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Dexamethasone suppression test in helpless rats
530
BIOL PSYCHIATRY 1989;26:53c-532
Dexamethasone Suppression Test in Helpless Rats Lawrence Greenberg, ~~~~~in~ Edwards, and Fritz A. Henn
Introduction As first described by Overmier and Seligman (1967), the learned helplessness (LH) model of depression is still one of the better animal models of a human psychiatric disorder (Willner 1984). In helpless models, animals are exposed to an uncontrollable aversive stimuli. Some of the auimals so treated will exhibit subsequent performance deficits. Those animals with performance deficits will also demonstrate phenomenology and treatment responses similar to human unipolar depression (Sherman and Petty 1982). Hy~ractivity and dysregulation of the hypothalmic-pituatary-adrenal axis (HPA) is a wellestablished biological abnormality in affective disorders (Sachar 1967), as well as other conditions. Clinically this is assessed by the Dexamethasone Suppression Test (DST). Though the dysregulation of the HPA axis is somewhat nonspecific for affective disorder, it does seem to be a characteristic seen in a majority of patients with most forms of affective disorder. Consequently, if an homolog of the DST in animals can be demonstrated in helpless animals, it would support the model as useful in understanding depression and might provide a system for examining the mechanism of dysregulation of cortisol production. This article reports on
From the Department of Psychiatry and Behavioral Sciences, SUNY at Stony Brook, Stony Brook, NY. Supported in part by Grant K I lMNOO543 from the National Institute of Mental Health, USPHS (L.C.). Address reprint requests to Dr. L. Greenberg, Department of Psychiatry and Behavioral Sciences, HSC TIO, Room 040, SUNY at Stony Brook, Stony Brook, NY 11794. Received May 19, 1987; revised November 2, 1988.
G 1989 Society of Biological
Psychiatry
our experience in measuring HPA function in helpless rats.
Methods Male Sprague-Dawley rats, weighing between 250 and 350 g, were housed three to a cage with free access to standard lab chow and water in a temperature/humidity-controlled facility. A 12:12 hr day/night cycle was maintained.
The established parameters for helplessness training and testing in our laboratory have recently been described (Edwards et al. 1986). Briefly, to produce the helpless condition, rats were given intermittent and random footshock through an electrified floor grid at 0.8 A for 40 min. A bar-press in the cage was inactive. Twenty-four hours later, the rats were tested in the same apparatus. This time, shock could be terminated by a bar-press. Failure to do so within 20 set counted as a failed trial. Shock continued for 1 min or until the bar was pressed. Intertrial latency was 45 sec. Each animal received a total of 15 trials. Ten or more trial failures were accepted as helplessness. The nonhelpless state was defined for the purposes of this experiment as four or fewer failed trials. Procedure Three groups of animals were tested with dexametbasone. There was a naive control group, which had received neither training nor testing.
O@X-3223189/$03.50
In addition, two groups of animals that had received training and testing were examined: one that showed helpless behavior and one that didn’t. At 4:00 PM on the day following testing, each rat was weighed and anesthesized with carbon dioxide. Two milliliters of blood was drawn into heparinized tubes via heart puncture. Each animal received a sub~u~a~~~usinjection of dex~e~ason~ (to0 &g/g). TW~~~-~O~~ hours Iat% ~~~~~~~~~~ was repeated. Plasma was separated in a Beckman reargue (at 8W mm, 4°C) and stored at -8CFC until analysis. Helpless and nonhelpless animals had undergone testing and training at the same time, 24 hr prior to the DST. Plasma corticosterone analysis was performed by radioimmunoassay (RIA) (lz51 Corticosterone Kit for rats, Radioassay Systems Laboratories, Inc.).
respectively. As would be predicted if helpless behavior rather fhan electrical stimulation was correlated, a r-test analysis comparing change in cort,icosterone between nonhelpless and naive animals was nonsignificant (t = 0.5, ~33 0.1).
Finding a stafisf~~~y significant di~~ere~~ein the degree of ~o~i&osfe~ne sup~s~i~n in response to dexame~a~one befween helpless and nonhelpless or naive animals in our DST hamo~og is true for populations. The variability among individual animals is entirely consistent with DST results from depressed patients (Arana et al. 19$5). Our finding of no significant difference in baseline corticosterone levels between helpless and nonhelpless animals is consistent with the findings of Maier et al. (1986). The results suggest that the majority of animals who became helpless develop im~ed There were 16 animals in the naive and and 17 feedback ~g~lation in the HPA. This defect is animals in the no~he~~less group. The helpless not universal, either in clinical pop~~ationa or in this animal model. Recently, Naracz and cogroup consisted of 42 animals. Corticosterone workers (1988) reported on their experience with levels are expressed in nanograms per milliliter dexamethasone suppression of corticosterone in +SD. Baseline corticostcrone h?Vek (prior to rats stmjected to inescapable (IS) and escapable injection of dexamethasone) for naive, non(ES) and no shock (NS) rats. In their analog of helpless, and helpless animals were 294 -+ 99, 31 I t 1.56, and 286 T?;75 n&ml, respectively. their DST, animals were pretreated with either There was no significant difference in baseline de~am~thasone or saline 2 hr prior to fmining. They found fhat rats freated with dexame~~s~ne corficosferone ievels among fhese groups. Dexame~aso~e injection produced sign%- exposed to inescapable shock had ~mpai~d cant effects on ~o~~~~~teronelevels in df three suppression of ~o~i~oste~ne ~ornp~~d to the groups. Figure I is a bar graph showing the de~amet~aso~-beak ES group. No difference percentage of animals per group that did not was found between the saline pretreated ES and suppress their corticosterone level below 150 IS groups. Their saline-pretreated NS group had mg/ml following tlexamethasone. Postdexa- lower baseline corticosterone levels than their methasone corticosteroue levels for naive, non- ES or 1S groups, and the dexamethasono-prehelpless, and helpless rats were 133 % 97, 132 treated NS group suppressed almost totally. Our + 124, and 195 it: 97 ng/ml, respectively. As results support their finding that helpless nnia group, helpless animals could be distinguished mals have an impaired ability to suppress corfrom nonhelpless and naive animals by their ticosterone in response to dexamethasone, impaired abihty to suppress ~o~~~osferone folRecent studies on the pa~way mediative the lowing d~xame~as~~e. A f-test anafysis of the befpless ~bavio~ in these animals (Henn et al. change in ~o~~~osfero~~levels bdween helpless 1985) suggesf that this altered behavior involves and nonhelpless and heipless and naive rats was ~f~~~~fion processed in hippocampus. This tract t = 2.5, p < 0.01 and c = 1.95, p < 0.05, is modulated by noradrenergic input from locus
532
100
m
r
References Arana GW, Baldessarini RJ, Omsteen M (1985): The Dexamethasone Suppression Test for diagnosis and prognosis in psychiatry. Arch Gen Psychiatr?, 42: 1193.
go-
% m 80it
a a
2 5 z
Brief Reports
BIOLPSYCHIATRY 1989:26:530-532
Edwards E, Johnson J, Anderson D, Turano P, Henn FA (1986): Neur~hemical and behavioral consequences of mild uncontrollable shock: Effects of PCPA. Pharmacol Biochem Behav 25:415.
7060-
Gray JA (1982): The Neuropsychology of Anxiety: An Enquiry into the Functions of the Septo-Hippocampal System. New York: Oxford University
50-
N
(n=16/
Figure 1. A comparison of the percentage of animals showing corticosterone suppression below a level of I.50 mg/ml 24 hr folIowing a dexamethasone cbailenge of 100 p&kg. N, animals in experimental group (0) Naive animals; (a) shocked but not helpless ) shocked helpless animals. and serotonin (5 = HT) input from raphe. This suggests that the bridge connecting HPA function to affective states may be the hippocampal hypothalamic connections. As the highest concentration of cortisol receptors centrally are found in hippocampus, this site is an attractive one for further study. The results presented here suggest that psychological stress can alter HPA function. This alteration may be mediated via the hippocampal formation and this coupling may depend on a combination of factors, including affective state. Further studies will assess this hy~thesis and examine why the relationship between stress and HPA dysfunction is variable among animals. This could improve our understanding of the meaning of DST results in human clinical populations. coerulus
Press. Haracz JL, Minor T, Wilkins JN, Z~~e~an EG (1988): Learned helplessness: An experimental model of the DST in rats. Biol Psychiatry 23:388. Henn FA, Johnson JO, Edwards E, Anderson D (1985): Melancholia in rodents: Neurobiology and pharmacology. P~chophar~co~ Bull 2 1:443. InseI Tr, Nina PT, Afoi J, Jimerson DC, Skolnick P, Paul SM ( 1984): A benzodiazepam receptor-mediated model of anxiety. Arch Gen Psychiatry 41:741. Kocsis JH, Davis JM, Katz MM, Koslow SH, Stokes PE, Caspar R, Redmond DE (1985): Depressive behavior and hy~ractive adren~o~ical function. Am J Psychiatry 142:1291. Maier SF, Ryan SM, Barksdale CM, Kahn NH (1986): Stressor controllability and the pituitary-adrenal symptom. Behav Neurosci 100:669. Minor TR, Lolordo VM (1984): Escape deficits following inescapable shock: The role of contextual odor. J Exp Psycho! 10:168. Overmier JB, Seligman MEP (1967): Effects of inescapable shock upon subsequent escape and avoidance learning. J Comp Physiol Psycho1 63:23. Sachar EJ (1967): Corticosteroids in depressive illness. II. A longitudinal psychoendocrine study. Arch Gen Psychiat~ 17:554.
Sherman AD, Petty F (1982): Specificity of the learned helplessness animal model of depression. Pharmacol Biochem Behav 16:449.
BIOL PSYCHIATRY 1989;26:53c-532
Dexamethasone Suppression Test in Helpless Rats Lawrence Greenberg, ~~~~~in~ Edwards, and Fritz A. Henn
Introduction As first described by Overmier and Seligman (1967), the learned helplessness (LH) model of depression is still one of the better animal models of a human psychiatric disorder (Willner 1984). In helpless models, animals are exposed to an uncontrollable aversive stimuli. Some of the auimals so treated will exhibit subsequent performance deficits. Those animals with performance deficits will also demonstrate phenomenology and treatment responses similar to human unipolar depression (Sherman and Petty 1982). Hy~ractivity and dysregulation of the hypothalmic-pituatary-adrenal axis (HPA) is a wellestablished biological abnormality in affective disorders (Sachar 1967), as well as other conditions. Clinically this is assessed by the Dexamethasone Suppression Test (DST). Though the dysregulation of the HPA axis is somewhat nonspecific for affective disorder, it does seem to be a characteristic seen in a majority of patients with most forms of affective disorder. Consequently, if an homolog of the DST in animals can be demonstrated in helpless animals, it would support the model as useful in understanding depression and might provide a system for examining the mechanism of dysregulation of cortisol production. This article reports on
From the Department of Psychiatry and Behavioral Sciences, SUNY at Stony Brook, Stony Brook, NY. Supported in part by Grant K I lMNOO543 from the National Institute of Mental Health, USPHS (L.C.). Address reprint requests to Dr. L. Greenberg, Department of Psychiatry and Behavioral Sciences, HSC TIO, Room 040, SUNY at Stony Brook, Stony Brook, NY 11794. Received May 19, 1987; revised November 2, 1988.
G 1989 Society of Biological
Psychiatry
our experience in measuring HPA function in helpless rats.
Methods Male Sprague-Dawley rats, weighing between 250 and 350 g, were housed three to a cage with free access to standard lab chow and water in a temperature/humidity-controlled facility. A 12:12 hr day/night cycle was maintained.
The established parameters for helplessness training and testing in our laboratory have recently been described (Edwards et al. 1986). Briefly, to produce the helpless condition, rats were given intermittent and random footshock through an electrified floor grid at 0.8 A for 40 min. A bar-press in the cage was inactive. Twenty-four hours later, the rats were tested in the same apparatus. This time, shock could be terminated by a bar-press. Failure to do so within 20 set counted as a failed trial. Shock continued for 1 min or until the bar was pressed. Intertrial latency was 45 sec. Each animal received a total of 15 trials. Ten or more trial failures were accepted as helplessness. The nonhelpless state was defined for the purposes of this experiment as four or fewer failed trials. Procedure Three groups of animals were tested with dexametbasone. There was a naive control group, which had received neither training nor testing.
O@X-3223189/$03.50
In addition, two groups of animals that had received training and testing were examined: one that showed helpless behavior and one that didn’t. At 4:00 PM on the day following testing, each rat was weighed and anesthesized with carbon dioxide. Two milliliters of blood was drawn into heparinized tubes via heart puncture. Each animal received a sub~u~a~~~usinjection of dex~e~ason~ (to0 &g/g). TW~~~-~O~~ hours Iat% ~~~~~~~~~~ was repeated. Plasma was separated in a Beckman reargue (at 8W mm, 4°C) and stored at -8CFC until analysis. Helpless and nonhelpless animals had undergone testing and training at the same time, 24 hr prior to the DST. Plasma corticosterone analysis was performed by radioimmunoassay (RIA) (lz51 Corticosterone Kit for rats, Radioassay Systems Laboratories, Inc.).
respectively. As would be predicted if helpless behavior rather fhan electrical stimulation was correlated, a r-test analysis comparing change in cort,icosterone between nonhelpless and naive animals was nonsignificant (t = 0.5, ~33 0.1).
Finding a stafisf~~~y significant di~~ere~~ein the degree of ~o~i&osfe~ne sup~s~i~n in response to dexame~a~one befween helpless and nonhelpless or naive animals in our DST hamo~og is true for populations. The variability among individual animals is entirely consistent with DST results from depressed patients (Arana et al. 19$5). Our finding of no significant difference in baseline corticosterone levels between helpless and nonhelpless animals is consistent with the findings of Maier et al. (1986). The results suggest that the majority of animals who became helpless develop im~ed There were 16 animals in the naive and and 17 feedback ~g~lation in the HPA. This defect is animals in the no~he~~less group. The helpless not universal, either in clinical pop~~ationa or in this animal model. Recently, Naracz and cogroup consisted of 42 animals. Corticosterone workers (1988) reported on their experience with levels are expressed in nanograms per milliliter dexamethasone suppression of corticosterone in +SD. Baseline corticostcrone h?Vek (prior to rats stmjected to inescapable (IS) and escapable injection of dexamethasone) for naive, non(ES) and no shock (NS) rats. In their analog of helpless, and helpless animals were 294 -+ 99, 31 I t 1.56, and 286 T?;75 n&ml, respectively. their DST, animals were pretreated with either There was no significant difference in baseline de~am~thasone or saline 2 hr prior to fmining. They found fhat rats freated with dexame~~s~ne corficosferone ievels among fhese groups. Dexame~aso~e injection produced sign%- exposed to inescapable shock had ~mpai~d cant effects on ~o~~~~~teronelevels in df three suppression of ~o~i~oste~ne ~ornp~~d to the groups. Figure I is a bar graph showing the de~amet~aso~-beak ES group. No difference percentage of animals per group that did not was found between the saline pretreated ES and suppress their corticosterone level below 150 IS groups. Their saline-pretreated NS group had mg/ml following tlexamethasone. Postdexa- lower baseline corticosterone levels than their methasone corticosteroue levels for naive, non- ES or 1S groups, and the dexamethasono-prehelpless, and helpless rats were 133 % 97, 132 treated NS group suppressed almost totally. Our + 124, and 195 it: 97 ng/ml, respectively. As results support their finding that helpless nnia group, helpless animals could be distinguished mals have an impaired ability to suppress corfrom nonhelpless and naive animals by their ticosterone in response to dexamethasone, impaired abihty to suppress ~o~~~osferone folRecent studies on the pa~way mediative the lowing d~xame~as~~e. A f-test anafysis of the befpless ~bavio~ in these animals (Henn et al. change in ~o~~~osfero~~levels bdween helpless 1985) suggesf that this altered behavior involves and nonhelpless and heipless and naive rats was ~f~~~~fion processed in hippocampus. This tract t = 2.5, p < 0.01 and c = 1.95, p < 0.05, is modulated by noradrenergic input from locus
532
100
m
r
References Arana GW, Baldessarini RJ, Omsteen M (1985): The Dexamethasone Suppression Test for diagnosis and prognosis in psychiatry. Arch Gen Psychiatr?, 42: 1193.
go-
% m 80it
a a
2 5 z
Brief Reports
BIOLPSYCHIATRY 1989:26:530-532
Edwards E, Johnson J, Anderson D, Turano P, Henn FA (1986): Neur~hemical and behavioral consequences of mild uncontrollable shock: Effects of PCPA. Pharmacol Biochem Behav 25:415.
7060-
Gray JA (1982): The Neuropsychology of Anxiety: An Enquiry into the Functions of the Septo-Hippocampal System. New York: Oxford University
50-
N
(n=16/
Figure 1. A comparison of the percentage of animals showing corticosterone suppression below a level of I.50 mg/ml 24 hr folIowing a dexamethasone cbailenge of 100 p&kg. N, animals in experimental group (0) Naive animals; (a) shocked but not helpless ) shocked helpless animals. and serotonin (5 = HT) input from raphe. This suggests that the bridge connecting HPA function to affective states may be the hippocampal hypothalamic connections. As the highest concentration of cortisol receptors centrally are found in hippocampus, this site is an attractive one for further study. The results presented here suggest that psychological stress can alter HPA function. This alteration may be mediated via the hippocampal formation and this coupling may depend on a combination of factors, including affective state. Further studies will assess this hy~thesis and examine why the relationship between stress and HPA dysfunction is variable among animals. This could improve our understanding of the meaning of DST results in human clinical populations. coerulus
Press. Haracz JL, Minor T, Wilkins JN, Z~~e~an EG (1988): Learned helplessness: An experimental model of the DST in rats. Biol Psychiatry 23:388. Henn FA, Johnson JO, Edwards E, Anderson D (1985): Melancholia in rodents: Neurobiology and pharmacology. P~chophar~co~ Bull 2 1:443. InseI Tr, Nina PT, Afoi J, Jimerson DC, Skolnick P, Paul SM ( 1984): A benzodiazepam receptor-mediated model of anxiety. Arch Gen Psychiatry 41:741. Kocsis JH, Davis JM, Katz MM, Koslow SH, Stokes PE, Caspar R, Redmond DE (1985): Depressive behavior and hy~ractive adren~o~ical function. Am J Psychiatry 142:1291. Maier SF, Ryan SM, Barksdale CM, Kahn NH (1986): Stressor controllability and the pituitary-adrenal symptom. Behav Neurosci 100:669. Minor TR, Lolordo VM (1984): Escape deficits following inescapable shock: The role of contextual odor. J Exp Psycho! 10:168. Overmier JB, Seligman MEP (1967): Effects of inescapable shock upon subsequent escape and avoidance learning. J Comp Physiol Psycho1 63:23. Sachar EJ (1967): Corticosteroids in depressive illness. II. A longitudinal psychoendocrine study. Arch Gen Psychiat~ 17:554.
Sherman AD, Petty F (1982): Specificity of the learned helplessness animal model of depression. Pharmacol Biochem Behav 16:449.