fold increase of central venous pressure (P = 0.002), a 25% increase in mean arterial pressure (P = 0.011), and a 20% reduction in packed red cell volume (P = 0.024). Accompanying these hemodynamic changes, plasma ANF concentrations doubled (17.0 +/- 3.0-41.0 +/- 7.8 pmol/liter at 60 min vs. 20.0 +/- 4.7-19.3 +/- 3.6 pmol/liter in controls) and remained elevated for 4 h, whereas CSF ANF concentrations showed a transient 3-fold increase (2.1 +/- 0.3-6.6 +/- 1.9 pmol/liter at 120 min vs. 2.5 +/- 0.5-3.8 +/- 0.7 pmol/liter in controls). The maximum increments of both plasma and CSF ANF were statistically significant (P = 0.002 and 0.03, respectively). Basal CSF levels were approximately 1/10 those in plasma, and no correlation was seen in either basal plasma and CSF levels, or the maximum increments of plasma and CSF ANF concentrations. In vol-expanded sheep, plasma cyclic GMP concentrations tended to increase, although not significantly different from controls, while CSF cyclic GMP was similar to controls throughout the experiments. The entry of ANF to cisterna magna CSF was studied in five additional anesthetized sheep. Infusion iv of ileu rat ANP (50 ng/kg.min over 60 min) raised plasma ANF levels 30-fold, but CSF ANF concentrations did not change. These experiments show that plasma vol expansion in sheep, in addition to stimulating cardiac ANF secretion, induces an increase in CSF ANF, which cannot be accounted for by transfer of ANF from blood into CSF.

Failure of angiotensin II to inhibit corticotropin-stimulated cortisol secretion.

In view of the reported inhibitory effect of angiotensin II on cortisol secretion in human subjects, the effect of local angiotensin infusions on steroid secretion maintained by ACTH was examined by using sheep with cervical autotransplanted adrenal glands. During sustained submaximal stimulation by exogenous ACTH (40--80 microunit/min), the addition of local infusions of angiotensin II (1.6--160.0 ng/min) caused increased aldosterone and smaller increments in cortisol secretion in most experients. There was no evidence of inhibition of cortisol secretion by angiotensin. When similar experiments were undertaken during maximum stimulation by ACTH (16.6 mU/min), increments in aldosterone, but not in cortisol secretion, were observed. These studies exclude an acute inhibitory effect of angiotensin on cortisol biosynthesis, at least in ovine adrenal glands, during stimulation by ACTH.

Central C-type natriuretic peptide but not atrial natriuretic factor lowers blood pressure and adrenocortical secretion in normal conscious sheep.

C-type natriuretic peptide (CNP) and its specific receptor (ANPRB) are richly distributed throughout the brain, especially the anterior pituitary and hypothalamus but not in tissue of nonneural origin. These findings suggest that the actions of CNP, unlike atrial natriuretic factor (ANF), are largely confined to the brain where CNP may participate in the central regulation of hemodynamics and salt and water balance. Therefore, we have studied the hemodynamic, renal, and hormonal effects of continuous intracerebroventricular infusions of CNP (5 micrograms/h for 4 h) in a vehicle-controlled study and compared the responses to those of ANF in normal conscious sheep. Hemodynamic and hormonal responses to ANF were not different from control infusions. There was a trend for urinary sodium and potassium excretion to increase throughout the control infusion but not on the ANF day. Water intake during control infusion (358 +/- 160 ml/4 h) was almost 3-fold that ingested during ANF (127 +/- 89 ml/4 h, NS). In contrast, CNP induced a prompt fall in mean arterial pressure (mean decrement 5 mm Hg), arterial pressure remaining below time control values for the remainder of the infusion (P = 0.006). Rises in both heart rate and PRA observed on the control day tended to be attenuated by CNP. Urine electrolyte response to CNP was similar to that observed with ANF. Compared with control infusions, the responses of both plasma aldosterone (P = 0.006) and cortisol (P = 0.043) were significantly different. Following CNP-induced hypotension, plasma cortisol and aldosterone increased abruptly at 30 min after which values fell to control or lower levels until the infusion was terminated. These studies show that intracerebroventricular CNP lowers arterial pressure without increasing heart rate and also suppresses the adrenocortical response, whereas ANF given under the same conditions has no significant effects. These data support the hypothesis that CNP plays an important role in the central regulation of blood pressure and hormones.

Lack of direct effect of dopamine on aldosterone secretion in vivo.

Previous work suggests that aldosterone is modulated by dopamine, which exerts an inhibitory effect at the level of the adrenal cortex. This study reports the effect of dopamine on aldosterone secretion in conscious sheep with cervical adrenal transplants in whom endogenous ACTH secretion was suppressed by dexamethasone. In control experiments (n = 7) local adrenal infusions of angiotensin II (AII) (1.6 ng/min for 120 min) increased aldosterone secretion to peak levels (47.8 +/- 6.8 ng/min. mean +/- SEM) at 20 min, after which secretion fell to stable levels (20-28 ng/min) at 60-120 min. On separate days, sheep were restudied (n = 5) during systemic dopamine infusions (4 microgram/kg . min for 90 min), commencing 30 min before AII stimulation. There was no significant difference, either in the pattern or the sensitivity of the aldosterone response to AII, with dopamine infusions. Large intraadrenal infusions of dopamine (10 microgram/min) also failed to alter the aldosterone response to AII. The possibility that aldosterone was already under maximum tonic inhibition by dopamine was studied in four additional experiments using the dopamine blocking drug, metoclopramide. Although the systemic (iv) administration of metoclopramide increased aldosterone in both intact and transplanted sheep, local infusions of metoclopramide (0.5-15 microgram/min intraarterially) had no consistent effect on the aldosterone response to AII, and the addition of dopamine during metoclopramide infusions also had no effect. These results indicate that local (adrenal) dopaminergic mechanisms play little or no part in the regulation of aldosterone secretion in the sheep. The mechanism whereby aldosterone secretion is increased by systemic metoclopramide remains to be explained.

The sites of gene expression of atrial, brain, and C-type natriuretic peptides in mouse fetal development: temporal changes in embryos and placenta.

Atrial (ANP), brain (BNP), and C-type natriuretic peptide (CNP) belong to a family of hormones important in blood pressure and sodium homeostasis. Expression of ANP has been reported in embryo hearts, but BNP and CNP expression during development has not been described. We used in situ hybridization to identify the sites of gene expression of ANP, BNP, and CNP during development in mouse embryos at daily intervals from midgestation. Very intense expression of ANP and BNP was visible in the heart from 9.5 days gestation; levels of expression of both peptides in the ventricle exceeded those in atria throughout gestation. There was a major peak of atrial and ventricular ANP and BNP expression at 12.5 days, attaining levels similar to those in adult heart and then declining until birth. Cardiac expression of CNP was undetectable. Expression of ANP and CNP was also observed in distinct sites in the brain, and all three peptides were expressed in the spinal cord. In mouse placenta, strong CNP expression was seen in the decidua basalis around the large maternal blood vessels, and BNP message was detected at the peripheral margin of the decidual layer. This pattern of expression indicates that ANP and CNP are present during development of the mouse central nervous system and suggests that CNP and BNP participate in regulating the maternal blood supply to the developing embryo.

Intracerebroventricular atrial natriuretic factor (ANF) antiserum inhibits volume-induced ANF in sheep: evidence for the brain's regulation of ANF secretion.

Plasma volume expansion stimulates cardiac secretion of atrial natriuretic factor (ANF) and also increases the ANF concentration in cerebrospinal fluid. In order to determine whether brain ANF is involved in the compensatory response to hypervolemia or the regulation of cardiac secretion of ANF, we have studied the integrated hemodynamic, renal, and hormonal response to acute volume expansion (15 ml/kg Dextran over 30 min) in five sheep given nonimmune serum (control) and ANF antiserum by intracerebroventricular (icv) injections on separate days. Dextran loading caused similar decreases in hematocrit and increases in central venous and mean arterial pressures on both study days. Heart rate was higher after antiserum injections (P less than 0.05). Dextran loading increased plasma ANF on the control (20 pmol/liter maximal mean increment above baseline) but not on the antiserum day (P less than 0.01). The diuresis (P less than 0.01) and natriuresis (P less than 0.05) observed on the control day was inhibited by icv antiserum. Plasma aldosterone and cortisol levels showed similar falls in response to the dextran load on both days. These experiments show that icv ANF antiserum inhibits both the increase in cardiac secretion of ANF and the renal response to plasma volume expansion without affecting hemodynamic status. These data support the hypothesis that the brain ANF system is important in the systemic responses to volume loading.

Hormone and hemodynamic responses to atrial natriuretic peptide in conscious sheep and effect of hemorrhage.

The effect of rat atrial natriuretic peptide (ANP) on basal hemodynamic and hormonal function and on the response to acute hemorrhage was studied in eight conscious sheep. ANP infusions (3 micrograms/kg BW bolus, followed by 50 ng/kg.min for 70 min) increased plasma immunoreactive ANP levels from less than 12 pmol/liter to steady state levels of 523 +/- 20 pmol/liter, reduced arterial pressure by 14% (P less than 0.002), increased heart rate by 26% (P less than 0.06), and increased plasma norepinephrine levels (P less than 0.015) compared to control values. These changes were associated with a significant increase in plasma cortisol (P less than 0.05) and smaller increases in plasma ACTH and arginine vasopressin (AVP), but plasma angiotensin II (AII) and aldosterone were unaffected. When hemorrhage (15 ml/kg BW over 10 min) was performed during ANP or control infusions, hypotension was greater (P less than 0.0004) during ANP treatment and the responses of plasma ACTH, AVP, catecholamines, and AII were enhanced compared with those to control hemorrhage. Plasma immunoreactive ANP during ANP infusions was significantly higher after hemorrhage (mean, 833 +/- 46; P less than 0.003), but the disappearance rate after the termination of ANP infusion was the same (3.1 min) with or without hemorrhage. These studies show that ANP infusions, achieving plasma levels observed in pathological states such as congestive heart failure, inhibit the expected responses of plasma AII and aldosterone to mild acute hypotension, but do not inhibit the responses of plasma AVP, ACTH, AII, and aldosterone associated with acute moderate hemorrhage in conscious sheep.

Stress hormones in blood and cerebrospinal fluid of conscious sheep: effect of hemorrhage.

Acute moderate hemorrhage (15 ml/kg withdrawn over 10 min) was used to study stress hormone changes in blood and cerebrospinal fluid (CSF) of conscious sheep with chronic indwelling intracerebroventricular catheters. Mean plasma arginine vasopressin (AVP) and ACTH rose 150- and 14-fold, respectively, above basal values to peak levels at 20 min after onset of hemorrhage. A smaller (4- to 5-fold) rise occurred in plasma angiotensin II (AII) to peak levels at 10 min. The corticosteroid response (cortisol and aldosterone) occurred later (peak at 45 min) and was consistent with the dependence of these steroids on plasma ACTH and AII changes. Increases in plasma epinephrine and norepinephrine were small and transient. Compared to changes in plasma, changes in CSF hormone levels after hemorrhage were small and independent of plasma concentrations. Mean CSF AVP increased to peak levels at 15 min whereas rises in CSF ACTH, AII-like immunoreactivity, and cortisol were slower and delayed in comparison with the patterns observed in plasma. Despite evidence of increased sympathetic activity, and rise in plasma catecholamines, CSF epinephrine fell after hemorrhage and CSF norepinephrine did not change. These results show that in conscious sheep rapid and major increases in plasma AVP, ACTH, and AII follow acute moderate hemorrhage. Concomitant changes in CSF hormone levels are small and delayed. With the possible exception of AVP it appears unlikely that the acute systemic hormone response to hemorrhage is determined by hormone changes in CSF.

Atrial (ANP) and brain natriuretic peptide (BNP) expression after myocardial infarction in sheep: ANP is synthesized by fibroblasts infiltrating the infarct.

Cardiac gene expression of atrial natriuretic peptide (ANP) and that of brain natriuretic peptide (BNP) are markedly elevated after myocardial infarction. The cellular distribution and temporal responses of ANP and BNP messenger RNA (mRNA) expression were compared by in situ hybridization for 5 weeks after left coronary artery ligation in sheep. Ligation resulted in highly reproducible, transmural, left ventricular infarcts. Within the infarct, ANP mRNA appeared from 7 days after ligation, whereas BNP expression was undetectable in the infarct at any time. The cells synthesizing ANP were shown by in situ hybridization and immunocytochemistry to be fibroblasts invading the infarct. The appearance of ANP expression coincided with the transition of these cells to the myofibroblast phenotype. Treatment of cultured cardiac fibroblasts with transforming growth factor-beta (10 ng/ml) induced the expression of alpha-smooth muscle actin, characteristic of the transformation to myofibroblasts, and raised ANP concentrations in the medium. In the surviving myocardium of the left ventricle, ANP and BNP expression increased in response to ligation, BNP mRNA was particularly strong at the lateral margins of the infarct. In both left and right atria, levels of BNP mRNA increased markedly over the first 18 h, whereas levels of atrial ANP mRNA decreased over 3 days after infarction. This is the first report of ANP expression and synthesis by cardiac fibroblasts invading the fibrotic scar, suggesting that ANP may be involved in regulating fibroblast proliferation during reparative fibrosis.

The hormonal actions of corticotropin-releasing factor in sheep: effect of intravenous and intracerebroventricular injection.

The 41-residue ovine corticotropin releasing factor (CRF) was administered iv and intracerebroventricularly (icv) to merino sheep. A significant rise in plasma ACTH, beta-lipotropin (beta LPH) and cortisol was demonstrated after the administration of 200 micrograms, iv. A highly significant correlation between the increments in plasma ACTH and beta LPH was observed. The plasma ACTH rise was evident within 5 min and was abolished by the prior administration of 0.4-4.0 mg dexamethasone. No significant rise in plasma GH, LH, PRL, insulin, glucagon, pancreatic polypeptide, met-enkephalin, angiotensin II, aldosterone, or vasopressin could be demonstrated. Although smaller doses of CRF (50 ng to 5 micrograms) were effective when given icv, the ACTH response was more delayed. It is concluded that CRF stimulates a rapid increase in the secretion of ACTH and beta LPH in sheep. Suppression of this response by dexamethasone indicates that glucocorticoids are capable of acting on the pituitary to inhibit the ACTH response to CRF. The delayed response when CRF is given icv may be due to diffusion. The action of CRF appears to be relatively specific, in that the plasma concentrations of the other pancreatic, pituitary, and adrenal hormones measured were not affected.

The plasma cortisol and corticotropin response to hypoglycemia following adrenal steroid and ACTH administration.

The plasma cortisol response to hypoglycemia is widely used as a test of hypothalamic-pituitary-adrenal function. It was the aim of this study to determine whether this test gives a reliable indication of pituitary corticotropin (ACTH) release in patients recovering from adrenocortical suppression due to corticosteroid or ACTH therapy. The 16 patients who were studied (6 on more than one occasion) had received in excess of 5 mg predinisone or equivalent daily for over 12 months. The insulin tolerance tests were carried out 18 h after stopping steroid therapy. The tests were then repeated three to four days after adrenal function had been restored (as indicated by urinary oxogenic steroid excretion of greater than 35 mg/24 h) by zinc tetracosactrin administration. The ACTH response to hypoglycemia was significantly impaired in the steroid-treated group. However with the exception of one patient who had persistently elevated resting ACTH levels there was a significant correlation (P less than 0.01) between the maximum increments in plasma cortisol and ACTH during hypoglycemia. No significant difference in sensitivity to endogenous ACTH could be demonstrated between the steroid-treated group and 12 normal control subjects. Following ACTH administration the plasma ACTH and growth hormone responses to hypoglycemia were significantly reduced, but the response in plasma cortisol was not significantly affected. It is concluded that the plasma cortisol response to hypoglycemia gives a useful indication of ACTH release in steroid-treated patients provided that they have not recently received exogenous ACTH.

Atrial natriuretic factor in hypertension: bioactivity at normal plasma levels.

To ascertain whether small shifts in plasma atrial natriuretic factor (ANF) exerted biological effects in hypertension, we studied the renal, hemodynamic, and hormonal effects of ANF [human ANF-(99-126)] infused at a dose (0.75 pmol/kg/min for 3 hours) that would induce changes in plasma ANF confined to the normal, resting range, in a group of six young men with uncomplicated, mild essential hypertension. During ANF infusions, the patients excreted 11.8 +/- 2.0 mmol (mean +/- SEM) sodium more than during the time-matched placebo phase natriuresis (p less than 0.001, mean increase of 53% above placebo values). Urinary excretion of cyclic guanosine monophosphate rose to more than double (212%, p less than 0.001) placebo values. Plasma renin activity (0.4 +/- 0.05 vs. 0.9 +/- 0.12 nmol/l/hr, p less than 0.0001) and aldosterone concentrations (102 +/- 4 vs. 184 +/- 47 pmol/l, p less than 0.05) were clearly suppressed during administration of ANF. Plasma norepinephrine also fell significantly below placebo values (268 +/- 17 vs. 439 +/- 35 pg/ml, p less than 0.05). Urine volume, the excretion of electrolytes other than sodium, hematocrit, effective renal plasma flow, glomerular filtration rate, and filtration fraction were unaffected by ANF. Similarly, plasma concentrations of epinephrine, arginine vasopressin, adrenocorticotropic hormone, and cortisol were unchanged. Blood pressure and heart rate were unchanged. Minor perturbations in plasma ANF concentrations exert clear biological effects in patients with mild essential hypertension. These data suggest that such minor shifts in plasma ANF are of physiological relevance in mild hypertension and probably contribute to volume homeostasis in this condition.