Alcohol-mediated Purkinje cell loss in the absence of hypoxemia during the third trimester in an ovine model system.

BACKGROUND: Although the mechanisms that underlie fetal alcohol-induced neuronal loss have not been determined, hypoxia/hypoxemia has been considered a leading candidate. This study was designed to test the hypothesis that neuronal loss could occur in the developing brain in the absence of fetal hypoxemia. METHODS: Three groups of pregnant sheep were used: a control group, a binge-drinking group, and a pair-fed group. The alcohol and pair-fed animals were anesthetized on day 113 of pregnancy, and the mothers and fetuses were instrumented with arterial and venous catheters. All animals were killed on day 133. Stereological cell counting techniques were used to estimate the total number of Purkinje cells in the fetal cerebellum. RESULTS: Peak maternal and fetal blood alcohol concentrations did not produce fetal hypoxemia. Nevertheless, there was a 25% loss of Purkinje cells of the cerebellum in the alcohol-exposed fetuses compared with that in the pair-fed controls. The loss of neurons was not accompanied by microencephaly or a concomitant decrease in either cerebellar weight or volume of the fetal cerebellum. CONCLUSIONS: Neuronal loss can be observed after alcohol exposure during the third trimester equivalent in fetal sheep in the absence of alcohol-induced hypoxemia. Furthermore, cell loss in the absence of deficits in gross brain weight or regional brain volume indicates that the lack of gross brain volume deficits from magnetic resonance imaging techniques is not a reliable indication that the brain is unaffected by the alcohol exposure.

Fetal and maternal sheep hypothalamus pituitary adrenal axis responses to chronic binge ethanol exposure during the third trimester equivalent.

BACKGROUND: We tested the hypothesis that in utero ethanol exposure results in changes in fetal and maternal adrenocorticotropin (ACTH) and cortisol during the third trimester equivalent, by using a chronically instrumented fetal sheep model. METHODS: Pregnant ewes received saline or ethanol intravenously 3 consecutive days per week from day 109 to day 132 of gestation. Fetal and maternal blood samples were collected on days 118 and 132. RESULTS: Maternal and fetal ACTH and cortisol values increased on days 118 and 132 of gestation in response to ethanol infusions that created blood ethanol concentrations (BECs) that are easily achievable by human drinkers. Peak ACTH and cortisol values were detected 30 to 60 min after peak BECs were achieved. CONCLUSIONS: Chronic ethanol exposure during the third trimester equivalent in sheep resulted in repeated activation of the hypothalamus-pituitary-adrenal axis in both the mother and fetus. Temporally, the patterns of maternal and fetal responses to ethanol infusion were similar. We conclude that ovine maternal ethanol exposure during the third trimester equivalent increases fetal ACTH and cortisol concentrations, hormonal responses that may play a role in mediating alcohol-related birth defects.

Third trimester binge ethanol exposure results in fetal hypercapnea and acidemia but not hypoxemia in pregnant sheep.

BACKGROUND: The mechanisms by which maternal ethanol abuse during pregnancy causes neurodevelopmental injury in the fetus are not well understood. The purpose of this study was to use a chronically instrumented fetal sheep model system to determine if a binge pattern of ethanol exposure administered throughout the third trimester reduced fetal arterial partial pressure of oxygen (PaO2); a positive finding would support the hypothesis that fetal hypoxemia may play a role in mediating ethanol-related birth defects. METHODS: Pregnant ewes received saline or 0.75, 1.25, 1.5, or 1.75 g/kg of ethanol intravenously over 1 hr beginning on day 109 of gestation (term = 145 days) for 3 consecutive days per week followed by 4 days without exposure. The fetuses were surgically instrumented on day 113, and experiments were performed on days 118 or 132, the 6th and the 12th ethanol exposure, respectively. RESULTS: Ethanol infusions resulted in peak blood ethanol concentrations of 80.8 +/- 6.5, 182.5 +/- 13.5, 224.4 +/- 13.9, and 260.6 +/- 20.0 mg/dl +/- SEM (maternal) and 70.0 +/- 5.9, 149.7 +/- 9.0, 216.9 +/- 14.0, and 233.3 +/- 19.8 mg/dl +/- SEM (fetal) in response to the 0.75, 1.25, 1.5, and 1.75 g/kg doses, respectively. Maternal and fetal heart rate and maternal blood pressure increased whereas fetal blood pressure decreased in a dose-dependent manner in response to ethanol infusions. Maternal and fetal arterial pH decreased and arterial partial pressures of carbon dioxide increased in response to ethanol infusions. Maternal PaO2 decreased whereas fetal PaO2 did not change in response to ethanol infusions. CONCLUSIONS: A binge ethanol exposure paradigm, three consecutive days per week throughout the third trimester at ethanol doses that created blood ethanol concentrations commonly achieved by human ethanol abusers, resulted in changes in maternal and fetal heart rate, changes in blood pressure, hypercapnea, acidemia, and maternal, but not fetal, hypoxemia. We conclude that in an ovine model system, ethanol doses that create blood ethanol concentrations as high as 260 mg/dl do not result in fetal hypoxemia. Remaining issues to address with this model system are whether neurodevelopmental injuries that are associated with maternal ethanol abuse are mediated by a reduction in fetal cerebral blood flow, fetal hypercapnea, or acidemia.

Brain high energy phosphate responses to alcohol exposure in neonatal rats: an in vivo 31P-NMR study.

BACKGROUND: The mechanisms that mediate fetal brain injury which results from maternal alcohol consumption are not well understood. Although fetal hypoxia is a popularly proposed mechanism, it has been difficult to assess brain oxygenation in vivo. We measured intracellular high energy phosphate concentrations and estimated intracellular pH (pHi) in brains of unanesthetized neonatal rat pups by using in vivo 31P-NMR spectroscopy. We reasoned that decreases in brain oxygenation sufficient to result in brain injury would also reduce high energy phosphates and pHi. METHODS: On postnatal day 4, before alcohol administration, pups were placed into a 20 mm diameter NMR probe, their heads were positioned carefully in the center of the 31P detection coil, and spectra were collected over 20 min. Animals were then fed diet with or without 4.5 g/kg of ethanol in two (in succession) of 12 daily feedings via artificial rearing methods. A second spectrum was collected at 90 min after the beginning of the second alcohol feeding, at the time that coincided with the peak blood alcohol concentration (BAC). Identical feedings were performed daily until day 9, when pre- and postfeeding spectra were again obtained. Positive control groups were fed control diet and were studied in atmospheres of 5% oxygen, 95% nitrogen or 0% oxygen, 100% nitrogen. RESULTS: Phosphocreatine (PCr), beta-adenosine triphosphate (ATP), and pHi decreased and inorganic phosphate (Pi) increased in day 4 animals subjected to 0% oxygen (20 min) compared with pretreatment and all other treatment groups. Day 9 animals did not tolerate these conditions. There were no significant changes in response to 5% oxygen on day 4, but Pi increased and beta-ATP decreased compared with pretreatment values and compared with alcohol and control groups on day 9. There were no changes in PCr, beta-ATP, or pHi in response to alcohol treatment at either age. PCr was significantly increased in the alcohol and 5% oxygen groups and apparently increased in the control group on day 9 compared with day 4, most likely due to increases in cranial muscle mass within the NMR coil. CONCLUSIONS: We conclude that acute alcohol exposure that results in peak BACs of 315 mg/dl does not alter brain high energy phosphate concentrations or pHi in neonatal rat pups, although these BACs are known to result in significant brain injury. These findings do not support hypoxia as a mechanism of alcohol-mediated brain injury during the third trimester equivalent in the rat pup model.

Biomedical device design discovery team approach to teaching physiology to undergraduate bioengineering students.

Teaching effectiveness is enhanced by generating student enthusiasm, by using active learning techniques, and by convincing students of the value of acquiring knowledge in the area of study. We have employed a technique to teach physiology to bioengineering students that couples students' enthusiasm for their chosen field, bioengineering, with an active learning process in which students are asked to design a biomedical device to enhance, replace, or create a new cellular or organ system function. Each assignment is designed with specific constraints that serve to direct students' attention to specific areas of study and that require students to create original designs. Preventing students from using existing designs spurred student invention and enthusiasm for the projects. Students were divided into groups or "design discovery teams" as might be done in a biomedical device industry setting. Students then researched the physiological issues that would need to be addressed to produce an acceptable design. Groups met with faculty to brainstorm and to obtain approval for their general design concepts before proceeding. Students then presented their designs to the instructors in a structured, written outline form and to the class as a 10-minute oral presentation. Grades were based on the outline, oral presentation, and peer evaluations (group members anonymously rated contributions of other members of their team). We believe that this approach succeeded in generating enthusiasm for learning physiology by allowing the students to think creatively in their chosen field of study and that it has resulted in students developing a more thorough understanding of difficult physiological concepts than would have been achieved with a traditional didactic lecture approach.

Cardiovascular, adrenocorticotropin, and cortisol responses to hypertonic saline in euvolemic sheep are altered by prostaglandin synthase inhibition.

Small volume intravenous infusions of hypertonic saline (HTS) increase blood pressure, heart rate, adrenocorticotropic hormone (ACTH), and cortisol by mechanisms that are not fully understood. We hypothesized that HTS infusions increase prostaglandin biosynthesis and that a prostaglandin synthase metabolite is responsible for mediating actions of HTS. We further hypothesized that thromboxane A2 (TxA2) is the specific metabolite responsible for mediating responses to HTS infusion. Adult female sheep (n=8) were chronically instrumented with vascular catheters and infused intravenously with 7.5% saline at a rate of 4 mL x kg(-1) over 5 min with or without pretreatment with the prostaglandin synthase inhibitor flunixin. Blood pressure, ACTH, and cortisol increased in response to HTS, and these responses were prevented by flunixin. Heart rate increased in response to HTS infusion, and flunixin reduced but did not prevent a heart rate response. Hematocrit decreased significantly in response to HTS but only following flunixin treatment. Arginine vasopressin increased but only modestly in response to HTS, and responses were not different following flunixin. Arterial pH, partial pressure of CO2, and partial pressure of O2 did not change. Circulating concentrations of thromboxane B2, a stable metabolite of TxA2 and an index of TxA2 formation, remained low and did not change in response to HTS. We conclude that heart rate, blood pressure, ACTH, and cortisol responses to HTS are mediated at least in part by a product of prostaglandin synthase metabolism. These responses were not due to increases in circulating concentrations of TxA2 but might involve local formation of TxA2 or some other prostaglandin synthase metabolite.

Thromboxane A2 acts on the brain to mediate hemodynamic, adrenocorticotropin, and cortisol responses.

Conditions that increase the formation of thromboxane A2 (TxA2) also result in activation of hemodynamic and adrenocortical responses. The purpose of this study was to test the hypothesis that TxA2 acts directly on the brain to mediate these responses. Adult sheep were chronically instrumented with vascular and intracerebroventricular catheters. The TxA2 analog U-46619 (0, 100, or 1,000 ng.kg-1.min-1) and artificial cerebrospinal fluid (CSF) were infused intracerebroventricularly for 30 min. Heart rate increased in response to 100 ng.kg-1.min-1 U-46619 infusions. Heart rate did not change over preinfusion values in response to the highest infusion rate, but values were elevated compared with the postinfusion period. Mean arterial pressure, ACTH, cortisol, hematocrit, and arterial pH (pHa) increased, and arterial partial CO2 pressure (PaCO2) fell in response to 1,000 ng.kg-1.min-1 infusions of U-46619. Plasma vasopressin concentrations and arterial partial O2 pressure did not change. In a second study, U-46619 or artificial CSF was infused intracerebroventricularly during prostaglandin synthase blockade. Blockade reduced but did not prevent blood pressure responses to U-46619 infusion, suggesting that the U-46619 infusions increased prostaglandin synthase metabolism to contribute de novo TxA2 or a second metabolite to augment the blood pressure response. Heart rate, pHa, PaCO2, ACTH, and cortisol responses to U-46619 were not different with blockade. We conclude that TxA2 acts on the brain to mediate blood pressure, heart rate, pHa, PaCO2, hematocrit, ACTH, and cortisol responses. These findings support the hypothesis that TxA2 acts directly on the brain to promote cardiovascular and hormonal responses that may serve a protective function during conditions when TxA2 formation is increased.

Thromboxane A2 does not act at the carotid sinus to mediate cardiovascular, adrenocorticotropin, cortisol, or blood gas responses.

Thromboxane A2 (TxA2), well known as a vasoconstrictor and activator of platelets, also stimulates reflex cardiovascular, pituitary, adrenocortical, and blood gas responses, although the site of action is unknown. Previously we determined that the site of these actions is perfused by the carotid vasculature. The purpose of the present study was to test the hypothesis that TxA2 stimulates these responses by acting at the carotid sinus. The TxA2 mimetic U46619 (1 microgram.kg-1.min-1) or saline was infused into the carotid artery (CA) or vena cava in conscious, chronically instrumented carotid sinus denervated (CSD) or sham-operated sheep. Mean arterial pressure increased in all groups receiving U46619. Heart rate increased only in the CSD group receiving CA infusions of U46619. Adrenocorticotropic hormone (ACTH) and cortisol increased in the sham and CSD groups receiving CA U46619, and responses were not different between sham and CSD groups. PaCO2 values were higher in all CSD treatment groups compared with sham treatment groups. Arterial pH increased and PaCO2 decreased in both the sham and CSD groups in response to CA U46619. Although PaCO2 values were higher overall in the CSD group, the magnitude of change in response to U46619 infusions was similar in sham and CSD animals. There was no difference in pHa between CSD and sham groups. Hematocrit and PaO2 did not change. We conclude that TxA2 does not act at the carotid sinus, as responses to U46619 infusions in CSD animals were not different in the cases of ACTH, cortisol, and blood gases, or were enhanced rather than diminished in the case of heart rate. These findings support a hypothesis that TxA2 acts at the brain to mediate cardiovascular, pituitary, adrenocortical, and blood gas responses.

Fetal pulmonary immunoreactive adrenocorticotropin: molecular weight and cellular localization.

The hypothalamus-pituitary-adrenal axis of the sheep fetus plays a critical role in fetal development, responsiveness to stress, and initiation of parturition. We have recently reported that the fetal lung contains and secretes significant amounts of immunoreactive adrenocorticotropin (iACTH). The present study was designed to identify the molecular weight profile and the cellular location of iACTH in this tissue. iACTH extracted from fetal lung was immunoprecipitated, electrophoresed, and immunoblotted. Pulmonary iACTH was found in several molecular forms. The largest peptides appeared as doublets, and had molecular weights similar to POMC (32, 33 kD). Smaller peptides appeared in molecular weights (17, 24, and 27 kD) which were not consistent with the post-translational processing of POMC in fetal pituitary, but which were consistent with known processing of POMC by chromaffin granule aspartyl protease. None of the molecular forms of iACTH were glycosylated. Immunohistochemistry revealed that the iACTH was contained within bronchial epithelium and within groups of cells within the parenchyma of the lung. Both of these types of cells are consistent with pulmonary neuroendocrine cells. The distribution of neuroendocrine cells and apparent concordance with the iACTH-positive cells was confirmed by immunostaining for neuron specific enolase, a marker for neuroendocrine cells within the lung. We conclude that the lung contains unprocessed and partially processed POMC within cells known to contain neuropeptides. We speculate that secretion of the POMC-related peptides from these cells is physiologically important in the late-gestation fetus.

Sheep model for study of maternal adrenal gland function during pregnancy.

Our goal was to develop a model for the study of maternal adrenal gland regulation and the effects of maternal cortisol secretion on fetal homeostasis. At about 108 days of gestation, before the time of rapid fetal growth or fetal adrenocortical maturation, ewes, under halothane anesthesia with controlled ventilation and positioned in sternal recumbency, were adrenalectomized. Ewes were treated with aldosterone by intravenous infusion (3 micrograms/kg of body weight per day) to induce normal late-gestation aldosterone concentration. Ewes were also treated with cortisol; for 2 postoperative days, this infusion (1 to 2 micrograms/kg per min) induced plasma concentration similar to that associated with stress. Thereafter, the dosage of cortisol was reduced to induce plasma values similar to normal late-gestation cortisol concentration in ewes (1 mg/kg per day), or to values in nonpregnant ewes (0.6 mg/kg per day). Administration of cortisol and aldosterone was required to prevent electrolyte imbalance and signs of hypoadrenocorticism. With steroid replacement, plasma protein, electrolyte, and glucose concentrations in adrenalectomized ewes were not different from those in sham-operated pregnant ewes. Of 11 adrenalectomized ewes, one died as a result of failure of the infusion pump, and one died as a result of inappropriate treatment for hypoglycemia. Of the remaining ewes, two aborted fetuses, three ewes each delivered one live and one dead fetus, two delivered live singleton fetuses, and two delivered twins. Therefore, this model of relative hypoadrenocorticism in pregnancy is feasible and practical for studying the influence of maternal cortisol concentration on maternal and fetal homeostasis.

Vagal blockade does not prevent adrenocorticotropin, cortisol, and cardiopulmonary responses to thromboxane A2.

Previously, we reported that thromboxane A2 (TxA2) mediates heart rate, adrenocorticotropin (ACTH), cortisol, and blood gas responses, although the specific site of action was not identified. In the present study, we interrupted vagal nervous transmission in chronically instrumented conscious sheep and infused the TxA2 mimetic U46619 or saline into the carotid artery or U46619 into the vena cava to determine whether TxA2 acts at vagal afferent nerves. Heart rate increased in all three groups during vagal blockade, and responses were not different between groups. Carotid artery and intravenous infusions of U46619 resulted in an increase in blood pressure, but responses were not different between groups. PaO2 decreased in response to vagal blockade in all groups, and responses were not different among groups. Arterial pH increased and PaCO2 decreased during vagal blockade in response to carotid artery U46619 infusions but not in response to vagal blockade alone or combined with carotid artery saline or intravenous U46619. ACTH, cortisol, and hematocrit increased significantly in response to carotid artery infusions of U46619 during vagal blockade but not in response to carotid artery saline or intravenous U46619 infusions. In summary, carotid artery infusions of TxA2 mimetic result in ACTH, cortisol, PaCO2, pHa, and hematocrit responses that are not prevented by vagal blockade. We conclude that these responses are mediated at a site perfused by the carotid vasculature and not at a site innervated by the vagal nerves, findings consistent with the hypothesis that TxA2 acts on the brain to mediate cardiopulmonary and pituitary-adrenal responses.

Immunoreactive thromboxane synthase is measurable in ovine fetal hypothalamus as early as 86 days' gestation.

Thromboxane A2 (TxA2) augments hypothalamus-pituitary-adrenal axis activity in both fetal and adult animals. We have proposed that TxA2 acts as a neuromodulator within the brain to stimulate the release of corticotropin releasing hormone (CRH) or arginine vasopressin (AVP) into the hypophyseal-portal blood. We performed the present experiments to identify immunoreactive thromboxane synthase (TxS) within fetal brain regions and to quantify developmental changes in the TxS immunoreactivity measurable within those regions. We found that immunoreactive TxS was present in fetal hypothalamus, pituitary, brainstem, and lung. In fetal hypothalamus, we found immunoreactive TxS in three identifiable molecular weights, approximately 65, 42, and 35 kD. In fetal pituitary and lung, we found the 65 and 35 kD forms, and in the brainstem we found only the 35 kD form. In fetal pituitary, there was a clear ontogenetic change in TxS immunoreactivity. The 42 kD TxS immunoreactivity was not present in the youngest fetal sheep studied (86-90 days' gestation), but was expressed in the other age groups (125-128, 135-139, 141-term, and postnatal ages). The other molecular weight forms appeared to increase in the older fetuses, but the changes were not significant. In the hypothalamus, all three forms of TxS were measurable at all ages, and there was no significant change in relative abundance. We conclude that immunoreactive TxS is present in the fetal brain throughout the last half of fetal gestation, but that the significance of multiple molecular weight forms is not clear.

Thromboxane A2 acts at a site perfused by the carotid vasculature to mediate cardiovascular and adrenocortical responses.

Increases in thromboxane A2 (TxA2) synthesis are associated with hemodynamic responses and activation of the hypothalamus-pituitary-adrenal axis. This study tested the hypothesis that TxA2 acts on a site perfused by the carotid vasculature to mediate these responses. The TxA2 mimetic U46619 was infused for 30 min into the carotid artery or the vena cava of chronically instrumented adult sheep at doses of 0, 0.25, 0.5, and 1.0 microgram.kg-1.min-1. Mean arterial pressure increased in response to carotid or vena cava U46619 infusions of 0.5 and 1.0 microgram.kg-1.min-1. Heart rate was elevated in response to carotid (0.5 and 1.0 microgram.kg-1.min-1) or vena cava (1.0 microgram.kg-1.min-1) infusions of U46619. Paco2 declined and pH2 increased significantly in response to carotid infusions of 0.5 and 1.0 microgram.kg-1.min-1 but did not change in response to vena cava infusions. Adrenocorticotropic hormone (ACTH) increased in response to carotid infusions of 0.5 microgram.kg-1.min-1, while cortisol increased in response to infusions of 0.25, 0.5, and 1.0 microgram.kg-1.min-1. ACTH and cortisol did not change in response to vena cava infusions. Pao2, hematocrit, and arginine vasopressin did not change significantly. Pulmonary artery pressure and total peripheral resistance increased while cardiac output decreased in response to carotid or vena cava U46619 infusions of 1 microgram.kg-1.min-1; carotid and vena cava responses were not different from one another. We conclude that increases in blood pressure are mediated by peripheral PGH2/TxA2 receptor activation and that pituitary adrenal, blood gas, and heart rate responses are mediated by PGH2/TxA2 receptor activation at a site perfused by the carotid vasculature.

Acute hemodynamic, pituitary, and adrenocortical responses to alcohol in adult female sheep.

Alcohol was infused intravenously into chronically cannulated adult female sheep as a 40% solution (w/v) at doses of 0.5, 1.0, 1.5, or 2.0 g/kg over 1 hr. Saline infusions, equal in volume to the highest dose, served as a control. Dose-dependent peak blood alcohol concentrations (BACs) were attained 60 min after the beginning of alcohol infusion for all doses (86.5 +/- 3.7, 213.2 +/- 11.0, 373.2 +/- 14.3, and 494.1 +/- 34.5 mg/dl +/- SE, respectively). Plasma cortisol concentrations increased in response to the 0.5 g/kg infusions (BACs less than 100 mg/dl), whereas both ACTH and cortisol concentrations increased in the 1.0 and 2.0 g/kg dose groups. Mean arterial pressure, heart rate, and Paco2 increased, whereas Pao2 decreased in response to the 1.5 and 2.0 g/kg infusions. Arterial pH declined in the highest dose group. Respiratory rate was lower in all groups receiving alcohol compared with that of the control group. Hematocrit did not change. We conclude that BACs in adult female sheep below 100 mg/dl (levels easily achieved by social drinkers) result in activation of the hypothalamus-pituitary-adrenal axis. At high BACs (> 350 mg/dl), pituitary adrenal responses are accompanied by increases in heart rate, blood pressure, and Paco2 and decreases in Pao2 and arterial pH. These findings support the hypothesis that alcohol acts directly on the brain to mediate pituitary adrenal responses and that the additional responses to high BACs (the blood gas and hemodynamic responses), might be mediated by direct actions of alcohol on the brain, by cerebral ischemia, or by alcohol-mediated suppression of ventilatory drive and hypoxemia.

Does thromboxane mediate the fetal ACTH response to acidemia?

Intravenous mineral acid infusions into fetal sheep stimulate increases in plasma adrenocorticotropic hormone (ACTH) and cortisol concentrations that correlate to the induced changes in arterial pH (pHa). We have recently demonstrated that ACTH and cortisol responses to mineral acid infusion in adult sheep are mediated by thromboxane A2 (TxA2). We designed the present experiments to test the hypothesis that fetal ACTH and cortisol responses are also mediated by TxA2. We infused chronically instrumented fetal sheep with 1 N HCl (0.5 ml/min i.v.) for 60 min, with or without pretreatment with the cyclooxygenase inhibitor flunixin-N-methylglucamine. HCl infusion significantly decreased pHa and significantly increased the arterial partial pressure of O2 (PaO2) and CO2 (PaCO2). Flunixin pretreatment significantly decreased fetal plasma thromboxane B2 (TxB2) concentrations but did not significantly alter the blood gas and pH response to HCl. TxB2 is a stable metabolite of TxA2 and was measured as an index of TxA2 generation. HCl increased fetal heart rate only in the flunixin group. Plasma ACTH and cortisol concentrations were increased significantly in both groups; flunixin did not significantly alter the responses. HCl infusion did not significantly alter plasma TxB2 concentrations. We conclude that the fetal ACTH and cortisol responses to HCl infusion are not mediated by TxA2 or other prostanoids whose synthesis depends on cyclooxygenase activity.

Secretion and clearance of immunoreactive ACTH by fetal lung.

We have previously reported the presence of immunoreactive adrenocorticotropic hormone (irACTH) in fetal and adult ovine lung and secretion of the irACTH into the pulmonary venous blood during surgical stress. The aim of the present experiments was to determine whether the fetal lung secretes irACTH during less stressful conditions. Seven fetal sheep were chronically prepared with systemic, pulmonary arterial, and pulmonary venous catheters. After recovery from surgery, paired samples were obtained from the pulmonary arterial and venous catheters. In 51 pairs of samples, we quantified the arteriovenous (A-V) difference in plasma irACTH concentration. Pulmonary A-V difference in plasma irACTH was significantly related to the arterial plasma concentration of irACTH but not to fetal blood gases, pH, or gestational age. Regression analysis revealed that, at arterial plasma concentrations of irACTH less than approximately 200 pg/ml, the lung releases irACTH into plasma and that, at higher concentrations, the lung clears irACTH from the plasma. We conclude that the lung independently secretes and clears irACTH into and from plasma.

Ontogeny and ultradian rhythms of adrenocorticotropin and cortisol in the late-gestation fetal horse.

Fetal maturation and the timing of parturition in both sheep and primates are thought to be controlled by the hypothalamic-pituitary-adrenal axis but little is known about the endocrinology of the equine fetus. We investigated the ontogeny of plasma concentrations of adrenocorticotropic hormone (ACTH), cortisol and corticosteroid binding capacity in the late-gestation fetal horse. We also wished to determine whether there is ultradian rhythmic release of ACTH and cortisol in fetal horses and we compared fetuses to maternal and non-pregnant adult horses. Six fetuses, 278-304 days gestation (term approximately 335), were catheterized and sampled daily until delivery. Mean (+/- S.E.M.) ACTH concentrations increased significantly from 159 +/- 21 to 246 +/- 42 pg/ml over the last 2 days before parturition. Fetal cortisol increased significantly from 3.1 +/- 1.0 to 13.4 +/- 3.7 ng/ml (mean +/- S.E.M.) over the last 9 days before delivery. The slope of regressions for ACTH and cortisol concentrations with respect to time were positive in all subjects and statistically significant in 3 of 6 for ACTH and 5 of 6 for cortisol. Fetal corticosteroid binding capacity declined from 49.5 +/- 20.5 to 16.1 +/- 2.2 ng/ml (mean +/- S.E.M.) over the last 10 days before parturition. However, the greatest changes in ACTH, cortisol and corticosteroid binding capacity occurred very late in gestation, during the last 48 to 72 h before parturition.(ABSTRACT TRUNCATED AT 250 WORDS)

Thromboxane A2 receptor antagonism prevents hormonal and cardiovascular responses to mineral acid infusion.

Small infusions of strong acid create large elevations in heart rate (HR), mean arterial pressure (MAP), adrenocorticotropic hormone (ACTH), cortisol, and thromboxane A2 (TxA2). We hypothesized that TxA2 is responsible for these hormonal and hemodynamic responses. Conscious sheep received HCl (1 N, 1 ml/min for 30 min) with or without receiving SQ-29548 [a TxA2/prostaglandin (PG) H2 receptor antagonist]. HCl increased TxB2 from 133 +/- 44 to 1,213 +/- 531 (SE) pg/ml while SQ-29548 + HCl increased TxB2 from 141 +/- 41 to 1,051 +/- 518 pg/ml. HCl decreased pH (7.464 +/- 0.015 to 7.413 +/- 0.011), arterial PCO2 (31.6 +/- 1.3 to 25.9 +/- 1.8 mmHg), and arterial PO2 (98.0 +/- 2.2 to 90.5 +/- 3.2 mmHg), and increased MAP (75 +/- 2 to 88 +/- 5 mmHg), HR (72 +/- 4 to 93 +/- 8 beats/min), hematocrit (25 +/- 1 to 29 +/- 2%), ACTH (154 +/- 41 to 549 +/- 217 pg/ml), and aldosterone (25 +/- 1 to 151 +/- 74 pg/ml) while these responses were prevented by SQ-29548. SQ-29548 reduced but did not prevent the cortisol response to HCl (9 +/- 2 to 23 +/- 10 ng/ml compared with 6 +/- 2 to 17 +/- 4 pg/ml after SQ-29548). K+ and aldosterone also increased after the end of SQ-29548 + HCl treatment (4.0 +/- 0.1 to 4.5 +/- 0.1 meq/l and 53 +/- 21 to 147 +/- 61 pg/ml, respectively). We conclude that TxA2 mediates the blood gas, MAP, HR, ACTH, and aldosterone responses to HCl infusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Content, in vivo release, and bioactivity of fetal pulmonary immunoreactive adrenocorticotropin.

We hypothesized that fetal lung contains and releases immunoreactive (ir) adrenocorticotropin (ACTH). Fetal sheep lung (89-145 days gestation, n = 13) irACTH content was 4,100 +/- 672 (SE) pg/g lung wet wt compared with 5,425 +/- 1,403 (SE) pg/g in adults (n = 5) when measured by radioimmunoassay (RIA). irACTH values normalized to protein content were 158 +/- 18 and 95 +/- 22 (SE) ng/g protein for fetuses and adults, respectively. Grouped by age, < 90 days (n = 4), 131-139 days (n = 5), and 143-145 days (n = 4), the values for irACTH were 184 +/- 28, 173 +/- 33, and 114 +/- 33 (SE) ng/g protein, respectively, and declined significantly from < 90 days to adulthood. RIA serial dilution curve slopes were different from standard in 7 of 13 fetuses and in all adults, suggesting that irACTH structure differs from ACTH-(1-39). Acute in vivo fetal pulmonary artery and vein sampling (n = 5, 120-128 days gestation) revealed a mean difference of -609 +/- 476 (SE) pg/ml, a statistically significant release of irACTH across the lung. Western blotting revealed that pulmonary irACTH migrates in multiple immunostaining bands between molecular weight 14,000 and 46,000 and does not colocalize with ACTH-(1-39). Adrenal cell bioassay revealed that pulmonary irACTH is not corticotropic. We concluded that ovine fetal lung contains large-molecular-weight irACTH, that content declines from 90 days gestation to adulthood, and that, under acute conditions, the fetal lung releases irACTH substances in sufficient quantity to contribute to circulating hormone concentrations of irACTH.