Dopaminergic control of 18-hydroxycorticosterone responses to posture, isometric exercise, and diuretic administration in normal man.

This study investigates dopaminergic mechanisms involved in the control of corticosteroid secretion in man. The responses of plasma 18-hydroxycorticosterone (18-OHB) and aldosterone levels to upright posture and isometric handgrip exercise and furosemide administration as well as PRA and catecholamine responses to posture and exercise were evaluated in six normal subjects with and without bromocriptine (BEC) treatment. To evaluate the role of PRL suppression by BEC in affecting corticosteroid responses, we evaluated the effect of BEC on plasma 18-OHB and aldosterone responses to 20 mg furosemide in two subjects with autoimmune hypoprolactinemia. BEC (2.5 mg, three times a day for 4 days) markedly suppressed basal levels of 18-OHB, but not aldosterone, in the six normal subjects as well as the two subjects with autoimmune hypoprolactinemia. BEC also suppressed the 18-OHB and aldosterone responses to upright posture, isometric exercise, and furosemide administration without altering electrolytes, PRA, or plasma cortisol levels. Additionally, BEC suppressed basal levels of PRL, norepinephrine, and epinephrine as well as norepinephrine and blood pressure responses to upright posture and isometric exercise in the normal subjects. These results offer additional evidence that dopaminergic mechanisms modulate the secretion of 18-OHB and aldosterone, perhaps indirectly via inhibitory effects of dopaminergic pathways on catecholamine secretion.

Dopaminergic modulation of 18-hydroxycorticosterone secretion in man.

This study was designed to investigate mechanisms of dopaminergic control of corticosteroid secretion and to determine on which step in the aldosterone biosynthetic pathway dopamine exerts its effects. Plasma concentrations of electrolytes, PRA, plasma cortisol, 11-deoxycorticosterone, corticosterone (18-OHB), and 18-hydroxy-11-deoxycorticosterone were not altered by the iv administration of 10 mg metoclopramide in six healthy male volunteers. Metoclopramide increased plasma aldosterone from 6.3 +/- 0.9 ng/dl to a maximum of 23.0 +/- 3.4 ng/dl, plasma 18-OHB from 11.4 +/- 1.1 ng/dl to a maximum level of 42.8 +/- 4.4 ng/dl, and PRL from 9.9 +/- 1.4 ng/ml to a maximum of 71.0 +/- 5.5 ng/ml. The aldosterone and 18-OHB responses displayed a parallel time course, with significant responses of both occurring with 5 min after metoclopramide administration. Dopamine infusions (3 micrograms/kg . min) begun 60 min before the administration of metoclopramide markedly decreased the 18-OHB as well as the aldosterone and PRL responses to the dopamine antagonist. A parallel time course of stimulation of 18-OHB and aldosterone secretion with no change in other aldosterone precursors suggests that dopamine may modulate the activity of the glomerulosa 18-hydroxylase enzyme. Thus, rather than simply affecting aldosterone secretion, dopaminergic mechanisms appear to modulate the biosynthesis of aldosterone.

Development and optimisation of a radioimmunoassay for plasma captopril.

A specific and sensitive radioimmunoassay has been developed to monitor plasma levels of captopril, the first orally active angiotensin-converting enzyme inhibitor. Because of the reactive nature of the captopril thiol group, captopril was measured as the captopril-N-ethylmaleimide complex (captopril-NEM). Accuracy studies, using samples with known added concentrations of captopril, were satisfactory (r = 0.95, p less than 0.001, y = 0.97x - 2.02), and the radioimmunoassay results compared well with those determined by a gas chromatography/mass spectrometric method (r = 0.98, p less than 0.0005, y = 1.2x - 19). The minimum detection limit of the assay was 2 micrograms captopril/litre plasma.

The development and application of a radioimmunoassay for 18-hydroxy-corticosterone.

A sensitive and specific radioimmunoassay has been developed for 18-hydroxy-corticosterone (18-OH-B) and applied to the measurement of this steroid in peripheral plasma. High specific activity label (3H-18-OH-B) was prepared using the incubation of 3H-corticosterone with duck adrenal mitochondria. Antisera were produced by immunisation with 18-OH-B gamma-lactone 3-oxime conjugated to bovine serum albumin. The antibodies examined showed 100% cross-reactivity with 18-hydroxy-deoxycorticosterone gamma-lactone (18-OH-DOC gamma-lactone), but minimal cross-reactivity with other steroids. Paper chromatography was used to separate 18-OH-DOC gamma-lactone from 18-OH-B gamma-lactone. The interassay precision was 7.6% and the intra-assay precision 11.0%. The accuracy of the method was confirmed by showing a linear relationship between amounts of 18-OH-B added and amounts of 18-OH-B gamma-lactone measured (y = 0.854 X +15.1, r = 0.9. p less than 0.001). The mean plasma level in normal subjects on an ad libitum sodium intake was 225 +/- 92.7 (SD) pg/ml (n = 17) when standing, and 99 +/- 38.3 (SD) pg/ml (n = 6) after lying down for 30 minutes.

Conformational changes of DNA in the presence of 12-s-12 gemini surfactants (s=2 and 10). Role of the spacer's length in the interaction surfactant-polynucleotide.

A multifaceted study on the interaction of calf-thymus DNA with two different cationic gemini surfactants alkanediyl-α-ω-bis(dodecyldimethyl-amonium)bromide, 12-s-12,2Br(-) (with s=2, G2, and 10, G10) was carried out. The measurements were done at different molar ratios X=[surfactant]/[DNA]. Results show two different conformational changes in DNA: a first compaction of the polynucleotide corresponding to a partial conformational (not total) change of DNA from an extended coil state to a globular state that happens at the lower molar ratio X. A second change corresponds to a breaking of the partial condensation, that is, the transition from the compacted state to a new more extended conformation (for the higher X values) different to the initial extension. According to circular dichroism spectra and dynamic light scattering measurements, this new state of DNA seems to be similar to a ψ-phase. Measurements confirm that interactions involved in the compaction are different to those previously obtained for the analog surfactant CTAB. X values at which the conformational changes happen depend on the length of the spacer in the surfactant along with the charge of the polar heads.

Effects of modulation of renal kallikrein-kinin system in the nephrotic syndrome.

Albuminuria (UAlbV) can be reduced by converting-enzyme inhibitors (CEI), but the hormonal mechanism responsible for this effect has not previously been defined. Since CEI increase kinin activity as well as reduce angiotensin II (ANG II) activity, experiments were performed to determine the effect of isolated alterations in kinin and ANG II metabolism on UAlbV in rats with passive Heymann pephritis. Phosphoramidon was used to potentiate kinin activity without altering ANG II synthesis. Aprotinin was utilized in combination with the CEI, enalapril, to prevent the increase in kinin activity caused by CEI. UAlbV and the fractional renal clearance of albumin (FCAlb) decreased significantly after either phosphoramidon or enalapril, although only enalapril reduced blood pressure. Glomerular filtration rate (GFR) was not affected by either drug. Phosphoramidon did not affect plasma renin activity (PRA) or the pressor response to angiotensin I (ANG I), indicating that ANG II synthesis was not altered. Aprotinin prevented the reduction in UAlbV and FCAlb produced by CEI but not the hypotension, elevated PRA, or ANG I pressor blockade produced by CEI. Aprotinin alone had no effect on UAlbV, GFR, PRA, or blood pressure. UAlbV can be reduced by increasing kinin activity by a mechanism that is not dependent on suppression of ANG II activity or reduction in GFR or blood pressure. CEI may reduce proteinuria as a result of their action on the kallikrein-kinin system rather than on the renin-angiotensin system.

Arginine augments neither albuminuria nor albumin synthesis caused by high-protein diets in nephrosis.

Dietary protein independently modulates albuminuria (U(Alb)V) and albumin synthesis (AlbSyn) in nephrotic rats. While some amino acids are without effect on renal hemodynamics, arginine (Arg) augments renal blood flow and glomerular filtration rate, increases AlbSyn in tissue culture and isolated perfused livers, and could be one specific amino acid causing both decreased glomerular permselectivity and increased AlbSyn. Nephrotic rats were fed 10% casein (LP); 30% casein (HP); 30% casein with the inhibitor of nitric oxide (NO) synthesis N omega-nitro-L-arginine methyl ester (HP + L-NAME); 10% casein supplemented with Arg and amino acids that are Arg precursors of or are derived from Arg (proline, glutamate, and aspartate) in an amount in the increment between 10 and 30% casein (ArgAA); ArgAA supplemented with NH4 acetate to provide a diet isonitrogenous to 30% casein (ArgAA + NH4); or 10% casein plus an incomplete mixture of amino acids (Inc) containing the increment in histidine, phenylalanine, tryptophan, tyrosine, lysine, glycine, alanine, serine, threonine, cysteine, and methionine provided when the diet was changed from 10 to 30% casein. U(Alb)V increased significantly in HP and by a significantly greater amount in HP + L-NAME, but did not change in LP, ArgAA, or ArgAA + NH4. U(Alb)V tended to increase in Inc, was significantly greater than in LP or in ArgAA + NH4, but less than in HP. AlbSyn ([3H]phenylalanine incorporation) was no different in Inc than in HP, and was significantly greater than in either ArgAA + NH4 or LP. Increased AlbSyn results from increased ingestion of one or more of amino acids in Inc, but not from Arg or its precursors or products or from total dietary nitrogen.(ABSTRACT TRUNCATED AT 250 WORDS)

Muscle protein synthesis is impaired in nephrotic rats.

Muscle protein synthesis is impaired in nephrotic rats: muscle and hepatic protein synthesis was measured as the incorporation of [3H]phenylalanine [3H]phe) into muscle and liver in male Sprague-Dawley rats with passive Heymann nephritis (HN) in comparison to both normal male (SDM) and female Sprague-Dawley rats (SDF). Incorporation of [3H]phe was significantly less muscle in HN (1.55 +/- 0.08 x 10(4) cpm/g muscle/h) than in SDM (2.55 +/- 0.14 x 10(4), p < 0.01) and no different than in SDF (1.64 +/- 0.23 x 10(4)). Growth rate was also significantly less in SDF and HN compared to SDM. Total [3H]phe incorporation and the % of [3H]phe incorporated into muscle correlated inversely with urinary albumin excretion in HN (p < 0.01) but not with serum albumin. In contrast, [3H]phe incorporation was increased in livers of HN (13.89 +/- 0.99 x 10(3) cpm/g rat/h) compared to either SDM (5.96 +/- 0.38 x 10(3), p < 0.01) or SDF (4.71 +/- 0.35 x 10(3), p < 0.01). Total liver mass, liver protein content, and the ratio of liver weight/body weight were all increased in HN. There were no differences in liver weight, liver protein content, or the ratio of liver weight/body weight between SDM and SDF. We have previously shown that decreased muscle protein accrual in HN cannot be overcome by increasing dietary protein intake. Hepatic protein synthesis and mass are increased in proteinuric rats while muscle protein synthesis is reduced. As a consequence growth rate and muscle protein accrual are diminished in nephrosis. The mechanism responsible for reduced muscle protein synthesis in the nephrotic syndrome is unknown.

Effects of dietary sodium on circadian rhythm and physiological responses of 18-hydroxycorticosterone.

1. The effects of dietary sodium intake on plasma 18-hydroxycorticosterone (18-OHB) responses to physiological stimuli and recumbent 24-h-plasma 18-OHB levels have been examined in nine normal male subjects. 2. Basal supine levels of 18-OHB during a 40 mmol of sodium intake period (62.5 +/- 6.0 ng/dl) were considerably greater (P less than 0.0001) than the levels during a 200 mmol of sodium intake period (9.8 +/- 1.2 ng/dl). Further incremental and percentage changes of 18-OHB in response to graded dose infusions of angiotensin II and adrenocorticotropic hormone (ACTH) were greater during the 40 mmol of sodium intake period. 3. Although the mean 24 h levels of plasma 18-OHB during the 40 mmol of sodium intake period (43.9 +/- 4.0 ng/dl) were greater (P less than 0.001) than those during the 200 mmol of sodium intake period (9.4 +/- 1.2 ng/dl), the circadian rhythm of 18-OHB secretion was similar under the two extremes of sodium intake. 4. Factors which increase angiotensin II levels, such as sodium restriction, isometric exercise and angiotensin infusion, selectively increase 18-OHB and aldosterone, suggesting that angiotensin II increases 18-OHB and aldosterone secretion, in part, by modulation of the 18-hydroxylation reaction involved in conversion of corticosterone into 18-OHB.

Branched-chain amino acids augment neither albuminuria nor albumin synthesis in nephrotic rats.

Both albuminuria (UalbV) and albumin synthesis (AlbSyn) are modulated by dietary protein in nephrotic rats, but the agent(s) linking diet to altered UalbV and AlbSyn is unknown. Others have reported that branched-chain amino acids (BCAA) cause neither increased renal blood flow nor glomerular filtration rate (GFR) normally induced by dietary protein nor increased blood glucagon thought to be necessary for protein-mediated effects on renal hemodynamics. The effect of BCAA on UalbV is unknown. Because BCAA increase AlbSyn in tissue culture and after a fast, it is possible that feeding BCAA may increase AlbSyn but not UalbV in nephrosis. Nephrotic rats were fed either 8.5% casein (LP); 21% casein (NP); 8.5% casein supplemented with valine, leucine, and isoleucine to the total amount provided by a 21% casein diet (2.37%) (LBC); or 8.5% casein plus 12.5% BCAA providing a diet isonitrogenous to 21% casein (HBC). UalbV and AlbSyn were significantly greater in NP compared with LP, LBC, or HBC and were the same in the latter three groups. Glucagon was infused into nephrotic rats fed 8.5% casein either subcutaneously or intraperitoneally in quantities sufficient to increase plasma levels to over 10 times control but had no effect on UalbV. The ability of dietary protein to increase AlbSyn or UalbV is not a result of total alpha-amino nitrogen intake but is a result of the specific amino acid composition of the diet and must result entirely from the effect of one or more non-BCAA. Increased blood glucagon alone has no effect on UalbV.

Plasma IgG pool is not defended from urinary loss in nephrotic syndrome.

In the nephrotic syndrome, the plasma level of some proteins of hepatic origin is partially defended by an increase in their synthetic rate. The plasma levels of several liver-derived proteins, as well as immunoglobulin G (IgG) are increased in one condition where plasma albumin concentration and oncotic pressure (II) are reduced, i.e., hereditary analbuminemia. To determine whether the urinary loss of IgG, a protein derived from the immune system, is compensated for by an increased synthetic rate, we measured IgG synthesis in normal Sprague-Dawley rats (SD); two models of the nephrotic syndrome: Heymann nephritis (HN) and Adriamycin treatment; and in Nagase analbuminemic rats (NAR), a model of decreased II without urinary protein loss. Plasma IgG and total IgG mass were significantly reduced in both HN and Adriamycin, yet IgG synthesis was nearly identical in HN, Adriamycin, and SD. In contrast, plasma and total IgG and IgG synthesis were all markedly increased in NAR. We derived a pathogen-free colony of NAR by Caesarean section and found that plasma IgG was not increased in pathogen-free NAR, despite reduced II. Thus, unlike proteins of hepatic origin (e.g., albumin) where synthesis increases following urinary loss, no compensatory increase in IgG synthesis occurs. Increased plasma IgG as well as IgG synthesis in the NAR is not a compensatory response to the absence of albumin or reduction in II, but rather is due to subclinical infection. Profound hypogammaglobulinemia of nephrotic syndrome occurs in part because no compensatory synthetic mechanisms balance urinary loss, and alteration in plasma II does not modulate IgG synthesis.

Evaluation of the mineralocorticoid activity of 18-hydroxycorticosterone.

1. The mineralocorticoid activity of 18-hydroxycorticosterone (18-OH-B) has been compared with that of aldosterone by using human bioassay in vivo with measurement of rectal potential difference and urinary log 10 Na+/K+ ratio. 2. A log-linear relationship was found between maximum change in rectal potential difference and increasing doses of aldosterone. 3. No mineralocorticoid activity could be demonstrated after an intravenous bolus and infusion of 18-OH-B. 4. The half-life of clearance of 18-OH-B was measured in three subjects and found to be 28, 48 and 24 min.

Differing actions of dietary protein and enalapril on renal function and proteinuria.

Albuminuria (UalbV) increases in proportion to dietary protein in rats with passive Heymann nephritis. To determine whether a similar relationship existed in normal animals, 14 normal rats were switched from an 8.5% protein diet (LP) to a 40% protein diet (HP). Initially UalbV and glomerular filtration rate (GFR) increased in parallel, but GFR ceased to increase after 48 h while UalbV continued to increase, causing a significant increase in fractional renal clearance of albumin (FCalb). In contrast, HP for 4 days caused only a transient increase in GFR in nephrotic rats but effected a threefold sustained increase in UalbV. Pretreatment of nephrotic rats with enalapril blunted but did not entirely prevent the increase in UalbV after switching to HP and did not affect the increase or subsequent decline in GFR. Treatment with enalapril for 10 days reduced UalbV and FCalb in nephrotic rats fed either LP or HP. The similar pattern of changes in urinary albumin excretion in normal and nephrotic rats after dietary protein augmentation suggests that dietary protein may modify UalbV by the same process in both normal and nephrotic animals. The increases in UalbV and GFR resulting from dietary protein augmentation represent parallel but independent processes, since only the proteinuric response is modified by converting enzyme inhibition. Dietary protein restriction and converting enzyme inhibition exert an additive effect to reduce UalbV.

Non-iron mediated alteration in hepatic transferrin gene expression in the nephrotic rat.

Both transferrin and the iron it carries are lost in the urine in the nephrotic syndrome. Patients may develop hypochromic microcytic anemia and synthesis of transferrin, a protein regulated in large part by iron availability, is increased. Transferrin synthesis has also been reported to be increased in liver slices from rats with hereditary analbuminemia, and their plasma transferrin levels are increased, suggesting that transferrin synthesis may be stimulated by processes other than iron depletion in this hypoalbuminemic condition. Transferrin metabolism was studied in rats with Heymann nephritis (HN), in a strain of Sprague-Dawley (SD) rats with hereditary analbuminemia [Nagase analbuminemic rats (NAR)], and in normal SD rats. Plasma transferrin concentration and mass was decreased significantly in HN, but increased in NAR. Transferrin synthesis was increased both in NAR (measured either as the disappearance of [125I] labeled transferrin or as the incorporation of [3H] phenylalanine) and in HN (incorporation of [3H] phenylalanine). The fractional rate of transferrin catabolism was unchanged in NAR. Thus transferrin mass was increased in NAR entirely as a consequence of increased synthesis. Transferrin and albumin synthesis correlated with one another in both HN and SD (P < 0.001). Transferrin mRNA was increased in both HN and NAR and was unaffected by administration of iron to HN. Hepatic transferrin and albumin mRNA levels were also correlated positively in HN and SD, suggesting that increased hepatic synthesis of both proteins might be responding to the same stimuli. Transferrin gene transcription was increased in both HN and NAR and was unaffected by administration of iron to HN. Transferrin mRNA was not increased in the testis in either HN or NAR, suggesting that augmentation in transferrin gene expression is driven by a non-iron dependent process and is confined to the liver.