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The effects of corticosterone implantation on schedule-induced wheelrunning and HPA function in intact rats
Life Sciences, Vol. 46, pp. 1451-1456 Printed in the U.S.A.
Pergamon Press
THE EFFECTS OF CORTICOSTERONE IMPLANTATION ON SCHEDULE-INDUCED WHEELRUNNING AND HPA FUNCTION IN INTACT RATS Wenjuan Lin and George Singer Department of Psychology Brain-Behaviour Research Institute La Trobe University, Bundoora, Australia, 3083 (Received in final form March 9, 1990)
Summary Previous research has shown that Schedule-induced wheelrunning (SIW) is dependent on the function of the nature of pituitary-adrenal axis and that the involvement is mainly through pituitary-adrenal circulating corticosterone levels. In the present study it was shown that a lo-20 day period of subcutaneous implantation of exogenous corticosterone in intact rats alters HPA function but does not influence SIW. Plasma corticosterone levels in corticosterone-treated rats significantly decreased when compared with either the vehicle treated or untreated controls, and atrophy of adrenals occurred in corticosterone-treated animals, suggesting that exogenous corticosterone implantation impairs HPA function. However, the behavioural results showed that corticosterone implantation to intact rats did not affect the acquisition and development of SIW. whether raise question of These results the corticosterone exerts a permissive action or not. of schedule-induced Since Falk's initial demonstration drinking (l), a host of other forms of schedule-induced behaviour (SIB) have been observed (2-5). To understand the nature of SIB, studies have been directed towards finding number of ihysiological substrates involved in this behaviour (6-9). We shown that both adrenalectomy and hypophysectomy have recently schedule-induced wheelrunning (SIW) the suppress and that suppressant effect under both conditions can be counteracted by corticosterone implantation (10, 11). These findings show that SIW is dependent on the function of the pituitary-adrenal axis and that the nature of pituitary-adrenal involvement is mainly through circulating corticosterone levels. The present study was designed to investigate the effect of In the corticosterone implantation on SIW in intact rats. hypothalamo-pituitary-adrenal (HPA) system there is a negative feedback mechanism which operates to control the levels of glucocorticoids in the blood. administration of Prolonged 0024-3205/90 $3.00+.oo Copyright (c) 1990 Pergamon Press plc
1452
CorticosteroneImplantationand SIW
Vol. 46, No. 20,199O
glucocorticoids to normal animals inhibits ACTH release and consequently causes the adrenal cortices to regress (12-16). It also been shown that HPA activity may be impaired in patients has on corticosteroid therapy (17-20). In the present study it was decided to investigate whether a lo-20 day period of subcutaneous implantation of corticosterone in intact animals would HPA alter function, and whether this treatment would influence SIW. The examination of the effects of exogenous corticosterone administration on HPA function as well as on SIB in intact animals will provide more information regarding the involvement of corticosterone in SIB. To monitor the function of the pituitary adrenocortex system, plasma corticosterone levels and adrenal gland weights of all rats under schedule conditions were measured at the termination of the experiment. Method Thirty-two male hooded Long-Evans Subjects an rats with initial body weight of about 300g and aged between 90 and 120 days were used. Rats were individually housed under temperature controlled conditions ( 2z'C + 1 ) with a 12 hour light/ 12 hour dark cycle. After a period of acclimatization to the laboratory, the rats were body-weight reduced, over a fourteen day period of restricted food intake, to 80% of their free feeding body weight, and were maintained at this weight throughout the experiment. Water was freely available at all times. Apparatus The test chamber was made of clear perspex with a stainless steel barred floor, and measured 33.5 x 28 x 42 cm. A food cup was located on one end wall of the chamber approximately 3.5 cm above the floor. Pellet delivery was automatically controlled by standard relay circuitry. Noyes standard formula 45 mg food pellets were used. The chamber was illuminated by a 40 watt globe, and ventilation fans provided masking against external sounds. A 26 cm diameter running wheel was at the rear of the chamber and turned in either or counter-clockwise a clockwise direction. The number of wheel revolutions was recorded on a five-digit electromechanical counter. Experimental sessions were conducted in eight chambers simultaneously. Procedure Twenty-four rats were used to assess SIW and were randomly assigned to one of 3 equal-sized groups: corticosterone treated (Intact(C)-Sch), vehicle treated (Intact(O)-Sch), or untreated (Intact-Sch). Rats in the Intact(C)-Sch group were subcutaneously implanted under ether anaesthesia with a pellet consisting of 50 mg of corticosterone combined with 50 mg of cholesterol (21). Rats in the Intact(O)-Sch group were implanted with with 100 mg cholesterol using the same procedure. a pellet Rats were allowed a 10 day post-implantation recovery period prior to testing, since corticosterone treated rats showed an immediate for the first food intake reduction and weight loss 5-6 days following implantation, after which food intake recovered and 80% of body weight was re-established. All rats in 3 schedule groups were tested daily for 1.5 hours at the same hour of day for 10 consecutive days in a FT 120 set non-reinforcement contingent schedule under which they received one 45 mg pellet every two minutes.
Vol. 46, No. 20, 1990
CorticosteroneImplantationand SIW
1453
The remaining eight rats served as non-schedule controls (Intact-Nsch). They were tested under non-schedule conditions in which the same number of pellets used for the schedule condition were given in a single food presentation at the start of each session. Biochemical Assay Plasma corticosterone levels were measured using a modification of the competitive protein binding globulin assay described by Murphy [22]. Horse serum provided the source of binding protein. The limit of sensitivity was approximately 0.50 ng corticosterone, intraand the and inter-assay coefficients of variation All were 10% and 13% respectively. subjects from the schedule groups were killed by decapitation at the completion of ten test sessions. Trunk blood was collected in heparinised tubes and centrifuged at 3200 rpm for 20 minutes. The plasma was stored at -80% until assayed. Measurement of Adrenal Gland Adrenal glands were removed from the animals immediately after the collection of blood. Each gland was dissected from the periadrenal tissue and weighed on an electric balance. Results The mean numbers of wheel revolutions (M + SE) over the lo-day period for the 4 experimental groups are shown in FIG.l. A two-way ANOVA with one repeated measures showed a significant treatment effect (F=3.17, df=3,28; p
MEAN NO. OF WHEEL
REVOLUTIONS
mo-
400 -
-5
INTACT
(C)-Sch
-A-
INTACT
(0)-Sch
+
INTACT
-Sch
-e-
INTACT
-Nach
300 -
200-
100-
01”“““”
0
1
2
3
4
5
8
7
8
910
DAYS FIG. 1 Mean number of wheel revolutions during 1.5 hours of exposure to schedule or non-schedule food for 4 groups of experimental rats.
1454
CorticosteroneImplantationand SIW
30
OORTICOSTERONE
Vol, 46, No. 20, 1990
(ug/lOOml)
n INTACT -Sch
INTACT (0) -Sch
INTACT (C) -Sch
FIG. 2 Plasma corticosterone levels (M + SE) (ng/100ml) groups of corticosterone-treatedvehicle-treated untreated intact rats after 10 daily wheelrunning under schedule conditions. ADRENAL
15
r
WEIGHT
INTACT -Sch
(mg/lOOg
in and testes
B.W.)
INTACT (0) -Sch
INTACT (C) -Sch
FIG. 3 Adrenal weights (M + SE) (mg/lOOg body wt) in groups of vehicle-treated and untreated corticosterone-treaded, under tests 10 daily wheelrunning intact rats after schedule conditions.
Vol. 46, No. 20, 1990
CorticosteroneImplantationand SIW
1455
Simple main effect tests revealed that only Intact-Nsch group had significant low levels of wheel revolutions and there was no day effect on wheelrunning in this group. Inspection of FIG.1 shows that all three schedule groups significantly increased wheelrunning over the 10 day testing period and had the same level of SIW. These results show that corticosterone treatment did not affect SIW in the intact rats.
Plasma corticosterone concentrations (M + SE) in the 3 schedule groups are shown in FIG.2. A one-way ANOVA showed a significant treatment effect (Fa4.42, df= 2,21; P < 0.05). A post hoc Newman-Keuls test at the 0.05 level of significance showed that the corticosterone-treated animals had significantly lower levels of corticosterone than did either the vehicle-treated or The the untreated animals which did not differ from each other. corticosterone-treated rats had an average level of 11.6 ug/lOO ml of plasma corticosterone, which is comparable to the level reported for rats with corticosterone adrenalectomized implantation (7). This suggests that the circulating levels of corticosterone observed in corticosterone-treated intact rats reflect exogenous levels of corticosterone dissolved from the than implanted corticosterone rather endogenous pellets, corticosterone secreted from the adrenal cortices. Mean adrenal weights for the 3 schedule groups are shown in FIG.3. A one-way ANOVA showed a significant treatment effect (~-7.83, df-2, 21; P
1456
Corticosterone Implantation and SIW
rapidity of impairment corticoid treatment.
of
HPA
function
Vol. 46, No. 20, 1990
following
exogenous
Of interest is that the impaired HPA function induced by exogenous corticosterone implantation did not influence SIW. This is because a basal physiological level of corticosterone (11.6 ug/lOO ml), which may have been dissolved from the corticosterone implants, was still in general circulation. These results are consistent with earlier findings in the which plasma corticosterone levels in adrenalectomized rats with corticosterone implantation were found to be significantly lower than those in sham operated rats, yet the SIW suppressed by adrenalectomy was completely reversed at that level (10). These findings raise the question of whether the action of corticosterone in SIW is permissive or regulatory. A further dose-response study is in progress. References 1.
2. 3. 4. 5. 6. 7. 8. 9.
10. 11.
12. 13. 14. 15. 16. 17. 18. 19.
20. 21. 22.
J.L. FALK, Science 133 195-196 (1961). J.L. FALK, Phsiol. Behav. 6 557-588 (1971). M.J. WAYNER, Physiol. Behav. -12 851-869 (1974). M. WALLACE, G.SINGER, Biochem Behav. -5 483-490 (1976). A. TAZI, R. DANTZER, P.MORMEDE, M. LEMOAL, Behav. Brain Res. 19 249-256 (1986). G. SINGER, S. ARMSTRONG, M.J. WAYNER, Pharmac. Biochem. Behav. 3 869-872 (1975). L.D. DEVENPORT, J. Comp. Physiol. Psychol. 92 651-660 (1978). J.W. WRIGHT, S.C. KELSO, Physiol. Behav. -26 l-5 (1981). M. WALLACE, G. SINGER, J. FINLAY, S. GIBSON, Pharmac. Biochem. Behav. 18 129-136 (1983). W. LIN, G. SINGER, M. PAPASAVA, Pharmal. Biochem. Behav. 30 101-106 (1988). w. LIN, G. SINGER, D. IRBY, Pharmacol. Biochem. Behav. 35 (1990). (in press) XD. DUNN, A.J. CARRILLO, J. Endocr. 76 63-66 (1978). J.R. HODGES, J. SADOW, Br. J. PharmaK Chemother. 30 385-391 (1967). D.T. KRIEGER, A.S. LIOTTA, H. HAUSER, M.J. BROWNSTEIN, Endocrinology 105 737-742 (1979). General Endocrinology C.D. TURNER, J.T. BAGNARA, p.324-359 6th ed. W.B. Saunders Company, (1976). K. YAMADA, T. SATOH, Res. Conunun.Chem.Pathol. Pharmacol. 47 441-444 (1985). k. KNOL, Stand. J. Resp.Dis. 68 41-48 (1969). J.B. WOOD, J. clin. J. LANDON, V. WYNN, V.H.T. JAES, Endocr. Metab. 602-611 (1965). P.A. SAMPSON, KE. WINSTONE, B.N. BROOKE, Lancet 2 322-324 (1962). B.L.J. TREADWELL, 0. SAVAGE, E.D. SEVER, W.S.C. COPEMAN, Lancet 1 355-358 (1963). J.S. METER, D.J. MICCO, B.S. STEPHENSON, L.C. KREY, B.S. MCEWEN, Physiol. Behav. 22 867-870 (1979). B.E. PERSON-MURPHY, J. Clin. Endocrinol. -27 973-990 (1967).
Pergamon Press
THE EFFECTS OF CORTICOSTERONE IMPLANTATION ON SCHEDULE-INDUCED WHEELRUNNING AND HPA FUNCTION IN INTACT RATS Wenjuan Lin and George Singer Department of Psychology Brain-Behaviour Research Institute La Trobe University, Bundoora, Australia, 3083 (Received in final form March 9, 1990)
Summary Previous research has shown that Schedule-induced wheelrunning (SIW) is dependent on the function of the nature of pituitary-adrenal axis and that the involvement is mainly through pituitary-adrenal circulating corticosterone levels. In the present study it was shown that a lo-20 day period of subcutaneous implantation of exogenous corticosterone in intact rats alters HPA function but does not influence SIW. Plasma corticosterone levels in corticosterone-treated rats significantly decreased when compared with either the vehicle treated or untreated controls, and atrophy of adrenals occurred in corticosterone-treated animals, suggesting that exogenous corticosterone implantation impairs HPA function. However, the behavioural results showed that corticosterone implantation to intact rats did not affect the acquisition and development of SIW. whether raise question of These results the corticosterone exerts a permissive action or not. of schedule-induced Since Falk's initial demonstration drinking (l), a host of other forms of schedule-induced behaviour (SIB) have been observed (2-5). To understand the nature of SIB, studies have been directed towards finding number of ihysiological substrates involved in this behaviour (6-9). We shown that both adrenalectomy and hypophysectomy have recently schedule-induced wheelrunning (SIW) the suppress and that suppressant effect under both conditions can be counteracted by corticosterone implantation (10, 11). These findings show that SIW is dependent on the function of the pituitary-adrenal axis and that the nature of pituitary-adrenal involvement is mainly through circulating corticosterone levels. The present study was designed to investigate the effect of In the corticosterone implantation on SIW in intact rats. hypothalamo-pituitary-adrenal (HPA) system there is a negative feedback mechanism which operates to control the levels of glucocorticoids in the blood. administration of Prolonged 0024-3205/90 $3.00+.oo Copyright (c) 1990 Pergamon Press plc
1452
CorticosteroneImplantationand SIW
Vol. 46, No. 20,199O
glucocorticoids to normal animals inhibits ACTH release and consequently causes the adrenal cortices to regress (12-16). It also been shown that HPA activity may be impaired in patients has on corticosteroid therapy (17-20). In the present study it was decided to investigate whether a lo-20 day period of subcutaneous implantation of corticosterone in intact animals would HPA alter function, and whether this treatment would influence SIW. The examination of the effects of exogenous corticosterone administration on HPA function as well as on SIB in intact animals will provide more information regarding the involvement of corticosterone in SIB. To monitor the function of the pituitary adrenocortex system, plasma corticosterone levels and adrenal gland weights of all rats under schedule conditions were measured at the termination of the experiment. Method Thirty-two male hooded Long-Evans Subjects an rats with initial body weight of about 300g and aged between 90 and 120 days were used. Rats were individually housed under temperature controlled conditions ( 2z'C + 1 ) with a 12 hour light/ 12 hour dark cycle. After a period of acclimatization to the laboratory, the rats were body-weight reduced, over a fourteen day period of restricted food intake, to 80% of their free feeding body weight, and were maintained at this weight throughout the experiment. Water was freely available at all times. Apparatus The test chamber was made of clear perspex with a stainless steel barred floor, and measured 33.5 x 28 x 42 cm. A food cup was located on one end wall of the chamber approximately 3.5 cm above the floor. Pellet delivery was automatically controlled by standard relay circuitry. Noyes standard formula 45 mg food pellets were used. The chamber was illuminated by a 40 watt globe, and ventilation fans provided masking against external sounds. A 26 cm diameter running wheel was at the rear of the chamber and turned in either or counter-clockwise a clockwise direction. The number of wheel revolutions was recorded on a five-digit electromechanical counter. Experimental sessions were conducted in eight chambers simultaneously. Procedure Twenty-four rats were used to assess SIW and were randomly assigned to one of 3 equal-sized groups: corticosterone treated (Intact(C)-Sch), vehicle treated (Intact(O)-Sch), or untreated (Intact-Sch). Rats in the Intact(C)-Sch group were subcutaneously implanted under ether anaesthesia with a pellet consisting of 50 mg of corticosterone combined with 50 mg of cholesterol (21). Rats in the Intact(O)-Sch group were implanted with with 100 mg cholesterol using the same procedure. a pellet Rats were allowed a 10 day post-implantation recovery period prior to testing, since corticosterone treated rats showed an immediate for the first food intake reduction and weight loss 5-6 days following implantation, after which food intake recovered and 80% of body weight was re-established. All rats in 3 schedule groups were tested daily for 1.5 hours at the same hour of day for 10 consecutive days in a FT 120 set non-reinforcement contingent schedule under which they received one 45 mg pellet every two minutes.
Vol. 46, No. 20, 1990
CorticosteroneImplantationand SIW
1453
The remaining eight rats served as non-schedule controls (Intact-Nsch). They were tested under non-schedule conditions in which the same number of pellets used for the schedule condition were given in a single food presentation at the start of each session. Biochemical Assay Plasma corticosterone levels were measured using a modification of the competitive protein binding globulin assay described by Murphy [22]. Horse serum provided the source of binding protein. The limit of sensitivity was approximately 0.50 ng corticosterone, intraand the and inter-assay coefficients of variation All were 10% and 13% respectively. subjects from the schedule groups were killed by decapitation at the completion of ten test sessions. Trunk blood was collected in heparinised tubes and centrifuged at 3200 rpm for 20 minutes. The plasma was stored at -80% until assayed. Measurement of Adrenal Gland Adrenal glands were removed from the animals immediately after the collection of blood. Each gland was dissected from the periadrenal tissue and weighed on an electric balance. Results The mean numbers of wheel revolutions (M + SE) over the lo-day period for the 4 experimental groups are shown in FIG.l. A two-way ANOVA with one repeated measures showed a significant treatment effect (F=3.17, df=3,28; p
MEAN NO. OF WHEEL
REVOLUTIONS
mo-
400 -
-5
INTACT
(C)-Sch
-A-
INTACT
(0)-Sch
+
INTACT
-Sch
-e-
INTACT
-Nach
300 -
200-
100-
01”“““”
0
1
2
3
4
5
8
7
8
910
DAYS FIG. 1 Mean number of wheel revolutions during 1.5 hours of exposure to schedule or non-schedule food for 4 groups of experimental rats.
1454
CorticosteroneImplantationand SIW
30
OORTICOSTERONE
Vol, 46, No. 20, 1990
(ug/lOOml)
n INTACT -Sch
INTACT (0) -Sch
INTACT (C) -Sch
FIG. 2 Plasma corticosterone levels (M + SE) (ng/100ml) groups of corticosterone-treatedvehicle-treated untreated intact rats after 10 daily wheelrunning under schedule conditions. ADRENAL
15
r
WEIGHT
INTACT -Sch
(mg/lOOg
in and testes
B.W.)
INTACT (0) -Sch
INTACT (C) -Sch
FIG. 3 Adrenal weights (M + SE) (mg/lOOg body wt) in groups of vehicle-treated and untreated corticosterone-treaded, under tests 10 daily wheelrunning intact rats after schedule conditions.
Vol. 46, No. 20, 1990
CorticosteroneImplantationand SIW
1455
Simple main effect tests revealed that only Intact-Nsch group had significant low levels of wheel revolutions and there was no day effect on wheelrunning in this group. Inspection of FIG.1 shows that all three schedule groups significantly increased wheelrunning over the 10 day testing period and had the same level of SIW. These results show that corticosterone treatment did not affect SIW in the intact rats.
Plasma corticosterone concentrations (M + SE) in the 3 schedule groups are shown in FIG.2. A one-way ANOVA showed a significant treatment effect (Fa4.42, df= 2,21; P < 0.05). A post hoc Newman-Keuls test at the 0.05 level of significance showed that the corticosterone-treated animals had significantly lower levels of corticosterone than did either the vehicle-treated or The the untreated animals which did not differ from each other. corticosterone-treated rats had an average level of 11.6 ug/lOO ml of plasma corticosterone, which is comparable to the level reported for rats with corticosterone adrenalectomized implantation (7). This suggests that the circulating levels of corticosterone observed in corticosterone-treated intact rats reflect exogenous levels of corticosterone dissolved from the than implanted corticosterone rather endogenous pellets, corticosterone secreted from the adrenal cortices. Mean adrenal weights for the 3 schedule groups are shown in FIG.3. A one-way ANOVA showed a significant treatment effect (~-7.83, df-2, 21; P
1456
Corticosterone Implantation and SIW
rapidity of impairment corticoid treatment.
of
HPA
function
Vol. 46, No. 20, 1990
following
exogenous
Of interest is that the impaired HPA function induced by exogenous corticosterone implantation did not influence SIW. This is because a basal physiological level of corticosterone (11.6 ug/lOO ml), which may have been dissolved from the corticosterone implants, was still in general circulation. These results are consistent with earlier findings in the which plasma corticosterone levels in adrenalectomized rats with corticosterone implantation were found to be significantly lower than those in sham operated rats, yet the SIW suppressed by adrenalectomy was completely reversed at that level (10). These findings raise the question of whether the action of corticosterone in SIW is permissive or regulatory. A further dose-response study is in progress. References 1.
2. 3. 4. 5. 6. 7. 8. 9.
10. 11.
12. 13. 14. 15. 16. 17. 18. 19.
20. 21. 22.
J.L. FALK, Science 133 195-196 (1961). J.L. FALK, Phsiol. Behav. 6 557-588 (1971). M.J. WAYNER, Physiol. Behav. -12 851-869 (1974). M. WALLACE, G.SINGER, Biochem Behav. -5 483-490 (1976). A. TAZI, R. DANTZER, P.MORMEDE, M. LEMOAL, Behav. Brain Res. 19 249-256 (1986). G. SINGER, S. ARMSTRONG, M.J. WAYNER, Pharmac. Biochem. Behav. 3 869-872 (1975). L.D. DEVENPORT, J. Comp. Physiol. Psychol. 92 651-660 (1978). J.W. WRIGHT, S.C. KELSO, Physiol. Behav. -26 l-5 (1981). M. WALLACE, G. SINGER, J. FINLAY, S. GIBSON, Pharmac. Biochem. Behav. 18 129-136 (1983). W. LIN, G. SINGER, M. PAPASAVA, Pharmal. Biochem. Behav. 30 101-106 (1988). w. LIN, G. SINGER, D. IRBY, Pharmacol. Biochem. Behav. 35 (1990). (in press) XD. DUNN, A.J. CARRILLO, J. Endocr. 76 63-66 (1978). J.R. HODGES, J. SADOW, Br. J. PharmaK Chemother. 30 385-391 (1967). D.T. KRIEGER, A.S. LIOTTA, H. HAUSER, M.J. BROWNSTEIN, Endocrinology 105 737-742 (1979). General Endocrinology C.D. TURNER, J.T. BAGNARA, p.324-359 6th ed. W.B. Saunders Company, (1976). K. YAMADA, T. SATOH, Res. Conunun.Chem.Pathol. Pharmacol. 47 441-444 (1985). k. KNOL, Stand. J. Resp.Dis. 68 41-48 (1969). J.B. WOOD, J. clin. J. LANDON, V. WYNN, V.H.T. JAES, Endocr. Metab. 602-611 (1965). P.A. SAMPSON, KE. WINSTONE, B.N. BROOKE, Lancet 2 322-324 (1962). B.L.J. TREADWELL, 0. SAVAGE, E.D. SEVER, W.S.C. COPEMAN, Lancet 1 355-358 (1963). J.S. METER, D.J. MICCO, B.S. STEPHENSON, L.C. KREY, B.S. MCEWEN, Physiol. Behav. 22 867-870 (1979). B.E. PERSON-MURPHY, J. Clin. Endocrinol. -27 973-990 (1967).