Increased pituitary activation following metyrapone administration in post-traumatic stress disorder

Psychoneuroendocrinology, Vol. 21, No. 1, pp. 1-16, 1996 Pergamon Copyright © 1996 Elsevier Science Ltd. All rights reserved

Printed in Great Britain 0306-4530/96 $15.1X1 + .iX)

0306-4530(95)00055-0

CURT P. RICHTER AWARD WINNER

INCREASED PITUITARY ACTIVATION FOLLOWING METYRAPONE ADMINISTRATION IN POST-TRAUMATIC

STRESS DISORDER

Rachel Yehuda, Robert A. Levengood , James Schmeidler , Skye Wilson, Ling Song Guo and Douglas Gerber

Traumatic Stress Studies Program, Psychiatry Department, Mount Sinai School of Medicine and Bronx Veterans Affairs Medical Center, New York, NY 10468, USA

(Received 9 August 1995; in final form 26 September 19951

SUMMARY

Our previous findings have demonstrated that individuals with post-traumatic stress disorder (PTSD) show lower basal cortisol levels, a larger number of lymphocyte glucocorticoid receptors, and an enhanced suppression of cortisol following the administration of dexamethasone compared to normals and patients with major depression. We have previously suggested that these alterations reflect an enhanced negative feedback inhibition of the hypothalamic-pituitary-adrenal (HPA) axis in PTSD. However, in the absence of direct knowledge of pituitary capability in this disorder, it has been equally likely that the alterations observed reflected either pituitary or adrenal insufficiency. In thc present study, we examined ACTH release from the pituitary gland in PTSD following the administration of metyrapone. Metyrapone resulted in a significantly greater increase of ACTH and l l-deoxycortisol in combat veterans with PTSD (n = 11) compared with normal male volunteers (n = 8). When seen in the context of other abnormalities observed in PTSD, the present demonstration of increased pituitary activity in the absence of negative feedback provides unequivocal support for the hypothesis of enhanced negative feedback.

Keywords--Hypothalamic-pituitary-adrenal axis; Post-traumatic stress disorder; Metyraponc stimulation test; Depression; Stress; Negative feedback.

I N T R O D U C T I O N

Post- t raumat ic stress d isorder (PTSD) is a condi t ion that can occur in individuals who have been exposed to ex t remely t raumat ic events (Amer ican Psychia t r ic Associa t ion , 1994). Al though classif ied as an anxiety disorder , many o f the symptoms of PTSD, such as insomnia, impai red concentrat ion, decreased interest and emot iona l numbing, are s imi lar to s y m p t o m s o f major depress ive disorder (MDD). Fur thermore , there is an unusual ly high incidence of comorb id i ty be tween PTSD and M D D (Fr iedman & Yehuda, 1995; Green et al., 1992; Kulka et al., 1991; McFar lane , 1992).

Address correspondence and reprint requests to: Rachel Yehuda, Ph.D., Psychiatry Department 116/A, Bronx VAMC, 130 West Kingsbridge Road, Bronx, NY 10468, USA (Tel: 718 584 9000, ext 5881; Fax: 718 933 2121).

2 R. Yehuda et al.

The hypothalamic-pituitary-adrenal (HPA) axis is one of the major neuroendocrine systems mediating responses to stress (Chrousos & Gold, 1992; Murick et al., 1984). HPA axis abnormalities are also well-documented in major depression (Gold et al., 1988; Owens & Nemeroff, 1993). Because of PTSD's association with both stress and depression, many investigators have hypothesized that the HPA axis would be altered in PTSD (Dinan et al., 1990; Halbreich et al., 1989; Hockings et al., 1993; Hoffman et al., 1989; Kosten et al., 1990; Kudler et al., 1987; Mason et al., 1986; Smith et al., 1989, Pitman & Orr, 1990; Yehuda et al., 1990a, 1991a).

Interestingly, however, the pattern of HPA axis disruption observed in PTSD is markedly different from that associated with both MDD and the classic stress response (e.g. as reviewed in Yehuda et al. (1991a, 1993a, 1995a). For example, basal 24 h urinary excretion of cortisol has been reported as lower in PTSD (Mason et al., 1986; Yehuda et al., 1990a, 1993b, 1995b), whereas concentrations in MDD are higher (Anton, 1987; Carroll et al., 1981; Mason et al., 1986; Yehuda et al., 1993b), compared with normals. Chronobiological analyses of plasma cortisol measures, obtained every 30 min over the diurnal cycle, have confirmed that cortisol levels are lower in PTSD, and that PTSD patients show a stronger circadian rhythm and enhanced "signal-to-noise" ratio of cortisol compared to normal and depressed subjects (Yehuda et al., 1994a).

Correspondingly, PTSD patients have more Type II glucocorticoid receptors (GR) on circulating lymphocytes compared to normals (Yehuda et al., 1991b, 1993b, 1995c), whereas depressed patients have fewer receptors (Gormley et al., 1985; Whalley et al., 1986). In response to dexamethasone (DEX), PTSD subjects have shown a substantially augmented cortisol suppression compared to normals (Yehuda et al., 1993c, 1995c); this finding is opposite to the classic cortisol non-suppression typically observed in MDD (Carroll et al., 1981; Evans & Nemeroff, 1983).

Some HPA axis abnormalities observed in PTSD, however, are similar to those in MDD. For example, a blunted ACTH response to CRF has been observed in combat veterans with PTSD compared to normals (Smith et al., 1989). More recently, CSF CRF has been shown to be increased in PTSD (Darnell et al., 1994). Both of these abnormalities are well- documented in MDD (Amsterdam et al., 1987; Arato et al., 1989; Banki et al., 1987; France et al., 1988; Gold & Chrousos, 1985; Gold et al., 1986; Kathol et al., 1989; Lesch et al., 1988; Nemeroff et al., 1984; Widerlov et al., 1988). The demonstration of both of these alterations in MDD has been used to support the idea of sustained HPA axis hyperactivity in depression (Gold et al., 1988; Owens & Nemeroff, 1993). Because it is difficult, at first glance, to resolve the findings of low cortisol, increased glucocorticoid receptors and enhanced DST suppression, with findings supporting CRF hypersecretion, many investigators have concluded that HPA axis abnormalities described in PTSD are conflicting (e.g. Hockings et al., 1993; Olivera & Fero, 1990; Pitman & Orr, 1990). However, the results obtained from studies to date are compatible with each other, and several biological models can accommodate this constellation of known facts.

It has been proposed, for example, that PTSD patients show an enhanced negative feedback sensitivity of the HPA axis (Yehuda et al., 1995a, 1995c), rather than the now well- described reduced negative feedback sensitivity in MDD (e.g. Lowy et al., 1989; Young et al., 1991). The enhanced negative feedback model postulates that PTSD is characterized by chronic increases in the release of hypothalamic and possibly extrahypothalamic CRF, which leads to an altered responsiveness of the pituitary (e.g. as suggested by a blunted ACTH response to CRF). However, because of a primary increase in GR responsiveness--

Pituitary Activation in PTSD 3

which is not a component of MDD or classic stress responses--there is a stronger negative feedback inhibition that results in attenuated baseline ACTH and cortisol levels and enhanced responsiveness to exogenous steroid (i.e. dexamethasone). The model of enhanced negative feedback sensitivity, therefore, is compatible with all the HPA axis alterations observed to date. However, no data have yet been gathered that directly address the source of pituitary responsiveness, which is a critical component of the model.

Indeed, the findings observed could also reflect two other types of HPA axis disturbances in addition to enhanced negative feedback inhibition. These are: (1) insufficient adrenal activity, and (2) insufficient ACTH production or release from the pituitary glands. If low cortisol levels in PTSD are due to either adrenal or pituitary failure, increases in lymphocyte GR would reflect a compensatory (i.e. secondary) change in response to low cortisol levels. In both situations, the cortisol response to DEX could be enhanced. Furthermore, both of these possibilities are consistent with the idea that stress-activated neuropeptides may stimulate hypothalamic release of CRF; however, because of a failure at either the level of the pituitary or adrenal, cortisol would be low.

In the present study, the metyrapone stimulation test was used in order to directly address the hypothesis of an enhanced negative feedback sensitivity in PTSD. Metyrapone prevents adrenal steroidogenesis by blocking the conversion of 11-deoxycortisol to cortisol, thereby unmasking the pituitary gland from the influences of negative feedback inhibition (Lisansky et al., 1989). Thus, metyrapone administration allows a direct examination of pituitary release of ACTH while eliminating the potentially confounding effects of differing ambient cortisol levels or glucocorticoid receptor responsiveness. When metyrapone is administered in the morning--when HPA axis activity is relatively high--maximal pituitary activity can bc achieved, making it possible to evaluate group differences in pituitary capability. If there is an enhanced negative feedback inhibition in PTSD, it would be predicted that removal of negative feedback by metyrapone administration would result in a relatively greater increase in ACTH release from the pituitary and an augmented accumulation of 11-deoxycortisol (i.e. a hyper-responsiveness) compared to the normal increase (usually between two- and five- fold higher than baseline) typically observed in healthy and non-psychiatric controls (Tepperman, 1981). Alternatively, if HPA abnormalities represent a pituitary insufficiency, increases in ACTH and 11-deoxycortisol levels following metyrapone administration would be lower than normal (i.e. hyporesponsive). On the other hand, if the HPA axis abnormalities in PTSD reflect an adrenal insufficiency, ACTH and 11-deoxycortisol levels following metyrapone would rise to a comparable extent to those of normals because metyrapone would not affect the adrenal gland. In conjunction with the information already obtained about HPA axis alterations in PTSD, the metyrapone challenge test can provide an unequivocal statement concerning suprapituitary activation by CRH in PTSD, and address the probable nature of the other underlying HPA axis alterations observed.

METHODS

Subjects PTSD subjects were 11 combat Vietnam veteran outpatients (age 45.82 __+ 0.72 years; age

range: 42-48 years) recruited from the Bronx Veterans Affairs PTSD program. Combat- related traumatic experiences in these veterans were confirmed using the Combat Exposure Scale (US Government Printing Office, 1981). Eight, age-comparable, normal male controls (age 41.25 _ 2.58 years; range: 36-54 years) with no personal or family history of major

4 R. Yehuda et al.

psychiatric disorder were also recruited. After providing written informed consent, all subjects were evaluated with the Structured Clinical Interview for the DSM-III-R (Spitzer & Williams, 1989). All subjects were medically healthy by history, physical examination and laboratory screens (including SMA-18, CBC with differential, glucose, blood urea nitrogen, creatinine, thyroid function tests, liver function tests, stool guaiac, chest X-ray, and EKG). Subjects were medication-free for at least 4 weeks prior to the start of the protocol. PTSD subjects with primary mood or anxiety disorders other than PTSD (i.e. syndromes that either predated the PTSD or were determined to occur independent of PTSD symptoms), psychotic disorders, or substance dependence within 6 months were excluded from this group. Patients were also evaluated with the Clinician Administered PTSD Scale (CAPS) (Blake et al., 1990) in order to determine the presence and severity of PTSD. The mean and range of CAPS subscale scores for the PTSD group were as follows: intrusive subscale score: 13.54___ 1.45 (highest possible score 24); avoidance subscale score: 34+_2.5 (highest possible score 56); hyper-arousal subscale score 30.64 _+ 1.30 (highest possible score 42). These scores reflect moderate to severe PTSD symptoms.

Procedure Subjects arrived at the Clinical Neuroendocrine Laboratory of the Bronx Veterans Affairs

Hospital on each of two mornings of the study after an overnight fast and were instructed to remain supine for the duration of the study. Vital signs and initial behavioral ratings were obtained at 0930h and were monitored every 30 rain until the end of the study. Behavioral ratings of mood, anxiety, and sedation were obtained every hour by asking subjects to self- rate subjective experiences in these areas on 100 mm lines. Blood samples were drawn from a forearm vein at 1000h, ll00h, 1300h and 1500h. On the metyrapone day, 2500 mg metyrapone (Ciba-Geigy, Tarrytown, NY, USA) was administered orally with 30 cc milk of magnesia at 1000h. The decision to administer metyrapone in the morning was based on our interest in examining differences in pituitary activity when the HPA axis is active rather than quiescent. The interval between the baseline and metyrapone day was 1-3 days. For 14 of the subjects the baseline day preceded the metyrapone day. However, for the remaining five subjects the baseline measurement occurred the day after metyrapone administration. Blood samples were drawn on ice into prepared heparinized tubes and then centrifuged. Plasma was separated, aliquoted and stored frozen at -30°C for subsequent determination of cortisol, ACTH and 11-deoxycortisol, the inactive precursor of cortisol that accumulates as a result of metyrapone blockade of l l-fl-hydroxylase.

Radioimmunoassay Cortisol was assayed using a double-antibody ~25I radioimmunoassay from Diagnostic

Products Corporation (Los Angeles, CA, USA), which provides high specificity cortisol antiserum with low cross-reactivity to other steroids, particularly 11-deoxycorticosterone, which is present in unusually high concentrations following treatment with metyrapone. The lowest detection limit of cortisol is 0.30/~g/dl. The antibody source for this assay was rabbit. lntra- and inter-assay coefficients of variation in our laboratory for this study were 6.3% and 11.6%, respectively.

ACTH was also radioassayed using a 125I antibody solution from Diagnostic Products Corporation. This assay uses mouse monoclonal antibody, which binds only to the N- terminal region of ACTH. This antibody is 125I-labeled for detection. Rabbit polyclonal antibody prepared by affinity chromatography binds only to the C-terminal region of ACTH.

Pituitary Activation in PTSD 5

25

B a s e l i n e day M e t y r a p o n e day

20 • P T S D (n = 11) • P T S D (n = 11) - - " , Y - - C o n t r o l (n = 8) - - " c ~ ' - - ( ' o n t r o l (n = 8)

E 5X 1 5 - •

z • i i

~-" •s

' I 4 / t 100(Jh 1100h ' 0 1 0 0 h ()300h

r i m e o f d a y

Fig. 1. Plasma cortisol levels in PTSD and normal subjects on baseline and metyrapone days.

The lowest measurable concentration (assay sensitivity) is 1.0 pg/ml. The intra- and inter- assay coefficients for this study were 5.5% and 7.6%, respectively.

1 l-deoxycortisol was radioassayed using a 125I antibody kit from Diagnostic Products Corporation, using rabbit anti-deoxycortisol. The assay sensitivity is greater than 0.50 ng/ml. The intra- and inter-assay coefficients, respectively, were 5.0% and 8.7%.

Data analysis Differences in plasma ACTH and l l-deoxycortisol response to metyrapone were

measured by the ratio of the maximum metyrapone day value and the baseline value at the same time. Comparison between the PTSD and the controls was by a t-test of these ratios. Comparisons are presented as mean _+ standard error of the mean. Patterns of drug response over time were compared by three-way analysis of variance, with day and time of day as repeated measures for each subject in each of the two groups. Patterns of peak response to cortisol, ACTH, and ll-deoxycortisol were examined by discriminant analysis predicting group membership.

RESULTS

Metyrapone was generally well-tolerated by the subjects. Almost all subjects reported an increased sleepiness during the first and second hour following metyrapone administration, resulting in significantly higher ratings of sedation for both groups during the metyrapone day. Six subjects (four PTSD, two controls) reported mild lightheadedness. Significant gastrointestinal side effects were reported by three individuals (two PTSD, one control), and one PTSD subject reported dizziness. These effects were transient, usually lasting between 1 and 2 h following the administration of metyrapone. One PTSD subject had a significant dissociative episode (e.g. in which there was lost awareness of his surroundings for a period of about 45 min) approximately 1 h after metyrapone administration. There were no

6 R. Yehuda et al.

250

200 ~m

150 F-

< =~ 100

5O

i I

Baseline day

• PTSD(,,=~I) I / - -q~-- -Contro l (n = 8)

Metyrapone dav / / T

• P T S D ( n = 11) /

i I / / / I , 1000h l l 00h / 0100h 0300h

Time of day

Fig. 2. Plasma ACTH levels in PTSD and normal subjects on baseline and metyrapone days.

significant group, day or time differences or interactions in mood, anxiety, blood pressure or

pulse. Administration of metyrapone resulted in an almost complete suppression of cortisol (Fig.

1). However, there were no differences between groups in peak suppression of cortisol ( t=0 .29; d f= 17; PTSD: 0.088_+0.013; control: 0.082_+0.016). Analysis of variance demonstrated that the group × day × time interaction was not significant ( F = 1.12;

df = 3,51). Metyrapone also caused a dramatic increase in ACTH in both groups (Fig. 2). The peak

ACTH response was significantly higher in the PTSD group compared to controls (t = 2.45, d f= 17, p = . 0 3 ; PTSD: 10.17_+ 6.22; control: 4 . 4 3 _ 2.60). There was a significant group × day × time interaction (F = 4.09; df= 3,51; p = .011).

70

6O

5(3

.,.~ 4<)

o 30

10

l Baseline day l

• PTSD (n = 8) ~ , ~ - - . 7 - - - C o n t r o l (n = 6) /

Metyrapone day

• PTSD (n = 8) - - ~ > - - C o n t r o l (n = 6) ±

IL

1000h 1100h / / 0100h 0300h

T i m e o f d a y

Fig. 3. Plasma ll-deoxycortisol in PTSD and normal subjects on baseline and metyrapone days.

Pituitary Activation in PTSD 7

60 u

5o

o 30

20

I0

PTSD = closed cl inic= Contro ls = open ciz¢l¢=

0 0

0

0 0

• O

i I I 0 5 10 15

P e a k A C T H r e s p o n s e

Fig. 4. Relationship between peak ACTH and peak ll-deoxycortisol response to metyrapone.

The plasma concentration of l l-deoxycortisol also increased in response to metyrapone (Fig. 3). Unlike cortisol and ACTH, the effect of metyrapone on 1 l-deoxycortisol persisted to the day after administration. (ll-deoxycortisol levels were significantly higher in the subjects in whom the metyrapone day preceded the baseline day, but there were no differences in cortisol and ACTH levels at 1000h on the baseline day based on whether the metyrapone day preceded or followed this day.) Therefore, the five subjects for whom the metyrapone day preceded the baseline day were excluded from the analyses. The peak response was significantly higher in the PTSD group compared to controls (t = 3.12, df = 12, p= .009 PTSD: 35.49_+3.14; control: 18.25 +4.8), indicating increased adrenal steroid production in this group. This was also reflected in a significant group × day × time interaction (F = 9.79; df = 3,36; p < .001).

Discriminant analysis predicting group membership from the three peak responses (cortisol, ACTH and 11-deoxycortisol) was statistically significant (Chi square = 123: df = 3; p = .(1051. This discriminant function correctly classified all 14 subjects. The correlations between peak ACTH, peak ll-deoxycortisol, and peak cortisol and the discriminant function were 0.62, 0.59, and 0.15, indicating that peak cortisol does not contribute so substantially to the prediction of group membership. Figure 4 demonstrates that the PTSD and control groups are well-distinguished by the combination of the other two variables, peak ACTH and peak ll-deoxycortisol. With the exception of the one PTSD subject with low peak ACTH, who had only moderate peak 11-deoxycortisol response, there was a clear pattern that PTSD subjects were relatively high on both variables. Interestingly, this was the PTSD subject mentioned above who had a dissociative experience during the metyrapone challenge. However, for each variable, there was one normal subject who was moderately high, but low on the other variable.

DISCUSSION

The present findings are consistent with an enhanced negative feedback sensitivity in PTSD in that PTSD patients showed a greater ACTH and l l-deoxycortisol augmentation following metyrapone administration compared to normal controls. The results do not support either adrenal or pituitary insufficiency since those conditions would predict either

8 R. Yehuda et al.

normal or hyporesponsiveness of the pituitary gland in PTSD. The multivariate analysis further demonstrated that when cortisol, ACTH and l l-deoxycortisol responses are considered, the metyrapone test clearly distinguishes between PTSD and normal controls.

Metyrapone treatment blocked cortisol release to the same extent in PTSD as in normals. Therefore, the results cannot be interpreted as an artifact of differential effects of metyrapone on the suppression of cortisol. Although the baseline day cortisol levels were somewhat higher, and metyrapone day cortisol levels at 1000h lower, in PTSD compared to normal controls, none of these differences were significant. The lack of difference in morning and early afternoon cortisol levels is consistent with our previous observation, from a chronobiological study evaluating cortisol release every 30 rain over a 24 h period, demonstrating that significantly lower cortisol levels in PTSD are seen primarily in the evening and early morning hours (Yehuda et al., 1994a).

An increased ACTH or l l-deoxycortisol response to metyrapone is consistent with hypothalamic CRF hypersecretion (Lisansky et al., 1989; Young et al., 1994). There is now abundant evidence for both hypothalamic (Licinio & Gold, 1991; Nemeroff et al., 1992; Owens & Nemeroff, 1993) and extrahypothalamic CRF hypersecretion in MDD (Nemeroff et al., 1988). However, only few studies have directly explored the effects of metyrapone on HPA axis parameters in MDD. Ur et al. (1992) found exaggerated rises in ACTH in depressed DST non-suppressors compared to depressed DST suppressors and normal controls following administration of metyrapone. Similarly, Young et al. (1994) reported an increased build up of l l-deoxycortisol in depressed patients following the evening administration of metyrapone. Therefore, the response to metyrapone in the present study is compatible with the hypothesis that hypothalamic CRF is hypersecreted in PTSD, and is consistent with earlier findings of a blunted ACTH response to CRF in PTSD (Smith et al., 1989).

CRF not only acts as a neuroregulator of pituitary hormones (Rivier & Plotsky, 1986; Rivier et al., 1982) but also as a neuromodulator and neurotransmitter within the central nervous system (Smith et al., 1986). Although the present study does not provide direct support for extrahypothalamic release of CRF in PTSD, this possibility is supported by the recent demonstration of increased CSF CRF concentrations in PTSD (Darnell et al., 1994). CRF neurons are concentrated in brain regions that are thought to be involved in the pathogenesis of PTSD, such as the amygdala (Fellman et al., 1982; Merchenthaler, 1984; Sawchenko & Swanson, 1985; Swanson et al., 1983) and locus coeruleus (Chappell et al., 1986; Merchenthaler, 1984; Swanson et al., 1983). Furthermore, intraventricular injection of CRF in animals produces behavioral effects that resemble hyperarousal symptoms of PTSD, such as sleep disruption and increased startle response (Swerdlow et al., 1986). Therefore, CRF may be important in directly producing some of the symptoms of PTSD via stimulation of these areas in addition to its role in stimulating the pituitary gland (e.g. reviewed in Yehuda & Nemeroff (1994)).

The present results, together with our previous findings, demonstrate that the HPA axis may be stimulated at a suprapituitary level even though cortisol levels are low. We have previously hypothesized that alterations in Type II glucocorticoid receptor responsiveness may underlie the lower cortisol levels observed in PTSD (Yehuda et al., 1995a, 1995c). Type II receptors are expressed in ACTH- and CRF-producing neurons of the pituitary, hypothalamus, and hippocampus, and mediate most systemic glucocorticoid effects, particularly those related to stress responsiveness (Beato, 1989; DeKloet et al., 1991; Herman et al., 1989; Patel et al., 1989; Keller-Wood & Dallman, 1984). PTSD patients have

Pituitary Activation in PTSD q

an increased number of Type II receptors on lymphocytes (Yehuda et al., 1991b, 1993b, 1995c). Lymphocyte and brain GR have been found to share similar regulatory and binding characteristics (Lowy, 1989, 1990). Therefore, an assessment of lymphocyte glucocorticoid receptor function may provide an estimate of peripheral and central cortisol regulation (Lowy et al., 1989).

In classic models of receptor-ligand interactions, changes in GR are conceptualized as reflecting compensatory responses to the concentration of hormone (McEwen et al., 1986; Sapolsky et al., 1984; Schlecte et al., 1982; Tornello et al., 1982). Chronic stress has been found to "downregulate" glucocorticoid receptors (Sapolsky et al., 1984). The decreased number of cytosolic lymphocyte GR in MDD has been interpreted as reflecting a secondary adaptation to the chronic hypercortisolism observed in this condition (Gormley et al., 1985; Whalley et al., 1986). Alternatively, however, in addition to being regulated by glucocorticoids, GR may also regulate hormonal release by modifying the strength of negative feedback (Lowy et al., 1989; Svec, 1985). Therefore, an increased number and/or sensitivity of GR may constitute a primary alteration in PTSD. Such is the case, for example, in animals who are subjected to "handling" during the neonatal period. Early handling results in a permanent upregulation of hippocampal glucocorticoid receptors (Meaney et al., 1985a, 1985b, 1988, 1989) and a resultant decrease in cortisol following exposure to subsequent stress (Ader, 1970; Hess et al., 1969; Haltmeyer et al., 1967; Levine et al., 1967), which is similar to that observed in PTSD. The "upregulation" of glucocorticoid receptors is somewhat unusual because, typically, GR are thought to "downregulate" in response to cortisol and stress. The increased GR number is thought to underlie the attenuated cortisol response to stress that is observed in adult animals who have received neonatal handling. The observations of altered HPA axis parameters in early handled rats nicely parallel many of the findings observed in PTSD (i.e. not only in regard to the HPA axis but also with respect to alterations in thyroid hormones: early handled animals show increases in T3 and T4 (Meaney et al., 1987) that are similar to those recently described in PTSD (Mason et al., 1994)). Thus, the model of early handling may provide an interesting basis for further hypothesis testing regarding the true etiology of HPA axis alterations and, in particular, GR abnormalities, in PTSD.

In contrast to some of the HPA axis abnormalities that have been observed (i.e. low cortisol and enhanced DST suppression), GR has been found to be significantly higher in trauma-exposed individuals who meet criteria for PTSD. In one study, the number of GR in the non-PTSD group was lower than in the PTSD group. However, most of the veterans in the "non-PTSD" group had met criteria for PTSD in the past. It is unclear, therefore, whether the increased GR number in traumatized individuals reflects the experience of trauma p e r se or rather whether it is a marker for the development of PTSD following trauma. In the present investigation, a trauma control group was not studied. Therefore, it is unclear whether the pituitary hyper-responsiveness reflects the presence of current PTSD-- as do most of the HPA axis alterations in PTSD--the experience of trauma exposure, or a risk factor for PTSD that may have predated the traumatic event. Future studies utilizing genetic approaches (e.g. family history studies, twin studies, intergenerational investiga- tions, molecular biological techniques) or longitudinal biological examination of individuals at risk for trauma exposure could clarify the extent to which any of the HPA axis abnormalities are present prior to, during, or subsequent to the index trauma.

Given the numerous modulatory influences on the HPA axis at the level of the hippocampus, hypothalamus and the pituitary, it is likely that biological events, other than

10 R. Yehuda et al.

those we have considered, contribute to the complex HPA axis profile observed in PTSD. For example, the synergistic effects of vasopressin and CRF suggests that altered modulation by arginine vasopressin may be a critical factor in the expression of HPA axis regulation at the pituitary (Gispen-de Wied et al., 1992). Furthermore, catecholamines also regulate (and are regulated by) the hormones of the HPA axis (Yehuda et al., 1990b). Both CRF and GR receptors are concentrated in catecholamine-rich areas (Fellman et al., 1982; H~irfstrand et al., 1986; Merchenthaler, 1984; Sawchenko & Swanson, 1985; Swanson et al., 1983). The strong evidence for catecholamine alterations in PTSD (Charney et al., 1993; Murburg, 1994; Southwick et al., 1995; Yehuda et al., 1992) makes it likely that these neuromodulators also contribute to the alterations observed at any of the target organs of the HPA axis.

Because PTSD is a disorder that occurs in response to extreme stress, the findings of HPA axis alterations in this disorder compel us to regard the human stress response as diverse and varied, rather than as conforming to a simple unidirectional pattern in which stress- related CRF activation culminates in hypercortisolism (Chrousos & Gold, 1992). The basic stress literature has for some time acknowledged that HPA axis responses to stress are far more heterogeneous than originally conceptualized by Selye (1956). This literature has demonstrated that the strength and direction of the pituitary-adrenocortical response to stress may depend on the interaction of several factors, including the nature of the stressor (i.e. type, chronicity, severity, controllability and predictability) (Mason, 1968; Mason et al., 1976), and other modifiers of the stress response, such as previous stress history, genetic vulnerabilities, social factors, and subsequent environmental stressors. In particular, the effect of these modifiers in humans may drastically alter the HPA axis stress response and contribute to a wider stress response spectrum. Thus, these modifiers increase an individual's risk for the subsequent development of PTSD following exposure to a traumatic event.

Indeed, the possibility of biological risk factors for PTSD is clearly suggested by results of epidemiological and longitudinal studies that demonstrate that the emergence of PTSD following exposure to trauma is not an inevitable outcome. Many individuals do not develop PTSD following exposure to a traumatic events (Breslau et al., 1991; Green et al., 1992; Helzer et al., 1987; Kulka et al., 1991). Among those who do develop PTSD, the majority experience a permanent remission of symptoms (Resnick et al., 1993; Shore et al., 1989). The chronic or intermittent state of PTSD occurs only in a distinct minority. The fundamental challenge of PTSD research, then, is to identify the distinguishing characteristics of those individuals from whom those atypical responses are produced. Seen in this context, the complex nature of the HPA axis stress response in PTSD described here is plausible.

Importantly, the unusual HPA axis profile observed in PTSD validates the idea that PTSD is a distinct disorder that can be differentiated from other psychiatric conditions. This supports efforts in the field of traumatology to develop specialized treatment strategies for PTSD patients. Studies of biological systems other than the HPA axis have also revealed alterations in PTSD that are distinct from those observed in most other psychiatric conditions and differentiable from normals (Pitman et al., 1987; Shalev et al., 1993, 1992). These findings suggest that the biological changes following exposure to a trauma are associated with the symptoms of PTSD, and can be differentiated from those observed simply as a result of stress exposure.

Pituitary Activation in PTSD 11

Finally, it should be noted that the model of enhanced negative feedback is consistent with clinical observations of PTSD patients as being unusually responsive to stress. PTSD patients often show exaggerated behavioral and biological responses to environmental challenge and react to non-traumatic events as if they were far more dangerous than they are in reality (Southwick et al., 1993, 1995; Yehuda et al., 1995d). The lower background (i.e. baseline cortisol levels) and the ability to hyper-respond to the environment (e.g. by showing exaggerated responses to neuroendocrine challenge) may reflect a type of adaptation to stress in which individuals become hypersensitive and hyperaroused. This pattern of hyper- responsiveness may be present not only in the HPA axis but also in the catecholamine system. Indeed, recent studies have demonstrated an enhanced MHPG secretion in response to yohimbine that is accompanied by behavioral symptoms of anxiety, panic attacks and flashbacks (Southwick et al., 1993).

In conclusion, the present findings demonstrate increased pituitary responsiveness in PTSD, and in the context of the other disturbances that have been identified, provide unequivocal support for the hypothesis of enhanced negative feedback. It can, therefore, be proposed that, in addition to the classic pattern of increased cortisol levels in response to stress, there may be a contrasting paradigm of cortisol abnormalities following stress, characterized by diminished cortisol levels as a result of a stronger negative feedback inhibition. This profile serves to distinguish PTSD from normals and other psychiatric disorders, particularly MDD. The next generation of biological studies in PTSD must focus on relating the complex neurobiology of PTSD to epidemiologically identified risk factors for this disorder.

Acknowledgements: The authors wish to acknowledge Ms. Abbie Elkin and Ms. Elizabeth Houshmand for data management skills. This work was supported by NIMH-49536 (RY), NIMH-49555 (RY) and a VA Merit Award (RY).

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