THE GRANULATIONOF JUXTAGLOMERULARCELLS IN RENALHYPERTENSION,DESOXYCORTICOSTERONEAND POST-DESOXYCORTICOSTERONEHYPERTENSION, ADRENAL

REGENERATIONHYPERTENSION, ANDADRENALINSUFFICIENCY 1, 2

By LOUIS TOBIAN,s JANET THOMPSON,ROBERTTWEDT, ANDJEANETTEJANECEK

(From the Department of Internal Medicine, University of Minnesota School of Medicine,and the University of Minnesota Hospitals, Minneapolis, Minn.)

(Submitted for publication November 20, 1957; accepted January 23, 1958)

The pathogenesis of renal hypertension remainsa mystery. Renal disease can be severe enough toproduce terminal uremia without producing hy-pertension. On the other hand, hypertension canbe induced by partial constriction of the renal ar-tery, and the kidney in question may retain goodexcretory function. The excretory activities ofthe kidney seem not to be directly related to itshypertensive or antihypertensive functions. It istherefore inviting to consider aspects of renal anat-omy other than those directly and obviously con-cerned with excretion. Such a structure is thejuxtaglomerular apparatus. Certain cells of thisjuxtaglomerular complex containing discrete se-cretory granules are found in the wall of the af-ferent glomerular arteriole, just before it joins theglomerulus. Goormaghtigh studied them manyyears ago in relation to renal hypertension (2).These cells are in a position which would be par-ticularly affected by changes in intraluminal pres-sure, such as occurs with Goldblatt clamping. Inthe classic Goldblatt experiment, the renal arteryis partially constricted, producing a decrement inblood pressure downstream. Such a decrementin pressure would reduce the stretch of the wallsof the entire renal arborization, including thejuxtaglomerular cells residing in the arteriolarwalls. These cells, then, could be stretch recep-

1 This investigation was supported by a grant fromthe American Heart Association and by Grant No.H2008 from the National Heart Institute, United StatesPublic Health Service.

2This article is a full report of studies presented atthe meeting of the American Federation for Clinical Re-search, May, 1957 (1).

3 This work was started during the tenure of an Es-tablished Investigatorship of the American Heart As-sociation.

tors which change their rate of secretion in re-sponse to changes in stretch. With this in mind,the juxtaglomerular apparatus was studied in re-lation to the Goldblatt type of experimental hyper-tension. If the juxtaglomerular apparatus hasany fundamental role in renal hypertension itmight also be altered in other forms of experi-mental hypertension. Therefore, rats with desoxy-corticosterone hypertension, postdesoxycorticos-terone hypertension, and adrenal regenerationwere also studied.

METHODS

This study was greatly facilitated by the use of thestaining technique developed by the Hartrofts for thejuxtaglomerular (JG) apparatus (3). Each granule. isdiscretely stained in this method. A 40x fluorite oil im-mersion objective enabled us to see the individual secre-tory granules much better than the usual 43x achromatic"high dry" objective, yet provided a large field for rapidscanning. Compensating eyepieces (12x) were used inconjunction with this objective. The degree of granu-lation in the juxtaglomerular apparatus was estimatedusing the system devised by the Hartrofts (3). Theirweighted juxtaglomerular index (JG index) was utilized,giving a total index number per 100 glomeruli. Thissystem is advantageous in that it counts each JG com-plex and weighs it according to its degree of granula-tion. Experience indicated that whenever a large numberof juxtaglomerular complexes were to be seen, the num-ber of granules per complex were usually also increased.Even though this is a semiquantitative method, it hasconsiderable precision. Two different observers scan-ning the same slide rarely differed by more than 10 percent. Figure 1 shows various groups of juxtaglomerularcells.

Renal hypertension was produced in our rats by par-tially constricting the renal artery, using a silver ribbonclip. This is the method developed by Wilson and By-rom (4) and used extensively in recent years by Floyer(5). In the rat, hypertension can be produced by con-stricting only one renal artery and leaving the other

660

GRANULATIONOF JUXTAGLOMERULARCELLS IN HYPERTENSION

c "' . d JY.:A d:FIG. 1. VARIOUS GROUPSOF JUXTAGLOMERULARCELLS

In picture a, eight juxtaglomerular (JG) cells loaded with granules are seen in thewall of the afferent arteriole. In b four JG cells laden with granules are seen in thewall of the afferent arteriole. The unstained nuclei of these cells are surrounded bythe black granules. In c, two cells are seen which are only moderately filled withgranules. Pictures a, b, and c are sections from rat kidney. Picture d shows onegranular JG cell in a section from the kidney of a patient with malignant hypertension.

kidney untouched (4, 5). This form of hypertension wasutilized in these studies. Floyer's useful terminology isalso employed. The "clipped" kidney is one with the re-nal artery partially constricted; the "untouched" kidneyis the unmanipulated kidney opposite to the "clipped"one (5).

The level of arterial blood pressure in rats was de-termined with the Friedman microphonic method (6).The rats were lightly anesthetized with U.S.P. ether dur-ing the blood pressure measurements.

Only male rats were utilized in these experiments.They were all of the Wistar strain except those used inthe adrenal insufficiency and adrenal regeneration ex-periments. Except in the postdesoxycorticosterone ace-tate (DCA) group, the rats usually weighed between200 and 400 Gm. Within this range, the JG counts didnot noticeably vary with weight or age. 'The rats at alltimes were either eating Purina laboratory chow (0.24per cent sodium) or the special "synthetic" diets men-tioned below.

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L. TOBIAN, J. THOMPSON,R. TWEDT, AND J. JANECEK

RESULTS

A. Renal hypertensionNormal rats. The average juxtaglomerular in-

dex for 12 normal Wistar rats weighing between200 and 300 Gm. is 36, with a standard deviationof + 8. The mean blood pressure for 90 normalWistar rats is 99 (S.D., + 7) mm. Hg (Table I).

Sham operated rats. In nine rats a sham opera-

tion on one kidney was performed. The renal ar-

tery of this one kidney was dissected out, a silverclip was applied to it, and the clip was then im-mediately removed and the incision sutured. Fiveweeks after this sham operation, the operated kid-ney had a mean juxtaglomerular index of 34(S.D., + 12) and the contralateral unmanipulated

kidney had a mean JG index of 35 (S.D., + 13)(Table I).

Rats wtith unilateral constriction of a renal

artery. In sixteen Wistar rats weighing between150 and 250 Gm., a clip was applied to one renalartery in such a way as to constrict it partially;the other kidney was "untouched." Ten weekslater the blood pressure of these rats was ascer-

tained and eight were found to be definitely hyper-tensive, with a blood pressure of over 140 mm. Hg(Table II). In these eight rats the "clipped" kid-neys had an average JG index of 76, which is sig-nificantly higher than that of normal or shamoperated kidneys (p = 0.0001). The "untouched"contralateral kidneys of these rats showed striking

ILE I

The mean juxtaglomerular indices for several groups of Wistar rats* with renal hypertension along withappropriate control groups

Mean blood Data concerning JGt indicespressure at

end of StandardNo. of experiment Range of error of

Group rats (mm. Hg) Mean values the mean

a. Normal Wistar rats 12 99 One kidney examined 36 23 to 53 1: 2.5b. Sham operated rats 9 Sham operated kidney 34 23 to 65 =113.0

Contralateral untouchedkidney 35 21 to 65 1:3.7

c. Hypertensive rats 8 172 Clipped kidney 76 40 to 129 4110.210 weeks after Contralateral untouchedunilateral clipping kidney 1 0 to 4 i 0.7

d. Hypertensive rats 11 159 Clipped kidney 44 21 to 76 =t 5.86 weeks after Contralateral untouchedunilateral clipping kidneyt

e. Hypertensive rats 7 145 Clipped kidney 46 28 to 63 i 4.17-14 days after Contralateral untouchedunilateral clipping kidney 9 4 to 14 i1 1.4

f. Rats with little or 13 96 Clipped kidney 48 26 to 80 + 4.3no hypertension Contralateral untouched7 weeks after kidney 6 0 to 20 1.9unilateral clipping

g. Normal rats on a 8 110 One kidney examined 48 29 to 62 4- 4.0basic diet containing0.002% Na

h. Normal rats on the 8 111 One kidney examined 30 19 to 54 + 3.9basic diet plus0.4% Na

i. Rats with unilateral 22 169 Clipped kidney 66 39 to 98 4t 3.5clipped kidney on a Contralateral untouchedbasic diet con- kidney 6 0 to 33 + 1.8taining 0.002% Na

j. Rats with unilateral 13 181 Clipped kidney 58 22 to 112 + 7.8clipped kidney on Contralateral untouchedthe basic diet plus kidney 1 0 to 4 + 0.30.4% Na

* Both kidneys were present in all rats.t Juxtaglomerular.t This kidney was left intact for subsequent studies (Group b, Table III).

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GRANULATIONOF JUXTAGLOMERULARCELLS IN HYPERTENSION63

degranulation of the juxtaglomerular apparatus.From a normal value of 36 the index fell to an

average of 1 in these kidneys, and the JG indexranged from 0 to 4, far below normal JG indices.These findings are consistent with observationsby others that renal ischemia may be associatedwith an increase in the number of granular JGcells (2, 7) and also with observations that thecontralateral "untouched" kidney has a great de-crease in the granular JG cells (8, 9).

In another group of hypertensive rats with a

similar operation, the ischemic kidney was ex-

amined six weeks after constricting the renalartery. The JG index averaged 44 in these kid-neys, a value not as high as was seen in constrictedkidneys 10 weeks after clipping (Table I).

Another group of seven hypertensive rats was

sacrificed 7 to 14 days after a similar constrictionof only one renal artery. The "clipped" kidneyshad an average JG index of 46 with a range of28 to 63, and the contralateral "untouched" kid-ney had a mean JG index of 9 with a range of4 to 14. Even this brief interval after clipping one

renal artery allowed for considerable degranulationof the "untouched" kidney and hypergranulationof the ischemic kidney (Table I).

Thirteen other rats had the same clipping pro-

cedure as the foregoing groups of rats. However,they failed to develop hypertension during seven

weeks of constriction of one renal artery and allhad blood pressures under 110 mm. Hg. Eventhese rats had significant hypergranulation of the

TABLE II

The juxtaglomerular indices for eight individual hyper-tensive rats 10 weeks after a clip had been

applied to one renal artery

Blood JG indexpressure of contra-at end of JG* index lateral

Rat experiment of ischemic untouchedno. (mm. Hg) kidney kidney

1 163 68 02 174 105 03 175 129 34 185 40 05 150 53 16 185 68 47 178 62 08 168 84 1

Mean valuesfor the group 172 76±27t 1 ±2

* Juxtaglomerular.t ± Indicates standard deviation.

ischemic kidney. In this group the mean JG in-dex of the constricted kidney was 48 (S.D., ± 15),with values ranging from 26 to 80. The "un-touched" kidney had a mean JG index of 6 (S.D.,± 6), with values ranging from 0 to 20 (Table I).Even though the "untouched" kidney here showedconsiderable degranulation, it was not nearly ascomplete as that seen in truly hypertensive ratswith a similar operation.

The degranulation of the "untouched" kidneysafter unilateral clipping cannot be the result ofrenal hypertrophy, since rats with a single fullyhypertrophied kidney have normal JG indices(Table III).

TABLE III

The mean juxtaglomerular indices for certain groups of Wistar rats with only one kidney present, including groupswith past or present hypertension and control groups

Mean blood Data concerning JG* indicespressure at

end of StandardNo. of experiment Range of error of

Group rats (mm. Hg) Mean values the mean

a. Normal rats with one kidney, on Purinachow 11 34 9 to 50 ±3.6

b. Previously hypertensive rats that be-came normotensive 6 weeks after theexcision of a clipped kidney (eatingPurina chow) 11 96 28 16 to 39 ±2.2

c. Rats with one clipped kidney on a basicdiet containing 0.002% Na 14 174 55 37 to 82 ±4.3

d. Rats with one clipped kidney on thebasic diet plus 0.4% Na 8 184 45 27 to 94 ±7.5

* Juxtaglomerular.

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L. TOBIAN, J. THOMPSON,R. TWEDT, AND J. JANECEK

Partial constriction of the renal artery of de-granulated kidneys. In five rats, one renal arterywas partially constricted with the subsequent de-velopment of hypertension. As noted above, the"untouched" contralateral kidney seven weekslater has almost complete degranulation of the JGcells with an average JG index of 1, and valuesranging from 0 to 4. At this time the renal arteryof this degranulated "untouched" kidney was also"clipped." Four weeks later both kidneys wereexamined. In two of the five rats, the kidneywhich may be assumed to have been previouslydegranulated was still degranulated despite theclipping procedure. However, in the three otherrats the previously degranulated kidney had defi-nitely regranulated as a result of the second clip-ping operation. The JG indices were 36, 28, and14 in the previously degranulated kidneys of theserats.

The effect of a low sodium diet on the granula-tion of the juxtaglomerular apparatus. Three ex-periments were done to assess the influence of theamount of sodium in the diet on the degree ofjuxtaglomerular granulation. In these studiesthe diet consisted of purified materials, including20 per cent casein, 70 per cent sucrose, 5 per centcorn oil, 5 per cent sodium-free salt mixture madewith reagent-grade chemicals, and vitamin sup-plementation. This constituted our "low sodiumdiet" and contained 0.002 per cent sodium. Ourcontrol diet with 0.4 per cent sodium was madeby adding pure sodium chloride to the basic lowsodium diet. Water was given ad libitum.

The two diets were offered to two groups ofnormal Wistar rats. A group of eight rats on thediet containing 0.4 per cent sodium for five weekshad a mean JG index of 30 (S.D., ± 10). Thegroup of eight rats on the diet containing 0.002 percent sodium for five weeks had a mean JG indexof 48 (S.D., + 10). Thus the rats on a very lowsodium intake had a significantly higher degreeof JG granulation (p = 0.001). This confirms theearlier observations of the Hartrofts (3) (TableI).

The next experiment deals with hypertensiverats with a clip on one renal artery, the other kid-ney remaining "untouched." Sixteen weeks afterthe clip was applied, these hypertensive rats weredivided into two groups which were evenly matchedfor blood pressure. These two matched groups

were placed on the experimental diets describedabove for an additional eight weeks. There was aslight reduction in blood pressure in the group of22 rats on the 0.002 per cent sodium diet. Beforegoing on the diet, the mean blood pressure of thisgroup was 177 mm. Hg. After eight weeks on thediet, this mean had fallen to 169 mm. Hg. Theother group of 13 hypertensive rats on the 0.4 percent sodium diet had an initial blood pressure of173 mm. Hg which rose to 180 mm. Hg while onthis diet. The group of rats on the 0.4 per centsodium diet had the usual almost complete JG de-granulation in the "untouched" kidney. Themean JG index of the "untouched" kidney in thisgroup was 1 and the highest index for an indi-vidual rat was 4. On the other hand, the averageJG index of the "untouched" kidney was 6 in therats on the 0.002 per cent sodium diet, and 7 ratsout of the 22 in this group had indices higher than4. Three rats of this group had indices higherthan 17 in the "untouched" kidney. This wouldbe a very unusual finding in operated hypertensiverats on higher sodium intakes. The differencein granulation of the "untouched" kidney betweenthese two groups is highly significant (p = 0.005).The diet containing very little sodium unquestion-ably increased the JG granulation in these "un-touched" kidneys. Moreover, the slight reduc-tion in blood pressure resulting from the lowsodium intake probably does not account for theincreased granulation, since all hypertensive ratson the low sodium diet had a blood pressure higherthan 140 at the end of the experiment (Table I).

Varying the amount of sodium in the diet didnot greatly influence the JG granulation of the"clipped" kidneys in these rats. The group on 0.4per cent sodium in the diet had a mean JG index of58 in the "clipped" kidney, and the group on the0.002 per cent sodium diet had a mean JG indexof 66. The "clipped" kidneys of both groups arehypergranulated compared to a normal kidney butare not significantly different from one another(Table I).

Other rats were made hypertensive by apply-ing a clip on one renal artery and excising the con-tralateral kidney. All the rats studied had bloodpressures over 140 mm. Hg. Sixteen weeks afterthe operations, they were divided into two groupsevenly matched for blood pressure. One groupwas placed on the 0.002 per cent sodium diet and

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GRANULATIONOF JUXTAGLOMERULARCELLS IN HYPERTENSION

TABLE IV

A comparison of the juxtaglomerular indices in adrenalectomized and in normal Sprague-Dawley rats

StandardNo. of Mean JG* Range of error of

Group rats index values the meant

a. Normal rats on Purina chow andtap water 12 30 18 to 40 :1.9

b. Adrenalectomized rats on Purinachow and 1%saline 12 52 7 to 83 :7.2

* Juxtaglomerular.t p value of the difference between these means: 0.004.

the other on the 0.4 per cent sodium diet for fiveweeks and they were then killed. The blood pres-sure of all the rats on either diet remained above140 mm. Hg at the end of the experiment. The8 rats on the 0.4 per cent sodium diet had a meanJG index of 45, while the 14 rats on the 0.002 percent sodium diet had a mean JG index of 55.This difference was not statistically significant(p = 0.3) (Table III).

The effect on the juxtaglomerular apparatus of"curing" unilateral renal hypertension with a uni-lateral nephrectomy. Eleven rats were made hy-pertensive with blood pressures over 140 mm. Hgby clipping one renal artery as described above.The average blood pressure of this group was159 mm. Hg seven weeks after the operation.From previous experience, we know that the JGapparatus of the "untouched" kidney is virtuallydegranulated seven weeks after the original clip-ping operation in hypertensive rats. At this timethe "clipped" kidney was excised. Every rat hada decided fall in blood pressure following thisprocedure. Ten out of the 11 rats had blood pres-sure levels under 115 mm. Hg two weeks after theremoval of the "clipped" kidney. By six weeksafter removal of the "clipped" kidney, all the bloodpressures were under 111 mm. Hg with an aver-age of 96 mm. Hg. Thus all rats eventually hada blood pressure well within the normal range.When the lone kidney of these normotensive ratswas examined, it had an almost complete regranu-lation of the JG apparatus with an average JG in-dex of 28 and values ranging from 16 to 39 (TableIII). The lone kidney of three of these rats thathad become normotensive was seen two weeksafter the "clipped" kidney had been removed. TheJG indices in these three were 32, 23, and 24,

showing that regranulation was almost completein this short interval.

While it is obvious that regranulation occurs inthe JG apparatus of these rats when they changefrom a hypertensive to a normotensive state, itis of interest to compare the granulation in theselone kidneys with the JG granulation of rats thathave had one kidney excised without having under-gone any clipping operations.

Eleven Wistar rats had one kidney excised andtwo months were allowed to elapse for a new equi-librium to be established. The lone hypertrophiedkidney of these rats was then studied. The aver-age JG index in this lone kidney was 34 (S.D., +

11). Thus the regranulated JG complexes of thepreviously hypertensive rats are not greatly dif-ferent from those of simple one kidney rats. More-over, these simple one kidney rats have a JG gran-ulation quite similar to that seen in completelynormal rats (Table III).

B. Adrenal insufficiency

A number of male Sprague-Dawley rats wererandomly divided into two groups with 12 rats ineach group. The control rats in Group A withintact adrenals ate Purina laboratory chow anddrank tap water ad libitum. The rats in Group Bunderwent a complete bilateral adrenalectomy andsubsequently were allowed to drink only a 1 percent saline solution. They were also eating Pu-rina laboratory chow. All of these rats were killed18 days after the adrenalectomy of the rats inGroup B. The mean JG index in the normal ratsof Group A was 30 (S.D., + 6). The mean JGindex in the adrenalectomized rats of Group B was52 (S.D., + 24). This difference was highly sig-nificant with a p value of 0.004. These results are

665

L. TOBIAN, J. THOMPSON,R. TWEDT, AND J. JANECEK

similar to those found by Dunihue in adrenalecto- on the low sodium diet became hypertensive dur-

mized cats (10) (Table IV). ing the course of the experiment, and the average

blood pressure for the group was 99 mm. Hg atC. Rats treated with desoxycorticosterone in con- the end of the experiment. These results confirm

junction with either high or low sodium diets the previous observations of the Hartrofts (11)

Twenty-nine rats were given a subcutaneous (Table V).injection of 125 mg. of desoxycorticosterone tri- The rats receiving DCTMAin conjunction withmethylacetate (DCTMA).4 This injection was a diet low in sodium had a much lower JG indexrepeated in four weeks. One group with 10 of than the control group receiving the diet only.these rats was fed a "synthetic" diet containing This was true in spite of the fact that their bloodonly 0.002 per cent sodium (described above), and pressure was lower than that of the sodium-de-water ad libitum. Another group with 19 of these ficient control group. This would seem to indicaterats was fed this same diet with 0.4 per cent so- that DCTMAalone has a significant ability todium added (described above) and was allowed cause degranulation of the JG cells independentto drink nothing but 1 per cent sodium chloride. of hypertension (Table V).A control group that was fed the sodium-deficientdiet but did not receive DCTMAcan also be com De Postdesoxycorticosterone hypertension

pared with the above groups. It is well known that desoxycorticosterone inAll rats were killed two and a half weeks after conjunction with a generous intake of sodium will

the second injection of DCTMA. The group on produce hypertension in rats. An example of thisDCTMAand a high sodium intake had a virtual effect is demonstrated above. It has also been

disappearance of the JG granulation. Their av- shown that in certain rats this hypertension may

erage JG index was 0.3 with a range of 0 to 3Stergen out ofde the 193ratshad Jindies of 0t persist long after completion of desoxycorticoster-Sixteen out of the 19 rats had JG indices of 0.n bopinadwietert r aigi h*one absorption and while the rats are taking in the

All of these rats had some elevation of blood pres-

sure at the end of the experiment with an average .a

final blood pressure of 142 mm. Hg. On the pertension has been called "postdesoxycorticoster-. . . ~one" hypreso n Sapieeapeo hother hand, the group of rats that received ypertension and is a prime example of the

DCTMAand a diet very low in sodium did not principle that a hypertension may persist longshow this extreme JG degranulation. The mean after its original cause has been removed.JG index was 17 for the group. None of these rats To produce this type of hypertension a 25 mg.index

ish17

to othank Dr. bert G un tof these

C bat desoxycorticosterone acetate pellet was im planted*Wwis to thn.roetGan fteC subcutaneously in each of 150 male weanling rats.Pharmaceutical Company for generously supplying the

desoxycorticosterone. Another similar pellet was implanted five weeks

TABLE V

A comparison of the juxtaglomerular indices in Wistar rats receiving desoxycorticosterone andeither a high or low sodium intake

Mean blood Data concerning JG* indicespressure at

end of StandardNo. of experiment Range of error of

Group rats (mm. Hg) Mean values the mean

a. Rats receiving DCTMAt and 1%NaCl in food and water 19 142 0.3 0 to 3 4-0.lt

b. Rats receiving DCTMAand a verylow sodium diet 10 99 17 0 to 51 46.2§

c. Control rats receiving a very lowsodium diet 8 110 48 29 to 62 :1:4.0

* Juxtaglomerular.t Desoxycorticosterone trimethylacetate.t p value of the difference between groups a and b: 0.009.§ p value of the difference between groups b and c: 0.00006.

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GRANULATIONOF JUXTAGLOMERULARCELLS IN HYPERTENSION

TABLE VI

A comparison of the juxtaglomerular indices in "postdesoxycorticosterone" rats with either elevated or normal blood pressure

Mean blood Data concerning JG* indicespressure at

end of StandardNo. of experiment Range of error of

Group rats (mm. Hg) Mean values the meant

a. Post-DCAt rats with hypertension 17 160 7 1 to 15 1.0b. Post-DCA rats with normal blood

pressures 16 104 25 8 to 50 4:3.1

* Juxtaglomerular.t p value of the difference between these means: 0.0000001.: Desoxycorticosterone acetate.

later. The rats were all drinking a 1 per cent so-dium chloride solution for six months after thefirst pellet implantation. Most of them becamehypertensive during this period. They were thenoffered tap water for the next four and a halfmonths, at which time the experiment was termi-nated. The blood pressure of each rat was de-termined twice during the last week of the experi-ment, and the average of these readings was con-sidered the blood pressure for each rat. Whenthe rat was killed, the entire subcutaneous areaof the back was examined to be certain that bothpellets had been completely absorbed. In tworats the remnant of a pellet was seen and theserats were excluded from the study. This wasdone to insure that all the rats were indeed in the"postdesoxycorticosterone" phase. Seventeen ratsremained moderately to severely hypertensive af-ter the pellets had completely disappeared. Theblood pressure was above 140 in every rat of thisgroup with a mean of 160 mm. Hg. Many morerats had very mild hypertension with blood pres-sures between 120 and 140 mm. Hg. Sixteen ratshad blood pressures in the normal range (below118 mm. Hg) in the postdesoxycorticosterone pe-riod. The mean blood pressure of this group was104 mm. Hg. The JG granulation of the hyper-tensive group and the normotensive group wascompared. In the normotensive group the meanJG index was 25 (S.D., ±12). In the hyperten-sive group the mean JG index was 7 (S.D., ±4).This difference was quite significant (p = 107)(Table VI).

E. Experimental hypertension in rats associatedwith adrenal cortical regenerationSkelton has recently demonstrated a form of

experimental hypertension in rats which appears

during the regeneration of adrenal cortical tissuefollowing adrenal enucleation (13). In creatingthis experimental situation, we removed one kid-ney and one adrenal in male weanling Holtzmannrats. The contralateral adrenal capsule wasnicked, and the medullary and cortical tissue wasgently pressed out through the nick leaving onlythe capsule and a thin layer of adrenal corticalcells subjacent to it. These rats were then given a1 per cent saline solution to drink. Many of themsubsequently developed hypertension. A controlgroup of weanling rats with intact adrenals wasalso included in the experiment. The rats in thiscontrol group had one kidney excised and weredrinking only a 1 per cent saline solution. Halfof the rats with the single regenerating adrenalhad a blood pressure over 140 mm. Hg six weeksafter their initial operation. The average bloodpressure of these nine rats at this time was 171mm. Hg. Their average JG index was 7. Thecontrol group had a mean blood pressure of 121mm. Hg (S.D., ±7), and an average JG index of20. The rats with the hypertension of adrenal'regeneration had a significantly lower JG indexthan the operated control rats (p = 0.0004)(Table VII). In many ways the hypertension ofadrenal regeneration resembles the hypertensionassociated with desoxycorticosterone. However,the JG granules in desoxycorticosterone hyper-tension practically disappear, whereas they aredefinitely present, even if in diminished numbers,in the hypertension of adrenal regeneration.

DISCUSSION

Unfortunately these studies do not delineate theexact function of the JG apparatus. The granularcells in it have all the appearances of secretorycells, even when seen with the electron microscope

667

L. TOBIAN, J. THOMPSON,R. TWEDT, AND J. JANECEK

TABLE VII

The juxtaglomerular indices in rats* with hypertension of adrenal regeneration and in control rats with intact adrenals

Mean blood Data concerning JGt indicespressure at

end of StandardNo. of experiment Range of error of

Group rats (mm. Hg) Mean values the mean t

a. Rats with adrenal regeneration hyper-tension 9 171 7 0 to 16 +1.7

b. Control rats with intact adrenals 9 121 20 10 to 33 d3.0

* All rats in this study had one kidney excised and were drinking a 1 per cent saline solution throughout the ex-periment.

t Juxtaglomerular.t p value of the difference between these means: 0.0004.

(14). It is reasonable to assume that the secretorystate associated with hypergranulation is the op-

posite of that associated with degranulation. Itis not even certain whether the degranulation ofthe JG apparatus is associated with rapid secretionor with a cessation of secretion. However, thestate of granulation varies greatly in certain hy-pertensive states as well as with variations in so-

dium intake and adrenal function. It has beenthought by some that the JG granules may con-

tain renin. No direct proof of this exists, butPeart, Gordon, Cook, and Pickering have shownin the rabbit that renin exists only in or aroundglomeruli and not in purely tubular areas (15).Hence it is possible that the hypothesis is correct.

These juxtaglomerular secretory cells are abun-dant in all mammalian species, including man (Fig-ure 1). They have also been seen in lower verte-brates such as the frog (16). These cells in thewall of the afferent arteriole are next to themacula densa, a specialized area of cells in thewall of the distal convoluted tubule which is al-ways near the vascular pole of the glomerulus(17). There is also a rich network of nerves

surrounding all the specialized juxtaglomerularstructures (18). It is well known that the mam-

malian kidney has an uncannily accurate ability tokeep renal blood flow and glomerular filtrationconstant in the face of widely varying perfusionpressures (19). Renal vascular resistance can

make tremendous adjustments to maintain thisautoregulation of blood flow and filtration, even

when all vasomotor nerves leading to the kidneyhave been severed and under conditions of local

adrenergic blockade with RegitineQ (20). Thesechanges in renal resistance are much more re-sponsive to changes in perfusion pressure ratherthan to changes in blood flow (20). The wholefabric of evidence indicates that the main changesin resistance which accomplish renal autoregula-tion can be reasonably assigned only to the pre-glomerular arteriole. Recent experiments indicatethat the cell separation theory cannot account formore than a small part of the changes in resistanceoccurring during autoregulation (20). As men-tioned above, the granular JG cells are in a perfectanatomic situation to sense changes in perfusionpressure. Being in the wall of the arteriole, theycould easily sense changes in the stretch of the walland could then secrete more or less of their hor-mone in response to this stimulus. Changes inperfusion pressure would also tend to alter theglomerular filtration rate. Such a change in fil-tration rate might somehow be sensed by the spe-cialized cells of the macula densa in the distal tu-bule; Since the cells of the macula densa arequite close to the JG secretory cells, without evenreticular fibers separating them (21), the JG cellsmight change their rate of secretion in response toa change in glomerular filtration. In either case,this substance might act primarily on the sphincterof small smooth muscle cells located in the afferentarteriole in the region between the JG secretorycells and the glomerular capillary tuft. BothClara (22) and Appelt (23) have described thissphincter situated at the entrance into the glomeru-lus. The state of contraction of the cells in thismicromuscular sphincter could provide the fine

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adjustment of resistance to blood flow that wouldhave to be present on the upstream side of theglomerular tuft in order to have autoregulation ofboth renal blood flow and filtration.

The data obtained in our study partly supportsome of this speculation. In situations where therenal arterial bed is fully exposed to hypertensiveblood pressures, the granulation of the JG cellstends to be decreased. Examples of this can befound in the "untouched" kidney of unilateral re-nal hypertension, in post-DCA hypertension, inacute DCTMAhypertension, and in the hyper-tension of adrenal regeneration. When the bloodpressure in the renal arterial bed is lowered, theJG cells tend to increase their granulation. Ex-amples of this can be found in the kidneys of com-pletely adrenalectomized rats, in the various"clipped" kidneys of rats where a clip on the renalartery of either a normal or an "untouched" kid-ney decreases the pressure distal to the clip, andespecially in the "untouched" kidney of rats inwhom renal hypertension has been "cured" byexcising a "clipped" kidney. Wide variations ofsodium intake could also conceivably operate inthis manner. High intakes of sodium tend to in-crease blood pressure and blood volume; exceed-ingly low intakes tend toward a low blood pressureand blood volume. Either extreme would be likelyto produce some change in the stretch of the JGcells.

Whether or not the substance in the granulesof the JG cells influences local renal circulation,the question also arises as to whether it influencesthe caliber of arterioles throughout the systemiccirculation. If it did, it could be very importantin producing the increased generalized peripheralresistance of arterial hypertension. In hyperten-sion it could act either by producing an over-abundance of a pressor substance or by failing tosecrete the normal amount of a depressor sub-stance. Since hypertension can be produced byremoving both kidneys (24, 25), the latter con-cept is possibly more attractive. If the substanceof the JG cells had a role in renal autoregulationas well as an effect on systemic arterioles, its ef-fect in the hypertension of renal "ischemia" wouldhave to be the opposite for the two different setsof arteriolar smooth muscle. In partial constric-tion of a renal artery, autoregulation would de-

mand a relaxation of the preglomerular sphincter;while at the same time a generalized constrictionof arteriolar lumina would be occurring through-out the systemic circulation.

It is difficult to explain the partial degranula-tion of the JG cells in the "untouched" kidney ofrats whose opposite kidney has been clipped with-out the development of hypertension. Ninety-fiveper cent of rats with unilateral clipping develop atleast a slight elevation of blood pressure, and theseparticular rats represent the exceptional 5 percent. Since one kidney has been clipped, these ratsprobably have a tendency to become hypertensive.It may be that the JG cells of the "untouched" kid-ney in this group of rats became degranulated inresponse to this tendency to hypertension; andtheir process of degranulation, if it represented therelease of a depressor substance or the failure torelease the usual amount of a pressor substance(see above), might actually be opposing this hy-pertensive tendency and thereby helping to main-tain the normotensive state. This would implythat the JG cells of the "untouched" kidney wereexerting some sort of antihypertensive activity.If this antihypertensive mechanism became ex-hausted, similar to exhaustion of the pancreaticfl-cells, the tendency to hypertension would beconverted to actual hypertension. Weneed muchmore evidence before this idea can be anythingmore than pure speculation.

With our present lack of information, we couldbe completely overlooking even more importantdeterminants of juxtaglomerular granulation. Inany case, the JG cells do undergo marked changesin granulation in various forms of experimentalhypertension. Their exact role in these experi-mental models deserves further study.

SUMMARY

The juxtaglomerular index, which indicates thedegree of granulation of cells in the juxtaglo-merular aparatus, averages 35 in normal Wistarrats. Whenhypertension is produced by partiallyconstricting one renal artery, the juxtaglomerulargranules double in the ischemic kidney and virtu-ally disappear in the "untouched" contralateralkidney. If the ischemic kidney is then excisedseven weeks later, the hypertension disappears

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and j uxtaglomerular granules reappear in normalabundance in the previously completely degranu-lated contralateral kidney. After partial con-striction of one renal artery, the juxtaglomerulargranules in the opposite kidney disappear; but ifa constriction is then placed on the artery of thisdegranulated contralateral kidney, the juxtaglo-merular granules reappear. Another group ofrats with one ischemic kidney and a contralateraldegranulated kidney was placed on a sodium de-ficient diet. In half the rats, this diet caused thejuxtaglomerular granules to reappear in the previ-ously degranulated kidney. This diet also in-creased granulation in normal rats. Partiallyconstricting the renal artery in a rat with onlyone kidney will produce hypertension, and thegranulation of the lone ischemic kidney remainsslightly above normal.

Adrenalectomy doubles the juxtaglomerular in-dex; desoxycorticosterone and salt cause thegranules to disappear while concomitantly produc-ing hypertension.

Some rats that had received desoxycorticos-,terone pellets remained hypertensive long afterall pellets had been absorbed. The mean juxta-glomerular index for this group was 7, comparedto a value of 25 in a similarly treated group thathad become normotensive following the absorp-tion of the desoxycorticosterone pellets.

Hypertension was produced in other rats byremoving one adrenal and enucleating the other.Both the hypertensive and the control groups hadone kidney removed and were drinking saline solu-tion. The hypertensive group with adrenal re-generation folowing enucleation had a juxtaglo-merular index of 7, while the control group withintact adrenals had an index of 20.

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