- Email: [email protected]
The effects of predictable and unpredictable schedules of physical restraint upon rats
1896
Brain Fatty Acids and Response
to Alcohol
Vol. 28, No. 17, 1981
Acknowledgements We thank Dr. E. I. Eger, 11, for reading the manuscript. This research was supported by NIH Anesthesia Research Grant GM 15571, and by a grant from the Anesthesia Research Foundation.
References 1.
2. 3. 4. 5.
6. 7. 8.
9. 10. 11. 12. 13. 14.
15. 16. 17. 18. 19. 20. 21. 22.
BARAONA E. and C.S. LIEBER, 1979, J. Lipid Res. 20 289-315 (1979) LITTLETON J.M. and G. JOHN, J. Pharm. Pharmacol. 29 579-580 (1977) LITTLETON J.M., G.R. JOHN and S.J. GRIEVE, Alcoholism: Clin. Exp. Res. 3 50-56 (1979) SUN G.Y. and A.Y. SUN, Res. Commun. Chem. Pathol. Pharmacol. 24 405-408 (1979) CHIN J.H., L.M. PARSONS and D.B. GOLDSTEIN, Biochim. Biophys. Acta 513 358-363 (1978) MOSCATELLI E.A. and P. DEMEDIUK, Biochim. Biophys. Acta 596 331-337 (1980) SUN G.Y., D.M. CREECH, A.Y. SUN, and T. S#JMORAJSKI, Res. Commun. Chem. Pathol. Pharmacol. 22 617-620 (1978) STARICH G.H, and R.C. REITZ, Toxicol. Letts. 4 1-5 (1979) HILL M.W. and A.D. BANGHAM, Adv. Exp. Med. Bi~l. 59 1-9 (1975) JOHNSON D.A., N.M. LEE, R. COOKE and H.H. LOH, MoI. Pharmacol. 15 739-746 (1979) JOHN G.R., J.M. LITTLETON and P.A. JONES, Life Sci. 27 545-555 (1980) WHITE H.B., C. GALLI and R. PAOLETTI, J. Neurochem. 18 869-882 (1971) LAMPTEY M.S° and B.L. WALKER, J. Nutr. 108 358-367 (1978) TINOCO J . , R. BABCOCK, I. HINCENBERGS, B. MEDWADOWSKI and P. MILJANICH, Lipids i__3 6-17 (1978) BONNICHSEN R.K. and H. THEORELL, Scand. J. Clin. Lab. Invest. 3 58-62 (1951) ZAR J.H., B i o s t a t i s t i c a l Analysis, pp. 151-156, Prentice Hall, Inc., Englewood c i i f f s , N.J. (1974). KOBLIN 0.D., D.E. DONG and E.I. EGER, 11, J. Pharmacol. Exp. Ther. 211 317-325 (1979) JOLY J.G. and C. HETU, Can. J. Physiol. Pharmacol. 55 34-41 (1977) SUN G.Y., H. WINNICZEK, J. GO and S.L. SHENG, Lipids i0 365-373 (1975) KOBLIN D.D., D.E. DONG, J.E. DEADY and E.I. EGER, 11, J. Pharmacol. Exp. Ther. 212 546-552 (1980) DHOPESHWARKARG.A. and J.F. MEAD, Adv. Lipid Res. I I 109-142 (1973) LAMPTEY M.S. and B.L. WALKER, J. Nutr. 106 86-92 (~76
Life Sciences, Vol. 28, pp. 1897-1902 Printed in the U.S.A.
Pergamon Press
THE EFFECTS OF PREDICTABLE AND UNPREDICTABLE SCHEDULES OF PHYSICAL RESTRAINT UPON RATS Carlos MI. Quirce, Mauricio Odio and Jose M. Solano Laboratorio De Neurociencias Facultad De Farmacia Universidad De Costa Rica San Jose, Costa Rica and Department of Pharmacology and Toxicology School of Pharmacy and Pharmacal Sciences Purdue University West Lafayette, Indiana 47907 (Received in final form February 2, 1981)
Summary Adult, male rats were subjected to 15, 25, or 36 r e g u l a r l y or i r r e g u l a r l y scheduled sessions of physical r e s t r a i n t . I r r e g u l a r l y scheduled r e s t r a i n t applied for 25 or 36 sessions, resulted in p o s t - t r i a l a l t e r a t i o n s in plasma levels ef corticosterone, glucose, and free f a t t y acids of greater magnitude and longer duration than did r e g u l a r l y scheduled trials. The r e s u l t s confirm a more pronounced e f f e c t for the unpredictable stress. The e f f e c t s of predictable and unpredictable stressful stimulation have been the subject of much controversy. Ader and Friedman (I) reported that b r i e f exposures to novel environments s i g n i f i c a n t l y elevated plasma l l - h y d r o x y c o r t i c o s t e r o i d levels. Psychological factors have been indicated (2) as being more s t r e s s f u l than physical factors as measured by the elevation in adrenocortical activity. Testing of t h i s behavioral hypothesis has led to studies of the e f f e c t s of signalled versus unsignalled shock, although consistent findings have not been reported. Some studies indicate that s i g n a l l i n g reduces aversiveness (3, 4); these observations contrast with studies u t i l i z i n g physiological studies of aversiveness (5, 6, 7) which suggest that signalled shock is more s t r e s s f u l . Bassett and Cairncross (8) have c r i t i c i z e d the d e f i n i t i o n of p r e d i c t a b i l i t y as i d e n t i c a l with the presence or absence of a signal preceding a r e g u l a r l y occurring shock. When these authors examined both the presence and absence of s i g n a l l i n g paired to e i t h e r regular or i r r e g u l a r shock, they observed maximum elevations of plasma corticosterone during signalled i r r e g u l a r applications. I r r e g u l a r exposure to both meal and intermeal durations has s i m i l a r l y been reported (9) to cause more glandular stomach lesions than corresponding regular treatments. Experiments showing that animals prefer predictable to unpredictable shock (I0, I I , 12, 13, 14) have lent supportive evidence to the greater aversiveness of i r r e g u l a r l y applied stress.
0024-3205/81/171897-06502.00/0 Copyright (c) 1981 Pergamon Press Ltd.
1898
Predictable and Unpredictable
Stress
Vol. 28, No. 17, 1981
The present study examines the e f f e c t s of r e p e t i t i v e sessions of physical r e s t r a i n t upon rats. Varying the number of sessions was examined in both pred i c t a b l e (regular) and unpredictable ( i r r e g u l a r ) schedules. Materials and Methods Male, Sprague Dawley rats (250-350 grams) were group housed (4 to 5 per cage) in metal cages f o r ten days p r i o r to commencement of r e s t r a i n t . The animals had free access to tap water and were fed Purina Laboratory Chow throughout a l l treatments, l l l u m i n a t i o n was on a 12/12 schedule with l i g h t onset at 0730 hours. All r e s t r a i n t sessions were carried out between 1300 and 1700 hours in i n d i v i d u a l metal boxes with a fixed f l o o r and a moveable upper l i d . Wire mesh side screens permitted adequate v e n t i l a t i o n throughout the four hour sessions. Restraint was administered according to two types of schedules: unpredictable ( i r r e g u l a r ) or predictable ( r e g u l a r ) . During i r r e g u l a r schedules, the session date was randomly varied. Thus, a 24 hour period might separate one given r e s t r a i n t t r i a l from the preceding, while several days could lapse before another session was administered. Regular schedules were designed so that r e s t r a i n t t r i a l s were applied every 48 hours f o r the duration of the experiment. Table I summarizes the type of schedule u t i l i z e d during i r r e g u l a r t r i a l s . Animals were subjected to e i t h e r 15, 25 or 36 r e s t r a i n t sessions of e i t h e r schedule. Upon termination of any given treatment the animals were given 48, 96, or 144 hours in t h e i r home cages p r i o r to s a c r i f i c e . Plasma samples were collected and assayed f o r corticosterone (22), free f a t t y acids (23), and glucose (24). Control groups were treated i d e n t i c a l l y but did not receive r e s t r a i n t sessions. Significance was assessed by Student's t - t e s t . Results Table I I summarizes the r e s u l t s obtained from a p p l i c a t i o n of 15, 25, or 36 sessions of r e s t r a i n t on a regular basis, i . e . at 48 hour i n t e r v a l s . When animals were s a c r i f i c e d at 48 hours a f t e r the l a s t stress session, the data indicate v i r t u a l l y no deviations from control values. Only s l i g h t decreases in l e v e l s of plasma corticosterone ( a f t e r 36 sessions of r e s t r a i n t ) and plasma f a t t y acids ( a f t e r 15 sessions of r e s t r a i n t ) were noted. In contrast, Table I I I summarizes the r e s u l t s obtained when animals were exposed to s i m i l a r numbers (15, 25, or 36) of r e s t r a i n t sessions but on an i r r e g u l a r schedule. When s a c r i f i c e d 48 hours a f t e r the l a s t of 15 such sessions, no s i g n i f i c a n t e f f e c t s were seen, although plasma l e v e l s of f a t t y acids and glucose were s l i g h t l y elevated. Application of 25 sessions of r e s t r a i n t on an i r r e g u l a r basis evoked s i g n i f i c a n t elevations of a l l three plasma markers; a f t e r 36 sessions of r e s t r a i n t on the i r r e g u l a r schedule, plasma l e v e l s of corticosterone and f a t t y acids were s i g n i f i c a n t l y elevated, although glucose had returned to control values. A more d e t a i l e d analysis of the persistence of altered plasma biochemistry a f t e r 25 i r r e g u l a r r e s t r a i n t sessions is presented in the r e s u l t s in Table IV. As can be seen, s i g n i f i c a n t l y elevated l e v e l s of plasma corticosterone persisted for at least four days, followed, on day s i x , by a s i g n i f i c a n t decrease below control l e v e l s . Both plasma f a t t y acids and plasma glucose changed from s i g n i f i c a n t l y elevated values on day two a f t e r the l a s t r e s t r a i n t session, to s i g n i f i c a n t l y decreased values on days four and s i x .
Vol. 28, No. 17, 1981
Predictable and Unpredictable Stress
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TABLE I DAYS OF 25 SESSION UNPREDICTABLE STRESS SCHEDULE STRESS
REST
STRESS
REST
STRESS
27 28 29 30 31
53
REST 54 55 56 57
32
58 59
33 34 35 36
8
I0 II 12 13
37 38 39 40 41
14
15 16 17 18 19 20 22
64 65 66 67 68
42 43
69
44
70 71 72 73
45 46 47
21 23
60 61 62 63
48 49
74 75
24 25
50 51 52
26
76 77 78
TERMINATE-
TABLE I I EFFECTS OF REGULAR SESSIONS OF RESTRAINT ON PLASMA LEVELS OF CORTICOSTERONE, FATTY ACIDS, AND GLUCOSE IN RATS Animals were s a c r i f i c e d 48 hours a f t e r the l a s t stress session. Values reported are mean + S.E.M.: Numbers of animals are in parenthesis. Values s i g n i f i c a n t l y d i f f e r e n t from corresponding c o n t r o l s (P < .05) are underlined. PLASMA LEVELS NUMBER OF CORTICOSTERONE FATTY ACIDS GLUCOSE SESSIONS: GROUP ~g/ml uEq/ml mg/dl CONTROL 15 25 36
0.13 + .02
(II)
0.28 +
Ol
12)
140 + 4
(13)
RESTRAINT
0.12 + .03
(g)
0.23 +
Ol
12)
145 + 8
(13)
CONTROL
0.12 + .01
(I0)
0.26 +
02
12)
146 + 4
(12)
RESTRAINT
0.12 + .01
(I0)
0.27 +
02
12)
140 + 3
(15)
CONTROL
0.12 + .02
(9)
0.37 +
02
I0)
119 + 2
(14)
RESTRAINT
0.07 + .01
(9)
0.37 +
03
9)
112 ÷ 6
(15)
1900
Predictable
and Unpredictable
Stress
Vol. 28, No. 17, 1981
TABLE I I I EFFECTS OF IRREGULAR SESSIONS OF RESTRAINT ON PLASMA LEVELS OF CORTICOSTERONE, FATTY ACIDS, AND GLUCOSE IN RATS Animals were s a c r i f i c e d 48 hours a f t e r the l a s t session. Values reported are mean + S.E.M.; Numbers of animals are in p a r e n t h e s i s . Values s i g n i f i c a n t l y d i f f e r e n t from corresponding c o n t r o l s (P < .05) are u n d e r l i n e d . PLASMA LEVELS NUMBER OF SESSIONS:
GROUP
CORTICOSTERONE ug/ml
FATTY ACIDS ~Eq/ml
GLUCOSE mg/dl
CONTROL
0.II
+ .01
(II)
0.25 + .01
(13
121 + 5
(13)
RESTRAINT
0.II
+ .03
(II)
0,30 + .03
(II
131 + 5
(II)
CONTROL
0.09 + .01
(I0)
0.32 + .02
(13
128 + 2
(13)
RESTRAINT
0.14 ± .01
(I0)
0,54 + .01
(14
148 + 4
(15)
CONTROL
0.II
(9)
0.29 + .01
(II
125 '+ 3
(12)
RESTRAINT
0.17 + .02
(I0)
0.35 + .01
(13
122 + 1
(12)
15
25 + .01
36 Discussion
Bassett and Cairncross (8, 15) have suggested that irregularly applied stress may elevate corticosterone concentrations through a central component acting upon the hypothalamic-hypophyseal-adrenocortical system. In situations involving extreme psychological stress, normal inhibitory feedback mechanisms acting on the hypothalamus would be bypassed through the involvement of higher brain centers (15). In this context, the four days following the terminal session would show corticosterone elevations due to the maintenance of this central bypass system. By day six this overriding effect would yield to normal regulatory hypothalamic-adenophypophyseal-adrenocortical mechanisms with the consequent feedback inhibition resulting.in decreases. The fact that only irregularly scheduled restraint results in persistent elevations of plasma corticosterone, f a t t y acids, and glucose suggests that psychological factors play an extremely important role in the development of an enduring stress-like syndrome. This agrees with observations regarding the effects of unpredictability formulated by Mason (17). Furthermore, the poststress persistence of this syndrome after either 25 or 36, but not 15 sessions, suggests that longterm irregular applications are necessary in order to develop lasting alterations. This may prove a valuable model for understanding the development of stress in humans. Certainly, humans need not be continuously subjected to stressful stimuli in order to show stresslike responses. Acutally, continual exposure to stressful situations such as overcrowding were shown by various authors (18, 19) to lead to habituation and adaptation. However, other human studies (20, 21) have suggested a relationship between unpredictable changes in l i f e style and the development of stresslike symptomatology.
Vol. 28, No. 17, 1981
Predictable and Unpredictable Stress
1901
TABLE IV DURATION OF EFFECTS OF IRREGULAR STRESS SESSIONS Animals were exposed to 25 sessions of i r r e g u l a r l y scheduled r e s t r a i n t stress and s a c r i f i c e d at 48, 96, or 144 hours following termination of the l a s t session. Values reported are mean + S.E.M.; Numbers of animals are in parenthesis. Values s i g n i f i c a n t l y d i f f e r e n t from corresponding controls (P < .05) are underlined. CORTICOSTERONE ug/ml
PLASMA LEVELS FATTY ACIDS uEq/ml
CONTROL
0.09 + .01
(13)
0.32 +
02
(13)
122 + 3
(13)
RESTRAINT
0.14 + .02
(I0)
0.58 +
Ol
(lO)
140 + 4
(I0)
CONTROL
0. II + .01
(13)
0.28 +
Ol
(13)
127 + 3
(13)
RESTRAINT
0.20 + .01
(I0)
0.19+
02
(10)
I15 + 5
(.I0)
CONTROL
0. II + .01
(13)
0.31 +
Ol
(13)
128 + 3
(13)
RESTRAINT
0.06 + .01
(I0)
0.21 +
Ol
(I0)
117 + 3
(I01
TIME AFTER LAST SESSION GROUP DAYS
GLUCOSE mg/dl
2
4
Traditional models of stress, based mainly upon the e f f e c t s of regular a p p l i c a t i o n s of s t r e s s f u l s t i m u l i , have tended to i d e n t i f y shortlived a l t e r a t i o n s in adrenocortical a c t i v i t y with stress i t s e l f . The data presented in t h i s paper indicates that l a s t i n g post-stimulus a l t e r a t i o n s may be a b e t t e r d e f i n i t i o n of stress than mere short-term a c t i v a t i o n of these markers. REFERENCES
I. 2. 3. 4. 5. 6. 7. 8. 9. I0. II. 12. 13. 14. 15. 16. 17.
R. ADER and S. B. FRIEDMAN, Neuroendocrinology 3 378-386 (1968). S. B. FRIEDMAN, R. ADER, L. J. GROTAand T. L~SON, Psychosom. Med. 29 323-328 (l 967). M. E. SELIGMAN, J. comp. physiol. Psychol. 66 402-411 (1968). J. M. WEISS, Psychosom. Med. 32 397-408 (19TO). J. P. BRADY, D. R. THORNTONand D. DeFISHER, Psychosom. Med. 24 590-595, (1962). W. P. PARE, J. Psychol. 57 143-151 (1964). W. P. PARE, J. comp. physi---ol. Psychol. 74 459-466 (1971). J. R. BASSETT and K. D. CAIRNCROSS, Physi--ol. Behav. I0 901-907 (1973). R. L. WELKER, J. GARBER and F. BROOKS, Physiol. Behav--/ 18 639-645 (1977) J. M. ARABIAN and O. DESIDERATO, Anim. Learn. Behav. 3 191--195 (1975). P. BADIA, J. HARSH and C. C. COKER, Learn. Motivat. 6-264-278 (1975). J. HARSH and P. BADIA, J. exp. Analysis Behav. 23 349-355 (1975). J. S. LOCKARD, J. comp. physiol. Psychol. 56 526---530 (1963). C. C. PERKINS, R. G. SEYMANN, D. J. LEVIS and H. R. SPENCER, J. exp. Psychol. 72 190-196 (1966). J. R. BASS~TT and F. D. CAIRNCROSS, Pharmac. Biochem. Behav. 3 139-142 (1975). D. F. GANONG, Review o f MedloaZ Physiology, pp. 587, Lange Med. Pub., Los Altos, CA, 7th Ed. (1975). J. W. MASON, Psycbosom. Med. 30 576-607 (1968).
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18. 19. 20. 21. 22. 23. 24.
Predictable
and Unpredictable
Stress
Vol. 28, No. 17, 1981
R. E. MITCHELL, Amer. Sociol. Rev. 36 18-29 (1971). P. M. INSEL and H. C. LINDGREN, Psyc--hology Today II lO0-106 (1977). N. A. SCOTCH, Amer. J. Public Health 53 1205-1213-~-1963). S. HOOBLER, C. TEJADA, M. GUZMAN and ~ PARDO, Circulation 32 1205-1213 (1965). R. GUILLEMIN, G. W. CLAYTON, H. S. LIPSCOMB and J. D. SMITH, J. Lab Clin. Med. 53 830-832 (1959). W. G. DUNCOMBE, Clin. Chim. Acta 9 122-125 (1964). A. H. KADISH, R. L. LITTLE and J.-C. STERNBERG, Clin. Chem. 14 116-131 (1968).
Brain Fatty Acids and Response
to Alcohol
Vol. 28, No. 17, 1981
Acknowledgements We thank Dr. E. I. Eger, 11, for reading the manuscript. This research was supported by NIH Anesthesia Research Grant GM 15571, and by a grant from the Anesthesia Research Foundation.
References 1.
2. 3. 4. 5.
6. 7. 8.
9. 10. 11. 12. 13. 14.
15. 16. 17. 18. 19. 20. 21. 22.
BARAONA E. and C.S. LIEBER, 1979, J. Lipid Res. 20 289-315 (1979) LITTLETON J.M. and G. JOHN, J. Pharm. Pharmacol. 29 579-580 (1977) LITTLETON J.M., G.R. JOHN and S.J. GRIEVE, Alcoholism: Clin. Exp. Res. 3 50-56 (1979) SUN G.Y. and A.Y. SUN, Res. Commun. Chem. Pathol. Pharmacol. 24 405-408 (1979) CHIN J.H., L.M. PARSONS and D.B. GOLDSTEIN, Biochim. Biophys. Acta 513 358-363 (1978) MOSCATELLI E.A. and P. DEMEDIUK, Biochim. Biophys. Acta 596 331-337 (1980) SUN G.Y., D.M. CREECH, A.Y. SUN, and T. S#JMORAJSKI, Res. Commun. Chem. Pathol. Pharmacol. 22 617-620 (1978) STARICH G.H, and R.C. REITZ, Toxicol. Letts. 4 1-5 (1979) HILL M.W. and A.D. BANGHAM, Adv. Exp. Med. Bi~l. 59 1-9 (1975) JOHNSON D.A., N.M. LEE, R. COOKE and H.H. LOH, MoI. Pharmacol. 15 739-746 (1979) JOHN G.R., J.M. LITTLETON and P.A. JONES, Life Sci. 27 545-555 (1980) WHITE H.B., C. GALLI and R. PAOLETTI, J. Neurochem. 18 869-882 (1971) LAMPTEY M.S° and B.L. WALKER, J. Nutr. 108 358-367 (1978) TINOCO J . , R. BABCOCK, I. HINCENBERGS, B. MEDWADOWSKI and P. MILJANICH, Lipids i__3 6-17 (1978) BONNICHSEN R.K. and H. THEORELL, Scand. J. Clin. Lab. Invest. 3 58-62 (1951) ZAR J.H., B i o s t a t i s t i c a l Analysis, pp. 151-156, Prentice Hall, Inc., Englewood c i i f f s , N.J. (1974). KOBLIN 0.D., D.E. DONG and E.I. EGER, 11, J. Pharmacol. Exp. Ther. 211 317-325 (1979) JOLY J.G. and C. HETU, Can. J. Physiol. Pharmacol. 55 34-41 (1977) SUN G.Y., H. WINNICZEK, J. GO and S.L. SHENG, Lipids i0 365-373 (1975) KOBLIN D.D., D.E. DONG, J.E. DEADY and E.I. EGER, 11, J. Pharmacol. Exp. Ther. 212 546-552 (1980) DHOPESHWARKARG.A. and J.F. MEAD, Adv. Lipid Res. I I 109-142 (1973) LAMPTEY M.S. and B.L. WALKER, J. Nutr. 106 86-92 (~76
Life Sciences, Vol. 28, pp. 1897-1902 Printed in the U.S.A.
Pergamon Press
THE EFFECTS OF PREDICTABLE AND UNPREDICTABLE SCHEDULES OF PHYSICAL RESTRAINT UPON RATS Carlos MI. Quirce, Mauricio Odio and Jose M. Solano Laboratorio De Neurociencias Facultad De Farmacia Universidad De Costa Rica San Jose, Costa Rica and Department of Pharmacology and Toxicology School of Pharmacy and Pharmacal Sciences Purdue University West Lafayette, Indiana 47907 (Received in final form February 2, 1981)
Summary Adult, male rats were subjected to 15, 25, or 36 r e g u l a r l y or i r r e g u l a r l y scheduled sessions of physical r e s t r a i n t . I r r e g u l a r l y scheduled r e s t r a i n t applied for 25 or 36 sessions, resulted in p o s t - t r i a l a l t e r a t i o n s in plasma levels ef corticosterone, glucose, and free f a t t y acids of greater magnitude and longer duration than did r e g u l a r l y scheduled trials. The r e s u l t s confirm a more pronounced e f f e c t for the unpredictable stress. The e f f e c t s of predictable and unpredictable stressful stimulation have been the subject of much controversy. Ader and Friedman (I) reported that b r i e f exposures to novel environments s i g n i f i c a n t l y elevated plasma l l - h y d r o x y c o r t i c o s t e r o i d levels. Psychological factors have been indicated (2) as being more s t r e s s f u l than physical factors as measured by the elevation in adrenocortical activity. Testing of t h i s behavioral hypothesis has led to studies of the e f f e c t s of signalled versus unsignalled shock, although consistent findings have not been reported. Some studies indicate that s i g n a l l i n g reduces aversiveness (3, 4); these observations contrast with studies u t i l i z i n g physiological studies of aversiveness (5, 6, 7) which suggest that signalled shock is more s t r e s s f u l . Bassett and Cairncross (8) have c r i t i c i z e d the d e f i n i t i o n of p r e d i c t a b i l i t y as i d e n t i c a l with the presence or absence of a signal preceding a r e g u l a r l y occurring shock. When these authors examined both the presence and absence of s i g n a l l i n g paired to e i t h e r regular or i r r e g u l a r shock, they observed maximum elevations of plasma corticosterone during signalled i r r e g u l a r applications. I r r e g u l a r exposure to both meal and intermeal durations has s i m i l a r l y been reported (9) to cause more glandular stomach lesions than corresponding regular treatments. Experiments showing that animals prefer predictable to unpredictable shock (I0, I I , 12, 13, 14) have lent supportive evidence to the greater aversiveness of i r r e g u l a r l y applied stress.
0024-3205/81/171897-06502.00/0 Copyright (c) 1981 Pergamon Press Ltd.
1898
Predictable and Unpredictable
Stress
Vol. 28, No. 17, 1981
The present study examines the e f f e c t s of r e p e t i t i v e sessions of physical r e s t r a i n t upon rats. Varying the number of sessions was examined in both pred i c t a b l e (regular) and unpredictable ( i r r e g u l a r ) schedules. Materials and Methods Male, Sprague Dawley rats (250-350 grams) were group housed (4 to 5 per cage) in metal cages f o r ten days p r i o r to commencement of r e s t r a i n t . The animals had free access to tap water and were fed Purina Laboratory Chow throughout a l l treatments, l l l u m i n a t i o n was on a 12/12 schedule with l i g h t onset at 0730 hours. All r e s t r a i n t sessions were carried out between 1300 and 1700 hours in i n d i v i d u a l metal boxes with a fixed f l o o r and a moveable upper l i d . Wire mesh side screens permitted adequate v e n t i l a t i o n throughout the four hour sessions. Restraint was administered according to two types of schedules: unpredictable ( i r r e g u l a r ) or predictable ( r e g u l a r ) . During i r r e g u l a r schedules, the session date was randomly varied. Thus, a 24 hour period might separate one given r e s t r a i n t t r i a l from the preceding, while several days could lapse before another session was administered. Regular schedules were designed so that r e s t r a i n t t r i a l s were applied every 48 hours f o r the duration of the experiment. Table I summarizes the type of schedule u t i l i z e d during i r r e g u l a r t r i a l s . Animals were subjected to e i t h e r 15, 25 or 36 r e s t r a i n t sessions of e i t h e r schedule. Upon termination of any given treatment the animals were given 48, 96, or 144 hours in t h e i r home cages p r i o r to s a c r i f i c e . Plasma samples were collected and assayed f o r corticosterone (22), free f a t t y acids (23), and glucose (24). Control groups were treated i d e n t i c a l l y but did not receive r e s t r a i n t sessions. Significance was assessed by Student's t - t e s t . Results Table I I summarizes the r e s u l t s obtained from a p p l i c a t i o n of 15, 25, or 36 sessions of r e s t r a i n t on a regular basis, i . e . at 48 hour i n t e r v a l s . When animals were s a c r i f i c e d at 48 hours a f t e r the l a s t stress session, the data indicate v i r t u a l l y no deviations from control values. Only s l i g h t decreases in l e v e l s of plasma corticosterone ( a f t e r 36 sessions of r e s t r a i n t ) and plasma f a t t y acids ( a f t e r 15 sessions of r e s t r a i n t ) were noted. In contrast, Table I I I summarizes the r e s u l t s obtained when animals were exposed to s i m i l a r numbers (15, 25, or 36) of r e s t r a i n t sessions but on an i r r e g u l a r schedule. When s a c r i f i c e d 48 hours a f t e r the l a s t of 15 such sessions, no s i g n i f i c a n t e f f e c t s were seen, although plasma l e v e l s of f a t t y acids and glucose were s l i g h t l y elevated. Application of 25 sessions of r e s t r a i n t on an i r r e g u l a r basis evoked s i g n i f i c a n t elevations of a l l three plasma markers; a f t e r 36 sessions of r e s t r a i n t on the i r r e g u l a r schedule, plasma l e v e l s of corticosterone and f a t t y acids were s i g n i f i c a n t l y elevated, although glucose had returned to control values. A more d e t a i l e d analysis of the persistence of altered plasma biochemistry a f t e r 25 i r r e g u l a r r e s t r a i n t sessions is presented in the r e s u l t s in Table IV. As can be seen, s i g n i f i c a n t l y elevated l e v e l s of plasma corticosterone persisted for at least four days, followed, on day s i x , by a s i g n i f i c a n t decrease below control l e v e l s . Both plasma f a t t y acids and plasma glucose changed from s i g n i f i c a n t l y elevated values on day two a f t e r the l a s t r e s t r a i n t session, to s i g n i f i c a n t l y decreased values on days four and s i x .
Vol. 28, No. 17, 1981
Predictable and Unpredictable Stress
1899
TABLE I DAYS OF 25 SESSION UNPREDICTABLE STRESS SCHEDULE STRESS
REST
STRESS
REST
STRESS
27 28 29 30 31
53
REST 54 55 56 57
32
58 59
33 34 35 36
8
I0 II 12 13
37 38 39 40 41
14
15 16 17 18 19 20 22
64 65 66 67 68
42 43
69
44
70 71 72 73
45 46 47
21 23
60 61 62 63
48 49
74 75
24 25
50 51 52
26
76 77 78
TERMINATE-
TABLE I I EFFECTS OF REGULAR SESSIONS OF RESTRAINT ON PLASMA LEVELS OF CORTICOSTERONE, FATTY ACIDS, AND GLUCOSE IN RATS Animals were s a c r i f i c e d 48 hours a f t e r the l a s t stress session. Values reported are mean + S.E.M.: Numbers of animals are in parenthesis. Values s i g n i f i c a n t l y d i f f e r e n t from corresponding c o n t r o l s (P < .05) are underlined. PLASMA LEVELS NUMBER OF CORTICOSTERONE FATTY ACIDS GLUCOSE SESSIONS: GROUP ~g/ml uEq/ml mg/dl CONTROL 15 25 36
0.13 + .02
(II)
0.28 +
Ol
12)
140 + 4
(13)
RESTRAINT
0.12 + .03
(g)
0.23 +
Ol
12)
145 + 8
(13)
CONTROL
0.12 + .01
(I0)
0.26 +
02
12)
146 + 4
(12)
RESTRAINT
0.12 + .01
(I0)
0.27 +
02
12)
140 + 3
(15)
CONTROL
0.12 + .02
(9)
0.37 +
02
I0)
119 + 2
(14)
RESTRAINT
0.07 + .01
(9)
0.37 +
03
9)
112 ÷ 6
(15)
1900
Predictable
and Unpredictable
Stress
Vol. 28, No. 17, 1981
TABLE I I I EFFECTS OF IRREGULAR SESSIONS OF RESTRAINT ON PLASMA LEVELS OF CORTICOSTERONE, FATTY ACIDS, AND GLUCOSE IN RATS Animals were s a c r i f i c e d 48 hours a f t e r the l a s t session. Values reported are mean + S.E.M.; Numbers of animals are in p a r e n t h e s i s . Values s i g n i f i c a n t l y d i f f e r e n t from corresponding c o n t r o l s (P < .05) are u n d e r l i n e d . PLASMA LEVELS NUMBER OF SESSIONS:
GROUP
CORTICOSTERONE ug/ml
FATTY ACIDS ~Eq/ml
GLUCOSE mg/dl
CONTROL
0.II
+ .01
(II)
0.25 + .01
(13
121 + 5
(13)
RESTRAINT
0.II
+ .03
(II)
0,30 + .03
(II
131 + 5
(II)
CONTROL
0.09 + .01
(I0)
0.32 + .02
(13
128 + 2
(13)
RESTRAINT
0.14 ± .01
(I0)
0,54 + .01
(14
148 + 4
(15)
CONTROL
0.II
(9)
0.29 + .01
(II
125 '+ 3
(12)
RESTRAINT
0.17 + .02
(I0)
0.35 + .01
(13
122 + 1
(12)
15
25 + .01
36 Discussion
Bassett and Cairncross (8, 15) have suggested that irregularly applied stress may elevate corticosterone concentrations through a central component acting upon the hypothalamic-hypophyseal-adrenocortical system. In situations involving extreme psychological stress, normal inhibitory feedback mechanisms acting on the hypothalamus would be bypassed through the involvement of higher brain centers (15). In this context, the four days following the terminal session would show corticosterone elevations due to the maintenance of this central bypass system. By day six this overriding effect would yield to normal regulatory hypothalamic-adenophypophyseal-adrenocortical mechanisms with the consequent feedback inhibition resulting.in decreases. The fact that only irregularly scheduled restraint results in persistent elevations of plasma corticosterone, f a t t y acids, and glucose suggests that psychological factors play an extremely important role in the development of an enduring stress-like syndrome. This agrees with observations regarding the effects of unpredictability formulated by Mason (17). Furthermore, the poststress persistence of this syndrome after either 25 or 36, but not 15 sessions, suggests that longterm irregular applications are necessary in order to develop lasting alterations. This may prove a valuable model for understanding the development of stress in humans. Certainly, humans need not be continuously subjected to stressful stimuli in order to show stresslike responses. Acutally, continual exposure to stressful situations such as overcrowding were shown by various authors (18, 19) to lead to habituation and adaptation. However, other human studies (20, 21) have suggested a relationship between unpredictable changes in l i f e style and the development of stresslike symptomatology.
Vol. 28, No. 17, 1981
Predictable and Unpredictable Stress
1901
TABLE IV DURATION OF EFFECTS OF IRREGULAR STRESS SESSIONS Animals were exposed to 25 sessions of i r r e g u l a r l y scheduled r e s t r a i n t stress and s a c r i f i c e d at 48, 96, or 144 hours following termination of the l a s t session. Values reported are mean + S.E.M.; Numbers of animals are in parenthesis. Values s i g n i f i c a n t l y d i f f e r e n t from corresponding controls (P < .05) are underlined. CORTICOSTERONE ug/ml
PLASMA LEVELS FATTY ACIDS uEq/ml
CONTROL
0.09 + .01
(13)
0.32 +
02
(13)
122 + 3
(13)
RESTRAINT
0.14 + .02
(I0)
0.58 +
Ol
(lO)
140 + 4
(I0)
CONTROL
0. II + .01
(13)
0.28 +
Ol
(13)
127 + 3
(13)
RESTRAINT
0.20 + .01
(I0)
0.19+
02
(10)
I15 + 5
(.I0)
CONTROL
0. II + .01
(13)
0.31 +
Ol
(13)
128 + 3
(13)
RESTRAINT
0.06 + .01
(I0)
0.21 +
Ol
(I0)
117 + 3
(I01
TIME AFTER LAST SESSION GROUP DAYS
GLUCOSE mg/dl
2
4
Traditional models of stress, based mainly upon the e f f e c t s of regular a p p l i c a t i o n s of s t r e s s f u l s t i m u l i , have tended to i d e n t i f y shortlived a l t e r a t i o n s in adrenocortical a c t i v i t y with stress i t s e l f . The data presented in t h i s paper indicates that l a s t i n g post-stimulus a l t e r a t i o n s may be a b e t t e r d e f i n i t i o n of stress than mere short-term a c t i v a t i o n of these markers. REFERENCES
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Predictable
and Unpredictable
Stress
Vol. 28, No. 17, 1981
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