Psychiafry Research. 37: 119-193 Elsevier

179

Growth Hormone and Other Hormonal Responses to Clonidine in Melancholic and Nonmelancholic Depressed Subjects and Controls

Philip Mitchell, George Smythe, Gordon Parker, Kay Wilhelm, Henry Brodaty, Philip Boyce, and Ian Hickie

Received June 4. 1990; revised version received September 10, 1990; accepted November 3, 1990.

Abstract. To study putative differences in central neurotransmitter function in depressive subtypes, growth hormone, adrenocorticotropic hormone (ACTH), cortisol, and prolactin responses to the cy,-noradrenergic receptor agonist clonidine (1.3 pg/ kg i.v.) were examined in 26 subjects with major depression, 13 of whom had melancholia. The responses of 10 of these endogenousi melancholic subjects were compared with those of 10 controls who were matched to the patients on age, sex, and menopausal status. In 15 of the depressed subjects, prolactin and cortisol responses to the putative serotonergic agonist fenfluramine were also examined to test for associations between these challenges. There were no significant differences in any of the responses between melancholic and nonmelancholic depressive subgroups after controlling for age and sex. With the exception of a greater reduction in ACTH in the endogenousimelancholic subjects, there were also no significant differences in hormonal responses between these patients and controls. There was, however, a significantly greater reduction in systolic blood pressure in the control subjects. There were no significant correlations between the responses to clonidine and fenfluramine. The findings suggest that clonidine at a dosage of 1.3 pg/ kg is neither able to differentiate reliably between depressive subtypes nor to differentiate reliably between depressed and control subjects.

Key Words. Clonidine, growth hormone, adrenocorticotropic hormone, depres- sion. melancholia.

Since at least the 1920’s (Gillespie, 1929), the subclassification of depression has been

extensively debated (Farmer and McGuffin, 1989). As noted recently, “an assumption

of psychiatric research is that there is at least one form of depression that constitutes a

disease with an underlying pathophysiological dysfunction” (Hsaio et al., 1989),

although it remains controversial as to whether the endogenousimelancholic “dis-

Phihp Mitchell, M.D.. M.R.C.Psych., F.R.A.N.Z.C.P., is Senior Lecturer, School of Psychiatry, Univer- slty of New South Wales, Sydney. Austraha. George Smythe, A.S.T.C.. B.Sc., Ph.D., is Head, C.E. Heath Neuroscience Laboratory, Garvan Institute of Medical Research, Sydney, Australia. Gordon Parker, M.D., Ph.D.. F.R.A.N.Z.C.P., is Professor, School of Psychiatry, University of New South Wales. Svdnev. Australia. Kay Wilhelm is Senior Staff Specialist, Divislon of Psychiatry, Prince Henry Hospital, Sidney: Australia. Ian B. Hickle, M.D., F.R.A.N.Z.C.P.. 1s Staff Specialist. Divislon of Psvchiatrv. Prince Henrv Hospital, Sydney, Australia. Henry Brodaty, M.D.. F.R. A.C.P., F.R.A.N.Z.C.P., is Seni& Staff Spe&- ist, Division of Psychiatry, Prince Henry Hospital, Sydney, Austraha. Philip Boyce, M.D., F.R.A.N.Z.C.P., is Senior Lecturer, School of Psychiatry, University of New South Wales, Sydney, Australia. (Reprint requests to Dr. P.B. Mitchell, Mood Disorders Unit, School of Psychiatry, Prince Henry Hospital, Little Bay, NSW. 2036, Australia.)

01651781/91/%03.50 @ 1991 Elsevier Scientific Publishers Ireland Ltd.

180

ease” differs from residual disorders as a distinct type of depression or by severity. To demonstrate the validity of subtypes, a number of approaches have been

adopted: (1) A search for necessary and sufficient clinical features (Foulds, 1973); (2) the use of multivariate statistical techniques such as factor analysis, discriminant analysis, cluster analysis, and latent class analysis; and (3) examination of biological markers such as the dexamethasone suppression test. Despite early enthusiastic claims (e.g., Carroll et al., 198 I). there is still no clinically or statistically derived system, or biological marker, yet accepted as a “gold standard.”

More recently, hormonal responses to pharmacological challenges have been used to examine central neurotransmitter function indirectly in depression and so indicate possible subtypes (Checkley, 1980). Differences between clinically diagnosed melan- cholic and nonmelancholic depressive patients have been investigated with various putative challenges of central noradrenergic neurotransmitter systems (Matussek et al., 1988). Although findings have been inconsistent, significant differences between these depressive subtypes have been reported for the insulin tolerance test (Czernik, 1982), the growth hormone (GH) response to amphetamine (Langer et al., 1976) the cortisol response to methylamphetamine (Checkley, 1979) and the GH response to desipramine (Laakmann et al., 1986).

There has been, however, considerable uncertainty about the specificity of these challenges for central noradrenergic systems. For this reason there has been particular interest in neuroendocrine responses to the specific a,-noradrenergic receptor agonist clonidine. The GH response to clonidine has been demonstrated to be reduced in depressed subjects compared to controls in most (Matussek et al., 1980; Checkley et al., 1981; Charney et al., 1982; Siever et al., 1982; Boyeretal., 1986; Amsterdamet al., 1989) though not all (Horton et al., 1986; Schittecatte et al., 1989) studies. This blunted GH response may also represent a trait abnormality in subjects with endogenous depression (Mitchell et al., 1988). The negative study of Horton et al. (1986) is of interest, being the only study to use a clonidine dosage of I .3 pgi kg body weight (i.v.), whereas the other studies used either 150 1.18 or 2 pg/ kg i.v., or 5 pg/ kg oral. Horton et al. also used an inadequate drug-free interval of only 7 days before testing. (Periods of at least 2 or 3 weeks without medications are usually considered necessary in such

studies.) As outlined in Table I, there have been seven published studies comparing the GH

response to clonidine in endogenous and nonendogenous subjects, including the original report by Matussek et al. (1980). The major limitations of that original study were: (1) the lack of a standardized assessment interview and operationalized diagnos- tic criteria; and (2) a fatlure to match or control for either age or sex--factors that significantly affect the GH response to this challenge (Checkley, 1980). Of the subse- quent studies, only two have accounted for these covariates in their comparisons (Checkley et al., 1984; Ansseau et al., 1988). Two other studies (Boyer et al., 1986; Horton et al., 1986) used inadequate drug-free intervals (< 2 weeks) before testing.

Because of the significance of the methodologically strict study of Checkley et al. (1984) which found a blunted GH response in endogenous compared with reactive depressed subjects, we aimed to extend that study by: ( I) confirming that the dosage of clonidine used by that group (1.3 pgi kg) was also able to demonstrate hormonal differences between closely matched melancholic and control subjects; and (2) investi-

181

1. in endogenous (ED) vs. non- endogenous depression (non-ED)

Drug- Age, Diagnostic Non- free Clonidine sex,

criteria ED ED interval dose/ matching/ Signifi- Study for ED 0~) 0-0 (days) route control cance

Matussek et al 119801

Checkley et al

119841

Dolan &

Calloway

(19861

Horton et al.

119861

Boyer et al

119861

Ansseau et al

119881

Amsterdam et al.

(19891

/CD-9

Newcastle

RDC

Catego/

RDC/

Newcastle

Newcastle

Newcastle1

RDC

Newcastle

DSM-//I

10

10

7

7

17

10

15

12

12 28

10 21

7

7 14

17 7

10 8

152 14

6 10

150 P9 i.v.

1.3 pg/kg

i.v.

5 pgfkg p.0.

1.3 /.rg/kg

i.v.

156 Pg i.v.

156 fig i.v.

2.5 pg/kg

i.v.

No f

Matched *

No NS

No NS

No f

Matched *

No ‘3

Note. Newcastle = Newcastle Scale (16 = endogenous, 55 = nonendogenous!. NS = no signiffcant difference in growth hormone iGW response. * = signlflcantly reduced growth hormone response In endogenous subjects

1. “Major depression” (classification system unspeclfled 1. 2. Minor depression by Research Dlagnostlc Criteria ~RDCI 3. Slgniflcant difference for mean cumulative response only

gating other hormonal responses to clonidine (adrenocorticotropic hormone [ACTH], cortisol, and prolactin) in addition to GH in the melancholic and nonmelan- cholic subtypes.

In an associated study, we examined relationships between the GH response to clonidine and hormonal responses to a putative serotonergic agonist, fenfluramine. Although abnormal responses to pharmacological challenges of both noradrenergic (Matussek, 1988) and serotonergic systems (Meltzer and Nash, 1988) have been identified in depressed subjects, there has not yet been a published investigation that has used putative challenges of both neurotransmitter systems in the same subjects. Both the prolactin (Siever et al., 1984~; Coccaro et al., 1989) and cortisol (Weizman et al., 1988) responses to fenfluramine have been reported to be blunted in depressed subjects as compared to controls, though there has been one negative report (Asnis et al., 1988). We have recently reported a study of 27 depressed and 14 control subjects, in which we found a significantly diminished prolactin response in endogenous subjects as compared with controls, though this difference was partially dependent on reduced baseline prolactin levels and increased baseline cortisol levels in the depressed subjects (Mitchell and Smythe, 1990). We also demonstrated a reduced prolactin response to fenfluramine in endogenous compared to nonendogenous subjects (Mitchell et al., 1990).

182

Methods

Subjects. Study 1. Hormonal responses to clonidine in melancholic and nonmelancholic

depressed subjects. Twenty-six depressed subjects were tested with clonidine. All subjects met DSM-III (American Psychiatric Association, 1980) criteria and Research Diagnostic Criteria (RDC; Spitzer et al., 1978) for major depression, derived from a semistructured interview (Brodaty et al., 1987). Because of recent evidence indicating that the number of patients allocated to depressive subtypes varies significantly depending on the classification system (Davidson et al., 1984) subjects were classified as having melancholic (endogenous) or nonmelancholic (nonendogenous) depression according to each of the following diagnostic systems: DSM-III, RDC, and ICD-9(World Health Organization, 1978). For the RDC system, subjects were divided into endogenous (definite endogenous) and nonendogenous (probable endogenous + nonendogenous) subtypes to examine a more clearly identified endogenous group. Likewise for [CD-9, subjects were allocated to endogenous (296.1,296.3) and nonendog- enous (300.4) subgroups.

No patient over the age of 65 was included in the study. Subjects with significant medical disorders or regular alcohol intake > 30 g daily were excluded. All subjects were required to have been free of antidepressant or other psychotropic medications for at least 2 weeks, although benzodiazepine hypnotics were allowed. Depression severity was assessed by both the observer-rated 2l-item Hamilton Rating Scale for Depression (Hamilton, 1967) and the Zung Self-rating Scale (Zung, 1986).

Study 2: Hormonal responses to clonidine in endogenous/melancholic subjects and matched controls. Ten of the depressed subjects from study 1 who fulfilled DSM-III, RDC, and ICD-9 criteria forendogenousimelancholicdepression wereable to be matched with IO controls for age (* 5 years), sex, and menopausal status. Control subjects were assessed by clinical interview to exclude previous or current psychiatric disorders. Controls with significant medical disorders (as assessed by interview and physical examination) or regular alcohol intake > 30 g daily were excluded. They were also required to have been free of antidepressant or other psychotropic medications for at least 2 weeks, and were also excluded if they were taking other medications that might affect the neuroendocrine responses to clonidine. such as steroids or thyroid hormone replacement.

Study 3: Hormonal responses to clonidine and fenfluramine in the samedepressed subjects. Fifteen of the 26 depressed subjects from study 1 consented to testing with both fenfluramine and clonidine. For 12 of the 15 subjects, the fenfluramine test was performed first. The average interval (* SD) between tests was 4.3 (* 3.7) days (range 1-14 days). Table 2 provides details of the depressed samples for each of the three studies.

Procedures. Clonidine test. Patients were fasted overnight. and at 8-8:30 a.m.. a catheter was inserted

into an antecubital vein, after which they rested for I hour. Three baseline samples were then taken at l5-min intervals. The clonidine (1.3 wg, kg body weight, as used by Checkley et al. [ 19841) was diluted in 10 ml of normal saline and injected slowly over IO min. For 90 min after the injection began, blood was withdrawn at 15min intervals for hormone estimation, Throughout the procedure, pulse, blood pressure, and observer ratings of sedation were made every 5 min.

Fenfluramine test. This test was carried out on 15 of the same patients on a different test day. The same initial procedure was carried out as for the clonidine test. However. after the three baseline samples had been taken, n.t.-fenfluramine. 60 mg, was administered orally. Blood samples were then drawn hourly for 5 hours while the subject remained supine. A light meal was provided 4 hours after fenfluramine administration

Assays. Growth hormone levels were measured using polyclonal antiserum with an intra-assay coefficient of variation (CL’) of 5.80/c and an interassay cv of 10.99: at 3.95 mum I. Serum cortisol

183

Table 2. Details of subjects in each study

Study 1 Study 2

Depressed Controls

Study 3

Sample size

Inpatients

Female Total Postmenopausal

Male

26 10 10 15

22 9 11

16 7 7 10 6 3 3 2

10 3 3 5

Age Mean 43.2 46.6 42.6 40.7 SD 11.2 11 .o 10.5 11 .o

Bipolar disorder, depressed phase

Delusional depression

DSM-//I melancholic

RDC endogenous (definite)

/CD-9 endogenous

Hamilton score Mean SD

Zung score Mean SD

4 3 - 3

4 2 2

13 10 6

14 10 - 8

12 10 6

21.8 25.2 - 22.1 7.0 6.9 6.5

57.7 60.3 - 57.4 6.3 5.4 5.3

Note. RDC = Research Diagnostic Criteria Hamilton = Hamilton Rating Scale for Depression. Zung = Zung Self-rating Scale.

levels were measured by radioimmunoassay (RIA), with an mtra-assay cv of 6.8% and an interassay cv of 7.9% at 375 nmol, I. Plasma ACTH levels were measured by RIA, with an intra-assay cv of 13.2% at 26.6 ngil and an interassay cv of 23.5% at 25.4 ng/ I. Serum prolactin levels were also measured using a double antibody RIA wtth an interassay cv of 5% and an intra-assay cv of 6.5% at 658 mlUi 1.

Cortisol assays were performed for 25 of the 26 depressed subjects and all of the matched controls, while ACTH levels were assayed for 17 of the depressed subjects and all 10 of the controls. However, only six of the matched pairs had complete ACTH assays for both subjects.

Data Analysis. Baseline concentrations of the various hormones were taken as the average of the three prechallenge levels. Hormonal responses were analyzed as the absolute mcrease to peak above baseline (A). The frequency distributions of the examined variables were assessed by means of the Shapiro and Wilk test for normality of data (as set out in Madansky, 1988). Apart

184

from subjects’ age and blood pressure responses, all data were nonnormally distributed. For nonnormally distributed data, variables were log,,,-transformed before use of I tests. Student’s I test was used to compare group means. Repeated measures analysis of variance (ANOVA). testing for both linear (FIUI) and quadrattc (F ~UUI) trends, was used to look for any significant change in hormonal levels after the clonidine challenge. Analysis of covariance (ANCOVA) was used to determine the effect of covartates such as age and sex. All dependent variable data were log,,,-transformed before ANOVA or ANCOVA. Spearman’s rank order correlations (r~) were used to assess interrelationships between variables.

Results

Study 1: Hormonal Responses to Clonidine in Melancholic and Nonmelan- cholic Depressed Patients.

Baseline levels. As indicated in Table 3, there were no significant differences m baseline levels between DSM-/I/ melancholic and nonmelancholic subjects for any of the four hormones. Similarly, there were no differences between subgroups defined according to either the RDC or ICD-Y diagnostic systems.

Responses to clonidine. Repeated measures ANOVA of the combined depressed groups demonstrated a significant increase In GH ( Fqrru,/ = 20.4; l/f= I. 25; /I < 0.001) and significant reductions in both ACTH (F/J,, = 5.0; @= I, 15; /> < 0.05) and prolactin (F/,,, = 8.0; &= I, 25; p < 0.0 1). There was no significant change in cortisol ( F/!jg= 0.55;

~/f q I, 24; p = 0.45).

Table 3. Hormonal baseline levels and responses to clonidine in melancholic and nonmelancholic major depression

Melancholic (n = 13) Nonmelancholic (n = 13)

Mean SD Mean SD

Age yr 47.6 12.7 38.8 7.51

Baseline levels GH mu/I, 1.1 1.1 2.2 26

Cortisol nmoVl12 412 3 96.6 401.1 178.9

ACTH rig/l 83 57.5 44.0 82 3 83.7

Prolactln ( mlU/I 139.5 90.1 210.5 173.1

Response A’ GH mu/I 6.5 10.0 9.8 12.3

Cortisol nmol/l,2 -101.8 66.3 - 82.0 130.3

ACTH rig/l 13 - 15.0 10.6 - 14.7 11.9

Prolactln mlU/I - 41.4 38.3 - 75.5 134.7

Note GH growth hormone ACTH adrenocortlcotroplc hormone Raw means are given Diagnoses are according to DSM-III

1 p I 0 05 t test 2 n 11.14 3n 8. 9

As indicated rn Table 3 and Fig. 1, there were no stgnificant differences in the responses of any of the hormones between DSM-III melancholic and nonmelancholic subjects (e.g., for A GH: I = -0.03, #= 24,~: 0.98). ANCOVAs covarying for either age (F = 0.00 I ; L//‘= 1, 25; p q 0.97) or sex (F = 0.002; df= I, 25; p = 0.97) also indicated no

185

Fig. 1. Growth hormone response to cionidine in melancholic and non- melancholic depressed subjects

-30. -35. 0. 15. 50. 45. 00. 75. 00.

Time (minutes)

Mean + SEM. DSM-//I melancholic. n = 13 [solid Ilne,; DSM-III nonmelancholvz. n = 13 #dotted line

significant differences m A GH between groups. Even after excluding the one nonme- lancholic subject with an elevated baseline level (10.0 mUil), there was still no significant difference between groups for A G H (f = 0.3 1, u”= 23, p = 0.76). There were likewise no differences between groups defmed by RDC or /CD-9.

To examine whether prior fenfluramine testing had an effect on responses to clonidine, the I2 subjects given prior fenfluramme were compared with the other 14. There were no significant differences between responses in these two groups. Within the sample of 12 subjects given prior fenfluramine, there was no significant correlation between the interval between the challenges and (GH or other hormonal) responses.

Blood pressure. Effects on blood pressure were used as an indicator of the biologtcal activity of clonidine at this dose. Clonidine significantly reduced both systolic(l= -12.7,df= 25,p<0.001)anddiastolic(t=-13.5,df=25,p<0.00l)levelsin the combined depressed groups. There were no significant differences between DSM- II/melancholic and nonmelancholic subjects with regard to the maximum reductions (mmHg f SD) of either the systolic (-19.7 f 8.3 vs. -18.6 + 7.3) or diastolic blood pressure (-12.9 f 4.3 vs. -12.2 f -5.3).

Study 2: Hormonal Responses to Clonidine in EndogenousIMelancholic Subjects and Matched Controls.

Baseline levels. As outlined in Table 4, there were no significant differences in the baseline levels of the four hormones between the endogenousi melancholic depressed subjects and controls.

Response to clonidine. Repeated measures ANOVA of the 10 control subjects demonstrated significant reductions in both cortisol (F/r,,= 13.1; df= 1,9;p < 0.0 1) and prolactin (FqOOd q 20.1; df q 1, 9; p < 0.01) levels, a strong trend toward a significant increase in GH (Fquod= 5.0; df= 1,9;p= 0.05), but no significant change in ACTH (F/,,, q

186

Table 4. Hormonal baseline levels and responses to clonidine in matched DSM-IIVRDWICD-9 endogenous/melancholic and control subjects

Endogenous depressed Controls (n = 10) (n = 10)

Mean SD Mean SD

Baseline levels

GH t mu/I 1 Cortisol t nmoVl11

ACTH 1 rig/l 12

Prolactin (mlU/I j

Response A GH ImU/Ii

Cortisol I nmol/l~I

ACTH i rig/l 12

Prolactin I mlU/L

1.2 1.2 2.1 3.6

412.6 113.0 377.7 309.1

71.5 41.9 37.4 37.0

146.5 101.9 185.6 82.5

7.9 11.2 4.0 5.2

-103.2 72.5 -151.9 145.9

- 19.3 8.2 - 6.0 4.13

- 42.8 42.1 - 43.8 29.6

Note. GH = growth hormone. ACTH = adrenocorticotropic hormone. RDC= Research DIagnosticCrIterIa. Raw means are gwen.

1 n=9.9 2. n = 6. 6. 3. p < 0.05 (r test after loglo-transformation,

1.43; df’= I, 9;~ = 0.26). When results from the matched depressed and control subjects were combined, however, there was a significant increase in GH ( Fquot/= 12.7; gf= 1, 19;

p = 0.002), and significant reductions in each of cortisol (FL,, = 7.5; df= I, 18;~ = O.Ol), ACTH (n,,, = 5.2; df= I, 14;p<O.O5)and prolactin(F:,,= 14.7; df= 1, 19;p= 0.001).

As detailed in Table 4 and Fig. 2, there was no significant difference in A GH between the depressed and control groups (t = 1.76, df’= 9, p = 0.11). In fact, the trend was toward an increased response In the depressed subjects. Even after exclusion of one control subject with an elevated baseline of I 1.7 mU/ 1, there was still no difference between groups (r = 1.71, df’= 8,~ = 0.13), with the trend in the same direction. (When the total group of 26 depressed patients was compared to the 10 controls, there was also no significant difference between groups: t = 0.83, df= 34, p = 0.41.) The only significant difference in hormonal responses between the groups was a greater reduc- tion in ACTH levels in the endogenousl melancholic depressed patients vs. the control subjects (t = -2.94, LIP= 5,~ < 0.05) (Fig. 3). When the o/o reduction in ACTH levels in these groups was compared, there was no significant difference between the groups (r =

-1.56, &‘= 5, p = 0.18). Blood pressure. Clonidine significantly reduced both systolic (t = -9.46, df= 9,p<

0.001) and diastolic (t = -7.21, df= 8, p < 0.001) blood pressure levels in the control subjects. There was a significantly greater reduction in systolic blood pressure (mm Hg

k SD) in controls than in endogenous depressed subjects (-24.9 + 8.3 vs. - 18.4 ?r 8.4; t = 3.3 1, df= 9, p < 0.0 1). When 96 reductions of systolic blood pressure were considered, there was still a trend toward a significant difference (-19.5 f 5.6~~. -15.8 + 5.7; t = 2.12. @= 9, p = 0.06). There were no significant differences between the groups with regard to reductions (mmHg + SD) In diastolic blood pressure (-12.7 + 5.1 vs. 14.9 + 6.2; I = 0.96, Q’= 8, p = 0.36). To ensure that the GH response was not merely a reflection of

187

Fig 2. Growth hormone response to clonidine in endogenous/melancholic depressed subjects and matched controls

I2 r

0’ I ’ ’ I ’ ’ I ’ ’

-30. -1s. 0. 15. 30. 45. 60. 75. 90.

Time (minutes)

Mean + SEM. Endogenous/melancholic, n = 10 (solid line); matched controls, n = 10 (dotted line)

clonidine-induced hypotension, we examined this association in the combined

depressed and control groups (n = 39). There was no significant correlation between the reduction in systolic blood pressure and GH response (I = -0.14, NS).

Fig. 3. ACTH response to clonidine in endogenous/melancholic depressed subjects and matched controls

100

i

? F

70

E *

101 1 I ’ ” ’ ’ I ’

-30. -15. 0. IS. 30. 45. 50. 7s. 90.

Time (minutes)

Mean k SEM. Endogenous/melancholIc, n = 6 / solld lineI, matched controls, n = 6 [dotted line ACTH = adrenocorticotroplc hormone.

Study 3: Hormonal Responses to Clonidine and Fenfluramine in the Same Depressed Subjects. As indicated in Table 5, there were no significant correlations

188

between hormonal responses to fenfluramme and clonidine. Similarly, when only the 12 subjects who had received fenfluramine initially were considered, there were no significant intercorrelations.

Table 5. Spearman’s rank order correlations between hormonal responses to fenfluramine and clonidine (total depressed sample; n = 15)

Clonidine

Fenfluramine A GH 1 Cortisol 1 Prolactin

1\ Prolactin 0.25 -0.32 -0.06

_L Cortisol -0.29 0.12 0.30

Discussion

Our study failed to confirm the important finding of Checkley et al. (1984) that there was a significant difference in the GH response to 1.3 pgi kg clonidine between melancholic and nonmelancholic subgroups. The results of Checkley et al. had suggested differences in noradrenergic function between these subtypes. We were also unable to demonstrate a difference in the GH response between closely matched endogenous; melancholic depressed subjects and normal controls. (In fact, there was a weak trend toward an increased response m the depressed subjects, the opposite of previous reports). As discussed in the introduction, the only other study to use this dosage when comparing depressed and normal subjects (Horton et al., 1986) also failed to demonstrate a difference in the GH response, though that study had used an inadequate drug-free interval and did not control for age or sex.

Two recent dose-response trials of clonidine in normal male subjects (Hoehe et al., 1988; Brown et al., 1990) are relevant to our findings. Hoehe et al., using dosages of 1.5 and 2.0 pg! kg, demonstrated a significant dose-dependent GH increase with cloni- dine. Although their GH response to clonidine. 1.5 p*g/ kg, was significantly greater than that with saline, at that dosage few subjects demonstrated peak GH responses of at least 5 ng/ml (10 mU;‘l), an arbitrary cutoff level used to differentiate blunted from normal GH responses (Sever et al., 1986). Similarly, Brown et al. (1990) demonstrated a greater GH release with 2. I pugi kg than with either 1.4 or 0.7 pgikg, though all doses did increase GH levels significantly. That study also found peak GH responses of at least 5 ng;‘ml in only 3 of their IO subjects when tested at doses of either 0.7 or 1.4 pug! kg. The results of these two studies are consistent with our own findings, with only 9 of our 26 depressed subjects and 4 of our IO controls demonstrating peak responses at or above this level. Our results therefore also suggest an unreliable GH release with clonidine at a dosage of I .3 /Igi kg. The variability of response at this dose is the most likely explanation for the unexpected trend toward a greater GH response in the depressed subjects.

The findings of our study did, however, indicate that clonidine was physiologically active at the dosage used: (1) There were significant reductions in both systolic and diastolic blood pressures. (2) A significant increase in GH was demonstrated by the use of repeated measures ANOVA. These results are also consistent with those of Brown

et al. (1990). The lack of differentiation between melancholic and nonmelancholic subgroups

189

with this dosage of clonidine, even after controlling for age and sex, is therefore potentially explicable on the basis of unreliable GH release. It is also probable that the difference between our results and those of Checkley et al. (1984) reflects the signifi- cant variability in the GH response at this dosage. It is unlikely that these conflicting findings are due to differences in clinical populations, as both studies used RDC diagnoses to distinguish depressive subgroups, in addition to other subclassification systems. Our results suggest that if there is indeed a difference in the GH response between melancholic and nonmelancholic depressives, then it is unlikely to be reliably elicited by this dose of clonidine. The trend toward a significant difference in the yc reduction of systolic blood pressures between depressed and control subjects was unlikely to have confounded the GH responses, as both our study and others (La1 et al., 1975) have found no association between blood pressure and GH responses to clonidine.

Our findings with regard to ACTH, cortisol, and prolactin responses are of interest as these have been examined in few studies of depressed subjects. Animal studies suggest that while clonidine has no effect on basal secretion of ACTH, it reduces increased secretion stimulated by agents such as the glucose analogue 2-deoxyglucose (2-DG) (Smythe et al., 1985) urethane (Smythe et al., 1987), and 5-hydroxytrypto- phan (Smythe et al., 1988). Studies of theeffect ofclonidine on basal ACTH in normal humans have been inconsistent, with reports of clonidine inhibiting (Lanes et al., 1983) having no effect on (Milson et al., 1986; Hunt et al., 1986), or stimulating these levels in man (Jackson et al., 1988). Jackson et al. (1988) did, however, demonstrate that clonidine reduced fenfluramine-stimulated ACTH levels-a finding analogous to the animal studies.

Cortisol in normal human controls has also been variously reported to be either reduced (Matussek et al., 1980; Lanes et al., 1983) or unaffected byclonidine (Siever et al., 19846; Hunt et al., 1986). There have been few studies investigating prolactin. Terry and Martin (1981) and others have reported that clonidine increases prolactin levels in animals. Lanes et al. (1983) and Hunt et al. (1986) using oral clonidine, found no change compared with placebo in normal human subjects. La1 et al. (1975) using i.v. clonidine in six normal male subjects, also reported no significant change, though inspection of their raw data suggests a reduction at 30 min.

In our study, repeated measures ANOVA indicated significant reductions in the levels of each of these three hormones. As we did not use a placebo control, however, we cannot exclude the possibility that these decreases may have represented normal circadian variation.

Our findings with regard to the ACTH and cortisol responses are of particular interest. Lesch et al. (1988) have reported that clonidine (2 pgi kg, i.v.) significantly increased ACTH levels, but their subjects were only required to have been drug-free for a minimum of 3 days. Our finding of a greater absolute reduction in the depressed subjects is of significance as it is consistent with both the animal and normal human data of the effect of clonidine on stimulated ACTH secretion. The animal studies by one of our group (Smythe et al., 1985, 1987, 1988) indicate that clonidine reduces the elevated central noradrenergic activity mediating increases in ACTH secretion. Our findings in study 2 are therefore consistent with increased noradrenergic activity in melancholia. An alternate explanation is that the difference reflects different baseline ACTH levels.

190

These ACTH findings are consistent with the study of Siever et al. (1984b), which demonstrated both a greater absolute and Yc reduction in cortisol levels in their depressed subjects versus controls. We did not, however, demonstrate any difference in the cortisol response-a finding in agreement with that of Matussek et al. (1980) and Amsterdam et al. (1989). There were no differences in the prolactin response between depressed and control subjects. This result was consistent with that of Siever and Uhde (1984) and Amsterdam et al. (1989).

As with the GH response, we found no evidence of differences in ACTH, cortisol, or prolactin responses between the melancholic and nonmelancholic subgroups. Matus- sek et al. (1980) and Amsterdam et al. (1989) have also reported no differences in cortisol or prolactin responses between these subgroups.

Although study 3 was the first examination of the relationship between endocrine responses to fenfluramine and clonidine in the same subjects, our negative results were invalidated by the inability of the dose of clonidine used to induce sufficient GH release to distinguish normal from blunted responses. Three previous studies testing for the association of GH responses to clonidine and hormonal responses to other challenges (Corn et al., 1984; Siever et al., 1985; Ansseau et al., 1988) did not undertake comparisons with normal controls to confirm abnormalities in these responses.

In summary, our study failed to demonstrate differences in hormonal responses to 1.3 pgi kg clonidine between melancholic and nonmelancholic subgroups. Moreover, the only difference between endogenousi melancholic subjects and closely matched normal controls was a greater reduction in ACTH levels in the former group. These findings suggest that the value of clonidine (at least at this dosage) in identifying neurotransmitter differences between melancholic and nonmelancholic subgroups must still be regarded as uncertain.

Acknowledgments. This research was supported by a grant from the Australian National Health & Medical Research Council. We thank Mr. Dusan Hadzi-Pavlovic for statistical advice. Ms. Christine Burke and Ms. Anna Zournazi for technical assistance, and Mrs. Zora Vuckovic for preparation of the manuscript.

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