An immunoassay to determine the effects of changing gravity and weightlessness on vasopressin control system is discussed. Seven extracts from persons known to have inappropriate ADH syndrome secondary to pulmonary oat cell cancer were examined. The extracts had previously been subject to bioassay in rats. The intent of the examination was to determine if the vasopressin content could be confirmed immunologically. The results compared favorably with the values obtained by biological assay

Moran, W. H.

English

NASA Technical Reports Server

NATIONAL AERONAUTICS AND SPACE ADMINISTRATION
PROGRESS REPORT
Report Prepared by: Walter H- Moran, Jr., M.D.
For Period: February 16, 1971 to February 15, 1972
Grant: NGR-^ 9-001-019
Title: The Effect of Changing Gravity
and Weightlessness of Vasopressin
Control Systs'Jis
A. TUMOR IMMUNOASSAY
Seven extracts were sent to Dr. Myron Miller's laboratory in Syracuse
for immunoassay. These extracts came from three patients known to have
the inappropriate ADH syndrome secondary to pulmonary oat cell carcinoma.
These extracts had previously been subject to bioassay in the rat and we
now wished to see if their vasopressin content could be confirmed immuno-,
logically. The following results were obtained from the immunoassay.
TABLE I
IMMUNOASSAY OF SELECTED EXTRACTS FROM PATIENTS WITH
INAPPROPRIATE ADH SYNDROME
# . ADH
(MU/ML)
3100 POOLED PLASMA - GM 1.4
3 - 0 1 " " - CN 0.98
. 3 1 0 2 C H R O M A T O G R A P H 1C PEAK, POOLED PLASilA - GM 0 .11
3103 " " > " " - WC 0.48
3104 " " , LIVER METASTASES - CN GT 10
3105 . " " , LUNG TUMOR - CN GT 10
3106 " " , LUNG TUMOR - CM 0.72
GT = GREATER THAN
These results compaired favorably with the values obtained by biological
assay. It appears that the active agent in these cases has the chromato-
graplic mobility of arginine vasopressin and also its.biologic and immuno-
chemical properties and therefore is probably identical to arginine vasopressin.
B. DEVELOPMENT OF ANTIBODIES TO OXYTOCIN AND VASOPRESSIN
Four rabbits are being immunized with "coupled" albumin-oxytocin complex
mixed in Freunds adjuvant. Six rabbits ar,e just beginning a course of similar
treatment using an albumin-arginine vasopressin complex. Serum is being collected
at monthly intervals and will be checked for antibody development shortly.
https://ntrs.nasa.gov/search.jsp?R=19720015420 2020-03-11T18:49:45+00:00Z
C. URINE VASOPRESSIN EXCRETION
Since 1962, aliquots of urine have been collected, frozen and
stored in our laboratory from patients undergoing studies of their
plasma ADH levels. Recently, a program of determining the ADH content
of the samples was undertaken to ascertain the range of urinary ADH
levels that might be encountered in pathological states and to see
if any relationship exists between urinary ADH excretion and plasma
ADH levels. There were no normal subjects in this group.
The urinary assay of ADH consisted of a modification of the plasma
ADH bioassay precedure currently used in this laboratory. The urine
samples which had been frozen without the addition of any preservatives
were rapidly thawed and extracted with 2 volumes of ether. The washed
urine was then adjusted to a pH of 4.2 and added to a column of CG-50
resin. From this point on the precedure was identical to the plasma
bioassay. The extracts of urine produced antidiuretic responses that
were similar to those produced by the arginine vasopressin reference
standards. The high levels in some of these samples indicated that
very little, if any, ADH had been lost during prolonged storage.
At present, 90 urine samples have been assayed, 57 of which had
plasma ADH levels so timed that ADH clearances could be calculated.
Five patients (table II) might be classified as having diabetes
insipidus (Dl). DI-1 and DI-2 had pulmonary carcinoma with metastases
to the brain. DI-3 had carcinoma of the thyroid with cerebral metastises
as a cause of her DI. DI-4 was a boy with DI and DI-5 was a diabetic
with severe retinopathy who was undergoing a pituitary stalk.section.
The urinary ADH levels were very low in this group ranging from not
detectable to 9.^juU/ml. Plasma samples were infrequently taken because
of the difficulty in detecting ADH levels in DI. The high plasma ADH
levfcl in DI-2 occurred following a dose of vasopressin and the high
plasma levels in DI-5 occurred during the surgical procedure. The mean
urinary ADH excretion rate in DI without an open loop stimulus was 9.93 i
(N = 12)juU/min. The mean urinary ADH excretion when injected pitressin
had achieved a clinically satisfactory response (DI-2) was 11.6 + .9
(N = ?) ;iU/rain.
Two patients had samples taken while they were undergoing surgical
procedures (table III). SURG-1 had a partial gastrectomy. SURG-2 had
a partial colectomy. Their mean urinary ADH excretions were 339 ± 58
(N = 6) and 241 +29 (N = 6)/iU/min., respectively.
Four patients had the Inappropriate Antidiuretic Hormone Secretory
Syndrome (table IV). IADHS-1 had severe pulmonary emphysema as a cause
of his inappropriately high ADH levels. While his urinary excretion
rate (8.6 ± 3 (N = 4) ;iU/min.) and plasma levels (2.6 + .4 (N = 4/iU/ml.)
might be considered to be within the normal range, they were difinitely
too high for his plasma osmolality of 240 mOsm/Kg. and were unresponsive
to water loading. IADHS-2, IADHS-3, and IADHS-4 had oat cell pulmonary
carcinomas. Their urinary ADH excretion rates were quite high (411,
50.1, and 224juU/min.) and were unresponsive to water loading.
The average values for urinary ADH excretion, plasma ADH concent-
ration and the renal clearance of ADH are listed in Table V. The ADH
clearance data of the DI patients was insufficient to warrant any
conclusion. The ADH clearance values of all of the other patients,
except IADHS-2 varied between 3.7 and 11.5 ml/min. but were not really
significantly different. The 3^.5 ml/min. clearance of IADHS-2 cannot
be explained at present.
Careful examination of the data in tables II, III and IV revealed
that small changes in urinary flow in patients with low urinary outputs
were associated with small changes in urinary ADH excretion that were
directly proportional and were probably related to small changes in renal
clearance. If the urinary output was high and not completely of osmotic
nature the urinary flow was inversely proportional to ADH excretion
(DI-2 and DI-5).
A plot of all 5? urinary ADH excretion rate - plasma ADH concen-
tration value pairs (figure 1) demonstrated a fairly linear relation-
ship between them. Five of the points from SURG-1 fell quite far above
the line and represented a period when both renal and hepatic clearance
could have been reduced because of events surrounding the surgical pro-
cedure. The other point of this patient fe!3. on this line and represented
the highest urinary ADH excretion rate observed (6l6 ;uU/min.). The
points from IADHS-2 fell below this line as mentioned earlier.
In general, the urinary ADH excretion rates followed plasma ADH
concentrations giving an average renal clearance of ADH of about
10 ml/min.
TABLE II - DIABETES INSIPIDUS
• URINE . PLASMA
•DURATION. RATE . C A D H 3
• CM IN) . (ML /WIN) . -CUU/ML) . (
V . U
DI
.
*
DI
.
.
.
.
.
.
*
.
DI
.
'*
•
.
*
*
*
.
DI
.
.
.
. •
.
.
*
DI
.
.
.
.
.
•
-1
720
720
-2
60
60
60
60
60
60
60
60
-3
60
15
15
15
5
5
5
5
5
-4
60
60
60
60
60
60
60
-5
240
120
120
120
360
360
0.67
1.39
18.92
. 1 0 . 8 3
6.33
4.25
3.00
2.67
2.33
1.65
0.93
1 . 53
1.40
2.67
4.33
4.80
4.80
5.40
4.67
1.50
1.80
1.08
. 0.50 •
0.83
1 .25
. 1.00
0.73
0.82
0.53
0.53
0.90
3.32
V
1.4 .
1.2 .
0.6
0.3 .
1.1
3.3 .
4.6
4.0
5.5 .
7.3
0.6
0.3 .
0.3 .
N.D.
N.D. .
N.D.
N.D.
N.D.
N.D*
1.4
0.3
0.7 .
1.4 .
1 .2
0.9 .
0.8 .
1.9 .
9.4 i
7.5 .
3.8
5.3 .
0.8 •
ADH
UU/MIN) .
UV
1.0 .
1.6
11.0 .
3.2
7.1 .
14.2 .
13.6
10.6
12.8
12.0 .
0.6
0. 5 .
0.3 •
*
•
*
.
.
.
2.1 .
0.6
0.8
0.7 .
1.0 .
1.1 •
0.8
1.4
7. 7 .
4.0 «
2.0 .
4.8
2.8
C A D H 3 . CADH
CUU/ML) . (ML/MIN) .
P . UV/P
0.0 . .
. • ---- .
. .
6.3 0* 5 .
.
•»•»•»» • » •• * — ^
•» — » — • ^ ^ • ••. ^
. - _ _ - .
. « .
• «_.*. e . — — «• — ^
• W K • * ** 0
. .... .
0.7 . 0.5
.... . .... .
. .
....
 9 - . . . . -0
, .... .
.. '_.._ .
. — . . _ _ _ - .
w~» • _ • » • • ^
. .... ' .'
.... . - — - .
.... ^ .... ^
. _ „ _ _ .
. .
12.8 • 0.1
6.8 • 1.1
.... . .... .
3.1 . 0.7 .
2.7 . 1.8
-» * _ *
 m •• — •»_ 9
* N.D. = NOT DETECTABLE
TABLE III - SURGICAL PROCEDURE
. DURATION.
. (MIN)
•
SURG-1
. 30 .
30 •
30 .
30 .
30 .
30 .
SURG-2
. ' 30 .
30 .
30 .
30 .
30 .
TOl
UR]
RATE
(ML/MIN)
V
1.30
2.47
4. 17
1 .90
1.83
1 .30
1.83
5.03
8.63
7.83
6.23
[N!
•
*
•
*
•
•
•
•
•
•
•
•
•
•
CADH]
(UU/ML) .
U .
474.
1 18.
81 .3
117.
135.
243.
149.
59.0 .
29. 1
29.2 .
47.0
ADH
(UU/MIN)
UV
616.
291.
339.
222.
248.
315.
273.
297.
251.
229.
293.
. PLASMA
CADH]
. (UU/ML)
P
41 .2
44.2
. . 41.0
54.2
54.5
45.7
21.8
24.7
17.5
. 23.7
20.8
CADK
. (ML/MIN) .
UV/P
15.0
6.6
8.3
4. 1
4.5
6.9
12.5
12.0
14.4
9.7
14. 1
30 1.12 96.9 108. 16.7 6.5
TABLE IV - INAPPROPRIATE ADH SYNDROME
.
9
I
.
.
.
.
I
*
.
.
.
.
.
.
.
.
.
I
.
.
.
.
,
.
.
.
.
•
.
.
.
.
.
.
.
.
.
.
DURATION.
CM IN) . (
.
ADHS- 1
1 440
1 440
1440
1440
ADHS- 2
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720
720 .
ADHS- 3
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 •
720 .
720 .
URINE
RATE
:ML/MIN> .
V
0. 44
0.29
0.34
0. 10
0.93
0.85
1.21
1 .46
1 .85
1.95 .
1.31
1.33
0.95
0.93
0.57
1 .04
0.74
1 .00
0.61
0.88
0.73
1.18
0. 46
1.15
0.57
1.13
1.29
1.25
2.08
1.46
0.82
0.96
0.46
0. 54
CADH]
(UU/ML) . <
U
17.5 .
48.9
33.3 •
12. 5 .
359.
340.
445.
341 .
283.
245.
339.
342.
295.
307.
.73.5 .
58.4 .
71.0 .
45.4 .
70.2 .
46.7
63.6
44. 5
56.4 .
54.6 .
58.4 .
47.7 .
39.7
43.6
30.3 «
51.0 .
68.6 .
64.3
104.
64.3 .
. f
ADH
:UU/MIN) . <
UV
7.6 .
14.3 .
11.3
1.3 .
334.
288.
537.
497.
522.
475.
442.
455.
278.
285.
41 .8
60.4 .
52-3 .
45.4 .
42.9 .
41.2 .
46.4 .
52.5 •
25.8 •
62.9 .
33.2 •
53.6 .
51.2 .
54.4 .
63.0 •
74.4 .
56.2 .
61 .6 •
47.6 .
34.8 .
3LASMA .
CADH]
: UU/MD . <
P
1.5 .
2. 4
3.6
2.7 .
11.7 .
9.7 .
9 . 1 .
13.8 .
8.5 .
• .
9.9 .
.
6.3 .
.
4.7
5.8 .
5.5 .
6.5 .
5.9 .
4.2 .
5.0 .
5.9 .
4.7 .
5. 1 .
6.3 .
5.3 .
6.9 .
8.7 .
8.3 .
9.7 .
6.5 .
6.5 .
7.0 .
.
CADH
:ML/MIN)
UV/P
5.0
6.0
3«2
0.5
28.5
29.7
58.9
36. 1
61.8
44. 7
44.8
8.9
10.5
9.6
7.0
7.3
9.8
9.3
8.9
5.5
12.3
5.3
10.2
7.4
6.3
7.6
7.7
8.7
9.5
6.8
•
.
o
o
.
e
o
.
•
.
*
.
o
.
.
. .
.
.
o
.
.
•
.
.
.
.
e
.
.
.
.
.
.
.
.
.
.
.
TABLE IV - INAPPROPRIATE ADH SYNDROME (CONT.)
•
» DURATION.
» (MIN)
* .
IADHS-4
720 .
720 .
720 .
720 .
720 .
720 .
720 .
720 .
. 720 .
• 720
720 .
720 .
UR
RATE
(ML/MIN)
V
0.80
1 .29
1 .04
1.53
2.64
3. 1 1
0.78
1. 10
0.61
0.83
0.60
0.78
IN
•
0
e
•
•
•
*
•
•
•
•
•
•
•
*
E
CADH]
(UU/ML)
U
230.
177.
181 .
174.
125.
1 16.
231 .
226.
198.
257.
244.
283.
ADH
. (UU/MIN)
UV
184.
228.
189.
265.
329. .
361.
180.
243.
1 19.
214.
146.
220.
. PLASMA
CADH 3
. (UU/ML)
P
13.7
16. 1
20. 1
17.3
18.7
.
21.0
. '
21. 1
. .-__
, 21.2
.
. •
CADH
. ( ML /M IN) .
UV/P
13.4
14.2
9.4
15.4
17.7
. • .
8.6
. .
5.7
. .
6.9
. .
TABLE V - AVERAGES FROM TABLES I, II, III, AND IV
NAME
DI-1
DI-2
DI-3
DI-4
DI-5
URINE ADH
(UU/MIN)
UV
1.3 + .3<N=2>
12.6 + .63CN=5)
0.5 + . 1(N = 3)
1.0 + «2(N=7)
3.8 + .9<N=6)
PLASMA ADH
. CUU/ML)
P
0.0
6.3
0. 7
6. 4 + 2. (N=4)
ADH
(t
0.5
0.5
0.9
SURG-1 339. + 58<N=6)
SURG-2 241. + 29CN=6)
I ADHS-1 8.6 + 3 . < N = 4 )
IADHS-2 411. + 3 3 ( N = 1 0 >
IADHS-3 50.1 + 2 . 6 ( N = 2 0 )
IADHS-4 224. + 20OM=12>
46.8 + 2 . 5 ( N = 6 >
20.9 + 1.3(N=6>
2.6
9.9
6.2
18.7
, 4CN=4)
.9CN=7)
.3CN=19)
.97(N=8)
 CLEARANCE
ML/MIN)
UV/P
. 6 < N = 4 )
7.6 + 1 . 6 ( N = 6 )
1 1 . 5 + 1 . 2 ( N = 6 )
3.7 + 1 . 2 C N = 4 )
43.5 * 5 . 0 ( N = 7 )
8.4 + . 4 C N = 1 9 >
1 1 . 4 + I .SCN-8 )
8
50
^^i
5 403
z:
(/) -in
uj 30
tr.
a.
o
^ 20
<
to
2 lo
r»
oo
-
0o
o
-
• .
Qo
O O C^ ° O
o
f. OO\J O
0 0
0
 0
00° ° ° o°
• ^o*^ °
^ o
E M ' 1 I I I I 1 I I I I I 1
0 100 200 300 400 500 600
URINE VASOPRESSIN (juU/min)
figure 1
D. PUBLICATIONS:
1. Moran, W. H., Jr.: CPPB and Vasopressin Secretion, Editorial
Views, J. of Anesthesiology, 3^ :501-50^  (1971).
10
APPENDIX I - PAGE 1
EXPERIMENTS SUPPORTED IN PART BY NGR-001-019
THROUGH 10/20/71
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ADH STABIL ITY I, EFFECT OF TEMPERATURE
EFFECT OF BLEED ON ADH AND CRF
ADH STABIL ITY II, REPEATED FREEZE- T H A W PLASMA
ADH STABIL ITY III, REPEATED FREEZE - T H A W STANDARD SOLUTION
ADH STABILITY IV, EFFECT OF TEMPERATURE
ANTIDIUhETIC A C T I V I T Y OF ANGIOTENSIN
ANGIOTENSIN RECOVERY ON CG-50 RESIN COLUMNS
ADH S T A B I L I T Y V, NA O X A L A T E AND EDTA AS INHIBITORS
ADH RECOVERY I, NO ETHER WASH
ADH STABIL ITY VI, NA OXALATE AND EDTA AS INHIBITORS
CRF ISOLATION I, TO AMES
CRF ISOLATION II, TO AMES
ADH RECOVERY II, NO ETHER WASH
ADH RECOVERY -I I I, PH 4.0 RESIN COLUMN
ADH RECOVERY IV, EE VS NEE /
ADH RECOVERY V, EE VS NEE .
ADH RECOVERY VI, EE VS NEE
CRF ISOLATION III, TO AMES
VEITNAM SAMPLES, CRF TO AMES
ADH ISOLATION I, DIURETIC PEAK
ADH ISOLATION II, NAOX AND EDTA ON DIURETIC PEAK
ADH RECOVERY VII, PLASMA
ADH ISOLATION III, NA ION IN E X T R A C T
ADH ISOLATION IV, FH IOGL Y C O L A T E CORD BLOOD
ADH ISOLATION V, THIOGL YCOLATE CORD BLOOD
ADH ISLOATION VI, PITRESSIN VS. USP POST PIT REF IN RAT
ADH STABILITY VII, STORAGE OF DILUTED EXTRACT INJECT SOL
ADH CHROMATOGRAPHY I, PITRESSIN
ADH CHROMATOGRAPHY II, ELUTION FROM PAPER
ADH CHROMATOGRAPHY III, USP PIT REF
ADH CHROMATO GRAPH IT IV, RESIN + PAPER WITH USP REF
ANGIOTENSIN CHROMATOGRAPHY I,
O X Y T O C I N CHROMATOGRAPHY I,
ADH CHROMATOGRAPHY VIII, PLASMA-LUNG CA POOL: GM
CRF ISOLATION IV, TO AMES
ADH CHROMATOGRAPHY V, CORD BLOOD
ADH RECOVERY VIII, RESIN PLUS PAPER CHROMATOGRAM
ADH CHROMATOGRAPHi ' VI, USP REF STD - DIST COEF
ADH CHROMATOGRAPHY VII, CG-50 ION EXCHANGE COL
BIOLOGICAL HALF-LIFE OF ADH I, DOG
BIOLOGICAL HALF-LIFE OF ADH II, DOG
EFFECT OF TILT ON ADH SECRETION
ADH CHROMATOGRAPHY IX, PLASMA-LUNG CA POOL: CN
ADH CHROMATOGRAPHY X, PLASMA-LUNG CA POOL: WC
ADH CHROMATOGRAPHY XI, TISSUE-NORMAL LUNG: CN
APPENDIX I - PAGE 2
EXPERIMENTS SUPPORTED IN PART BY NCR-001- 019
THROUGH 10/20/71
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ADH CHROMATOGRAPHY
ADH CHROMATOGRAPHY
ADH CHROMATOGRAPHY
XII, TISSUE-LUNG CA: CM
XIII, TISSUE-LIVER METAST.:
XIV, PL ASM A-HEAD TRAUMA: FJ
COSM VS. DOSE
CM
ADH RESPONSE I, URATE, CHOH,
APOLLO 9 PLASMA SAMPLES
ADH IMMUNIOASSAY-B IOASSAY COMPARISON I
ADH IMMUNIOASSAY-B IOASSAY COMPARISON II
BIOLOGICAL HALF-LIFE OF ADH III, DOG
ADH IMMUNIOASSAY-B IOASSAY COMPARISON III
SHOCK PLASMA ULTRAFILTRATE I
BIOLOGICAL HALF-LIFE OF ADH IV, DOG
ADH PLASMA, DIABETES INSIPIDUS
ADH IMMUNIOASSAY-B IOASSAY COMPARISON IV
ADH PLASMA, INAPPROPRIATE SECRETION SYNDROME
ADH PLASMA, HEAD TRAUMA I
BIOLOGICAL HALF-LIFE OF ADH V, DOG
DIABETES INSIPIDUS I, RAT URINE
ADH REFERENCE STANDARDIZATION I, G VS V STASTICAL STUDY
ADH REFERENCE STANDARDIZATION II, G VS V STASTICAL STUDY
ADH INFUSION STANDARD, DOG
BIOLOGICAL HALF-LIFE OF ADH VI, DOG
DIABETES INSIPIDUS II, HUMAN URINE
ADH REFERENCE STANDARDIZATION III, G VS V STASTICAL STUDY
ADH REFERENCE STANDARDIZATION IV, G VS V STASTIAL STUDY
ADH PLASMA, IADH SYNDROME
ADH PLASMA, IADH SYNDROME-HEAD INJURY
ADH IMMUNIOASSAY-BIOASSAY COMPARISON V
ADH REFERENCE STANDARIZATION V
ADH REFERENCE STANDARDI TiATI ON-SYNTHETI C ARGININE VASOPRESSIN
EFFECT OF ADH INFUSION ON RENAL BLOOD FLOW
EFFECT OF ADH INFUSION ON RENAL BLOOD FLOW
COLUMN RECOVERY EXPERIMENT
COLUMN RECOVERY EXPERIMENT
COLUMN RECOVERY EXPERIMENT
SAMPLES FROM TUMOR TISSUE OF PATIENTS WITH IADH SYNDROME
SENT FOR
URINE ADH,
URINE ADH,
URINE ADH,
URINE ADH,
URINE ADH,
URINE ADH,
URINE ADH,
URINE ADH,
URINE ADH,
IMMUNOASSAY
IADHS
IADHS
IADHS
PARTIAL GASTRECTOMY
HYPOPHSECTOMY
LUNG CA
DIVERTICULITIS
DI
APPENDIX I - PAGE 3
E X P E R I M E N T S S U P P O R T E D IN PART BY N G R - 0 0 1 - 0 1 9
T H R O U G H 10/20/71
H 179091 U R I N E ADH,
H 1 58561 U R I N E ADH, DI
E 107 S I Z E M O R E STDS
H I 65291 U R I N E ADH.. D I
H127365 U R I N E ADH, I A D H S


The effect of changing gravity and weightlessness of vasopressin control systems

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