John G. McDougall

Action of atrial natriuretic peptide in the immature ovine kidney.

ABSTRACT: This study examines the effects of infused human atrial natriuretic peptide 1–28 (hANP) on ovine fetal renal function. hANP was infused into chronically cannulated ovine fetuses of 103–128 days gestation (term 142–152 days) at 1.1 (4), 2.2 (5), and 4.4 (5) μg/h for 2 h. Isotonic saline was infused in 10 control experiments. The fractional reabsorption of infused lithium was used as a marker of proximal tubular sodium reabsorption. Glomerular filtration rate and urinary water and electrolyte excretion were assessed. The blood pressure and heart rate were unaltered by any dose of hANP. The lowest dose (1.1 μg/h) did not produce any significant changes in glomerular filtration rate, urinary electrolyte or water excretion, or fractional reabsorption of lithium. hANP, at 4.4 μg/h, caused a significant (p < 0.001) 5-fold increase in the excretion of Na, CI, and Ca, doubled the excretion rate of K and free water clearance, and significantly increased glomerular filtration rate. Fractional sodium reabsorption and fractional reabsorption of lithium were significantly decreased (p <0.001, < 0.01, respectively). The results show that the fetal kidney, at this stage, is as responsive to hANP as is the adult kidney. The natriuretic action of hANP is related to increases in glomerular filtration rate and proximal tubular rejection of sodium (as assessed by fractional reabsorption of lithium). The excessive salt loss of the premature or low birth weight neonate, which also involves increased delivery of filtrate to the distal tubule, may be due to endogenous hANP, circulating at high concentrations.

Gestational Changes in Renal Responsiveness to Cortisol in the Ovine Fetus

ABSTRACT: The ontogenetic renal responsiveness to exogenous cortisol was examined in the chronically cannulated ovine fetus. The contribution of effects at proximal and distal tubule of the kidney were studied also. Cortisol (81.5 µg/h) was infused into immature ovine fetuses (mean gestational age —113.9 days) on five occasions and increased blood cortisol from 0.8 ± 0.5 to 21.3 ± 6.2 nmol/liter. This dose of cortisol produced a highly significant diuresis and natriuresis, in part due to an increase in GFR and in part due to a significant decrease in proximal tubular reabsorption of sodium. Cortisol (107.2 ± 4.7 µg/h) was infused into mature fetuses (mean gestational age 133.4 days) and produced an increase in blood cortisol concentration from 11.4 ± 5.6 to 33.7 ± 6.8 nmol/liter. No natriuresis or diuresis was seen in the mature fetuses. Cortisol caused a significant depression of proximal tubular sodium reabsorption in mature fetuses, but this extra load was reabsorbed in the distal tubule in these fetuses. The inability of the premature or very low birth wt baby to maintain normal sodium balance on a standard salt intake may be due, at least in part, to a “fetal” renal response to the high plasma cortisol concentrations found in such babies. As the kidney matures it becomes capable of increasing distal tubular sodium reabsorption to compensate for any increased distal tubular fluid delivery.

The effect of corticotrophin (ACTH) administration on the pressor action of angiotensin II, noradrenaline and tyramine in sheep.

1. Pressor responses to angiotensin II, noradrenaline and tyramine were examined in sheep prior to and during the development of corticotrophin‐induced hypertension.

Differential blockade of the renal vasoconstrictor and diuretic responses to endothelin-1 by endothelin antagonist

1. The effect of cyclo(d‐Trp‐d‐Asp—Pro‐d‐Val—Leu) (or BQ123), a selective ETa receptor antagonist, on the vasoconstrictor and diuretic responses elicited by endothelin‐1 (ET‐1) was examined in conscious sheep with chronic indwelling renal arterial cannulae.

RENAL EFFECTS OF ATRIAL NATRIURETIC FACTOR (99–126) IN CONSCIOUS SODIUM-REPLETE SHEEP

1. The effect of renal arterial infusion of synthetic human atrial natriuretic factor (ANF(99–126)) on renal function in the conscious euvolaemic sheep was characterized. ANF (99–126) was infused for 2 h at 5 and 50 μg/h into the renal artery of crossbred Merino ewes with chronically indwelling cannulae inserted in the renal artery. The effect on absolute and fractional excretion of Na, K, Ca, Cl and HCO3, glomerular filtration rate (GFR), effective renal plasma flow (ERPF) and free water clearance (CH2O) were measured.

Hemodynamic Effects of Atrial Natriuretic Peptide in Conscious Sheep

The present study examines the effects of intravenous infusion of atrial natriuretic peptide (ANP) on blood pressure, heart rate, cardiac output, sodium excretion and urine output in conscious, chronically instrumented sheep. Human ANP (1-28) was infused into the jugular vein (I.V.) for 60 min at 20, 50, 100 and 500 micrograms/h. ANP caused a decrease in blood pressure at all doses which was associated with a reduction in stroke volume and cardiac output. There was also a decrease in right atrial pressure. At the two higher rates of infusion an increase in both heart rate and calculated total peripheral resistance was observed. These data are consistent with ANP acting on the venous side of the circulation to produce venodilatation, and a reduction in venous return, stroke volume and cardiac output. The increases in urinary sodium excretion and urine output observed when ANP was infused I.V. at 100 micrograms/h for 60 min were small. The data suggest that the minimum dose for effects on the cardiovascular system (20 micrograms/h) is less than that required to produce renal effects (100 micrograms/h). ANP has potent effects on the cardiovascular system in conscious sheep, exerting its effect on blood pressure primarily by its action on the venous circulation and on cardiac output.

Processing and bioactivity of the corpuscles of Stannius protein of the Australian eel.

The primary structure of the major protein from the Corpuscles of Stannius (CS) of the Australian eel was elucidated from the cDNA sequence and was found to bear close similarity to the N-terminal amino acid sequence of the presumably homologous salmon hormone, teleocalcin (TC). The cDNA sequence predicted a preproprotein of 263 amino acids. Following removal of a 17 amino acid signal peptide, specific monobasic cleavage at an Arg-Phe bond generates the 231 amino acid mature form of the protein. The isolation and sequence determination of the prosequence confirms that the precursor contains a prosegment of 15 residues. Various fragments of the protein have been synthesized chemically and their biological activity assessed. The N-terminal 1–20 fragment of the mature protein inhibits calcium uptake in fingerling trout, the effect being similar, but not equipotent to salmon teleocalcin. Further, infusion of either the N-terminal 1–20 or the 81–94 fragment at 50 μg/h into the renal artery of conscious sheep, caused significant decreases in systemic plasma potassium concentration and in potassium excretion. The 1–20 fragment also gave rise to a small but significant increase in sodium excretion. Infusion of TC at the same rate results in a significant decrease in plasma potassium and phosphate concentration as well as a significant decrease in potassium excretion. Bovine PTH (1–34) at 100 μg/h causes a small decrease in plasma potassium and phosphate and an increase in plasma calcium concentration, and was the only peptide to cause a significant decrease in calcium excretion.

Functional and expression analysis of ovine steroid 11ß-hydroxylase (Cytochrome P45011ß)

In this study, the ovine steroid 11 beta-hydroxylase (P450(11 beta) or CYP11B) cDNA previously reported by us (1) was transfected into COS-7 cells. Using 3H-11-deoxycorticosterone (3H-DOC) as the substrate, and paper partition chromatography for separation of steroid products, the expressed enzyme was shown to catalyse the conversion of DOC to corticosterone (B), 18-hydroxy-11-deoxycorticosterone (18-OH-DOC), 18-hydroxy-corticosterone (18-OH-B), and aldosterone (ALDO). These results suggest that the expressed ovine cDNA exhibited 11 beta-hydroxylase, 18-hydroxylase and aldosterone synthesis activities. The enzymatic activity of the enzyme was further analysed by adding unlabelled steroids to compete with 3H-DOC. The conversion of 3H-DOC to 3H-ALDO was inhibited by the addition of excess DOC, B and 18-OH-DOC, indicating that all these steroids were potential substrates of the enzyme. The results also demonstrated that 18-hydroxylation could occur before 11 beta-hydroxylation with this enzyme. However, the addition of excess cold 18-OH-B had no significant effect on the level of 3H-ALDO that was synthesised. This result could imply that 18-OH-B is not an intermediate involved in the conversion of DOC to aldosterone, or, more likely, the enzyme substrate site is not accessible readily. Our results also indicated that DOC was preferred to 18-OH-DOC as a substrate for the enzyme. We have demonstrated by hybridisation histochemistry using specific oligonucleotide probes that the corresponding P450(11 beta) RNA transcript was present in all zones in the sheep adrenal cortex. In summary, we have shown that the enzyme encoded by the predominant P450(11 beta) cDNA isolated from the sheep adrenocortical cDNA library has all the enzymatic activities to biosynthesise ALDO from DOC. The corresponding transcript of this ovine P450(11 beta) cDNA was located throughout the adrenal cortex and thus the inability of the zonae fasciculata-reticularis to secrete ALDO remains to be understood.

Aldosterone Secretion: A Molecular Perspective

The major mineralocorticoid hormone aldosterone is secreted from the zona glomerulosa of the adrenal cortex. Aldosterone is synthesized from cholesterol via a series of hydroxylations and oxidations. The enzymes involved in these reactions are mostly members of the cytochrome P450 superfamily. The final steps of this pathway, the conversion of 11-deoxycorticosterone (DOC) to aldosterone, require conversion via the intermediates 18-hydroxy-DOC or corticosterone and 18-hydroxycorticosterone. There are significant differences between species in the number of genes that encode the P450(11beta)-related enzymes (CYP11B) involved in these steps and the zonal distribution of their expression. One enzyme is capable of 11-hydroxylation, 18-hydroxylation, and 18-oxidation of DOC to aldosterone. The genetic basis of four diseases-congenital adrenal hyperplasia due to 11beta-hydroxylase deficiency, glucocorticoid-remediable aldosteronism, aldosterone synthase deficiency type I and type II-is explicable by mutations in these cytochrome P450(11beta)-related genes.

THE HAEMODYNAMIC EFFECTS OF CYCLOSPORIN A IN SHEEP

SUMMARY