Innervation of the sheep adrenal cortex: an immunohistochemical study with rat corticotrophin-releasing factor antiserum

Using indirect immunohistochemistry and an antiserum raised against rat corticotrophin-releasing factor (CRF) we have outlined an asymmetric network of cells and varicose fibers in sheep adrenal cortex. This network was not associated with the larger splanchnic nerves, but was occasionally found in small bundles or with blood vessels; in most instances fibers were found weaving independently through cortical parenchyma. A plexus of fibers was found in the zona reticularis, with a few fibers ramifying into adjacent medulla. Uni or bipolar cells were found throughout the cortex, with the greatest frequency at the corticomedullary junction; a multipolar-type cell was also found in this area. Staining of varicose structures and most cells was abolished by incubation with excess rat CRF 1-41, but not by ovine CRF or a range of other peptides. Though the immunoreactive species has not as yet been identified, it may thus share homology with sequences present in rat but not ovine CRF.     

Stress-Related Changes in β-Endorphin Processing: The Limitations of Slaughterhouse Material

In the sheep, unlike many other species, a significant proportion (>25%) of immunoreactive β-endorphin in the anterior pituitary is post-translationally modified to opioid-inactive, α-N-acetylated forms. In a study to determine the precise molecular nature of α-N-acetylated β-endorphin immunoreactivity, we noted a striking difference in high-performance liquid chromatography profiles of anterior pituitary extracts between sheep killed on the farm, and age-, sex- and strain-matched slaughterhouse animals. These altered patterns of a-N-acetylated β-endorphin processing were reproduced in farm animals by chronic (≤ 4 days) treatment with the synthetic glucocorticoid dexamethasone; in contrast dexamethasone had no effect on a-N-acetylated β-endorphin processing in hypothalamo-pituitary disconnected sheep. These data suggest that (1) the change in processing is a stress response, mediated by prolonged glucocorticoid exposure, (2) this effect is central, rather than a direct effect on the pituitary, and (3) the relative abundance of various peptide sequences in slaughterhouse-derived material may not reflect their abundance under more physiological conditions.     

Turbulent stress measurements downstream of three bileaflet heart valve designs in pigs

Objective: Mechanical heart valves can cause thromboembolic complications, possibly due to abnormal flow patterns that produce turbulence downstream of the valve. The objective of this study was to investigate whether three different bileaflet valve designs would exhibit clinically relevant differences in downstream turbulent stresses. Methods: Three bileaflet mechanical heart valves (Medtronic Advantage^®, CarboMedics© Orbis™ Universal and St. Jude Medical^® Standard) were implanted into 19 female 90 kg pigs. Blood velocity was measured during open chest conditions in the cross sectional area downstream of the valves with 10 MHz ultrasonic probes connected to a modified Alfred^® Pulsed Doppler equipment. As a measure of turbulence, Reynolds normal stress (RNS) was calculated at three different cardiac output ranges (3–4, 4.5–5.5, 6–7 L/min). Results: Data from 12 animals were obtained. RNS correlated with increasing cardiac outputs. The highest instantaneous RNS observed in these experiments was 47 N/m², and the mean RNS taken spatially over the cross sectional area of the aorta during systole was between 3 N/m² and 15 N/m². In none of the cardiac output ranges RNS values exceeded the lower critical limit for erythrocyte or thrombocyte damage for any of the valve designs. Conclusions: Reynolds normal stress values were below 100 N/m² for all three valve designs and the difference in design was not reflected in generation of turbulence. Hence, it is unlikely that any of the valve designs causes flow induced damage to platelets or erythrocytes.     

Pro-opiomelanocortin messenger ribonucleic acid and posttranslational processing of beta endorphin in spleen macrophages.

We have previously demonstrated low levels of immunoreactive (ir)-beta-endorphin (beta-EP) and ir-ACTH in a subpopulation of mouse spleen macrophages, which is consistent with an involvement of opioid peptides in modulation of immune responses. Gel chromatography studies suggested the presence of an approximately 3.5,000-molecular weight (mol wt) species, putatively beta-EP, an approximately 11.5,000-mol-wt species, putatively beta-lipotropin, and a higher molecular weight species (putative beta-EP precursor, pro-opiomelanocortin (POMC). In this study we have extended our original findings by demonstrating the presence of messenger RNA for POMC by the use of a complementary DNA probe and Northern blot analysis of extracts of mouse and rat spleen. In addition, using high performance liquid chromatography (HPLC), we have shown that the major endorphin species in mouse spleen macrophages is beta-EP1-31, and that there are smaller amounts of each of the acetylated forms, N-acetyl-beta-EP1-16 (alpha-endorphin), N-acetyl-beta-EP1-17 (gamma-endorphin), N-acetyl-beta-EP1-27, and N-acetyl-beta-EP1-31. We interpret these studies as showing that (a) the spleen is an organ of POMC synthesis and that (b) the predominant COOH-terminal product of macrophage POMC is the opiate-receptor active species beta-EP1-31.     

Evidence that the central noradrenergic and adrenergic pathways activate the hypothalamic-pituitary-adrenal axis in the sheep.

These studies were undertaken to test the hypothesis that stimulation of the central noradrenergic and adrenergic pathways activates the hypothalamic-pituitary-adrenal axis in vivo in the conscious sheep. Blood samples were taken at 10-min intervals over 4 h to establish the baseline state, and then each animal received an intracerebroventricular (icv) injection of NaCl (control animals) or catecholamine [norepinephrine (NE) or epinephrine (EPI)]. A more frequent rate of venous sampling was used for the 30-min period after the icv injection, after which time the 10-min rate of blood sampling was continued for another 3.5 h. NaCl (n = 4) caused no change in pituitary-adrenal secretion. In contrast, 10 micrograms NE (n = 4) caused acute 1.9- and 3.2-fold increases in mean plasma ACTH and cortisol levels over the 1 h period post injection, and 1.6- and 2.3-fold increments in their concentrations over the 4 h postinjection period. Although 10 micrograms EPI (n = 4) did not elevate mean plasma ACTH, it produced significant 1.7- and 1.5-fold increases in plasma cortisol during the 1- and 4-h periods post injection. However, when 100 micrograms EPI was injected (n = 4), acute 9.5- and 5.5-fold increases in plasma ACTH and cortisol were seen over the 1 h period post injection, and 6.1- and 4.2-fold increments in their plasma concentration were noted during the entire post-injection period. To determine the predominant site of action of the catecholamines, we also examined the ability of NE and EPI to release ACTH from cultured ovine anterior pituitary cells. NE and EPI (10(-9)-10(-6) M) stimulated the release of ACTH in a dose-dependent manner, but with maximal increments only 1.5-fold greater than the basal secretion. NE and EPI also increased the maximal ACTH response to CRF, but did not alter the maximal ACTH release induced by arginine vasopressin.(ABSTRACT TRUNCATED AT 400 WORDS)     

Luteinizing hormone-beta mRNA levels are regulated primarily by gonadotropin-releasing hormone and not by negative estrogen feedback on the pituitary

Long-term effects of gonadotropin-releasing hormone (GnRH) and/or estrogen on pituitary mRNA levels for the beta-subunit of luteinizing hormone (LH-beta) were determined in anterior pituitary glands from ovariectomized (OVX) ewes. The relative roles of these two factors were assessed by studying hypothalamopituitary disconnected (HPD) ewes with appropriate hormonal treatments. Levels of LH-beta mRNA were increased by ovariectomy and substantially reduced by HPD. Treatment of OVX-HPD ewes with pulses of GnRH (250 ng each 2 h) for 1 week restored LH-beta mRNA levels to OVX levels, whereas treatment with estrogen alone did not alter the low levels found in OVX-HPD ewes. Combined GnRH and estrogen treatment for one week produced LH-beta mRNA levels that were similar to those found in OVX-HPD ewes given GnRH alone; plasma LH pulse amplitudes were also similar in these two groups. From these data we conclude that the long-term negative feedback effect of estrogen to reduce LH secretion is due to a primary inhibition of GnRH secretion and is not a pituitary effect of estrogen. Long-term regulation of LH-beta mRNA is thus primarily regulated by GnRH.