Goldblatt Units Research Articles

Neuronal Nitric Oxide Synthase-Deficient Mice Have Impaired Renin Release But Normal Blood Pressure

Nitric oxide deficiency is involved in the development of hypertension, but the mechanisms are currently unclear. This study was conducted to further elucidate the role of neuronal nitric oxide synthase (nNOS) in blood pressure regulation and renin release in relation to different sodium loads. Blood pressure and heart rate were measured telemetrically and assessed during periods of physical activity and inactivity. Urinary solute excretion was measured by metabolism cages and plasma renin concentration (PRC) was determined by radioimmunoassay; all in nNOS knockout (nNOS(-/-)) and wild-type (nNOS(+/+)) mice after 10 days of low (0.01% NaCl) and high (4% NaCl) sodium diets. The resting heart rate was reduced in nNOS(-/-) mice, but the two genotypes had similar blood pressure during the low (nNOS(+/+) 104 +/- 2 mm Hg; nNOS(-/-) 103 +/- 2 mm Hg) and high (nNOS(+/+) 107 +/- 3 mm Hg; nNOS(-/-) 108 +/- 2 mm Hg) sodium diets. During the high sodium diet, PRC did not differ between the genotypes (nNOS(+/+) 743 +/- 115 10(-5) Goldblatt units; nNOS(-/-) 822 +/- 63 10(-5) Goldblatt units), but during the low sodium diet, nNOS(-/-) mice failed to increase PRC (nNOS(+/+) 2164 +/- 220 10(-5) Goldblatt units; nNOS(-/-) 907 +/- 101 10(-5) Goldblatt units) and renal renin mRNA. On the low sodium diet, nNOS(-/-) mice also showed increased urine flow rate and osmolar excretion, observations not made during a high sodium diet. Our results show that nNOS is necessary for stimulation of renin in response to sodium restriction. Furthermore, nNOS(-/-) mice are normotensive, and their blood pressure responds normally to an increased dietary sodium intake, indicating that nNOS deficiency does not cause salt-sensitive hypertension.

ANG II-induced downregulation of RBF after a prolonged reduction of renal perfusion pressure is due to pre- and postglomerular constriction.

Previous experiments from our laboratory showed that longer-lasting reductions in renal perfusion pressure (RPP) are associated with a gradual decrease in renal blood flow (RBF) that can be abolished by clamping plasma ANG II concentration ([ANG II]). The aim of the present study was to investigate the mechanisms behind the RBF downregulation in halothane-anesthetized Sprague-Dawley rats during a 30-min reduction in RPP to 88 mmHg. During the 30 min of reduced RPP we also measured glomerular filtration rate (GFR), proximal tubular pressure (P(prox)), and proximal tubular flow rate (Q(LP)). Early distal tubular fluid conductivity was measured as an estimate of early distal [NaCl] ([NaCl](ED)), and changes in plasma renin concentration (PRC) over time were measured. During 30 min of reduced RPP, RBF decreased gradually from 6.5 +/- 0.3 to 6.0 +/- 0.3 ml/min after 5 min (NS) to 5.2 +/- 0.2 ml/min after 30 min (P < 0.05). This decrease occurred in parallel with a gradual increase in PRC from 38.2 +/- 11.0 x 10(-5) to 87.1 +/- 25.1 x 10(-5) Goldblatt units (GU)/ml after 5 min (P < 0.05) to 158.5 +/- 42.9 x 10(-5) GU/ml after 30 min (P < 0.01). GFR, P(prox), and [NaCl](ED) all decreased significantly after 5 min and remained low. Estimates of pre- and postglomerular resistances showed that the autoregulatory mechanisms initially dilated preglomerular vessels to maintain RBF and GFR. However, after 30 min of reduced RPP, both pre- and postglomerular resistance had increased. We conclude that the decrease in RBF over time is caused by increases in both pre- and postglomerular resistance due to rising plasma renin and ANG II concentrations.

Salt Wasting in Simple Virilizing Congenital Adrenal Hyperplasia

To evaluate possible derangement in sodium balance in patients with the simple virilizing (SV) form of congenital adrenal hyperplasia (CAH) which might have implications for therapeutic procedures. Patients were sodium loaded throughout the protocol and studied after interruption of cortisone therapy for 4 days, after treatment with dexamethasone 1 mg/m2/d for 3 days and after additional therapy with 9alpha-fluorocortisone (9alphaF) 0.1 mg/m2 for 3 days and 9alphaF 0.2 mg/m2 for 3 days. After each phase, basal and stimulated (2 h in an upright position), aldosterone and plasma renin concentrations (PRC) were evaluated. Nine children aged 5.0 to 12.8 years with the clinical classification of SV CAH were studied. Diagnosis was established at the age of 2.9 +/- 1.9 years (mean +/- SD) and the patients were treated with oral hydrocortisone at a mean dose of 22.5 mg/m2/d, given in two or three daily doses. Seven patients were heterozygous for the Ile172Asn point mutation in exon 4, and two for the Pro30Leu mutation in exon 1 of the CYP21 gene. All of them had a more severe mutation or deletion in the second allele. PRC was determined by RIA and expressed as Goldblatt units (GU). Aldosterone was determined by RIA. Genotyping for disease-causing deletions and mutations was performed by Southern blot analysis, PCR and direct sequencing of CYP21. PRC was significantly higher in patients off hydrocortisone replacement therapy than in age matched control subjects (basal 3.3 +/- 0.5 vs 1.2 +/- 0.2 GU 10(-4)/ml [mean +/- SEM], p<0.001; stimulated 8.6 +/- 0.5 vs 2.4 +/- 0.4 GU 10(-4)/ml; p<0.05). Upon treatment with dexamethasone, patients with CAH demonstrated a decrease in basal (2.1 +/- 0.5 GU 10(-4)/ml) but not in stimulated PRC (8.8 +/- 2.6 GU 10(-4)/ml). When dexamethasone treatment was supplemented by 9alphaF, both supine (0.9 +/- 0.1 GU 10(-4)/ml) and stimulated (1.6 +/- 0.3 GU 10(-4)/ml) PRC were suppressed into the normal range. Aldosterone concentrations were elevated after interrupting hydrocortisone treatment only under basal conditions. Dexamethasone caused a decrease below the reference level and 9alphaF resulted in further suppression of aldosterone concentration. All patients were hemizygous for a CYP21 mutation that is usually considered not to be associated with clinically relevant salt loss. However, we demonstrated an aldosterone secretion disturbance in patients with SV CAH which cannot be corrected by glucocorticoid treatment alone. Additional mineralocorticoid therapy should be considered in order to suppress PRC and reduce the glucocorticoid dose required for adequate control.

Purification and characterization of human truncated prorenin.

Posttranslational processing of enzymatically inactive prorenin to an active form participates in the control of the activity of a key system involved in blood pressure regulation, growth, and other important functions. The issue is complicated because renin can be produced by a number of tissues throughout the body, in addition to the kidney, but the mechanism by which they process prorenin to renin is unknown and difficult to determine because of the small amounts of renin present. In the juxtaglomerular cell of the kidney, a 43 amino acid prosegment is cleaved from the amino terminus of prorenin to generate renin of molecular weight 44,000 [Do, Y. S., Shinagawa, T., Tam, H., Inagami, T., & Hsueh, W. A. (1987) J. Biol. Chem. 262, 1037-1043]. Using human uterine lining or a recombinant human prorenin system, we employed the same approach as that used in kidney, ammonium sulfate precipitation at pH 3.1 followed by pepstatin and H-77 affinity chromatography or gel filtration, to purify to homogeneity a 45,500-MW totally active renin. The specific activity of the active truncated prorenin was 850 Goldblatt units (GU)/mg of protein for chorion-decidua renin and 946 GU/mg of protein for recombinant renin, both similar to that reported for pure human renal renin. Both forms of renin cross-reacted with an antibody generated against 44,00-MW pure human renal renin and with an antibody generated against a peptide identical to the carboxy-terminal one-third of the prosegment.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenalectomy causes a delayed increase of inactive plasma renin in rats

Using a new assay for inactive renin in rat plasma, inactive renin was unchanged 24 h after bilateral adrenalectomy but increased about threefold, from 0.43 Goldblatt units (GU)/l (range 0.27-0.62 GU/l; n = 14) to 1.24 GU/l (0.74-1.94 GU/l; n = 14) after 48 h. The increase was abolished by substitution with the mineralocorticoid deoxycorticosterone acetate. The results are in accordance with the high plasma concentration of inactive renin, previously reported in humans with adrenal insufficiency, and do not support activation of inactive renin in the circulation.

Action of endogenous atrial natriuretic peptide in calves with experimental acute central venous congestion and low cardiac output

The aim of the study was to investigate plasma concentrations of atrial natriuretic peptide, aldosterone, and renin during experimentally induced acute central venous congestion. Two experimental calf models were used: (1) right heart failure due to pulmonary artery obstruction; (2) inferior vena cava syndrome produced by inferior vena caval obstruction. Hormonal responses and haemodynamic variables were measured over 6 h. Experiments were performed on three female "Schwarzbund" calves, age 3 months, weight 92 +/- 8 kg. In the pulmonary artery obstructed group there was an increase of plasma aldosterone from 6.5(SEM 1.6) to 22.1(3.2) ng.dl-1 (p less than 0.05), of renin from 0.7(0.1) to 2.5(0.3) Goldblatt units x 10(-4).ml-1 (p less than 0.05), and of atrial natriuretic peptide from 22.1(4.5) to 141.4(27.8) pmol.litre-1 (p less than 0.05). During inferior vena caval obstruction, aldosterone increased from 2.4(0.4) to 20.9(2.0) ng.dl-1 (p less than 0.05), and renin increased from 0.4(0.05) to 2.0(0.20) Goldblatt units x 10(-4).ml-1 (p less than 0.05). In this experiment, atrial natriuretic peptide remained unchanged. Cardiac output decreased in both groups. There was significant fluid and electrolyte retention during both experiments, with urine volume decreasing from 87.7(11.6) to 35.0(1.2) ml-h-1 in experiment (1), and from 185(14) to 95.7(8.6) ml.h-1 in experiment (2). The study suggests (1) that in an experimental acute state of reduced cardiac output due to pulmonary artery stenosis with constantly increased right heart pressures, raised endogenous atrial natriuretic peptide failed to induce diuresis and natriuresis; (2) that in acute right heart failure, renin and aldosterone secretion could not be suppressed by raised atrial natriuretic peptide concentrations; and (3) atrial natriuretic peptide secretion seemed to be exhausted after 6 h continuous atrial distension.

Sexual Dimorphism of Inactive Renin in Rat Plasma

The plasma concentration of inactive renin was higher (p less than 0.002) in male (0.48 Goldblatt units (GU)/l, range 0.28-0.67 GU/l; n = 20) than in female rats (0.19 GU/l, range 0.09-0.30 GU/l; n = 20). This new finding indicates a sexual dimorphism of inactive plasma renin similar to that recently reported in mice and humans. Castration of male rats (n = 10) decreased (p less than 0.01) inactive plasma renin from 0.45 GU/l (range 0.22-0.65 GU/l) to 0.26 GU/l (range 0.15-0.37 GU/l) over a period of seven weeks. Similarly, inactive renin decreased (p less than 0.01) from 0.50 GU/l (range 0.28-0.67 GU/l) to 0.30 GU/l (range 0.19-0.47 GU/l) during treatment with the anti-androgen flutamide (n = 10). In castrated male rats (n = 10), inactive plasma renin was unaffected of substitution with testosterone. In intact female rats (n = 10), inactive renin increased (p less than 0.05) from 0.18 GU/l (range 0.12-0.30 GU/l) to 0.27 GU/l (range 0.15-0.40 GU/L) during testosterone treatment. Our findings indicate that androgens may be involved in the regulation of the plasma concentration of inactive renin in rats, but also suggest that other factors may be of importance.

Absence of Processing Enzymes for Inactive Renin in Mouse Blood but Polymorphonuclear Neutrophils Contain Active Renin

The possible role of enzymes in plasma and blood cells for the conversion of inactive renin was investigated in mice of the Balb C strain. The concentrations of both inactive and active renin were unchanged in unfractionated plasma and whole blood during incubation at 37 degrees C for 5 h, indicating, that inactive renin had not been activated. Purified platelets, mononuclear cells (monocytes and lymphocytes), and polymorphonuclear leukocytes were incubated separately with partially purified inactive renin at 37 degrees C for 1 h. Inactive renin was neither activated nor degraded by whole blood cells. Polymorphonuclear neutrophils were also obtained in mice after i.p. injection of fluid thioglycollate medium. Extracts of the polymorphonuclear neutrophils were incubated with partially purified, inactive renin. During incubation at pH 7.5 for 1 h at 37 degrees C, inactive renin was stable and no activation was registered, whereas degradation, without a concomitant increase in active renin, occurred at pH 4.5. Extracts of polymorphonuclear neutrophils were found to contain active, but no inactive renin. The mean amount of renin per cell was 0.5 X 10(-11) Goldblatt units (GU) (range 0.3 X 10(-11) GU - 0.9 X 10(-11) GU; n = 6). These findings argue against a role for the blood cells and plasma in a physiological activation of inactive renin, and do not support the concept that activation of inactive renin in the circulation may participate in homeostasis.

Quantitative activation and determination of inactive renin by high performance liquid chromatography.

Assay of inactive renin in unfractionated mouse plasma is difficult and often impossible because of high concentrations of active renin and plasma protease inhibitors. Therefore, 0.025 to 0.05 ml of plasma or amniotic fluid from mice was separated by high performance liquid chromatography (HPLC) using a silica-based size exclusion column. The eluates were examined for enzymatically active renin before and after limited proteolysis with trypsin. Since inactive renin eluted as a single peak corresponding to a molecular weight of 38,000 daltons and the elution of active renin was markedly retarded, inactive and active renin were partially separated. compared with plasma, inactive renin in amniotic fluid eluted as a broader and sometimes diphasic peak, suggesting heterogenicity. The rapid and reliable separation by HPLC provided a more than 300-fold purification of inactive renin. Despite low concentrations of plasma protease inhibitors, a 1 000 000-fold molar excess of trypsin (1 mg/ml) was needed for optimal activation. The necessity for high trypsin concentrations for activation may partly be explained by enzyme kinetic considerations. By combining HPLC with trypsin activation, inactive renin was readily measured and found to be 9.4 (5.1-13.2) GU (Goldblatt units)/l in normal and 8.0 (5.1-12.2) GU/l in sialoadenectomized and nephrectomized mouse plasma, which is higher than previously determined in our laboratory. The concentration was 12.4 (8.8-16.1) GU/l in amniotic fluid. Thus, the concentration of inactive renin in plasma is almost as high as active renin in normal mice (17.6 GU/l) and is uninfluenced by the decrease of active renin to 0.3 GU/l after sialoadenectomy and nephrectomy.

Renin release from different parts of rat afferent arterioles in vitro.

A technique was designed to study renin release from superfused rat glomeruli with short attached arterioles (SAG), from single glomeruli with long attached arterioles (LAG), and from single afferent arterioles (AA). The preparations obtained by magnetic isolation and microdissection were superfused individually, and the renin release was measured by an ultramicroradioimmunoassay with a detection limit of 3 X 10(-9) Goldblatt units. The renin content of one SAG was about one-fifth of that contained in one AA. Isoprenaline (10(-5) M) did not change renin release from SAG, whereas renin release from AA and LAG increased threefold (P less than 0.01). A 30-mosmol/kg reduction in medium sodium chloride concentration increased renin release from SAG 50% (P less than 0.01). This challenge caused no change in renin release from AA. It is concluded that the isoprenaline-sensitive juxtaglomerular (JG) cells are located in the afferent arteriole only at some distance from the glomerulus, whereas those cells sensitive to sodium chloride are located within and/or close to the glomerulus.

Episodic release of renin from single isolated superfused rat afferent arterioles.

Doubts have been raised about the involvement of an exocytotic event in the renin release process. This motivated the development of a technique which permitted the study of renin release from one single superfused rat afferent arteriole with a time resolution of 20 seconds. By using this technique it is shown in 43 experiments that the undisturbed renin release is episodic with a renin discharge of 45.2 +/- 3.3 (SEM) nano Goldblatt units per episode (n = 114) and a frequency of one episode per 5 min. The total renin content of one arteriole was about 30 microGU. The renin discharge and frequency correspond to calculated values for the renin content of single juxtaglomerular cell granules and the release rates in vivo, respectively. Release activity could be stimulated by an acute decrease in the osmolality of the superfusion medium (-20 mOsm sucrose, n = 14) indicating that an osmotic water movement is involved in the secretory process. This study provides functional evidence that renin release is exocytotic. In addition it reports what appears to be the first direct measurement of release of secretory material compatible with secretion of single granules from any secretory system.

Increased plasma vasopressin in low renin essential hypertension.

Baseline plasma vasopressin concentrations were measured in 48 men (all 50 years old) with decreased plasma renin concentration and untreated, sustained essential hypertension and in 29 healthy normotensive men. Mean hypertensive plasma vasopressin concentration was more than twice as high as the corresponding normotensive level (15.7 +/- 2.2 [SE] vs 7.5 +/- 1.0 pg/ml; p less than 0.001). Plasma renin concentration in the hypertensive group was reduced compared with that in the normotensive group (0.28 +/- 0.04 vs 0.46 +/- 0.06 Goldblatt units X 10(-4)/ml). These differences appeared despite virtually identical serum osmolality, creatinine clearance, and urinary sodium excretion in the two groups. In the first 38 hypertensive subjects, arterial plasma epinephrine concentrations were significantly increased over those of the first 28 control subjects (99 +/- 12 vs 68 +/- 6 pg/ml; p less than 0.025). In contrast to those with low renin essential hypertension, 35 men with normal renin essential hypertension (all 40 years old) had normal plasma vasopressin levels that were not significantly different from those in a comparable normotensive control group (3.7 +/- 0.8 vs 3.5 +/- 0.4 pg/ml). Arterial epinephrine concentrations were not significantly different between normal renin subjects and the control group. After 6 weeks of treatment with the nonselective beta-adrenergic receptor blocker oxprenolol in 11 subjects with low renin hypertension, blood pressure was reduced and the plasma vasopressin concentration fell from 27.6 +/- 6.4 to 13.5 +/- 4.2 pg/ml (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Hemodynamic Effects of Chronic Intraventricular Administration of Renin in Awake Dogs

A possible role of brain angiotensin II (A-II) and angiotensin receptors within the central nervous system in the development of hypertension was first suggested in 1961 (1) and since then, investigators have identified every component of the renin-angiotensin system to be present within the brain (2). The chronic intraventricular (IVT) administration of A-II to awake dogs for 2 to 4 weeks has been reported to produce sustained hypertensive effects only if the daily intake of sodium is increased (3). This present study was undertaken to investigate the effects of chronic administration of renin, 0.15 Goldblatt units (Gu) IVT twice daily to awake instrumented dogs and to determine if an increase in the daily intake of sodium is necessary for physiological and/or pathological effects to be produced by the endogenous increase of A-II within the cerebrospinal fluid.

Aggression-provoked huge release of submaxillary mouse renin to saliva

In a normal male mouse, which had been kept isolated for some time, confrontation with another male mouse, resulted in aggressive behaviour, including fighting. The aggressive behaviour resulted in secretion of a few microlitre of saliva with vast renin concentration. The renin concentration in saliva reached values of 6000 Goldblatt units X ml-1, corresponding to 15 mg/ml of renin. This concentration is 6 orders of magnitude higher than normal plasma renin concentrations. The secretory pattern could be mimicked by ip injection of pharmacological doses (2-18 mg/kg) of the alpha-adrenergic agonist phenylephrine. This is probably the first demonstration of a physiological secretion of renin to saliva which may be of importance since renin most likely is injected into the opponent or prey through bites.

Rat kidney renin and cathepsin D: purification and comparison of properties.

Renin and cathepsin D were purified by seven-step procedures involving five steps common to both enzymes. These common five steps were extraction of freeze-dried kidney powder in 30% methoxyethanol-water, diethylaminoethyl-cellulose (DEAE-cellulose) batch absorption and elution, pepstatin-aminohexyl-Sepharose chromatography, Sephadex G-100 chromatography, and DEAE-cellulose chromatography. The renin component was purified further by passage through an anti-rat spleen cathepsin D immunoglobulin G-Sepharose (IgG-Sepharose) column followed by carboxymethyl-Sephadex (CM-Sepharose) chromatography which separated two renin components. Cathepsin D activity obtained by the fifth step was purified by passage through an anti-rat kidney renin IgG-Sepharose column followed by DEAE-Sephacel chromatography which separated three cathepsin D components. The homogeneity of renin and cathepsin D preparations was demonstrated by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. The two components of renins showed molecular weights of 42 000 and 36 000 by gel filtration and 38 000 and 36 000 by SDS gel electrophoresis, respectively. They showed isoelectric points of 5.35 and 5.65 by electrofocusing in 5% polyacrylamide gels. Their optimum pHs of enzyme activity were 6.5 as determined by using nephrectomized rat plasma as a substrate. Their specific angiotensin I (Ang I) generation activities were 158 and 146 micrograms of Ang I (microgram of protein)-1 h-1, respectively, which correspond to 1100 and 1020 Goldblatt units (mg of protein)-1 h-1. The three cathepsins showed molecular weights of 41 000, 43 000, and 41 000 by gel filtration and 46 000, 45 000, and 46 000 by SDS gel electrophoresis.(ABSTRACT TRUNCATED AT 250 WORDS)

Vast release of submaxillary mouse renin to saliva after stimulation with cholinergic, beta-adrenergic but especially alpha-adrenergic agonists.

alpha-adrenergic stimulation with phenylephrine causes release of vast amounts of mouse submaxillary gland renin to saliva. The amount released is about 100-fold higher than that released after beta-adrenergic stimulation with isoproterenol and about 1000-fold higher than the amount released after cholinergic stimulation with pilocarpine. alpha-blockade with phenoxybenzamine before phenylephrine stimulation strongly inhibits the renin release. However, phenoxybenzamine or beta-blockade with propranolol before isoproterenol stimulation have no effect on the amount of renin released. Thus, the release of submaxillary mouse renin to saliva is triggered mainly by activation of alpha-adrenergic receptors. Renin in cholinergic and alpha- and beta-adrenergic stimulated saliva has the same molecular weight and the same specific enzymatic activity as renin in the submaxillary gland. This indicates that renin is secreted to saliva, and probably also to the blood, in its fully enzymically active 40000 mol.wt. form regardless of the stimulus used. The vast amounts of renin released to saliva (up to 600 Goldblatt Units) demonstrate that submaxillary mouse renin is secreted predominantly to saliva and much less to the blood. The mouse submaxillary gland therefore seems to have an exocrine rather than an endocrine role for release of renin.

High levels of active 40 000-dalton renin in mouse saliva, but no evidence of inactive or high molecular weight forms

The presence of renin in parasympathetically elicited mouse saliva was demonstrated by using both the antibody trapping method, which measures renin's enzymatic activity, and a direct radioimmunoassay, which detects the renin molecule by its antigenic properties. Crossed immunoelectrophoresis of saliva samples using an antiserum elicited against pure submaxillary renin showed only one precipitation line, indicating the presence of only one form of renin. The position of the line was similar to that found when submaxillary gland extract was subjected to crossed immunoelectrophoresis. Tandem crossed immunoelectrophoresis showed complete identity between antigenic determinants in submaxillary and salivary renin. An apparent molecular weight of about 35 000 and 38 000 was found when saliva samples were subjected to gel filtration on Ultrogel AcA 44 and Sephadex G-100, respectively. No high molecular weight forms were present and no inactive forms could be demonstrated after limited pepsin or trypsin proteolysis. The specific enzymatic activity of renin in pilocarpine saliva was 0.37 Goldblatt Units (G.U.) · μg −1, which is identical to that of pure submaxillary gland renin (0.41 G.U. · μg −1) and to that of the storage form of renin in the submaxillary gland (0.4 G.U. · μg −1). An identical K m value was found for salivary renin, 1.01 μM, and for pure submaxillary renin, 0.98 μM. It is concluded that renin in pilocarpine-elicited saliva is similar to the storage form of renin in the submaxillary gland with respect to molecular weight, enzymatic and immunological properties.

Purification of rat renal renin from crude kidney extracts by diaminohexamethylene-Sepharose chromatography

Renin from rat kidney extracts was adsorbed to diaminohexamethylene-sepharose columns at extremely low ionic strength and neutral pH. Renin was retarded while the column was developed in 1 mM sodiumpyrophosphate and extraneous proteins were removed. Elution of renin was performed using a linear gradient of sodiumpyrophosphate, 1 – 17 mM at pH 6.8. Renin was purified in a yield up to approx. 60 per cent of the applied activity and a purification factor between 5 – 122 depending on the specific activity of the applied sample. The specific activity after this single chromatography of crude rat kidney homogenate on diaminohexamethylene-sepharose showed a median of 11.3 Goldblatt units per mg protein in a range of 5.3 – 42.0 Goldblatt units per mg protein. The renin binding capacity of the column was 1 Goldblatt unit per ml wet gel. The purified renin was subjected to G-100 Sephadex chromatography demonstrating two molecular weight forms of 44000 and 50000 dalton. Polyacrylamide gel electrophoresis demonstrated three separate fractions of renin.

DIRECT RADIOIMMUNOASSAY OF HUMAN RENIN

Antibodies were raised in rabbit against pure human renin. The antisera obtained are highly specific for human renin versus hog, dog and rat renin. They do not cross-react with acid proteases such as pepsin and human cathepsin D. A direct radioimunoassay is described for human renin in plasma and kidney extracts. 30 to 50 pg of enzyme (2.5 to 4 x 10(-5) Goldblatt units) are detected.

In mice aggressive behavior provokes vast increase in plasma renin concentration, causing only slight, if any, increase in blood pressure.

In mice aggressive behaviour causes a vast release of renin, which can result in about 600-fold increase in plasma renin concentration, reaching 6 Goldblatt Units, corresponding to 15 microgram renin per ml. This increase is mainly due to release of submaxillary renin, but there is also a significantly increased renal renin release. The degree of renin release is influenced by the duration of the aggression and by previous contact with other mice. Contrasting with the vast increase in plasma renin the blood pressure is normal or only moderately increased. This disproportion is not due to the depletion of renin substrate, caused by the increased renin, as shown by the increased calculated renin activity, as well as by decrease in blood pressure elicited by blockade of the renin system. Nor is the disproportion due to change in the sensitivity of the vessels to angiotensin II, the cause of this lack of tachyphylaxis being unknown. By way of exclusion the lack of pronounced increase in blood pressure can be explained by homeostatic function of the cardiovascular reflexes, which may also account for the fact that the pressor response after injection of pure submaxillary renin is only short, contrasting with a prolonged marked increase in plasma renin concentration.