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The effect of the cold pressor test on vasopressin secretion in man
Psychoneuroendocrinology, Vol. 11, No. 3, Lap.3 0 7 - 316, 1986.
0 3 0 6 - 4530/86 $3.00 + 0.00 Pergamon Journals Ltd.
Printed in Great Britain.
THE EFFECT OF THE COLD PRESSOR TEST ON VASOPRESSIN SECRETION IN MAN JONATHAN T . EDELSON* a n d GARY L. ROBERTSONt:~ *University of Chicago Pritzker School of Medicine and Charles A. Dana Clinical Scholars Program, University of Pennsylvania School of Medicine, U.S.A.; and i"Department of Medicine, University of Chicago Pritzker School of Medicine, U.S.A.
(Received 1 April 1985; in finalform 19 August 1985) SUMMARY Stress has been thought to induce the release of arginine vasopressin (AVP). We evaluated this claim by studying the effects of a modified cold pressor test on plasma AVP, plasma cortisol, blood pressure, pulse rate, and a number of variables known to affect AVP secretion. In a crossover study design, test and control values were obtained in seven male subjects. The pressor test was found to induce painful stress as evidenced by subjective reports and the objective findings of increased mean arterial pressure 03.9 • 3.1 mm Hg; p < 0.004), pulse rate (9.2 +_ 2.8 beats/min; p < 0.02), and plasma cortisoi (3.5 +_ 0.8 p.g/dl; p < 0.005). In contrast, there were no significant changes in plasma AVP that could be attributed to the cold pressor test. There also were no changes in plasma osmolality, measured plasma solutes, hematocrit or body temperature. An unexpected finding was a premonitory drop in plasma AVP occurring just prior to the pressor test (2.5 _+ 2.0 pg/ml; p < 0.04) and at the comparable time point in the control study (3.1 +_ 1.2 pg/ml; p < 0.001). There were no changes in any of the other measured variables which could account for this drop. We conclude that the cold pressor test is not a stimulus to AVP release and that anticipation of stress may inhibit secretion of this hormone. INTRODUCTION
SINCB Rydin and Verney's (1938) pioneering study in dogs, it has been almost axiomatic that pain, anxiety and other forms of emotional stress cause the release of vasopressin (AVP). However, this concept has been challenged by modern studies that have employed sensitive radioimmunoassays to directly measure the hormone. In rats, a variety of noxious stimuli have been shown to have no effect on vasopressin secretion unless they also induce hyperosmolality, hypovolemia, or other recognized non-osmotic stimuli (Keil & Severs, 1977; Robertson, 1977; Husain et al., 1979). These observations suggest that stress p e r se does not affect vasopressin secretion or that it stimulates secretion directly in some species but not in others. In this regard, Kendler et ai. (1978) suggested that in man pain induces vasopressin release, and Weidler et al. (1981) and v Borman et al. (1981) used the incremental increase in plasma vasopressin after surgery as an indicator of the efficacy of two anesthetics in relieving pain. However, none of these studies provide the kind of ancillary data needed to ascertain whether the observed increase was truly due to pain p e r se or was simply secondary to changes in other recognized stimuli such as hypovolemia, hypotension, or nausea (Robertson, 1977). Clarification of this issue is essential for the proper interpretation of any study of vasopressin function in acutely ill patients with clinical disturbances of salt and water ~/Correspondence to be addressed to: Dr Gary L. Robertson at University of Chicago, 5841 S. Maryland Avenue, Box 131, Chicago, IL 60637, U.S.A. 307
308
JONATHAN T. El)El.SON and GARY L. ROBERT~ON
balance. Therefore, we undertook a systematic study of the effect of stress on vasopressin secretion in man using a variant of the standard cold pressor test (Hines & Brown, 1936). This procedure is associated with a modest degree of pain and results in an acute rise in blood pressure and pulse rate, which is thought to be mediated by the sympathetic nervous system. In addition to these hemodynamic variables, we monitored the activity of the p i t u i t a r y - a d r e n a l axis as well as other variables known to directly affect vasopressin secretion (Moore, 1971; Robertson, 1977, 1985). In this study, we operationally defined "stress" as the induction of elevations in hemodynamic variables and in adrenal activity attributable to the cold pressor test. METHODS
Study design In a cross-over study design, seven subjects were randomized into one of two cells, one of which had the lest protocol first and the control protocol second, and the other of which had the protocols in the opposite order. The test and control protocols were of equal length, distinguished only by the fact thai a specific stress was applied in the test protocol but not in the control protocol. As a control for the possible influence of circadian rhythms, each subject participated in the control and test protocols at the same time of day one week apart.
Sltbjec+ls Seven healthy male volunteers in their third decade gave their written consent to participate in Ihis study, which was approved by the Clinical Investigations Committee at the University of Chicago Pritzker School of Medicine and which was carried out according to the principles of the Declaration of Helsinki. Female subjects ~ere excluded to avoid hormonal variations concomitant with the menstrual cycle (Forsling et aL, 1981). Each ~ubject received financial remuneration for participating in the study.
])1+olO('O] Experimental trials were run at the Outpatient Research Center at Billings Hospital between 0930 h and 1530 h. Subjects tested at 0930 h had no food or liquids after 2400 h the preceding night. Subjects tested at 1330 h had no food or liquids after 0830 h the day of the trial. No drugs, especially tobacco or alcohol, were used within 24 h of the trials. On the day of the first test, written informed consent was obtained, and the subject x~as told whether he would participate in the test or control protocol. The subject emptied his bladder and was x~eighed and placed in a semi-recumbent position. His blood pressure and pulse rate were monitored manually for 15 rain, and then his oral temperature was obtained. At the same time, a history of any adverse reactions to phlebotomy was determined, and the purpose, nature and possible discomforts of the procedure were explained in detail. The subject was asked to report any sensations he felt, especially nausea, faintness or pain, during the ¢c,ttrse of Ihe protocol. With standard sterile technique, a #21 gauge butterfly needle catheter was inserted and secured in an antecubilal vein. One ml of blood was withdrawn and discarded, after which another 10 ml of blood were x~ilhdraw n, seven of which were transferred to a pre-chilled N a - heparin vacutainer (Becton-Dickinson). The remaining three ml were transferred to a plain, unchilled vacutainer. A syringe containing heparin solution (100 units/ml) was attached to the catheter and one ml was injected. This moment was considered as t = - 60 min. The oral temperature' was taken, and every 5 min thereafter pulse and blood pressure were recorded. The lemperature measurement was repeated, and additional blood samples were collected and processed 30 min (t 30 min) and 59 min (t = - 1 min) after the first blood sample. Then, while the subject remained passively in a ,emi-recumbent position, one of his feet was placed ankle deep in a pan of ice water for 1 min. If the subject was in the control protocol, nothing was done to the subject in the corresponding minute. At the end of the I min, thc subject's foot was removed from the water and was wrapped in a dry towel. Blood pressure and pulse rate v, ere recorded at t = + 0.5, 1,2, 3, and 5 rain, and every 5 min thereafter, following initiation of the cold pressor tesl. Comparable measurements were made during the control protocol. Oral temperature was retaken and additional blood samples were drawn at t = + 1, 3, 5, 10, 15, 30, and 60 min after initiation of the pressor test, and at comparable times in the control protocol. After the last blood sample was drawn, the subject was instructed to report back the next week at the same lime for the remaining test or control prolocol.
.I.s:says Within 30 rain of venipuncture, aliquots of heparinized blood were drawn into micropipettes for determinatkm of hematocrit. Then each blood sample was centrifuged at 3000 rev/min for 15 rain at 4"C to
VASOPRESSIN SECRETION AND THE COl.l) PRESSOR TEST
309
separate plasma from the cells-. Osmolality was determined on a 2.5 ml aliquot of fresh plasma by freezing point depression {Advanced instruments Inc., Hi-Precision Osmometer, Model 3R). An aliquot of plasma was set aside for measurement of plasma solutes, and the remaining plasma samples were stored in glass vials at - 20°C until assayed for plasma vasopressin (Robertson et al., 1973) or cortisol (Diagnostic Products Corporation, No Predilution '2'i Double Antibody RIA). Plasma sodium and potassium were determined by flame photomelry (Corning 460 Automatic Flame Photometer). Plasma urea was determined with an enzymatic-conductivimetric assay {Beckman BUN Analyzer 2). Plasma creatinine was determined with a creatinine-alkaline-picrate complex assay (Beckman Creatinine Analyzer 2). Plasma glucose was determined with a glucose oxidase assay (Beckman Glucose Analyzer 2). All assays were performed by technicians blind to the protocols undergone by the subjects. I)ala analysis
Mean arterial pressure was calculated by adding one-third of the pulse pressure to the diastolic pressure. Data stability during the basal period (t = - 6 0 min to t = - I min) was evaluated in each of the test and control protocols by a distribution-free trend test based on the ranks of the data (Lehmann, 1975). Since establishing good baselines was critical to the validity of the experiment, and since the small number of subjects in the study inflates the p values that the trend test yields, significance was considered at p < 0.10 in order to avoid making a type II error in the analysis of the data. If a significant trend was found, the last measurement taken before the administration of the cold pressor test was used as a representative baseline value. If no trend was present in the basal period, a simple average of the values taken in the basal period was used as a baseline value. Differences between baseline values in test and control protocols were evaluated with student's paired t-test. The paired t-test also was used to evaluate the difference found in each subject between the baseline value and 1he peak value following administration of the pressor test or the corresponding control period. The peak value was identified by inspection of the summary graph, which is a plot of the average value of the variable across all subjects for the specific time points in the protocols (Fig. l). The difference between the maximal deviation from baseline following administration of the pressor lest in the test protocol and the comparable deviation from baseline in the control protocol was also evaluated with the paired t-test. Significance for all t-tests was considered at p < 0.05. Because of the physiologic expectation that plasma hormone levels follow trends rather than simply varying minute-to-minute in random fashion, plasma vasopressin and cortisol also were evaluated for overall trends. Trend tests were performed on the data collected from t = - l min to t = 60 min. Significance again was considered at p < 0.05. If the trend test demonstrated the same trend or no trend in both the test and control dala sets, it was taken as evidence that the cold pressor test did not affect the variable in question. If a trend was present in the test protocol data set but not in the control (or vice versa), then the pressor test was judged to have a specific effect on that variable. All values are given as the mean plus-or-minus the standard error of the mean. In a further effort to uncover a possible effect of stress on vasopressin secretion, correlation analysis was employed to determine if the maximum fractional change in plasma vasopressin (defined as the maximum increase divided by the basal value) correlated significantly with the maximal fractional change in mean arterial pressure, pulse rate, or plasma cortisol during the test and control protocols. Again, to avoid a type II error, significance was set at p < 0.10. RESULTS All s u b j e c t s r e p o r t e d t h a t t h e c o l d p r e s s o r t e s t w a s a p a i n f u l p r o c e d u r e , a n d it w a s o n l y w i t h e f f o r t t h a t e a c h w a s a b l e t o k e e p his f o o t in t h e w a t e r f o r t h e full m i n u t e . E x c e p t f o r s l i g h t d i s c o m f o r t d u r i n g i n s e r t i o n o f t h e i n t r a v e n o u s c a t h e t e r at t h e b e g i n n i n g o f e a c h p r o t o c o l , n o s u b j e c t s r e p o r t e d p a i n d u r i n g a n y o t h e r p a r t o f t h e e x p e r i m e n t . A t n o t i m e in a n y o f the trials did a n y o f the subjects r e p o r t feeling n a u s e a t e d or faint. F i g u r e 1 p o r t r a y s t h e d a t a in g r a p h i c f o r m . N o n e o f t h e v a r i a b l e s d i f f e r e d s i g n i f i c a n t l y d u r i n g t h e test a n d c o n t r o l b a s e l i n e s . F o l l o w i n g a d m i n i s t r a t i o n o f t h e c o l d p r e s s o r t e s t , all t h r e e s t r e s s p a r a m e t e r s i n c r e a s e d significantly compared to control values. At t = 1 min, mean arterial pressure increased 13.9 ± 3.1 m m H g ( p < 0 . 0 0 4 ) c o m p a r e d t o c o n t r o l v a l u e s . L i k e w i s e , p u l s e r a t e i n c r e a s e d 9.2 _+ 2.8 b e a t s / m i n ( p < 0.02) a t t = 0.5 m i n , a n d p l a s m a c o r t i s o l r o s e 3.5 ± 0.8 I.tg/dl ( p < 0.005) a t t = 15 m i n ( T a b l e I). N o s i g n i f i c a n t i n c r e a s e o v e r b a s e l i n e o c c u r r e d in t h e s e stress parameters during the control protocol.
310
.IONATHAN T . EDEI.SON a n d G A R Y L . ROBERTSON
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TIME, MIN. Fig. 1. Summary graph of the time course of mean arterial pressure, pulse rate, serum cortisol and vasopressin, plasma solutes, hematocrit and temperature before and after the cold pressor test at t = 0 min. The solid line represents test protocol results, while the dashed line represents control protocol results. Significant changes from previous baseline levels of each variable are indicated by *p < 0.05, **p < 0.005, and ***p < 0.001. (See Methods sections of text for determination of baselines and calculations of significant changes from baselines.)
Trend tests corroborated the cortisol finding. T h e m a x i m u m increase in plasma cortisol occurred at t = 15 min and then decreased s o m e w h a t through t = 60 min. This particular trend was f o u n d to be statistically significant in the test protocol data ( p < 0.0001) but was not present during the same period o f the control protocol. During the basal period, there were significant d o w n w a r d trends irt plasma vasopressin levels in both the control protocol ( p < 0.004) and the test protocol ( p < 0.008). In the control protocol, m e a n p l a s m a vasopressin levels did not change from t = - 60 min to t = - 30 min, but they fell significantly ( p < 0.001) by an average o f 3.1 _+ 1.2 p g / m l from t = - 30 min to t = - 1 min. D u r i n g the test protocol, plasma vasopresSin levels f o l l o w e d
VASOPRESSIN SECRETION AND THE COI.D PRESSOR TEST
311
a similar pattern of little or no change from t = - 60 min to t = - 30 min, followed by a significant decrease ( p < 0.04) o f 2.5 _+ 2.0 p g / m l from t = - 30 min to t = - 1 min. The plasma vasopressin level at t = - 1 min was used as a representative baseline value for both the control and test protocols. Following the cold pressor test, mean plasma vasopressin appeared to rise to a m a x i m u m at t = 5 min. However, the average increase, 3 . 2 _ 1.6 pg/ml, was not significant by paired t-test. During the same period of the control protocol, there was a comparable increase in plasma vasopressin (2.7 _+ 1.4 pg/ml) that also was insignificant. Further, the magnitude of the increases over baseline in the test and control protocols did not differ significantly (0.5 _+ 1.6 pg/ml). Trend tests yielded similar results. During the test protocol, mean plasma vasopressin values increased from a nadir at t = - 1 min to a peak at t = 5 rain and then decreased somewhat through t = 60 min. This particular trend was found to be significant for both the test protocol (p < 0.005) and the control protocol (p < 0.001). The maximal fractional deviations in plasma vasopressin did not correlate significantly with the maximal fractional deviations in mean arterial pressure in either the test protocol (r = 0.14) or the control protocol (r = 0.13); with pulse rate in either the test protocol (r = 0.28) or the control protocol (r = 0.06); nor with plasma cortisol in either the test protocol (r = 0.06) or the control protocol (r = 0.18). Plasma osmolality, sodium, potassium, urea, creatinine, and glucose, and hematocrit and temperature, did not change significantly from baseline in either the test or control protocols. DISCUSSION
The results of this study do not support the concept that stress and pain p e r se, induced here by the cold pressor test, have a physiologically significant stimulatory effect on vasopressin secretion in man. The stimulus used in this study clearly induced stress and pain, as evidenced by the subjects' symptoms and by the objective findings of increased arterial pressure, pulse rate, and plasma cortisol. The pulse rate and blood pressure response, which presumably was mediated by increased sympathetic activity, was of a magnitude comparable to that when the hand instead of the foot was immersed in ice water (Hines & Brown, 1936). The rise in plasma cortisol, which presumably was mediated by increased A C T H secretion, has not been reported previously for the cold pressor test, and was probably another manifestation of the well-known sensitivity of the p i t u i t a r y - a d r e n a l axis to stressful stimuli (Habener, 1981). Those hormonal and hemodynamic changes were clearly due to the cold pressor test, since they did not occur during the comparable period of the control protocol. Unlike these other variables, plasma vasopressin showed no change attributable to the cold pressor test. Trend analysis indicated that plasma vasopressin tended to increase immediately after immersion of the foot in ice water; but exactly the same tendency was observed during the comparable period of the control protocol when the foot was not immersed. Moreover, the increases in plasma vasopressin detected by trend analysis in both the test and control protocols were not significant by paired t-test. This failure might be attributed to the relatively small number of subjects in the study. However, even if
T Ct T-Ct T CI T-Ct T C T-Ct T Ct T-Ct T Ct T-Ct T Ct T-Ct T Ct T-Ct
M.A.P.
Pulse rate
Cortisol
AVP
Osmolality
Hematocrit
Temperatt|re
none none
none d o w n w a r d ( p < 0.05)
none d o w n w a r d ( p < 0.005)
d o w n w a r d ( p < 0.008) d o w n w a r d ( p < 0.004)
d o w n w a r d ( p < 0.09) none
none u p w a r d ( p < 0.05)
none none
Trends
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3.3 -, 1.6 p g / m l ( N S ) , t = 5 rain 2.7-+ 1.4 p g / m l ( N S ) , t = 5 rain 0.5 *. 1.6 p g / m l (NS) I rain 1 rain
3.7 ~. 1.1 ~ g / d l ( p < 0.02), t = 15 rain 0.2 + 0.8 ~.g/dl (NS), t - 15 rain 3.5 -+ 0.8 ~ g / d l ( p < 0.005)
9.9 + 3.4 b e a l s / m i n ( p < 0.03), t = 0.5 rain 0.6 + 1.1 b e a t s / r a i n (NS), t - 0.5 rain 9.2 + 2.8 b e a t s / r a i n ( p < 0.02)
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15.6 + 3.0 m m H g ( p < 0.002), t - I rain 1.7 +_ 1.0 m m H g (NS), ! = I min 13.9 ± 3.1 m m H g ( p <" 0.004)
81.0 + 3.4 m m H g 81.7 + 2.4 m m H g
M e a n baseline levels
T h e r e were no significant differences b e t w e e n test a n d c o n t r o l baselines for any o f lhe o b s e r v e d variables. T = t e s t , Ct = ' c o n t r o I , M . A . P . = m e a n arterial pressure, A V P = arginine v a s o p r e s s i n , a n d NS = not significant.
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Variable
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VASOPRFSSlN SE( RETION AND THE C t ) l D PRENN,OR TEST
313
increasing the number of subjects made the test protocol results significant, it would be equally likely to increase the significance of the changes in the control protocol. Hence, the relatively small increase in plasma vasopressin observed under these conditions could not be attributed to the pain and/or stress produced by the cold pressor test. The foregoing analysis also might have failed to detect a subtle effect on vasopressin secretion if the intensity of the stress or the response to it varied markedly from subject to subject. In either case, however, one might expect to have found some correlation between the magnitude of the individual changes in plasma vasopressin and the magnitude of the changes in the other, more stress-sensitive variables like blood pressure, pulse rate or plasma cortisol. The fact that no such correlations were found in either the test or control protocols in our study militates against the possibility that individual differences in the intensity of the stimulus obscured a subtle effect on vasopressin secretion. The lack of stimulatory effect of the cold pressor test also cannot be explained by offsetting changes in recognized stimuli (Moore, 1971; Robertson, 1977; 1985). We observed no decrease in plasma osmolality, plasma sodium, or body temperature, nor any increase in blood volume, at least as reflected by hematocrit. However, blood pressure did increase transiently during the cold pressor test, and acute hypertension induced by infusion of norepinephrine has been reported to inhibit vasopressin release in man (Shimamoto & Miyahara, 1976). Hence, the lack of rise in plasma vasopressin in our study might be attributable to inhibition of the stress stimulus by the associated rise in blood pressure. Even if this were the case, however, the possible stimulatory effect of stress on vasopressin release would be of less general concern, since it is likely that many other forms of acute stress evoke similar increases in blood pressure. The results of the present study are consistent with the findings of other studies conducted on rats. Brennan et al. (1975) and Husain et al. (1975; 1979) found that the stress of swimming, forced exercise, noise, ether vapor, and plastic restraint failed to elevate plasma vasopressin. The studies of Husain et al. showed that manual restraint, abdominal compression, and electric shock significantly increased plasma vasopressin. However, as originally noted, these procedures could produce transient changes in blood pressure and/or effective blood volume which were not measured in these studies; moreover, electric shock could act directly on neural pathways that release vasopressin. Keil & Severs (1977) found that the stress of ether, swimming, and rotation not only did not increase but actually suppressed plasma vasopressin. The present findings differ from those of several other studies in humans which concluded that plasma vasopressin was increased by the stress of exercise (Dessypris et ai., 1980; Wade & Claybaugh, 1980; Geyssant et al., 1981), difficult delivery (DeVane & Porter, 1980), surgery (Moran et al., 1964; Ukai et al., 1968; Oka et al., 1981), and the pain that accompanies various kinds of acute injuries (Kendler et al., 1978). However, in none of these studies was there adequate control of other factors known to affect vasopressin secretion (Moore, 1971; Robertson, 1977; 1985). In fact, in half these studies, known stimuli such as hypotension or hypovolemia were shown to have accompanied the increases in plasma vasopressin (Dessypris et ai., 1980; Wade & Claybaugh, 1980; Geyssant et al., 1981; Oka et al., 1981). Even Kendler et ai. (1978), who attempted to control for stimulatory factors such as osmolality, posture, blood pressure, blood
314
JONAI-HAN T. EDEI,SON and GARY L. ROBERT~SON
volume, and drug ingestion, failed to take into account the possible stimulatory effect of nausea, glucopenia, or antecedent hypotensive episodes. For all these reasons, previous conclusions that stress and pain p e r se increase vasopressin secretion in man must be regarded with caution. A provocative and unexplained finding of our study was the apparent drop in plasma vasopressin that occurred in both the control and test protocols at the very end of the basal period. This change could have represented a return to physiologic levels from initial elevations induced by orthostasis or other stimuli present at the start of the experimental protocols. This interpretation is dubious, however, since with a hormone that has an intravascular half-life of about 20 min (Robertson, 1977), the effect of any prior stimulus present at the start of a protocol should have begun to dissipate by t = - 30 min. In fact, plasma vasopressin levels were nearly identical at t = - 60 min and t = - 30 min and did not fall significantly until t = - l min, immediately before application of the stress or sham stimulus. It is also unlikely that this delayed drop in plasma vasopressin was due to circadian variation, since half the subjects were studied in the morning and the other half in the afternoon. An alternate hypothesis to account for the delayed drop in plasma vasopressin is that it was a psychically mediated anticipatory effect generated by observation of the investigators readying themselves for the numerous tasks required during the coming minutes of the next phase of the protocol. The exact nature of the anticipatory effect is somewhat elusive. It cannot properly be attributed to stress, since none of the other observed physiologic variables manifested any changes comparable to those observed in plasma vasopressin. Moreover, this premonitory dip in plasma vasopressin did not intensify in the minutes following the cold pressor test. If anything, it tended to reverse, and this reversal occurred in the control as well as the test protocol. Hence, the modest rise in plasma vasopressin observed after the cold pressor test and at the comparable time in the control protocol cannot be attributed to overriding stimulation by the stress of the procedure, but rather could have been due to the release of inhibitory influences attendant to the anticipation of stress. The mechanism by which such anticipation might inhibit vasopressin release is unclear. However, it could be mediated by endogenous opioids, which are known to be released by anticipation of pain (Wilier et al., 1981) and are present in the posterior pituitary (Robertson, 1985), where they could act to inhibit the release of vasopressin (Van Wimersma Greidanus et al., 1979; Kamoi et al., 1979; Rossier et al., 1979; Aziz et al., 1980; Brownell et al., 1980; Clarke et al., 1980; Grossman et al., 1980; Iverson et al., 1980; Knepel et al., 1980; Summy-Long et ai., 1981; Robertson, 1985). Further studies of the effect of opiate antagonists on the anticipatory drop in plasma vasopressin may shed light on this interesting possibility. One must be cautious in extrapolating our findings to forms of stress other than the cold .pressor test. It is conceivable that a noxious influence more severe, sustained, emotionally disconcerting, or unassociated with an increase in blood pressure would have different effects upon vasopressin release. Indeed, the very vagueness of the term and the ethical constraints on human and animal studies o f this type make it virtually impossible to prove conclusively that vasopressin secretion is refractory to stimulation by all forms of
VASOPRESSIN SECRETIONAND THE COI+DPRESSOR TEST
315
"stress". However, the present study clearly indicates that the concept that stress in general stimulates vasopressin release should be abandoned in favor of more limited statements which specify the exact nature and intensity of the stimulus involved. We gratefully acknowledge the statistical consultations given us by David Draper of the Department of Statistics and by Richard Blough of the Biomedical Computation Facilities at the University of Chicago. We thank Annette Miller for her assistance in the Clinical Research Center, Mary Beth Gaskill for her support in the laboratory, and Lynn Nelson for helping to prepare this manuscript for publication. Supported by National Institutes of Health grant AM26212.
REFERENCES Aziz L A, Forsling M L, Woolf C J (1980) The action of morphine on vasopressin release in the rat. J Physiol (Lond) 300: 24P - 25P. Brennan T C, Shelton R L, Robertson G L (1975) Effect of stress on plasma vasopressin and corticosterone in rats. Clin Res 23: 243A. BrowneU J, del Pozo E, Donatsch P (1980) Inhibition of vasopressin secretion by met-enkephalin (FK 33-824) in humans. Acta Endocrinol 94:304 - 308. Clarke G, Lincoln D W, Wood P (1980) Inhibition of vasopressin neurones by intraventricular morphine. J Physiol (LontO 303: 59P - 60P. Dessypris A, Wagar G, Fyhrquist F, Makinen T, Welin M G, Lamberg B A (1980) Marathon run: effects on blood cortisol-ACTH, iodothyronines-TSH and vasopressin. Acta Endocrinol 95:151 - 157. DeVane G W, Porter J C (1980) An apparent stress-induced release of arginine vasopressin by human neonates. J Clin Endocrinol Metab 51: 1412- 1416. Forsling M L, Akerlung M, Stromberg P (1981) Variations in plasma concentrations of vasopressin during the menstrual period. J Endocrinol 89:263 - 266. Geyssant A, Geelen G, Allevard A M, Vincent M, JarsaiUin E, Bizollon C A, Lacour J R, Gharib C (1981) Plasma vasopressin, renin activity and aldosterone: effect of exercise and training. Eur J Applied Physiol 46:21 - 30. Grossman A, Besser G M, Milles J J, Baylis P H (1980) Inhibition of vasopressin release in man by an opiate peptide. Lancet 8204:1108 - 1110. Habener J F (1981) Hormone biosynthesis and secretion. In: Felig P, Baxter J D, Broadus A E, Frohman L A (Eds) Endocrinology and Metabolism. McGraw-Hill, New York, pp 29 - 59. Hines E A, Brown G E (1936) The cold pressor test for measuring the reactibility of the blood pressure: data concerning 571 normal and hypertensive subjects. A m Heart J 1 1 : 1 - 9 . Husain K, Manger W M, Rock T W, Weiss R J, Bickner J, Dufton S, Hart C, Frantz A G (1975) Plasma vasopressin (VP) in rats--effect of stressful stimuli. Clin Res 23: 573A. Husain K, Manger W M, Rock T W, Weiss R J, Frantz A G (1979) Vasopressin release due to manual restraint in the rat" role of body compression and comparison with other stressful stimuli. Endocrinology 104: 641 - 6 4 4 . Iverson L L, Iverson S D, Bloom F E (1980) Opiate receptors influence vasopressin release from nerve terminals in rat neurohypophysis. Nature 284: 350-351. Kamoi K, White K, Robertson G L (1979) Opiates elevate the osmotic threshold for vasopressin (VP) release in rats. Clin Res 27: 254A. Keii L C, Severs W B (1977) Reduction in plasma vasopressin levels of dehydrated rats following acute stress. Endocrinology 100: 3 0 - 38. Kendler K S, Weitzman R E, Fisher D A (1978) The effect of pain on plasma arginine vasopressin concentrations in man. J Clin Endocrinol 8:89 - 94. Knepel W, Nutto D, Anhut H, Hertting G (1980) Naloxone promotes stimulus-evoked vasopressin release in vivo. Eur J Pharmacol 65: 449-450. Lehmann E L (1975) Nonparametrics: Statistical Methods Based on Ranks. Holt & Day, New York, pp 287 - 297. Moore W W (1971) Antidiuretic hormone levels in normal subjects. Fed Proc 30:1387 - 1394. Moran W H, Mitenberger F W, Shuayb W A, Zimmerman B (1964) The relationship of antidiuretic hormone secretion to surgical stress. Surgery 56: 9 9 - 108. Oka Y, Wakayama S, Oyama T, Orkin L R, Becker R M, Blaufox M D, Frater R M (1981) Cortisol and antidiuretic hormone responses to stress in cardiac surgical patients. Canad Anaesth Soc J 28: 3 3 4 - 338.
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JONATHAN T. EDEJ SON and GARV L. ROBERTSON
Robertson G L (1977) The regulation of vasopressin function in health and disease. Rec Prog Horm Res 33: 333-385. Robertson G L (1985) Vasopressin. In: Ingbar S (Ed) Contemporary Endocrinology, Vol. 2, Plenum, New York, pp 403 - 450. Robertson G L, Mahr E A, Athar S, Sinha T (1973) Development and clinical application of a new method for the radioimmunoassay of arginine vasopressin in human plasma. J Clin Invest 52: 2340- 2352. Rossier J, Battenberg E, Pittman Q, Bayon A, Koda L, Miller R, Guillemin R, Bloom F (1979) Hypothalamic enkephalin neurones may regulate the neurohypophysis. Nature 277:653 - 655. Rydin H, Verney E B (1938) The inhibition of water diuresis by emotional stress and by muscular exercise. Q J Exp Physiol 27:343 - 374. Shimamoto K, Miyahara M (1976) Effect of norepinephrine infusion on plasma vasopressin levels in normal human subjects. J Clin Endocrinol Metab 43:201 - 204. Summy-Long J Y, Keil L C, Deen K, Severs W B (1981) Opiate regulation of angiotensin-induced drinking and vasopressin release. J Pharmacol Exp Ther 217: 630- 637. Ukai M, Moran W H, Zimmerman B (1968) The role of visceral afferent pathways on vasopressin secretion and urinary excretory patterns during surgical stress. Ann Surg 168: 16-28. Van Wimersma Greidanus T B, Thody T J, Verspaget H, de Rotta G A, Goedemans H H, Croiset G, Van Ree J M (1979) Effects of morphine and ~-endorphin on basal and elevated levels of ¢t-MSH and vasopressin. Life Sci 24:579 - 586. v Bormann B, Weidler B, Denhardt R, Frings N, Lennhartz H, Hempelmann G (1981) Plasma-antidiuretic hormone level as an indicator of postoperative stress (part II). A nasth Intensivther Notfallmed 1 6 : 3 1 9 - 322. Wade C E, Claybaugh J R (1980) Plasma renin activity, vasopressin concentration, and urinary excretory responses to exercise in men. J Appl Physiol 49:930 - 936. Weidler B, v Bormann B, Lennhartz H, Denhardt R, Hempelmann G (1981) Plasma-antidiuretic hormone as indicator of perioperative stress (part I). A nasth lntensivther Notfallmed 16:315 - 318. Wilier J C, Dehen H, Cambier J (1981) Stress-induced analgesia in humans: endogenous opioids and naloxonereversible depression of pain reflexes. Science 212: 689-691.
0 3 0 6 - 4530/86 $3.00 + 0.00 Pergamon Journals Ltd.
Printed in Great Britain.
THE EFFECT OF THE COLD PRESSOR TEST ON VASOPRESSIN SECRETION IN MAN JONATHAN T . EDELSON* a n d GARY L. ROBERTSONt:~ *University of Chicago Pritzker School of Medicine and Charles A. Dana Clinical Scholars Program, University of Pennsylvania School of Medicine, U.S.A.; and i"Department of Medicine, University of Chicago Pritzker School of Medicine, U.S.A.
(Received 1 April 1985; in finalform 19 August 1985) SUMMARY Stress has been thought to induce the release of arginine vasopressin (AVP). We evaluated this claim by studying the effects of a modified cold pressor test on plasma AVP, plasma cortisol, blood pressure, pulse rate, and a number of variables known to affect AVP secretion. In a crossover study design, test and control values were obtained in seven male subjects. The pressor test was found to induce painful stress as evidenced by subjective reports and the objective findings of increased mean arterial pressure 03.9 • 3.1 mm Hg; p < 0.004), pulse rate (9.2 +_ 2.8 beats/min; p < 0.02), and plasma cortisoi (3.5 +_ 0.8 p.g/dl; p < 0.005). In contrast, there were no significant changes in plasma AVP that could be attributed to the cold pressor test. There also were no changes in plasma osmolality, measured plasma solutes, hematocrit or body temperature. An unexpected finding was a premonitory drop in plasma AVP occurring just prior to the pressor test (2.5 _+ 2.0 pg/ml; p < 0.04) and at the comparable time point in the control study (3.1 +_ 1.2 pg/ml; p < 0.001). There were no changes in any of the other measured variables which could account for this drop. We conclude that the cold pressor test is not a stimulus to AVP release and that anticipation of stress may inhibit secretion of this hormone. INTRODUCTION
SINCB Rydin and Verney's (1938) pioneering study in dogs, it has been almost axiomatic that pain, anxiety and other forms of emotional stress cause the release of vasopressin (AVP). However, this concept has been challenged by modern studies that have employed sensitive radioimmunoassays to directly measure the hormone. In rats, a variety of noxious stimuli have been shown to have no effect on vasopressin secretion unless they also induce hyperosmolality, hypovolemia, or other recognized non-osmotic stimuli (Keil & Severs, 1977; Robertson, 1977; Husain et al., 1979). These observations suggest that stress p e r se does not affect vasopressin secretion or that it stimulates secretion directly in some species but not in others. In this regard, Kendler et ai. (1978) suggested that in man pain induces vasopressin release, and Weidler et al. (1981) and v Borman et al. (1981) used the incremental increase in plasma vasopressin after surgery as an indicator of the efficacy of two anesthetics in relieving pain. However, none of these studies provide the kind of ancillary data needed to ascertain whether the observed increase was truly due to pain p e r se or was simply secondary to changes in other recognized stimuli such as hypovolemia, hypotension, or nausea (Robertson, 1977). Clarification of this issue is essential for the proper interpretation of any study of vasopressin function in acutely ill patients with clinical disturbances of salt and water ~/Correspondence to be addressed to: Dr Gary L. Robertson at University of Chicago, 5841 S. Maryland Avenue, Box 131, Chicago, IL 60637, U.S.A. 307
308
JONATHAN T. El)El.SON and GARY L. ROBERT~ON
balance. Therefore, we undertook a systematic study of the effect of stress on vasopressin secretion in man using a variant of the standard cold pressor test (Hines & Brown, 1936). This procedure is associated with a modest degree of pain and results in an acute rise in blood pressure and pulse rate, which is thought to be mediated by the sympathetic nervous system. In addition to these hemodynamic variables, we monitored the activity of the p i t u i t a r y - a d r e n a l axis as well as other variables known to directly affect vasopressin secretion (Moore, 1971; Robertson, 1977, 1985). In this study, we operationally defined "stress" as the induction of elevations in hemodynamic variables and in adrenal activity attributable to the cold pressor test. METHODS
Study design In a cross-over study design, seven subjects were randomized into one of two cells, one of which had the lest protocol first and the control protocol second, and the other of which had the protocols in the opposite order. The test and control protocols were of equal length, distinguished only by the fact thai a specific stress was applied in the test protocol but not in the control protocol. As a control for the possible influence of circadian rhythms, each subject participated in the control and test protocols at the same time of day one week apart.
Sltbjec+ls Seven healthy male volunteers in their third decade gave their written consent to participate in Ihis study, which was approved by the Clinical Investigations Committee at the University of Chicago Pritzker School of Medicine and which was carried out according to the principles of the Declaration of Helsinki. Female subjects ~ere excluded to avoid hormonal variations concomitant with the menstrual cycle (Forsling et aL, 1981). Each ~ubject received financial remuneration for participating in the study.
])1+olO('O] Experimental trials were run at the Outpatient Research Center at Billings Hospital between 0930 h and 1530 h. Subjects tested at 0930 h had no food or liquids after 2400 h the preceding night. Subjects tested at 1330 h had no food or liquids after 0830 h the day of the trial. No drugs, especially tobacco or alcohol, were used within 24 h of the trials. On the day of the first test, written informed consent was obtained, and the subject x~as told whether he would participate in the test or control protocol. The subject emptied his bladder and was x~eighed and placed in a semi-recumbent position. His blood pressure and pulse rate were monitored manually for 15 rain, and then his oral temperature was obtained. At the same time, a history of any adverse reactions to phlebotomy was determined, and the purpose, nature and possible discomforts of the procedure were explained in detail. The subject was asked to report any sensations he felt, especially nausea, faintness or pain, during the ¢c,ttrse of Ihe protocol. With standard sterile technique, a #21 gauge butterfly needle catheter was inserted and secured in an antecubilal vein. One ml of blood was withdrawn and discarded, after which another 10 ml of blood were x~ilhdraw n, seven of which were transferred to a pre-chilled N a - heparin vacutainer (Becton-Dickinson). The remaining three ml were transferred to a plain, unchilled vacutainer. A syringe containing heparin solution (100 units/ml) was attached to the catheter and one ml was injected. This moment was considered as t = - 60 min. The oral temperature' was taken, and every 5 min thereafter pulse and blood pressure were recorded. The lemperature measurement was repeated, and additional blood samples were collected and processed 30 min (t 30 min) and 59 min (t = - 1 min) after the first blood sample. Then, while the subject remained passively in a ,emi-recumbent position, one of his feet was placed ankle deep in a pan of ice water for 1 min. If the subject was in the control protocol, nothing was done to the subject in the corresponding minute. At the end of the I min, thc subject's foot was removed from the water and was wrapped in a dry towel. Blood pressure and pulse rate v, ere recorded at t = + 0.5, 1,2, 3, and 5 rain, and every 5 min thereafter, following initiation of the cold pressor tesl. Comparable measurements were made during the control protocol. Oral temperature was retaken and additional blood samples were drawn at t = + 1, 3, 5, 10, 15, 30, and 60 min after initiation of the pressor test, and at comparable times in the control protocol. After the last blood sample was drawn, the subject was instructed to report back the next week at the same lime for the remaining test or control prolocol.
.I.s:says Within 30 rain of venipuncture, aliquots of heparinized blood were drawn into micropipettes for determinatkm of hematocrit. Then each blood sample was centrifuged at 3000 rev/min for 15 rain at 4"C to
VASOPRESSIN SECRETION AND THE COl.l) PRESSOR TEST
309
separate plasma from the cells-. Osmolality was determined on a 2.5 ml aliquot of fresh plasma by freezing point depression {Advanced instruments Inc., Hi-Precision Osmometer, Model 3R). An aliquot of plasma was set aside for measurement of plasma solutes, and the remaining plasma samples were stored in glass vials at - 20°C until assayed for plasma vasopressin (Robertson et al., 1973) or cortisol (Diagnostic Products Corporation, No Predilution '2'i Double Antibody RIA). Plasma sodium and potassium were determined by flame photomelry (Corning 460 Automatic Flame Photometer). Plasma urea was determined with an enzymatic-conductivimetric assay {Beckman BUN Analyzer 2). Plasma creatinine was determined with a creatinine-alkaline-picrate complex assay (Beckman Creatinine Analyzer 2). Plasma glucose was determined with a glucose oxidase assay (Beckman Glucose Analyzer 2). All assays were performed by technicians blind to the protocols undergone by the subjects. I)ala analysis
Mean arterial pressure was calculated by adding one-third of the pulse pressure to the diastolic pressure. Data stability during the basal period (t = - 6 0 min to t = - I min) was evaluated in each of the test and control protocols by a distribution-free trend test based on the ranks of the data (Lehmann, 1975). Since establishing good baselines was critical to the validity of the experiment, and since the small number of subjects in the study inflates the p values that the trend test yields, significance was considered at p < 0.10 in order to avoid making a type II error in the analysis of the data. If a significant trend was found, the last measurement taken before the administration of the cold pressor test was used as a representative baseline value. If no trend was present in the basal period, a simple average of the values taken in the basal period was used as a baseline value. Differences between baseline values in test and control protocols were evaluated with student's paired t-test. The paired t-test also was used to evaluate the difference found in each subject between the baseline value and 1he peak value following administration of the pressor test or the corresponding control period. The peak value was identified by inspection of the summary graph, which is a plot of the average value of the variable across all subjects for the specific time points in the protocols (Fig. l). The difference between the maximal deviation from baseline following administration of the pressor lest in the test protocol and the comparable deviation from baseline in the control protocol was also evaluated with the paired t-test. Significance for all t-tests was considered at p < 0.05. Because of the physiologic expectation that plasma hormone levels follow trends rather than simply varying minute-to-minute in random fashion, plasma vasopressin and cortisol also were evaluated for overall trends. Trend tests were performed on the data collected from t = - l min to t = 60 min. Significance again was considered at p < 0.05. If the trend test demonstrated the same trend or no trend in both the test and control dala sets, it was taken as evidence that the cold pressor test did not affect the variable in question. If a trend was present in the test protocol data set but not in the control (or vice versa), then the pressor test was judged to have a specific effect on that variable. All values are given as the mean plus-or-minus the standard error of the mean. In a further effort to uncover a possible effect of stress on vasopressin secretion, correlation analysis was employed to determine if the maximum fractional change in plasma vasopressin (defined as the maximum increase divided by the basal value) correlated significantly with the maximal fractional change in mean arterial pressure, pulse rate, or plasma cortisol during the test and control protocols. Again, to avoid a type II error, significance was set at p < 0.10. RESULTS All s u b j e c t s r e p o r t e d t h a t t h e c o l d p r e s s o r t e s t w a s a p a i n f u l p r o c e d u r e , a n d it w a s o n l y w i t h e f f o r t t h a t e a c h w a s a b l e t o k e e p his f o o t in t h e w a t e r f o r t h e full m i n u t e . E x c e p t f o r s l i g h t d i s c o m f o r t d u r i n g i n s e r t i o n o f t h e i n t r a v e n o u s c a t h e t e r at t h e b e g i n n i n g o f e a c h p r o t o c o l , n o s u b j e c t s r e p o r t e d p a i n d u r i n g a n y o t h e r p a r t o f t h e e x p e r i m e n t . A t n o t i m e in a n y o f the trials did a n y o f the subjects r e p o r t feeling n a u s e a t e d or faint. F i g u r e 1 p o r t r a y s t h e d a t a in g r a p h i c f o r m . N o n e o f t h e v a r i a b l e s d i f f e r e d s i g n i f i c a n t l y d u r i n g t h e test a n d c o n t r o l b a s e l i n e s . F o l l o w i n g a d m i n i s t r a t i o n o f t h e c o l d p r e s s o r t e s t , all t h r e e s t r e s s p a r a m e t e r s i n c r e a s e d significantly compared to control values. At t = 1 min, mean arterial pressure increased 13.9 ± 3.1 m m H g ( p < 0 . 0 0 4 ) c o m p a r e d t o c o n t r o l v a l u e s . L i k e w i s e , p u l s e r a t e i n c r e a s e d 9.2 _+ 2.8 b e a t s / m i n ( p < 0.02) a t t = 0.5 m i n , a n d p l a s m a c o r t i s o l r o s e 3.5 ± 0.8 I.tg/dl ( p < 0.005) a t t = 15 m i n ( T a b l e I). N o s i g n i f i c a n t i n c r e a s e o v e r b a s e l i n e o c c u r r e d in t h e s e stress parameters during the control protocol.
310
.IONATHAN T . EDEI.SON a n d G A R Y L . ROBERTSON
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TIME, MIN. Fig. 1. Summary graph of the time course of mean arterial pressure, pulse rate, serum cortisol and vasopressin, plasma solutes, hematocrit and temperature before and after the cold pressor test at t = 0 min. The solid line represents test protocol results, while the dashed line represents control protocol results. Significant changes from previous baseline levels of each variable are indicated by *p < 0.05, **p < 0.005, and ***p < 0.001. (See Methods sections of text for determination of baselines and calculations of significant changes from baselines.)
Trend tests corroborated the cortisol finding. T h e m a x i m u m increase in plasma cortisol occurred at t = 15 min and then decreased s o m e w h a t through t = 60 min. This particular trend was f o u n d to be statistically significant in the test protocol data ( p < 0.0001) but was not present during the same period o f the control protocol. During the basal period, there were significant d o w n w a r d trends irt plasma vasopressin levels in both the control protocol ( p < 0.004) and the test protocol ( p < 0.008). In the control protocol, m e a n p l a s m a vasopressin levels did not change from t = - 60 min to t = - 30 min, but they fell significantly ( p < 0.001) by an average o f 3.1 _+ 1.2 p g / m l from t = - 30 min to t = - 1 min. D u r i n g the test protocol, plasma vasopresSin levels f o l l o w e d
VASOPRESSIN SECRETION AND THE COI.D PRESSOR TEST
311
a similar pattern of little or no change from t = - 60 min to t = - 30 min, followed by a significant decrease ( p < 0.04) o f 2.5 _+ 2.0 p g / m l from t = - 30 min to t = - 1 min. The plasma vasopressin level at t = - 1 min was used as a representative baseline value for both the control and test protocols. Following the cold pressor test, mean plasma vasopressin appeared to rise to a m a x i m u m at t = 5 min. However, the average increase, 3 . 2 _ 1.6 pg/ml, was not significant by paired t-test. During the same period of the control protocol, there was a comparable increase in plasma vasopressin (2.7 _+ 1.4 pg/ml) that also was insignificant. Further, the magnitude of the increases over baseline in the test and control protocols did not differ significantly (0.5 _+ 1.6 pg/ml). Trend tests yielded similar results. During the test protocol, mean plasma vasopressin values increased from a nadir at t = - 1 min to a peak at t = 5 rain and then decreased somewhat through t = 60 min. This particular trend was found to be significant for both the test protocol (p < 0.005) and the control protocol (p < 0.001). The maximal fractional deviations in plasma vasopressin did not correlate significantly with the maximal fractional deviations in mean arterial pressure in either the test protocol (r = 0.14) or the control protocol (r = 0.13); with pulse rate in either the test protocol (r = 0.28) or the control protocol (r = 0.06); nor with plasma cortisol in either the test protocol (r = 0.06) or the control protocol (r = 0.18). Plasma osmolality, sodium, potassium, urea, creatinine, and glucose, and hematocrit and temperature, did not change significantly from baseline in either the test or control protocols. DISCUSSION
The results of this study do not support the concept that stress and pain p e r se, induced here by the cold pressor test, have a physiologically significant stimulatory effect on vasopressin secretion in man. The stimulus used in this study clearly induced stress and pain, as evidenced by the subjects' symptoms and by the objective findings of increased arterial pressure, pulse rate, and plasma cortisol. The pulse rate and blood pressure response, which presumably was mediated by increased sympathetic activity, was of a magnitude comparable to that when the hand instead of the foot was immersed in ice water (Hines & Brown, 1936). The rise in plasma cortisol, which presumably was mediated by increased A C T H secretion, has not been reported previously for the cold pressor test, and was probably another manifestation of the well-known sensitivity of the p i t u i t a r y - a d r e n a l axis to stressful stimuli (Habener, 1981). Those hormonal and hemodynamic changes were clearly due to the cold pressor test, since they did not occur during the comparable period of the control protocol. Unlike these other variables, plasma vasopressin showed no change attributable to the cold pressor test. Trend analysis indicated that plasma vasopressin tended to increase immediately after immersion of the foot in ice water; but exactly the same tendency was observed during the comparable period of the control protocol when the foot was not immersed. Moreover, the increases in plasma vasopressin detected by trend analysis in both the test and control protocols were not significant by paired t-test. This failure might be attributed to the relatively small number of subjects in the study. However, even if
T Ct T-Ct T CI T-Ct T C T-Ct T Ct T-Ct T Ct T-Ct T Ct T-Ct T Ct T-Ct
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Osmolality
Hematocrit
Temperatt|re
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none d o w n w a r d ( p < 0.05)
none d o w n w a r d ( p < 0.005)
d o w n w a r d ( p < 0.008) d o w n w a r d ( p < 0.004)
d o w n w a r d ( p < 0.09) none
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none none
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9.9 + 3.4 b e a l s / m i n ( p < 0.03), t = 0.5 rain 0.6 + 1.1 b e a t s / r a i n (NS), t - 0.5 rain 9.2 + 2.8 b e a t s / r a i n ( p < 0.02)
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8.4 + 1.2 ~.g/dl 7.4 * 1.0 lag/dl
15.6 + 3.0 m m H g ( p < 0.002), t - I rain 1.7 +_ 1.0 m m H g (NS), ! = I min 13.9 ± 3.1 m m H g ( p <" 0.004)
81.0 + 3.4 m m H g 81.7 + 2.4 m m H g
M e a n baseline levels
T h e r e were no significant differences b e t w e e n test a n d c o n t r o l baselines for any o f lhe o b s e r v e d variables. T = t e s t , Ct = ' c o n t r o I , M . A . P . = m e a n arterial pressure, A V P = arginine v a s o p r e s s i n , a n d NS = not significant.
T or Ct
Variable
P o s t - i n t e r v e n l i o n d e v i a t i o n s f r o m baseline a n d time point o f d e v i a t i o n . For each v a r i a b l e , differ~ ence b e t w e e n test a n d c o m r o l responses shoran below
"]r'ABI /- 1, R I S P O N S f ¢)1 N O R M A l '41JB.IE('TS TO t H E COl I) PRES;SOR TEY,I
-v-
¥
z
VASOPRFSSlN SE( RETION AND THE C t ) l D PRENN,OR TEST
313
increasing the number of subjects made the test protocol results significant, it would be equally likely to increase the significance of the changes in the control protocol. Hence, the relatively small increase in plasma vasopressin observed under these conditions could not be attributed to the pain and/or stress produced by the cold pressor test. The foregoing analysis also might have failed to detect a subtle effect on vasopressin secretion if the intensity of the stress or the response to it varied markedly from subject to subject. In either case, however, one might expect to have found some correlation between the magnitude of the individual changes in plasma vasopressin and the magnitude of the changes in the other, more stress-sensitive variables like blood pressure, pulse rate or plasma cortisol. The fact that no such correlations were found in either the test or control protocols in our study militates against the possibility that individual differences in the intensity of the stimulus obscured a subtle effect on vasopressin secretion. The lack of stimulatory effect of the cold pressor test also cannot be explained by offsetting changes in recognized stimuli (Moore, 1971; Robertson, 1977; 1985). We observed no decrease in plasma osmolality, plasma sodium, or body temperature, nor any increase in blood volume, at least as reflected by hematocrit. However, blood pressure did increase transiently during the cold pressor test, and acute hypertension induced by infusion of norepinephrine has been reported to inhibit vasopressin release in man (Shimamoto & Miyahara, 1976). Hence, the lack of rise in plasma vasopressin in our study might be attributable to inhibition of the stress stimulus by the associated rise in blood pressure. Even if this were the case, however, the possible stimulatory effect of stress on vasopressin release would be of less general concern, since it is likely that many other forms of acute stress evoke similar increases in blood pressure. The results of the present study are consistent with the findings of other studies conducted on rats. Brennan et al. (1975) and Husain et al. (1975; 1979) found that the stress of swimming, forced exercise, noise, ether vapor, and plastic restraint failed to elevate plasma vasopressin. The studies of Husain et al. showed that manual restraint, abdominal compression, and electric shock significantly increased plasma vasopressin. However, as originally noted, these procedures could produce transient changes in blood pressure and/or effective blood volume which were not measured in these studies; moreover, electric shock could act directly on neural pathways that release vasopressin. Keil & Severs (1977) found that the stress of ether, swimming, and rotation not only did not increase but actually suppressed plasma vasopressin. The present findings differ from those of several other studies in humans which concluded that plasma vasopressin was increased by the stress of exercise (Dessypris et ai., 1980; Wade & Claybaugh, 1980; Geyssant et al., 1981), difficult delivery (DeVane & Porter, 1980), surgery (Moran et al., 1964; Ukai et al., 1968; Oka et al., 1981), and the pain that accompanies various kinds of acute injuries (Kendler et al., 1978). However, in none of these studies was there adequate control of other factors known to affect vasopressin secretion (Moore, 1971; Robertson, 1977; 1985). In fact, in half these studies, known stimuli such as hypotension or hypovolemia were shown to have accompanied the increases in plasma vasopressin (Dessypris et ai., 1980; Wade & Claybaugh, 1980; Geyssant et al., 1981; Oka et al., 1981). Even Kendler et ai. (1978), who attempted to control for stimulatory factors such as osmolality, posture, blood pressure, blood
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JONAI-HAN T. EDEI,SON and GARY L. ROBERT~SON
volume, and drug ingestion, failed to take into account the possible stimulatory effect of nausea, glucopenia, or antecedent hypotensive episodes. For all these reasons, previous conclusions that stress and pain p e r se increase vasopressin secretion in man must be regarded with caution. A provocative and unexplained finding of our study was the apparent drop in plasma vasopressin that occurred in both the control and test protocols at the very end of the basal period. This change could have represented a return to physiologic levels from initial elevations induced by orthostasis or other stimuli present at the start of the experimental protocols. This interpretation is dubious, however, since with a hormone that has an intravascular half-life of about 20 min (Robertson, 1977), the effect of any prior stimulus present at the start of a protocol should have begun to dissipate by t = - 30 min. In fact, plasma vasopressin levels were nearly identical at t = - 60 min and t = - 30 min and did not fall significantly until t = - l min, immediately before application of the stress or sham stimulus. It is also unlikely that this delayed drop in plasma vasopressin was due to circadian variation, since half the subjects were studied in the morning and the other half in the afternoon. An alternate hypothesis to account for the delayed drop in plasma vasopressin is that it was a psychically mediated anticipatory effect generated by observation of the investigators readying themselves for the numerous tasks required during the coming minutes of the next phase of the protocol. The exact nature of the anticipatory effect is somewhat elusive. It cannot properly be attributed to stress, since none of the other observed physiologic variables manifested any changes comparable to those observed in plasma vasopressin. Moreover, this premonitory dip in plasma vasopressin did not intensify in the minutes following the cold pressor test. If anything, it tended to reverse, and this reversal occurred in the control as well as the test protocol. Hence, the modest rise in plasma vasopressin observed after the cold pressor test and at the comparable time in the control protocol cannot be attributed to overriding stimulation by the stress of the procedure, but rather could have been due to the release of inhibitory influences attendant to the anticipation of stress. The mechanism by which such anticipation might inhibit vasopressin release is unclear. However, it could be mediated by endogenous opioids, which are known to be released by anticipation of pain (Wilier et al., 1981) and are present in the posterior pituitary (Robertson, 1985), where they could act to inhibit the release of vasopressin (Van Wimersma Greidanus et al., 1979; Kamoi et al., 1979; Rossier et al., 1979; Aziz et al., 1980; Brownell et al., 1980; Clarke et al., 1980; Grossman et al., 1980; Iverson et al., 1980; Knepel et al., 1980; Summy-Long et ai., 1981; Robertson, 1985). Further studies of the effect of opiate antagonists on the anticipatory drop in plasma vasopressin may shed light on this interesting possibility. One must be cautious in extrapolating our findings to forms of stress other than the cold .pressor test. It is conceivable that a noxious influence more severe, sustained, emotionally disconcerting, or unassociated with an increase in blood pressure would have different effects upon vasopressin release. Indeed, the very vagueness of the term and the ethical constraints on human and animal studies o f this type make it virtually impossible to prove conclusively that vasopressin secretion is refractory to stimulation by all forms of
VASOPRESSIN SECRETIONAND THE COI+DPRESSOR TEST
315
"stress". However, the present study clearly indicates that the concept that stress in general stimulates vasopressin release should be abandoned in favor of more limited statements which specify the exact nature and intensity of the stimulus involved. We gratefully acknowledge the statistical consultations given us by David Draper of the Department of Statistics and by Richard Blough of the Biomedical Computation Facilities at the University of Chicago. We thank Annette Miller for her assistance in the Clinical Research Center, Mary Beth Gaskill for her support in the laboratory, and Lynn Nelson for helping to prepare this manuscript for publication. Supported by National Institutes of Health grant AM26212.
REFERENCES Aziz L A, Forsling M L, Woolf C J (1980) The action of morphine on vasopressin release in the rat. J Physiol (Lond) 300: 24P - 25P. Brennan T C, Shelton R L, Robertson G L (1975) Effect of stress on plasma vasopressin and corticosterone in rats. Clin Res 23: 243A. BrowneU J, del Pozo E, Donatsch P (1980) Inhibition of vasopressin secretion by met-enkephalin (FK 33-824) in humans. Acta Endocrinol 94:304 - 308. Clarke G, Lincoln D W, Wood P (1980) Inhibition of vasopressin neurones by intraventricular morphine. J Physiol (LontO 303: 59P - 60P. Dessypris A, Wagar G, Fyhrquist F, Makinen T, Welin M G, Lamberg B A (1980) Marathon run: effects on blood cortisol-ACTH, iodothyronines-TSH and vasopressin. Acta Endocrinol 95:151 - 157. DeVane G W, Porter J C (1980) An apparent stress-induced release of arginine vasopressin by human neonates. J Clin Endocrinol Metab 51: 1412- 1416. Forsling M L, Akerlung M, Stromberg P (1981) Variations in plasma concentrations of vasopressin during the menstrual period. J Endocrinol 89:263 - 266. Geyssant A, Geelen G, Allevard A M, Vincent M, JarsaiUin E, Bizollon C A, Lacour J R, Gharib C (1981) Plasma vasopressin, renin activity and aldosterone: effect of exercise and training. Eur J Applied Physiol 46:21 - 30. Grossman A, Besser G M, Milles J J, Baylis P H (1980) Inhibition of vasopressin release in man by an opiate peptide. Lancet 8204:1108 - 1110. Habener J F (1981) Hormone biosynthesis and secretion. In: Felig P, Baxter J D, Broadus A E, Frohman L A (Eds) Endocrinology and Metabolism. McGraw-Hill, New York, pp 29 - 59. Hines E A, Brown G E (1936) The cold pressor test for measuring the reactibility of the blood pressure: data concerning 571 normal and hypertensive subjects. A m Heart J 1 1 : 1 - 9 . Husain K, Manger W M, Rock T W, Weiss R J, Bickner J, Dufton S, Hart C, Frantz A G (1975) Plasma vasopressin (VP) in rats--effect of stressful stimuli. Clin Res 23: 573A. Husain K, Manger W M, Rock T W, Weiss R J, Frantz A G (1979) Vasopressin release due to manual restraint in the rat" role of body compression and comparison with other stressful stimuli. Endocrinology 104: 641 - 6 4 4 . Iverson L L, Iverson S D, Bloom F E (1980) Opiate receptors influence vasopressin release from nerve terminals in rat neurohypophysis. Nature 284: 350-351. Kamoi K, White K, Robertson G L (1979) Opiates elevate the osmotic threshold for vasopressin (VP) release in rats. Clin Res 27: 254A. Keii L C, Severs W B (1977) Reduction in plasma vasopressin levels of dehydrated rats following acute stress. Endocrinology 100: 3 0 - 38. Kendler K S, Weitzman R E, Fisher D A (1978) The effect of pain on plasma arginine vasopressin concentrations in man. J Clin Endocrinol 8:89 - 94. Knepel W, Nutto D, Anhut H, Hertting G (1980) Naloxone promotes stimulus-evoked vasopressin release in vivo. Eur J Pharmacol 65: 449-450. Lehmann E L (1975) Nonparametrics: Statistical Methods Based on Ranks. Holt & Day, New York, pp 287 - 297. Moore W W (1971) Antidiuretic hormone levels in normal subjects. Fed Proc 30:1387 - 1394. Moran W H, Mitenberger F W, Shuayb W A, Zimmerman B (1964) The relationship of antidiuretic hormone secretion to surgical stress. Surgery 56: 9 9 - 108. Oka Y, Wakayama S, Oyama T, Orkin L R, Becker R M, Blaufox M D, Frater R M (1981) Cortisol and antidiuretic hormone responses to stress in cardiac surgical patients. Canad Anaesth Soc J 28: 3 3 4 - 338.
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Robertson G L (1977) The regulation of vasopressin function in health and disease. Rec Prog Horm Res 33: 333-385. Robertson G L (1985) Vasopressin. In: Ingbar S (Ed) Contemporary Endocrinology, Vol. 2, Plenum, New York, pp 403 - 450. Robertson G L, Mahr E A, Athar S, Sinha T (1973) Development and clinical application of a new method for the radioimmunoassay of arginine vasopressin in human plasma. J Clin Invest 52: 2340- 2352. Rossier J, Battenberg E, Pittman Q, Bayon A, Koda L, Miller R, Guillemin R, Bloom F (1979) Hypothalamic enkephalin neurones may regulate the neurohypophysis. Nature 277:653 - 655. Rydin H, Verney E B (1938) The inhibition of water diuresis by emotional stress and by muscular exercise. Q J Exp Physiol 27:343 - 374. Shimamoto K, Miyahara M (1976) Effect of norepinephrine infusion on plasma vasopressin levels in normal human subjects. J Clin Endocrinol Metab 43:201 - 204. Summy-Long J Y, Keil L C, Deen K, Severs W B (1981) Opiate regulation of angiotensin-induced drinking and vasopressin release. J Pharmacol Exp Ther 217: 630- 637. Ukai M, Moran W H, Zimmerman B (1968) The role of visceral afferent pathways on vasopressin secretion and urinary excretory patterns during surgical stress. Ann Surg 168: 16-28. Van Wimersma Greidanus T B, Thody T J, Verspaget H, de Rotta G A, Goedemans H H, Croiset G, Van Ree J M (1979) Effects of morphine and ~-endorphin on basal and elevated levels of ¢t-MSH and vasopressin. Life Sci 24:579 - 586. v Bormann B, Weidler B, Denhardt R, Frings N, Lennhartz H, Hempelmann G (1981) Plasma-antidiuretic hormone level as an indicator of postoperative stress (part II). A nasth Intensivther Notfallmed 1 6 : 3 1 9 - 322. Wade C E, Claybaugh J R (1980) Plasma renin activity, vasopressin concentration, and urinary excretory responses to exercise in men. J Appl Physiol 49:930 - 936. Weidler B, v Bormann B, Lennhartz H, Denhardt R, Hempelmann G (1981) Plasma-antidiuretic hormone as indicator of perioperative stress (part I). A nasth lntensivther Notfallmed 16:315 - 318. Wilier J C, Dehen H, Cambier J (1981) Stress-induced analgesia in humans: endogenous opioids and naloxonereversible depression of pain reflexes. Science 212: 689-691.