Jornalo Clinical InvestigationVol. 46,Oko. 5, 1967

Renin Release during Acute Reduction of Arterial Pressurein Nornotensive Subjects and Patients with

Renovascular Hypertension *YOSHIHIRo KANEKO,t TAKAOIKEDA, TADANAOTAKEDA, ANDHIDEO UEDA(From the Second Department of Internal Medicine, Faculty of Medicine, University of

Tokyo, Tokyo, Japan)

Summary. In normotensive subjects, acute reduction of mean arterial pres-sure to from 60 to 75 mmHg by infusion of sodium nitroprusside caused sig-nificant increase in renin activity of renal venous plasma and also in the renal-systemic difference of renin activity. At the same time, the products of therenal-systemic difference of renin activity and renal plasma flow increasedsignificantly, whereas renin substrate activity of plasma was unchanged, indi-cating that there was an increase in renin release during reduction in pressure.Renin activity of renal venous plasma, expressed in logarithms, showeda significant correlation with the degree of reduction in pressure; an increasein renin activity became significant when mean arterial pressure was reducedto below a level of 70 to 75 mmHg.

There was a striking difference in the renal response to reduction in pres-sure between patients with renovascular hypertension and normotensive sub-jects. In 10 renovascular hypertensive patients, significant increase in reninrelease occurred from the involved kidney at mean arterial pressures rangingfrom 90 to 137 mmHg; the threshold at which renin release increased wasshifted to a range much higher than that in normotensive subjects. Further-more, the magnitude of renin release from the involved kidney was signifi-cantly greater when compared to that in normotensive subjects. In contrast,in the contralateral uninvolved kidney, no significant release of renin wasdetected during reduction in pressure. The renal mechanism controllingrenin secretion appears to be operative at higher systemic arterial pressurelevels and with enhanced responsiveness in the involved kidney of renovas-cular hypertensive patients; the findings are consistent with the hypothesisthat the renin-angiotensin system participates in maintaining hypertensionin this disease.

Introduction

Recently, Skinner, McCubbin, and Page (1, 2)showed that reduction in mean renal perfusionpressure causes the kidney to increase renin se-

* Submitted for publication September 16, 1966; ac-cepted January 4, 1967.

This study was supported in part by U. S. PublicHealth Service research grant HE 09060 from the Na-tional Heart Institute.

A preliminary report was presented at the Twenty-third Meeting of the International Congress of Physio-logical Sciences, Tokyo, September 8, 1965.

cretion in normotensive and hypertensive dogs andsuggested that the renal mechanism controllingrenin secretion may be a homeostatic one thatparticipates in control of normal arterial pressureand that its dysfunction might be involved in thegenesis and maintenance of renal hypertension.It is as yet unknown, however, whether the hu-man kidney can similarly increase renin secretionin response to reduction in pressure and whether

t Address requests for reprints to Dr. YoshihiroKaneko, Second Department of Internal Medicine, TokyoUniversity Hospital, Hongo 7, Bunkyo-ku, Tokyo, Japan.

705

KANEKO, IKEDA, TAKEDA, ANDUEDA

there is any change in this renal mechanism in hu-man hypertensive patients. The present investi-gation examines the effect of acute reduction of ar-terial pressure on renin release in normotensivesubjects and in patients with renovascular hyper-tension.

Methods

Clinical material and procedureStudies were performed on hospitalized normotensive

and renovascular hypertensive patients on a normal diet.Normotensive patients, four women and nine men rang-ing in age from 19 to 47 years, showed no evidence ofsignificant cardiovascular disease, and their arterial bloodpressures ranged from 111/70 (average mean arterialblood pressure, 84) to 140/83 mmHg (average mean,102) at the time of the study. Patients with reno-vascular hypertension consisted of six women and fourmen ranging in age from 16 to 53 years, and their ar-terial pressures ranged from 150/78 (average mean, 102)to 225/151 mmHg (average mean, 176). Clinical find-ings are shown in Table I. The diagnosis of renovascu-lar hypertension was established by demonstration of se-vere stenotic lesions of the main renal artery by aortog-raphy and of an ischemic pattern in split renal functionstudies (Howard or Rapoport test), except in one pa-tient who had segmental ischemia due to severe stenosisof one of the double main renal arteries and showeda negative split function test and another from whomurine was not collected. In six patients, the diagnosiswas confirmed later by reduction of arterial pressure af-ter surgical treatment (Table I). Eight patients hadunilateral and one patient bilateral renal arterial stenosis.One patient had severe stenosis on one side and slightstenosis on the other. None showed clinical evidence ofheart or renal failure.

The purpose and procedures of the study were explainedto all subjects, and their free consent was obtained. Allmedications except for small amounts of sedatives hadbeen discontinued at least 2 weeks before study, and thepatients were fasting and supine at the time of measure-ments performed in the afternoon. To collect renal ve-nous blood, we introduced a polyethylene catheter (KIFAred-i) into a femoral vein with the patient under localanesthesia and advanced it under fluoroscopic control intoa renal vein of normotensive subjects or into the vein ofthe involved kidney of patients with renovascular hyper-tension. Position of the catheter was confirmed by ob-serving extraction of p-aminohippurate. In some patients,two catheters were introduced into both femoral veinsand passed into the bilateral renal veins at the same time.Systemic venous blood was collected from a vein of onearm or leg. Brachial arterial blood pressure was mea-sured by auscultation every 2 or 3 minutes along withheart rate, and mean arterial pressure (diastolic plusone-third of pulse pressure) was calculated. To lowersystemic arterial pressure, we infused sodium nitroprus-side, dissolved in physiologic saline in concentrations of

100 to 150 ,ug per ml, into a vein of one arm or leg ata rate of 30 to 240 ,&g per minute. Sodium nitroprussideis known to exert its depressor effect by direct action onvascular smooth muscle (3).

Control samples of renal and systemic venous bloodwere collected from the patients after they had restedat least 30 minutes in the supine position. Thereafter,mean arterial pressure was reduced progressively, by in-fusion of sodium nitroprusside, from 10 to 37 mmHg innormotensive subjects and 11 to 55 mmHg in patientswith renovascular hypertension, and repeated samplingsof renal and systemic venous blood were made whenarterial pressure had been stabilized for 10 to 15 min-utes. If symptoms of cerebral ischemia such as yawningappeared, the rate of infusion was decreased until thesymptoms disappeared; there were no serious reactionsin any subjects tested. In some patients, samplings wererepeated when arterial pressure had returned to almostthe preinfusion level after termination of the infusion.

Assay of plasma renin activityThe procedure used for determination of renin activity

of plasma was based on methods of Skinner and asso-ciates (1) and Helmer and Judson (4) and on the prin-ciple that the sensitivity of renin measurement can be in-creased by prolonging the time of incubation (5).

Approximately 8 ml of renal and systemic venous bloodwas collected simultaneously in polyethylene test tubes andmixed with EDTA(2 X 10' M) to inhibit angiotensinase(6) and with heparin (5 to 10 U per ml blood) as an ad-ditional safeguard against coagulation. The tubes wereplaced in an ice bath, the blood samples were centrifugedat 5,000 rpm for 10 minutes at below 40 C, and the sepa-rated plasma was kept frozen. The next morning, eachplasma sample was adjusted to pH 5.5 with 0.2 N HCland divided into three 1-ml portions. One portion wasdiluted with 0.5 ml of physiologic saline and immediatelyboiled for 10 minutes. To each of the other two por-tions was added one drop of 0.5% neomycin sulfate toprotect against possible bacterial contamination (7).To one of the two portions we added two drops of 0.1%diisopropyl fluorophosphate (DFP) as a further pre-caution against angiotensinase, according to Pickens andhis co-workers (5). The two portions were then incu-bated for 24 hours at 370 C without agitation and there-after diluted with 0.5 ml of saline and boiled for 15 min-utes in a chamber with a draft. The supernatant wasseparated by centrifugation at 5,000 rpm for 10 minutes.

The supernatant samples were assayed for pressoractivity in 180- to 300-g male rats treated at least 13hours before with intraperitoneal sodium pentobarbital(5 mg per 100 g as an initial dose and 1 to 0.5 mg per100 g as supplemental doses) and pentolinium tartrate(2 mg per 100 g initally and 0.4 mg per 100 g supple-mentally). Mean arterial pressure was measured froma cannulated carotid artery and recorded on a directwriting oscillograph. The supernatant samples from thethree portions were injected into a femoral vein in vol-umes of 0.1 to 0.3 (usually 0.2) ml. The unincubatedsample gave only a slight rise or no measurable change

706

RENIN RELEASE IN RENOVASCULARHYPERTENSION

mmHg

110

90

70

A A At A 1 t 3 4A A A A A 1 2 3 4

0.5 1.25 2.5ng ng ng

5

ng

10ng

5 6 7 A2.5ng

FIG. 1. TRACING FROMASSAY OF PLASMA RENIN ACTIVITY ON RAT. A indicates standard synthetic angiotensin. 1to 7 indicate plasma from a patient with renovascular hypertension (0.1 ml of the supernatant). 2 and 3 representincubated systemic and renal (involved side) venous plasma collected simultaneously during the control period, and4 and 5 incubated systemic and renal venous plasma collected during reduction in arterial pressure. 1, 6, and 7 areunincubated counterparts of 3, 4, and 5, respectively.

in pressure; the other two incubated samples served toconfirm the pressor activity generated by incubation(Figure 1). Under usual conditions, addition of DFPdid not produce significant change in the pressor activitygenerated in plasma. A series of synthetic angiotensinsolutions 1 were injected at frequent intervals and servedas standards for measuring pressor activity of the testsamples (Figure 1). Pressor activity generated inplasma as a result of 24-hour incubation was deter-mined, expressed in terms of nanograms angiotensinequivalent per milliliter of the original plasma, and desig-nated as plasma renin activity.

Evidence that the substances assayed are renin and angio-tensin

Incubated renal venous plasma collected during theexperimental period demonstrated greater pressor ac-tivity than incubated systemic venous plasma (Figure 1).Incubated renal and systemic venous plasma showedmuch greater pressor activity than their unincubatedcounterparts (Figure 1). The pressor response to in-cubated plasma was identical to that produced by syn-thetic angiotensin (Figure 1). In control experiments,pressor activity of the supernatant of incubated activeplasma was heat stable, was destroyed by incubation withchymotrypsin or trypsin, and disappeared after 24-hourdialysis. The mobility of the pressor substance in thesupernatant of incubated plasma on Whatman no. 1 paperchromatography at pH 2.1 was tested; 2 it was similar tothat of synthetic angiotensin. To determine the optimalpH in the reaction, we incubated three samples of activeplasma (EDTA and DFP added) at 370 C for 24 hoursat pH 4.5 to 7.5 and assayed them on the rat. The av-erage pressor activity generated after incubation at pH4.5, 5.0, 5.5, 6.0, 6.5, 7.0, and 7.5 was 7, 63, 148, 117, 64,

'Valyl-5 octapeptide, Ciba Pharmaceutical Co., Sum-mit, N. J.

2 In collaboration with Dr. Y. Takabatake.

14, and 7 ng per ml, respectively; the activity was great-est at pH 5.5, which is in accord with the results ob-tained by others (4, 5).

Examinations of the assay method

Effect of heparin. Since heparin used as an anticoagu-lant might affect angiotensin formation in incubatedplasma, its effect was checked. Heparin was added tothree samples of active plasma that had been collectedwithout it in concentrations of 0, 5, 10, 20, 40, 100, 200,and 400 U per ml, and the samples were incubated in thepresence of EDTAand DFP for 24 hours at pH 5.5. Inconcentrations below 40 U per ml, addition of heparin didnot produce significant change in the pressor activitygenerated in plasma, and, in concentrations of 100, 200,and 400 U per ml, the average pressor activity decreasedto 89, 78, and 64% of the control, respectively. The ef-fect of the doses of heparin used in the present methodwas considered insignificant.

Angiotensin formation during incubation and renin sub-strate activity of plasma. Each of four samples of activeplasma was incubated for 4, 8, 14, 20, and 24 hours.Angiotensin produced in plasma was roughly proportionalto the length of incubation time when the yield was below400 ng per ml. In plasma of higher renin activity, thereaction was slowed considerably (Figure 2). Sinceplasma renin activity measured by our method is below400 ng per ml under usual conditions, renin substrateconcentration in the reaction system was usually con-sidered excessive for 24 hours of incubation.

Human renin solution was prepared from cadaver kid-neys by the method of Haas and Goldblatt (8) ,3 andrenin substrate activity of plasma was assayed in fivenormotensive subjects and four patients with renovascularhypertension. To 0.5 ml plasma were added 0.2 ml of0.01 M EDTA, 0.1 ml of diluted human renin solution,and 0.3 ml saline. After pH adjustment to 5.5, 1 drop of

3 In collaboration with Drs. M. Ishii and H. Tagawa.

+ MINA5

ng

707

KANEKO, IKEDA, TAKEDA, ANDUEDA

TABLE I

Clinical data on 10 patients with renovascular hypertension*

Aortog-Known raphydura- Howardt (site of

tion of or renalAdmission hyper- Ocular RapoportJ arterial

Patient Age Sex BP tension fundi BUN RBF test stenosis) Remarks

years mmHg years grade§ mg/ ml/min100ml

S.U. 38 M 260/150 2.5 1 17 740 + Bilateral Had surgery. Postoperative BP, 134/84.H.I. 23 M 186/124 0.5 1 13 736 + Right Died at surgery.A.K. 27 F 240/130 2 2 15 700 + Right Had surgery. Postoperative BP, 134/92.H.I. 23 F 220/130 0.5 1 13 656 + Right Had surgery. Postoperative BP, 128/80.A.O. 29 M 250/126 1 1 14 1,140 + Left Surgery was not done.R.S. 53 M 210/120 1.5 2 16 547 + Leftil Had surgery. Postoperative BP, 130/80.A.U. 16 F 220/116 1 4 13 652 + Right¶ Had surgery. Postoperative BP, 160/94.K.M. 28 F 180/120 8 2 18 921 + Left Had surgery. Postoperative BP, 124/84.K.T. 29 F 164/ 90 3 1 11 1,180 Failed Right Surgery is pending.N.G. 19 F 170/100 2.5 1 12 963 + Right Surgery is pending.

* Abbreviations: BP = blood pressure; BUN = blood urea nitrogen; RBF = renal blood flow measured by p-aminohippurate clearance.t The test was considered positive when the affected side showed reductions of 50%or more in urine flow and 15% or more in the sodium con-

centration compared to the unaffected or less affected side.The test was considered positive when the tubular rejection fraction ratio was below 0.6 or above 1.6.Keith-Wagener classification.Severe stenosis of left main renal artery with slight stenosis of right main renal artery.Stenosis of one of the double main renal arteries (segmental ischemia).

0.1% DFP and 1 drop of 0.5% neomycin sulfate wereadded. The mixture was incubated for 24 hours at 370 Cand thereafter diluted with 2 ml of saline and boiled for15 minutes. The separated supernatant was assayed forpressor activity, and substrate activity was expressed asnanograms angiotensin produced per 1 ml plasma.Plasma substrate activity measured by this method rangedfrom 470 to 800 ng per ml in normotensive subjects.

Recovery. Human crude angiotensin was produced by

600-

400

0-8

zz

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i 200-

100-

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0 4 8 12 16 20 24

INCUBATION TIME IN HOURS

FIG. 2. ANGIOTENSIN FORMATIONDURING 24-HOUR IN-CUBATION. The upper two curves were obtained fromplasma incubated with human renin and the lower onesfrom active plasma of patients. See text.

incubation of human renin with human plasma. Possibleloss of angiotensin by the remaining angiotensinase inthe reaction system was determined by adding humanangiotensin in doses of about 10 to 400 ng to 1 ml ofplasma and carrying through the complete procedure.In eight experiments, the recovery after a 24-hour in-cubation ranged from 70 to 92%, mean 83%.

600 -

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RENIN RELEASE IN RENOVASCULARHYPERTENSION

Various amounts of human renin were added to 1 mlof human plasma and incubated for 24 hours. The angio-tensin yield after incubation was proportional to theamount of renin added when the yield was below 400 ngper ml (Figure 3); the mean recovery was 98 + 9.7(SD) %in those samples.

Sensitivity and reproducibility. The lower limit ofsensitivity of this method varied according to the as-say rats. Usually 0.2 to 0.5 ng of standard angiotensingave a measurable rise, and plasma renin activity of 1 to2 ng per ml or more was detected.

Renin activity of 50 different plasma samples was mea-sured in duplicate; the average difference between pairedmeasurements was 14.7 ± 10.7 (SD) %o. A difference inrenin activity between individual plasma samples wasconsidered significant when it exceeded 14.7 ± 21.3 (2 SD)or 36%o and also 5 ng per ml.

Statistical analyses. In all experiments, the differ-ences between the means were tested by Student's t test(9).

Results

Renin activity in plasma of normotensive sub-jects and renovascular hypertensive patients dur-ing control period. Renin activity of renal venousplasma of 13 normotensive subjects ranged from4 to 14 ng per ml in the control period, with amean of 8.1 ± 0.8 (SE); that of systemic venousplasma ranged from 3 to 10 ng per ml, with amean of 6.4 + 0.6. The difference in renin ac-tivity between renal and systemic venous plasmaranged from 0 to 5 ng per ml, mean 1.7 ± 0.5(Table II). The normal ranges of renin activityof renal and systemic venous plasma calculated bythe present method were considered to be 8.1 +5.6 (2 SD) and 6.4 + 4.6 (2 SD) ng per ml,respectively.

Renin activity of venous plasma from the in-volved kidneys of 10 renovascular hypertensivepatients (Table IV) ranged from 5 to 76 ng perml, with a mean of 40.4 + 8.0; the value was ele-vated beyond the normal range in 8 of the 10 pa-tients. Renin activity of systemic venous plasmaranged from 4 to 55 ng per ml, with a mean of21.7 + 5.2; elevated activity was found in 7 pa-tients. The difference in mean renin activity be-tween the normotensive and renovascular hyper-tensive groups was significant both with renal(p < 0.001) and with systemic (p < 0.005) ve-nous plasma. The difference in renin activity be-tween renal and systemic venous plasmas in therenovascular hypertensive group ranged from 1 to48 ng per ml, with a mean of 18.7 + 5.3, which

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was significantly (p < 0.005) greater than thatin the normotensive group.

Effect of reduction of arterial pressure on reninactivity in plasma of normotensive subjects. Af-ter control samplings of renal and systemic ve-nous blood were made, graded doses of sodium ni-troprusside were infused into 13 normotensivesubjects. Mean arterial pressure was reducedprogressively by from 10 to 37 mmHg in 1 to 3steps, and samplings were repeated during eachhypotensive period. Reduction of mean arterialpressure to 60 to 75 mmHg caused significant in-crease of renin activity in renal venous plasma in8 subjects and no significant change in 5; above75 mmHg, no subjects showed significant changein renin activity. Table II shows the results ob-tained during reduction in pressure to 60 to 75mmHg in 13 subjects, and Figure 4 illustrates atypical example of the response to reduction inpressure. On the average, renin activity of renalvenous plasma increased from 8.1 to 37.5 ng per mlwhen arterial pressure decreased from 93 to 68mmHg; the increase was significant (p < 0.01).The increase in renin activity of renal venous

mmHg140 -

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ARTERIALPRESSURE

RENIN ACTIVITY 40-OF

RENAL M ANDSYSTEMIC 3 10 -

VENOUSPLASMA ,,

SODIUM NITROPRUSSIDE

80 120 pg/monSYSTOLIC

r- r ri-20 0 20 40 60

TIME IN MINUTES

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FIG. 4. EFFECT OF REDUCTION OF ARTERIAL PRESSUREBY SODIUM NITROPRUSSIDE ON RENIN ACTIVITY OF RENALAND SYSTEMIC VENOUSPLASMAIN A 21-YmR-oLD NORMO-TENSIVE SUBJECT. Renin activity was unchanged at meanarterial pressure of 79 mmHg but increased at 69 mmHg.

plasma was accompanied by one in systemic venousplasma, which was significant but less in magni-tude. The renal-systemic difference in renin ac-tivity increased in 6 of the 13 subjects; the meandifference increased from 1.7 to 20.1 ng per ml(p < 0.05, Table II).

The increase in renal venous renin activity wasdetected after 10 to 15 minutes of reduction inmean arterial pressure to below 75 mmHg. Itpersisted for the hypotensive period and then re-turned almost to the control value 15 to 30 min-utes after arterial pressure recovered to near thepreinfusion level (Figure 4). The return ofsystemic venous renin activity was delayed com-pared to that of renal venous renin activity. Incontrol observations, renin activity was measuredseveral times without reduction in pressure, andthere, were no significant changes between mea-surements.

In three normotensive subjects, renal venousblood was collected from both kidneys simultane-ously. There was no significant difference inrenin activity between them, either in the controlor hypotensive periods.

Effect of reduction of arterial pressure on reninsubstrate activity of plasma and on renal plasmaflow in normotensive subjects. In five subjects,renin substrate activity of renal venous plasmawas measured at the same time, using human reninsolution. The plasma substrate activity was623 ± 61 (SE) ng per ml in the control periodand 614 + 65 in the hypotensive period; therewas no significant change during reduction inpressure.

In four subjects, total renal plasma flow (RPF)was measured simultaneously by the Fick prin-ciple, using p-aminohippurate. When averagemean arterial pressure was reduced from 93 to65 mmHg, RPF decreased from 653 ± 66 (SE)to 500 ± 82 ml per minute, and renal-systemicdifference of renin activity (A RA) increased from2.8 + 1.0 to 24.5 ± 7.8 ng per ml (Table III).A RAx RPF was then calculated as an index ofthe amount of renin released from the kidney. Itincreased from 1.64 ± 0.59 (SE) to 10.38 + 3.06jug per minute during the hypotensive period; theincrease was significant (p < 0.05, Table III).

Correlation between renin activity of renal ve-nous plasma and mean arterial pressure level dur-ing reduction in pressure. Renin activity of renal

710

RENIN RELEASE IN RENOVASCULARHYPERTENSION

TABLE III

Effect of reduction of arterial pressure on renal-systemic difference ofrenin activity and renal plasma flow in four normotensive subjects*

Renal-systemicSystolic and difference ofdiastolic BP MAP RA (A RA) RPF A RA X RPF

Subject Control Exp. Control Exp. Control Exp. Control Exp. Control Exp.

mmHg mmHg xg/sm mi/min ug/minK.G. 122/78 84/48 93 60 5 37 547 396 2.74 14.65K.I. 136/72 104/51 93 69 3 33 736 431 2.21 14.22T.M. 111/70 91/56 84 68 '0 2 795 745 0 1.49A.H. 132/84 82/54 100 63 3 26 532 429 1.60 11.15

Mean 125/76 90/52 93 65 2.8 24.5 653 500 1.64 10.38SE ±3.3 42.1 1.0 i7.8 ±t66 482 ±0.59 ±3.06P 0wou z 5-~

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TABLE IV

Effect of reduction of arterial pressure on renin activity of renal and

Systolic and Renin activity of systemicRate of diastolic BP Mean arterial pressure Heart rate venous plasmainfusion

Patient of SN Control Exp. Control Exp. Change Control Exp. Control Exp. Change

jug/min mmHg mmHg beats/min Ig/nmlS.U. 150 225/151 146/109 176 121 55 76 91 20 79 59HI.I. 100 172/115 114/ 78 134 90 44 72 82 15 73 58A.K. 130 220/130 190/110 160 137 23 84 98 20 43 23H.I. 48 186/120 118/ 76 142 90 52 84 100 7 32 25A.O. 175 190/ 98 146/ 76 129 99 30 78 82 24 214 190R.S. 233 190/ 94 140/ 73 126 103 23 57 70 4 17 13A.U. 104 221/103 182/, 72 142 109 33 88 114 4 5 1K.M. 73 180/100 128/ 80 127 96 31 74 94 40 65 25K.T. 53 150/ 78 130/ 71 102 91 11 90 112 55 81 26N.G. 68 150/100 125/ 79 117 94 23 86 104 28 87 59

Mean 113 188/109 142/ 82 136 103 33 79 95 21.7 69.6 47.9SE 46.7 :14.9 44.4 4:3.1 44.4 4±5.2 ±18.4 i17.0p

RENIN RELEASE IN RENOVASCULARHYPERTENSION 713

TABLE IV

systemic venous plasma in 10 patients with renovascular hypertension

Involved kidney Uninvolved kidney

Renin activity of renal Renal-systemic difference of Renin activity of renal Renal-systemic differencevenous plasma renin activity venous plasma of renin activity

Control Exp. Change Control Exp. Change Control Exp. Change Control Exp. Change

ng/ml ng/ml ng/ml ng/ml68 198 130 48 119 7123 252 229 8 179 17165 191 126 45 148 10334 64 30 27 32 5 5 12 7 -2 -20 -1839 353 314 15 139 124 17 189 172 -7 -25 -18

6 162 156 2 145 143 5 23 18 1 6 55 13 8 1 8 7 5 7 2 1 2 1

55 121 66 15 56 41 40 70 30 0 5 576 128 52 21 47 26 55 76 21 0 -5 -533 228 195 5 141 136 28 91 63 0 4 4

40.4 171 131 18.7 101 82.7 22.1 66.9 44.7 -1.0 -4.7 -3.748.0 ±30.7 +30.5 ±t5.3 418.8 ±19.2 +7.5 ±24.0 ±22.5 ±1.1 ±4.8 ±3.9

7KANEKO,IKEDA, TAKEDA, ANDUEDA

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Si Ri Lit t

52 R2

I I I , I II I I I I I I I II I I f I I

L2 S3 R3 L3 A A A2ng 4ng I Ong

+ MINA

20ng

FIG. 7. TRACING FROMASSAY OF RENIN ACTIVITY OF PLASMAFROMINVOLVED ANDUNINVOLVEDKIDNEYS AND OF SYS-TEMIC VENOUSPLASMAIN A 19-YEAR-OLD RENOVASCULARHYPERTENSIVEPATIENT. The patient had severe stenosis ofthe right main renal artery. The left artery was not affected. S, R, and L indicate incubated systemic venousplasma and right and left renal venous plasma (0.2 ml of the supernatant). 1, 2, and 3 represent samples collectedduring the control period, 20 minutes after reduction of mean arterial pressure from 117 to 94 mmHg, and 15 minutesafter recovery of pressure, respectively.

are consistent with the observations of Skinnerand associates in dogs (1). Acute reduction ofarterial pressure caused significant increase inrenin activity of renal venous plasma and in therenal-systemic difference of renin activity. Sincethere was no significant change in renin substrateactivity of plasma measured at the same time, andsince it has been reported that there is little differ-ence in renin activity between systemic arterialand venous plasma (11), the products of the re-nal-systemic difference of renin activity and RPFwere assumed to represent indexes of the amountof renin released from the kidney, although theproducts may give a minimal estimate of the truerenal secretion of renin, since it has been knownthat renin leaks out also in the renal lymphatics(12) and urine (13). There was a significantincrease in the calculated products parallel to in-increase in renin activity. Accordingly, markedincrease in renin activity of renal venous plasmaor in the renal-systemic difference of renin activityduring reduction in pressure appears to indicatean increase in renin secretion from the kidney.The results suggest that the human kidney actsas a baroreceptor, in the sense that it can respondto change in arterial pressure by altering the rateof renin secretion; they support the view thatthis function of the kidney controlling renin secre-tion could be a homeostatic mechanism which par-ticipates in control of normal arterial pressure(1). This sensing mechanism seems capable ofresponding to reduction in pressure in a quanti-tative manner; renin activity of renal venous

plasma, expressed in logarithms, showed a sig-nificant correlation with the degree of reductionin pressure and also with the level of mean ar-terial pressure. An increase in renin release be-came significant when mean arterial pressure wasreduced to below a level of 70 to 75 mmHg,whereas, above 75 mmHg, there was no signifi-cant change. The findings suggest a threshold forincreased secretion of renin at a mean arterialpressure level of 70 to 75 mmHg in normotensivesubjects.

The present study does not clarify what finalmechanism initiates increased secretion of reninwhen arterial pressure decreases, but there are atleast three possibilities. 1) Decrease in mean re-nal perfusion pressure may directly stimulate thejuxtaglomerular cells acting as baroreceptors(14, 1). A significant correlation between reninactivity of renal venous plasma and the degree ofreduction in pressure might support this theory,but there was no evidence which suggested thatreduction in pressure acted directly. 2) Decreasein tubular sodium concentration or other changein tubular fluid caused by decrease in renal bloodflow and glomerular filtration rate due to reduc-tion in pressure may stimulate the macula densaand the juxtaglomerular cells (15). 3) In-creased sympathetic discharge caused reflexly byreduction in pressure may exert a direct effect onthe juxtaglomerular cells or an indirect effect onrenal hemodynamics, resulting in increased secre-tion of renin. Stimulation of the renal nerve hasbeen shown to cause increased release of renin in

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dogs (16). Increased sympathetic discharge dur-ing reduction in pressure was apparent from anincrease in heart rate (17); the average heartrate increased from 75 to 96 beats per minute dur-ing the hypotensive period (Table II). On theother hand, a threshold of 70 to 75 mmHg dem-onstrated in normotensive subjects is near thelevel under which renal autoregulation is known tofail, and this failure might be associated with anincreased secretion of renin.

The differences in response between the kidneysof normotensive subjects and the involved kidneysof renovascular hypertensive patients were strik-ing. In renovascular hypertensive patients, sig-nificant increase in renin activity of renal venousplasma and the renal-systemic difference of reninactivity occurred in the involved kidney at meanarterial pressures ranging from 90 to 137 mmHg.Although blood flow of the involved kidney wasnot measured, marked increase in the renal-sys-temic difference appears to indicate an increase inrenin release from the involved kidney, sincethere was concomitantly a marked increase in reninactivity in systemic plasma in 9 of the 10 patients(Table IV) and there was no significant changein renin substrate activity of plasma. The find-ings, then, suggest that the threshold of arterialpressure for increased renin release in the involvedkidney was shifted to a range much higher thanin normotensive subjects. Also, the magnitude ofrenin release from the involved kidney was sig-nificantly greater compared to that in normoten-sive subjects, whereas the decrement in pressureor the degree of reduction in pressure in terms ofA MAP/MAPwas not significantly different. Inone patient -(A.U.), the magnitude of renin re-lease was considerably less than in other patients(Table IV); this might have been associated withthe segmental ischemia of this patient's kidney(Table I).

The finding that, in the involved kidneys of re-novascular hypertensive patients, the mechanismcontrolling renin secretion operates at much highersystemic arterial pressure levels with more en-hanced responsiveness than in normotensive sub-jects appears to suggest that the renin-angiotensinsystem participates in maintaining high arterialpressure in this disease. The altered response ofthe involved kidney could tend to oppose a fallof systemic arterial pressure to normotensivelevels. However, the participation of the renin-

angiotensin system in this disease is not due to itsdirect vasoconstrictor action alone. Circulatingrenin levels in renovascular hypertensive patientshave been reported to be variable (18-20) andwere unrelated to levels of arterial pressure; reninactivity of systemic venous plasma was in the nor-mal range in three patients, whereas arterial pres-sures measured at the same time were hyperten-sive (Table IV).

The upward shift of the threshold of systemicarterial pressure at which renin release increasedin renovascular hypertension does not imply re-setting of the sensitivity of the renal mechanismthat controls release, since renal perfusion pres-sure beyond the renal arterial stenosis could bemuch lower than systemic arterial pressure (21),and the renal arterial pressure sensed by themechanism could be the pressure level at whichrenin release normally increases. Skinner andco-workers (2) found, in dogs made hypertensiveby constriction of a renal artery, that renin re-lease occurred at elevated systemic arterial pres-sure, but actual renal perfusion pressure beyondthe constriction was in the same range as in nor-mal dogs. The enhanced responsiveness of theinvolved kidney might also be explained by thepressure drop across the stenosis of the renalartery. The magnitude of renin secretion wasshown to be correlated with the degree of reduc-tion in pressure, and Figure 5 shows, at lowerpressure levels, reduction in pressure of the samedegree could cause secretion of more renin thanat higher pressure levels.

The involved kidney, even when renin activityin the control period was normal, could cause sig-nificant increase in renin release at higher thannormal pressure when arterial pressure was re-duced. The finding that the effect of pressure re-duction was unrelated to renin activity in the con-trol period suggests that it may be preferable, fordiagnosis of renovascular hypertension, to measurethe threshold at which there is an increase in reninactivity.

In the contralateral uninvolved kidneys of reno-vascular hypertensive patients, no significant re-lease of renin was detected in the control or hypo-tensive periods; the renal-systemic difference ofrenin activity during reduction in pressure wassignificantly less, even when compared to that innormotensive subjects.

The mechanism of the different response of the

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KANEKO, IKEDA, TAKEDA, AND UEDA

uninvolved kidney was not known, but three pos-sibilities exist. 1) Reduction in pressure to levelsof 90 to 99 mmHg may not have reached thethreshold for renin release. 2) The renin con-tent of the uninvolved kidney may have been re-duced to the extent that no renin was available.It has been shown, in rats (22) and dogs (23)made hypertensive by unilateral renal arterialconstriction, that the renin content of the un-touched contralateral kidney is markedly reducedcompared to that in the normal kidney. 3)Large amounts of circulating renin and angiotensinreleased from the involved kidney may have ex-erted an inhibitory effect on renin secretion fromthe uninvolved kidney through a negative feedbackmechanism. The infusion of angiotensin isknown to suppress renin secretion in dogs (24)and plasma renin activity in man (25). In tworenovascular hypertensive patients, renin activityof plasma from the uninvolved kidney was lessthan that of systemic venous plasma during re-duction in pressure. This was unexplainable, butthe leakage of renin into the renal lymphatics andurine may have been responsible.

Acknowledgment

Wewish to express our appreciation to Dr. James W.McCubbin for his encouragement and valuable advice.

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Control of renin secretion. Circulat. Res. 1964,15, 64.

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17. Glick, G., and E. Braunwald. Relative roles of thesympathetic and parasympathetic nervous systemsin the reflex control of heart rate. Circulat. Res.1965, 16, 363.

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